LOCAL REGULATION OF SKELETAL MUSCLE BLOOD VESSELS: INFLUENCE OF PULSE PRESSURE AND VASOMOTOR TONE Dissertation for the Degree of Ph. D. MICHIGAN STATE UNIVERSITY BRIAN JOHN LaLONE 1975 This is to certify that the thesis entitled Local Regulation of Skeletal Muscle Blood Vessels: Influence of Pulse Pressure and Vasomotor Tone presented by Brian John LaLone has been accepted towards fulfillment of the requirements for Ph.D. , Physiology degree in mum/J Major professor/ 0-7639 F BINDING BY 3“ HUAE & SUNS’ 300“ “mm" INC. -lBRARY BYNOERS Ill g- famaeogr, ”‘1‘“! ABSTRACT LOCAL REGULATION OF SKELETAL MUSCLE BLOOD VESSELS: INFLUENCE OF PULSE PRESSURE AND VASOMOTOR TONE BY Brian John LaLone The vascularly isolated gracilis muscles from 18 dogs anesthetized with sodium pentobarbital were exposed to alterations in local perfusion pressure before and during hemorrhage induced elevations of vasomotor tone to evalu- ate the influence of reflex constriction on muscle blood flow autoregulation. Using the same preparation, another series of 10 experiments were designed to examine the influ- ence of pulse pressure on muscle blood flow autoregulation and the vascular responses to venous pressure elevation and increased mean vascular distending pressure. In one group of natural flow experiments (Series I; n = 10), the gracilis nerve was left intact and muscle vascular responses to 4 sequential step reductions of’ gracilis artery pressure were studied while the animals were normovolemic and normotensive and during two periods of hemorrhage induced systemic arterial hypotension. Hemorrhage significantly reduced muscle blood flow and Brian John LaLone elevated muscle vascular resistance. However, blood flow autoregulation was not affected since local reductions of gracilis artery pressure from approximately 120 to 70 mm Hg elicited statistically similar percent reductions in vascu- lar resistance during both normovolemia and hypovolemia. These data indicate that steady state muscle blood flow autoregulation is maintained when vascular tone is elevated by hemorrhage. In Series II, the innervated gracilis muscles of 8 dogs were perfused with the animals femoral arterial blood by a servocontrolled pump which maintained gracilis artery pressure at any set level by continually adjusting pump flow rate. Transient and steady state vascular responses to step changes in perfusion pressure to or from 140 mm Hg were examined while the animals were normovolemic and normo- tensive and while systemic arterial pressure was reduced to 100 mm Hg by hemorrhage. During normovolemia, all altera- tions in gracilis artery pressure below 140 mm Hg were associated with proportionately smaller changes in steady state blood flow. When vascular tone was significantly elevated by hemorrhage, the same local alterations in per- fusion pressure elicited slightly smaller percent changes in flow. However, the rate of development of these auto- regulatory responses were significantly prolonged during Brian John LaLone hemorrhage. These data indicate that, when vascular tone is elevated by hemorrhage, steady state blood flow auto— regulation is slightly improved, but some competition or interaction between remote vasoconstrictor influences and local control mechanisms delays the develOpment of the autoregulatory response during hypovolemia. To determine the influence of pulse pressure on local control of skeletal muscle blood vessels, vascular responses to local, graded hypotension, venous pressure elevation, and increased mean vascular distending pressure were examined using both pulsatile and non—pulsatile perfusion in the naturally perfused, denervated gracilis muscles of 10 dogs (Series III). Muscle vascular resistance and the vasoconstriction observed in reSponse to venous and mean vascular distending pressure elevations were all signifi— cantly greater during pulsatile compared to non-pulsatile perfusion. These data indicate that myogenic mechanisms of blood flow control are present within the muscle vascu— lature and are sensitive to the transmural pressure changes associated with pulse pressure distension. When mean gracilis artery pressure was progressively lowered from 140 to 60 mm Hg during pulsatile perfusion both pulse pressure and muscle vascular resistance decreased significantly, whereas during non-pulsatile perfusion the same maneuver Brian John LaLone elicited a progressive rise in vascular resistance. These data indicate that muscle blood flow autoregulation is mediated to a large extent by alterations in pulse pressure induced myogenic activity. LOCAL REGULATION OF SKELETAL MUSCLE BLOOD VESSELS: INFLUENCE OF PULSE PRESSURE AND VASOMOTOR TONE BY Brian John LaLone A DISSERTATION Submitted to Michigan State University in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY Department of Physiology 1975 To Joan and Aimee ii ACKNOWLEDGMENTS I am deeply indebted to my advisor, Dr. James M. Schwinghamer, for his patient assistance in the prepara- tion of this manuscript and for his friendship, support, and guidance throughout my graduate training. I would also like to extend my appreciation to Drs. James G. Cunningham, Erik D. Goodman, Robert P. Pittman, and Lester F. Wolterink for serving on my guidance committee and reviewing this dissertation. A Special thanks is due my wife, Joan, who has given of herself completely and unselfishly so that I might complete my graduate training. iii TABLE OF CONTENTS Page LSIT OF TABI‘ES000000000000000000000000000000000000000 Vii LIST OF FIGURES00000000000.00000000000000000000000000 ix INTRODUCTION00000000000000000000000000000000000000000 1 LITERATURE RBVIEW00000000000 00000000000000.00000 00000 9 I. Metabolic Mechanisms of Skeletal Muscle Blood Flow Autoregulation.......................... 10 A. Metabolically Active Vasoactive Chemicals. ll 10 oxygen.00000000000000000000000000000000 ll 20 HYdrOgen0000000000000000000000000000000 16 30 PotaSSi‘ml000000000000000000000000000000 l7 4. Adenosine and the Adenine Nucleotides.. 18 II. Myogenic Regulation of Vascular Tone......... 18 A. Electrical and Mechanical Properties of Vascular Smooth Muscle.................... 19 B. Vascular Smooth Muscle Responses to StretCh00000000000000000000000000000000000 21 C. Microvascular Responses to Transmural Pressure ChangeSoo 000000 0 00 000 0 00000000 0 00 22 D. Transmural Pressure Effects on Vascular ReSiStance00000000000000000000000000000000 24 METHODS0000000000000000000000000000000000000000000000 27 DATA ANALYSIS0000000000000000000000000000000000000000 54 RESULTSO000000000000000000000000000000000000000000000 56 I. Series I: Naturally Perfused, Innervated Gracilis Muscle; Effects of Local Hypotension During Normovolemic and Hypovolemic Periods.. 55 iv TABLE OF CONTENTS—-continued II. III. A0 MUSCle BlOOd FlOWI000000000000000000000000 B. Muscle Vascular Resistance................ Series II: Pump Perfused, Innervated Gracil- is Muscle; Effects of Alterations in Mean Arterial Pressure During Normovolemic and Hypovolemic Periods.......................... A. Muscle Blood Flow......................... B. Muscle Vascular Resistance................ C. Vascular Responses to Denervation During Hypovolemia............................... Series III: Naturally Perfused, Denervated Gracilis Muscles; Vascular Responses to In- creased Venous and Mean Distending Pressures and to Local Hypotension During Pulsatile or Non-pulsatile Perfusion...................... A. Vascular Responses to Alterations in Perfusion Mode............................ B. Vascular Responses to Increasing Mean Distending Pressure....................... C. Vascular Responses to Increased Venous Pressure.................................. D. Vascular Responses to Graded, Local Hypotension............................... DISCUSSION0000000000000000000000000000000000000000000 I. II. Series I and II: Naturally Perfused and Pump Perfused, Innervated Gracilis Muscles; Gracilis Artery Pressure Alterations During Normovolemic and Hypovolemic Periods......... A. Steady State Autoregulatory Responses to Local Alterations in Perfusion Pressure... B. Transient Vascular Responses to Altera- tions in Perfusion Pressure............... Series III: Naturally Perfused, Denervated Gracilis Muscles; Vascular Responses During Pulsatile and Non-pulsatile Perfusion........ A. Vascular Responses to Alterations in Perfusion Mode............................ B. Vascular Responses to Increased Venous Pressure.................................. C. Vascular Responses to Increasing Mean Distending Pressure....................... Page 56 65 7O 7O 92 98 101 101 107 111 116 124 124 124 125 129 129 132 135 TABLE OF CONTENTS—-continued D. Vascular Responses to Graded, Local Hypotension......................... SUWRY AND CONCLUSIONS000000000000000000000000 APPENDICESO000000000000000000000000000000.00000 A. Pressure, Flow, and Resistance Data.... B0 Statistical MethOdSoooooooooooooooooooo BIBLIOGRAPHY00000000000000000000000000000000000 vi Page 137 139 141 141 146 155 LIST OF TABLES TABLE Effects of papaverine infusions on gracilis muscle vascular responses to graded infusions of norepinephrine and acetylcholine............. Comparison of intravascular pressures, muscle blood flows, and vascular resistances observed prior to and after local, graded hypotension during both normovolemia and hypovolemia........ Gracilis muscle blood flow responses to step changes in perfusion pressure................... Effects of denervation during hemorrhage on gracilis muscle blood flow and vascular resist- ance00000000000000000000000000000000000000000000 Gracilis muscle vascular responses to perfusion mode changes in normal and papaverine treated muSCleS00000000000000000000000000000000000000000 Gracilis muscle vascular responses to elevations in mean vascular distending pressure during pulsatile and non—pulsatile perfusion in normal and papaverine treated muscles.................. Gracilis muscle vascular responses to 20 mm Hg elevations of venous pressure during pulsatile and non—pulsatile perfusion in normal and papa— verine treated muscles.......................... Gracilis muscle vascular responses to 20 mm Hg reductions in gracilis artery pressure during non-pulsatile perfusion in normal and papaverine treated muscles................................. Effects of local hypotension during normovolemic and hypovolemic periods on mean systemic arteri- al and gracilis artery and vein pressures, gracilis vein flow, and vascular resistance..... vii Page 50 69 81 99 105 108 112 123 141 LIST OF TABLES--continued TABLE Page A-2. Effects of local hypotension during normovolemic and two hypovolemic periods on mean systemic arterial and gracilis artery and vein pressures, gracilis vein flow, and vascular resistance..... 143 A-3. Effects of local gracilis artery pressure alter- ations during normovolemic and hypovolemic periods on gracilis muscle vascular resistance.. 144 A-4. Gracilis muscle vascular responses to graded, local hypotension during pulsatile and non- pulsatile perfusion............................. 145 viii LIST OF FIGURES FIGURE Page 1. Changes in mean systemic arterial pressure, gracilis artery pressure, and gracilis vein flow in response to local hypotension during normovolemia and during two levels of systemic arterial hypotension........................... 31 2. Schematic of the servosystem used to control gracilis artery pressure....................... 36 3. Gracilis artery blood flow responses to step changes in perfusion pressure.................. 40 4. Responses of servocontrolled pressure and pump flow rate to step changes in load.............. 42 5. Gracilis arterial and venous pressures and gracilis vein flow as functions of time from the initial portion of a selected Series III experiment..................................... 48 6. Gracilis arterial and venous pressure and venous flow responses to changing the mode of perfusion from non-pulsatile to pulsatile and vice versa..................................... 53 7. Effects of local hypotension during normovol- emic and hypovolemic periods on gracilis vein flow0000000000000000000000000000000000000000000 58 8. Effects of local hypotension during normovol- emic and two hypovolemic periods............... 61 9. Effects of local hypotension during normovol- emic and hypovolemic periods on gracilis vein flow and gracilis muscle vascular resistance... 64 10. Effects of local hypotension during normovol- emic and hypovolemic periods on gracilis muscle vascular resistance............................ 67 ix LIST OF FIGURES-~continued FIGURE 11. 12. 13. 140 15. 16. 17. 18. 19. 20. Gracilis artery blood flow responses to step changes in perfusion pressure during normo- volemic and hypovolemic periods................ Muscle blood flow responses to sequential gracilis artery pressure alterations from 140 to 80 to 140 mm Hg obtained during both normo- volemic and hypovolemic periods................ Effects of gracilis artery pressure alterations during normovolemic and hypovolemic periods on gracilis vein flow............................. Effects of gracilis artery pressure alterations during normovolemic and hypovolemic periods on gracilis vein flow normalized to % of control.. Effects of gracilis artery pressure alterations during normovolemic and hypovolemic periods on gracilis muscle vascular resistance............ Effects of gracilis artery pressure alterations during normovolemic and hypovolemic periods on gracilis muscle vascular resistance normalized to percent of control.......................... Gracilis muscle vascular responses to changing the mode of perfusion from non—pulsatile to pulsatile and baCk t0 non-pUlsatileo o o o o o o o o o o o Gracilis muscle vascular responses to eleva- tions in mean vascular distending pressure during pulsatile or non-pulsatile perfusion in normal and papaverine treated muscles.......... Gracilis muscle resistance responses to 20 mm Hg elevations of venous pressure during pulsa- tile or non-pulsatile perfusion in normal and papaverine treated muscles..................... Gracilis muscle vascular responses to graded, local hypotension during pulsatile and non- pulsatile perfu8i0n0000000000000000000000000000 Page 72 75 88 91 94 96 103 110 114 118 LIST OF FIGURES--continued FIGURE Page 21. Effects of graded, local hypotension during pulsatile and non—pulsatile perfusion on gracilis vascular resistance ........ . ..... ..... 121 xi INTRODUCTI ON Blood flow distribution in the systemic circulation is regulated by control systems which adjust the contractile state of vascular smooth muscle. Alterations in smooth muscle tone are mediated by inhibitory and excitatory stim- uli originating within and outside of the various tissues. In general, remote regulatory systems maintain a stable sys- temic arterial blood pressure whereas local circulatory control mechanisms establish optimal conditions for trans- vascular exchange. It is currently believed that mainte- nance of optimum exchange conditions results from local adjustments in arteriole and precapillary sphincter radius so that an appropriate regional blood supply and perfused capillary surface area is achieved. One example of local blood flow control involves those factors which stabilize an organ's blood supply during variations in arterial pressure. When perfusion pressure to certain systemic vascular networks is varied over the range of approximately 70 to 180 mm Hg, there is a less than proportionate change in blood flow. This ability to maintain a relatively constant blood flow in the face of varying perfusion pressure is termed autoregulation and has been defined by Johnson (64) as "the intrinsic tendency of an organ to maintain constant blood flow despite changes in arterial perfusion pressure". The results of extensive investigation over the last 25 years indicate that some com— bination of at least three different mechanisms accounts for blood flow autoregulation: 1) a tissue pressure mechan- ism; 2) a metabolic mechanism; and 3) a myogenic mechanism. Each of these is described below with emphasis on the experimental criteria used to determine which may be the predominant mediator of autoregulation in a given vascular bed. The tissue pressure hypothesis states that alterations in vascular caliber responsible for blood flow autoregula- tion result from the effect of tissue fluid pressure on veins. An elevation of arterial pressure is hypothesized to cause net efflux of fluid from capillaries into the interstitial space so that tissue pressure increases, especially in organs surrounded by relatively incompliant capsules. The increased tissue pressure allegedly compres— ses veins causing mean vascular radius to decrease so that flow increases less than perfusion pressure. Several aspects of the tissue pressure hypothesis can be tested experimentally. Because veins have the lowest internal pressure and wall rigidity, changes in tissue pressure should elicit correspondingly greater changes in venous as opposed to arterial resistance, Furthermore, an increase in arterial pressure should, unless the capsule surrounding the organ is absolutely rigid, increase extra— vascular volume and consequently organ weight. Finally, since elevation of venous pressure should cause net efflux of fluid from capillaries into the interstitial space, the tissue pressure hypothesis predicts an increased precapil- lary resistance in response to venous pressure elevation (i.e., the so-called venous arteriolar response). Because the tissue pressure hypothesis does not involve active vasomotion, neither the venous arteriolar response nor autoregulation should be abolished by pharmacological agents which paralyze vascular smooth muscle. The metabolic hypotheses of blood flow autoregulation state that a decreased blood flow results in vascular relaxation mediated either by dilator metabolites released from the surrounding tissues or by a decreased nutrient supply to these tissues. According to the metabolic hypothe- sis, reducing arterial pressure will decrease blood flow and cause accumulation of dilator metabolites, thereby eliciting a vascular relaxation which helps to restore flow. Decreased blood flow will also decrease interstitial p02 and conceivably alter tissue metabolism in a way that in- creases the production of vasodilator substances. In addi— tion, a lowered p0 may reduce vascular tone directly. 2 Therefore, the various metabolic mechanisms of autoregula- tion are characterized as being flow dependent responses which act to maintain a constant flow to metabolism ratio. The identification of a vasodilator substance in the venous effluent from organs perfused at low arterial preSv sure would provide strong experimental support for the metabolic hypothesis of blood flow autoregulation. Reduc— ing the pressure head in a vascular network by elevating venous pressure should decrease flow and, according to the metabolic hypothesis, cause an accumulation of dilator substances. Organs exhibiting metabolically mediated auto- regulation should therefore display a decreased vascular resistance when venous pressure is elevated (i.e., should not display a venous-arteriolar response). The myogenic hypothesis of blood flow autoregulation states that intrinsic mechanisms in smooth muscle cells of arteries, arterioles, and precapillary phincters allows these vessels to respond to a decrease in wall tension with relaxation or to an increase in wall tension with contrac- tion. Evidence has been presented to suggest that stretch of vascular smooth muscle evokes a contractile response by increasing pacemaker activity and eliciting more frequent bursts of action potentials which are propagated to neigh- boring cells. When vascular transmural pressure is in- creased, by increasing either arterial or venous pressure, or by decreasing pressure around an organ, the myogenic hypothesis predicts that the involved vessels will con- strict. Studies of the response to venous pressure elevation represent a critical experiment for demonstration of myogenically mediated autoregulatory responses. Eleva- tion of venous pressure will produce precapillary constric- tion according to both the myogenic and tissue pressure hypotheses, whereas the metabolic hypothesis predicts dilation in response to venous pressure elevation. Precapil- lary constriction resulting from venous hypertension will be abolished by agents which paralyze vascular smooth muscle if this constriction is myogenic but not if it is due to in— creased tissue pressure. The available data seem to indicate that myogenic. mechanisms are primarily responsible for blood flow auto- regulation in the intestine (48, 52, 62, 63, 65, 66, 69, 109), mesentery (67, 71), and liver (50, 53, 123), whereas .metabolic mechanisms are probably most important in the myocardium (9, 10, 27, 41, 100, 103). The mechanism(s) responsible for cerebral (30, 81, 94), renal (16, 44, 46, 54, 89, 90, 120, 126), and particularly skeletal muscle blood flow autoregulation are less well defined. Since agents that paralyze vascular smooth muscle abolish both autoregulation and the venous arteriolar response in skele- tal muscle (58, 84, 85, 111, 117), and since autoregulatory resistance responses to changes in perfusion pressure occur almost exclusively in precapillary vessels rather than veins (51, 58, 70, 88, 121), the tissue pressure mechanism for blood flow autoregulation appears to be relatively unimportant in skeletal muscle vasculature. Rather, skele- tal muscle blood flow autoregulation is probably mediated by myogenic and metabolic factors. The uncertainty regarding the relative importance of these two autoregulatory mechanisms results in part from conflicting reports of the resistance responses to venous pressure elevation. In the skinned hindlegs (80% skeletal muscle) of reserpinized cats Folkow and Oberg (36) observed 16 to 42% increases in vascular resistance when venous pres- sure was elevated 10 mm Hg. Similar results were reported by Nagle gt a1. (88) in the denervated canine gracilis muscle where 30 mm Hg elevations in venous pressure con- sistently increased vascular resistance by about 40%. Hanson (49) found equivocal alterations in vascular re— sistance when venous pressure was elevated 20 mm Hg in the isolated canine hindlimb (50% muscle; 20% skin; 30% bone). In half of Hanson's experiments resistance at elevated venous pressure was slightly below control, while in others it either did not change or increased slightly. Jones and Berne (73) obtained similar results in a skinned canine thigh preparation; 20 mm Hg elevations of venous pressure elicited only slight increases in vascular resistance in about one third of the preparations. These disparate find— ings may be related to preparation differences. The canine hindlimb preparations of Hanson (49) and Jones and Berne (73) were removed from the animals by sectioning the muscles J under study whereas the skinned cat hindlimbs used by Folkow and Oberg (36) were isolated from the animals at the level of the lower abdominal aorta so that the muscles being stud- ied were not sectioned. Furthermore, in the canine hindlimb preparations of Jones and Berne (73) resting blood flow per unit tissue weight was considerably lower than that reported by Folkow and Oberg (36) and Nagle gt a}. (88). This differ— ence appears to be especially important since, in the few preparations which displayed relatively high control blood flows, Jones and Berne (73) did observe an increase in vascu- lar resistance with elevated venous pressure. These results suggest that elevated vascular tone caused by preparation deterioration, surgical trauma, or hemorrhage may alter local control of skeletal muscle vasculature. Indeed, hemor- rhage has been suggested by several investigators (63, 72, 80, 126) to be a factor contributing to the somewhat labile nature of blood flow autoregulation. It has been found, for example, that hemorrhage abolishes renal blood flow auto- regulation (25, 126). However, Bond and Green (13, 43) reported that skeletal muscle blood flow autoregulation was more pronounced during hemorrhage. Because alterations in arterial perfusion pressure will cause qualitatively similar changes in blood flow and vascu- lar wall tension, it is difficult to determine whether the resistance responses elicited by this maneuver are meta- bolically or myogenically mediated. However, if autoregulation is primarily a myogenic response, one might expect a different degree of autoregulation depending on the magnitude of pulse pressure. Two separate investiga— tions (85, 99) have shown that pulse pressure distension contributes to the total basal vascular tone present in skeletal muscle. Since it is unlikely that variations in pulse pressure about a given mean pressure would alter the nutrient supply to skeletal muscle, potentiation of blood flow autoregulation by increased pulse pressure would sug— gest that myogenic mechanisms are important mediators of blood flow control in this tissue. The purpose of this study is two—fold. Because the conflicting results described above may be due to elevation of vascular tone, experiments were designed to evaluate the effects of hemorrhage induced increases in vascular tone on skeletal muscle blood flow autoregulation. The effect of pulse pressure distension on muscle blood flow autoregula— tion was also studied to evaluate the contribution of myogenic mechanisms to autoregulatory responses in this tissue. LITERATURE REVIEW Local regulation of blood flow in skeletal muscle is believed to result from both metabolic and myogenic mechan- isms (68). Because it is difficult to design experiments which clearly distinguish metabolic from myogenically medi- ated vascular responses, little definitive evidence exists regarding the relative importance of each of these mechan- isms. The following sections analyze the role of these two mechanisms in muscle blood flow control. The first section discusses literature pertinent to the metabolic hypothesis, particularly as it pertains to autoregulation. Most inves— tigations of metabolic blood flow control have focused on functional hyperemia (the blood flow increase accompanying skeletal muscle exercise) and reactive hyperemia (the blood flow increase after release of arterial occlusion), whereas relatively little attention has been directed at muscle blood flow autoregulation. Thus, in some instances it has been necessary to draw inferences from studies of function— al and reactive hyperemia when assessing the role of cer- tain vasodilator chemicals in autoregulation. The second section discusses vascular myogenic mechanisms and their possible importance in autoregulation. Since only a few investigators have examined myogenic mechanisms in skeletal 10 muscle vascular beds, some pertinent studies in other vascu— lar networks have been included. I. Metabolic Mechanisms of Skeletal Muscle Blood Flow Autoregulation The metabolic hypothesis of blood flow autoregulation proposes that a decrease in flow caused by a fall in perfu- sion pressure will decrease the concentration of oxygen or increase the concentration of vasodilator metabolites in the tissue fluids, thereby resulting in active arteriolar dilation. Although active vasodilation has been observed in response to decreases in skeletal muscle blood flow (14, 36, 46, 49, 51, 58, 73-75, 105, 116, 117, 125). this response is adequately explained by either the metabolic or myogenic hypothesis. Other experimental designs have pro- vided more definitive evidence to implicate metabolically linked chemicals in blood flow autoregulation. Some support for the View that metabolically linked chemicals play an important role in muscle blood flow auto- regulation comes from studies of the vasoactivity of venous blood. Folkow (34) has shown that intra-arterial injection of venous blood elicits vasodilation in the cat hindlimb. Similar results have been obtained by Haddy and Scott (46) in the dog forelimb. Scott gt 31. (105) found that perfu— sion of the dog forelimb with the venous effluent from the hindlimb produced an almost immediate fall in vascular 11 resistance in the assay forelimb. They employed this same bioassay preparation to demonstrate that a reduction in hindlimb blood flow, produced by compressing the femoral artery, led to vasodilation not only in the hindlimb, but also in the forelimb. Opposite responses were observed when active vasoconstriction was induced in the hindlimb by elevating flow rate. Although these bioassay studies provide evidence to support the metabolic hypothesis of skeletal muscle blood flow autoregulation, they do not identify a specific chemical mediator. As described below, the metabolically linked chemicals that are potential medi- ators of skeletal muscle blood flow control include oxygen, hydrogen, potassium, and adenosine and its nucleotides. A. Metabolically Linked Vasoactive Chemicals 1. Oxygen Several different lines of experimental evidence indi- cate that oxygen may participate in autoregulation of skele- tal muscle blood flow. These include studies of: a) changes in venous blood oxygen content associated with skeletal muscle blood flow autoregulation; and b) the effects of alterations in arterial blood oxygen content on skeletal muscle vascular resistance, blood flow autoregulation, arteriole diameter, and small artery conductance. A fall in the oxygen content or partial pressure of venous blood draining skeletal muscle has been shown to 12 accompany autoregulatory responses to decreased arterial perfusion pressure (14, 51, 73-75, 118). Jones and Berne (73 , 75) , using a pump perfused canine hindlimb preparation, reported that resistance responses to step changes in flow were accompanied by qualitatively similar changes in venous oxygen content. These investigators also observed that blood flow autoregulation was present in dog hindlimbs when venous blood oxygen content was low but not when oxygen con— tent of the venous effluent was high (74) . Using the canine gas trocnemius-plantaris muscle group, Stainsby and Otis (118) have shown that muscle venous blood oxygen content decreases with reductions in perfusion pressure which elicit blood flow autoregulation. Similar results have been ob- tained by Bond e1: a_l. (l4) and Hanson and Johnson (51) in canine hindlimb preparations. From such observations it has been suggested that autoregulatory changes in vessel Caliber result from changes in tissue oxygen tension. Such a hypothesis is tenable only if it can be shown that rela— tiVely small changes in tissue fluid p02 elicit significant changes in vascular resistance. Several investigations indicate that a fall in oxygen delivery will elicit vasodilation (19, 24, 57, 96, 110). when the hemoglobin saturation of blood perfusing the dog hindlimb was reduced by 10%, Ross e1; a1. (96) observed a 25% increase in flow. Further reduction of arterial blood hemoglobin saturation to 0% elicited a 3-4 fold increase in 13 flow. Similar results were obtained by Skinner and Powell (110) in canine gracilis preparations where 70% reductions in arterial blood oxygen content elicited 60% decreases in vascular resistance. Hutchins et 31. (57) reduced arterial p0 by 15% and observed a 2 to 18% increase in the diameter 2 of small precapillary vessels in the rat cremaster muscle. Carrier 32 £1. (19) reported an inverse relationship between vascular diameter and p0 in small arteries (0.5-1.0 mm 2 O.D.) from the dog hindlimb. The smallest vessels examined in this study displayed a 15% decrease in resistance when p02 was reduced from 100 to 90 mm Hg, whereas with the larger vessels it was necessary to reduce p02 from 100 to 50 mm Hg in order to obtain an equivalent change in resistance. Detar and Bohr (24) found that the active tension developed by aortic strips exposed to epinephrine was diminished when p02 in the bathing fluid was decreased from 100 to 70 mm Hg. The reports cited above suggest that vasomotor tone in skeletal muscle is modulated by oxygen so that a given fall in 02 delivery elicits a corresponding fall in vascular resistance. There is also evidence that supernormal oxygen levels in arterial blood cause vasoconstriction. Bachofen 23 El. (4), using a hyperbaric chamber, observed an 18% in- crease in canine hindlimb vascular resistance when arterial blood p0 was elevated from 92 to 1660 mm Hg. 2 Not all available data support the view that small changes in oxygen delivery elicit vasomotor responses. 14 Daugherty et a1. (21) failed to observe a change in dog forelimb vascular resistance when brachial artery p02 was reduced from 120 to 30 mm Hg. Only when arterial p02 was reduced to very low levels (from 30 to 8 mm Hg) did these investigators observe significant vasodilation in the fore- limb. Chalmers et 31, (20) were unable to show dilation in the hindlegs of rabbits perfused with hypoxic arterial blood until the oxygen tension was reduced below 30 mm Hg. These studies suggest a threshold for hypoxia—induced vaso— dilation which might reflect a shift to anaerobic metabolism in either the vascular smooth muscle or the parenchymal cells at low p0 While there is agreement that oxygen can 2. effect skeletal muscle vascular tone, the question of whether or not the oxygen effect has a threshold is diffi- cult to resolve since evidence has been presented to support each view. There are two investigations which specifically address the effects of altered blood oxygen content and p02 on skeletal muscle blood flow autoregulation. Using the canine hindlimb, Walker and Guyton (125) found that flow was auto- regulated at progressively higher levels when the arterial blood oxygen saturation was progressively decreased from 98 to 30%. It is interesting to note that even though vascular resistance fell with decreasing arterial blood oxygen satura— tion, autoregulatory behavior was essentially unchanged; that is, perfusion pressure reductions from 150 to 70 mm Hg 15 elicited similar percent reductions in blood flow and vascu- lar resistance regardless of the arterial blood oxygen content. Similar results have been reported by Bond et 31. (14) in the dog hindlimb when blood p0 was elevated by 2 changing the inspired gas from room air to 2 atmospheres of oxygen. Even though hindlimb venous blood p02 was increased from 50 to 240 mm Hg by this maneuver, no alterations were observed in either the resting level of blood flow or blood flow autoregulation. The results from these two studies indicate that although vascular tone is diminished by local hypoxia it is not effected by hyperoxia and neither maneuver appreciably changes the extent to which blood flow is auto- regulated in skeletal muscle. The mechanism by which hypoxia elicits dilation is not known. Many investigators believe that oxygen alters vascu- lar smooth muscle tone directly (19, 24, 96, 110). The work of Detar and Bohr (24) and Carrier gt_al. (19) on isolated small arteries and aortic strips provide strong support for this view. Honig (56) has reported in_yitrg_evidence to suggest that hypoxia may lower vasomotor tone directly through the inhibition of myosin ATPase by inorganic phos- phate and cyclic AMP elaborated within the vascular smooth muscle cells. However, other investigators have proposed that the dilator effect of hypoxia is secondary to an alter- ation in parenchymal tissue metabolism which releases vaso— active substances (ll, 26, 28, 29). Berne (11) reports 16 that oxygen lack in the myocardium leads to release of the potent vasodilator adenosine, and suggests a similar type of control for skeletal muscle vascular beds. Large amounts of adenosine are produced by skeletal muscle during severe ischemic exercise and lesser amounts may also be present in resting muscle (26). 2. Hydrogen Although the hydrogen ion is clearly vasoactive (21, 23, 32, 87, 92, 130), experimental evidence suggests that its role in the mediation of skeletal muscle blood flow autoregu- lation is probably minor. The difficulty in assigning a major role to the hydrogen ion results primarily from two observations: 1) induced changes in the pH of blood perfus- ing skeletal muscle must be very large to produce measurable changes in resistance (21, 86, 87, 104, 125); and 2) large decreases in vascular resistance associated with skeletal muscle exercise cause only small alterations in venous blood pH (97, 102, 120). For example, Daugherty gt_al. (21) ob— served that a severe local hypercapnic acidosis which re- duced forelimb blood pH from 7.58 to 7.19 only lowered vascular resistance 24% below control. In addition, periods of skeletal muscle exercise that produce reductions in re— sistance ranging from 25 to 70% below control levels are associated with only 0.03 to 0.07-unit reductions in efflu— ent blood pH (97, 102, 120). Since tissue fluid pH probably falls less during local hypotension than during exercise, 17 the contribution of altered hydrogen ion alone to blood flow autoregulation appears to be very small. The hydrogen ion may however, exert effects in combina- tion with hypoxia. Stowe et 31. (120) perfused a canine gracilis muscle (assay gracilis) with the venous effluent from the contralateral, exercising gracilis and studied the effects of restoring assay gracilis blood p02 and pH to pre- exercise levels. Correction of either pO alone or pH alone 2 only slightly reduced the vasodilator activity of venous blood from the exercising muscle. However, when both p02 and pH were restored to pre—exercise levels, dilation in the assay gracilis was abolished. These findings indicate that the steady state vasodilator activity of venous blood from exercising skeletal muscle appears to be mainly the combined result of decreased p0 and pH. Although the mechanisms for 2 maintenance of a constant flow to metabolism ratio during exercise may be different than those which mediate autoregu- lation, these results suggest that hydrogen ion could be important in skeletal muscle blood flow autoregulation through a combined effect with the hypoxemia which accompan- ies reductions in perfusion pressure. 3. Potassium Although the potassium ion is vasoactive (31, 42, 45, 77, 110) and is probably involved in the initiation of ex- ercise dilation (1, 93, 106, 107), it is not believed to be important in skeletal muscle blood flow autoregulation. 18 This conclusion is based on the observation that the marked reactive hyperemia which follows periods of circulatory arrest in skeletal muscle occurs without a measurable change in venous potassium concentration (102, 106, 107). However, Skinner and Powell (110) have shown that the hyperemia accompanying reduction of gracilis arterial blood oxygen saturation is enhanced when the arterial potassium ion con- centration is elevated above normal. 4. Adenosine and the Adenine Nucleotides Adenosine and its nucleotides appear to be important in the regulation of coronary blood flow (9, 76, 95, 101) and may be involved in exercise hyperemia (26, 37, 105, 108). However, evidence that these agents are involved in skeletal muscle blood flow autoregulation is lacking because they do not appear to be present in the venous effluent from muscles after release from long periods of flow reduction. Scott gE_§l. (108) reported that femoral venous blood contains adenosine and/or adenosine monophosphate during hindlimb exercise but not during reactive hyperemia. II. Myogenic Regulation of Vascular Tone The myogenic hypothesis of blood flow autoregulation states that vascular smooth muscle cells have an intrinsic ability to respond to changes in transmural pressure with contraction or relaxation. Support for this hypothesis has 19 been derived from studies of: a) the electrophysiological properties of isolated vascular smooth muscle; b) the elec- trical and mechanical responses of isolated vascular smooth muscle to stretch; c) the responses of intact microvascular networks to change in transmural pressure; and d) the resistance responses of whole vascular networks to changes in transmural pressure. A. Electrical and Mechanical Properties of Vascular Smooth Muscle There appear to be two distinct types of smooth muscle, "visceral" or single unit and multiunit smooth muscle (15). In the single unit variety, the cells act as a functional syncytium so that current spreads from cell to cell via low resistance pathways. In the multiunit type current does not pass from cell to cell and conduction is dependent upon nerves. Some cells of the single unit type appear to act as pacemakers since they are able to spontaneously depolarize to threshold. These cells are characterized as having a low and unstable membrane potential which slowly rises through some intrinsic pacemaker mechanism initiating complete depolarizations when threshold is reached. Periods of spike activity are followed by quiescent periods giving rise to phasic contractures. Cells of the multiunit variety show no tendency for spontaneous electrical activity and are thought to be driven by neural influences. 20 Prior to 1960 it was generally believed that vascular smooth muscle was of the multiunit type, incapable of myo— genic activity, since large arteries and veins failed to conduct action potentials in response to direct electrical stimulation (18, 38). It has recently been established that smooth muscle cells of small arteries and veins possess myogenic activity (3, 21, 39, 40, 60, 82, 114, 119). Funaki (39, 40) was the first to obtain intracellular electrical recordings from small blood vessels. He inserted microelec- trodes into smooth muscle cells of small pre- and post- capillary vessels of the frog tongue and observed a low, phasic, resting membrane potential. Local potentials were observed to gradually build until a depolarization occurred which was propagated to neighboring cells. Axelsson gt a1. (3) reported similar behavior in smooth muscle cells of the rat portal vein. During the quiescent period between bursts of action potentials he described a gradual rise in membrane potential suggestive of pacemaker activity. Reports have also appeared documenting the spread of induced activity in isolated blood vessels. In response to electrical or chemi- cal stimulation of rat portal vein strips, Johansson (60), Ljung and Stage (82), and Bevan and Ljung (12) observed a longitudinal spread of electrical and mechanical activity. Bevan and Ljung (12) obtained similar results in rabbit arteries, with propagation being especially pronounced in small arteries. These studies indicate that vascular smooth 21 muscle cells possess myogenic automaticity and behave as a functional syncytium capable of conducting electrical activity. B. Vascular Smooth Muscle Responses to Stretch Several studies have shown that vascular smooth muscle, like other single unit smooth muscles responds to passive stretch with contraction (7, 8, 22, 59, 61, 112, 113, 124). Using an isolated segment of carotid artery, Bayliss (8) was the first to observe contraction in response to elevation of internal pressure. From this and other less conclusive experiments on reactive hyperemia, he originated the myogen- ic hypothesis of blood flow control. The conclusions of Bayliss were criticized by Anrep (2) who could find no evidence of large vessel contraction with elevation of in- ternal pressure. In support of Bayliss' observation, Wachholder (124) reported rhythmic contraction of isolated carotid segments after elevation of internal pressure. Similar results were obtained twenty years later by Burgi (17) in his experiments on isolated segments of mesenteric arteries. More recently, Davignon e£_al. (22) perfused norepinephrine free human umbilical arteries and observed constriction when transmural pressure was elevated by either increasing perfusion pressure or applying vacuum outside the arteries. 22 Similar results have been obtained with helically cut strips of isolated arteries and veins. Using strips from canine paw arteries (200-500 u O.D.), Johansson and Bohr (59) observed slow spontaneous rhythmic contractions which increased in frequency with passive stretch. Sparks (112) found that the active tension development of human umbilical artery strips in response to passive stretch was related to the rate and increment of stretch, and to resting tension of the strip. Similar results were obtained by Sparks and Bohr (113) using helical strips from small branches of the canine superior mesenteric artery. In a recent study, Johansson and Mellander (61) observed that the electrical and contractile activity of portal vein strips depended on muscle length and were strongly influenced by variation in the rate of change in length. Active force and spike fre- quency showed graded increases with increasing rates of stretch. These studies on whole arterial segments and venous and arterial strips provide support for the view that vascular smooth muscle is myogenically active, respond- ing to increased stretch with contraction. C. Microvascular Responses to Transmural Pressure Changes A number of studies on intact microvascular networks provide support for the myogenic hypothesis (5, 33, 71, 91, 127-129). Fog (33) observed dilation of pial arteries with arterial pressure reduction and contraction with pressure 23 elevation. Similar results were obtained by Nicoll and Webb (91) in bat wing arterioles. Weideman (127, 129) studied the bat wing microvasculature and observed that venous pressure elevation elicited an increased frequency of rhythmic motion in venules. When pressure was elevated in the bat wing microvasculature by injection of saline, Weideman (128) found that the frequency of arteriole vaso- motion was increased. There is also evidence that precapil- lary sphincters respond to changes in transmural pressure. Johnson and Wayland (71) observed a periodic flow pattern in individual capillaries of cat mesentery that appeared to be caused by contraction and relaxation of the precapillary sphincters. Reduction of arterial pressure removed the periodic flow pattern but, when intravascular pressure was restored by increasing venous pressure, vasomotion was also restored. The responses described above do not appear to result from neural reflexes since bat wing vasomotion is still present after denervation and the periodic flow patterns in mesenteric capillaries were observed in a surgically iso- lated 100p of intestine. However, in each of these studies the induced changes in transmural pressure were accompanied by simultaneous changes in flow which might account for the responses via metabolic mechanisms. Evidence against this explanation is provided by Baez's (5) studies on rat meso— appendix arteriole diameter under conditions of no flow. 24 He found that intravascular pressure elevation caused some arterioles to constrict, with vessel radius actually becom— ing smaller at elevated intravascular pressure. D. Transmural Pressure Effects on Vascular Resistance There is a substantial amount of less direct evidence to suggest that the resistance vessels respond actively to changes in transmural pressure. The rise in precapillary resistance observed in many organs upon venous pressure ele- vation (venous-arteriolar response) is presently believed to be myogenically mediated (36, 49, 53, 62, 74, 88). Johnson (62) observed a strong venous arteriolar response in the intestine which was not dependent upon neural mechanisms nor due to purely physical factors. Hanson and Johnson (53) observed increases in hepatic arterial resistance in re- sponse to hepatic venous pressure elevation. Since the metabolic hypothesis predicts that these maneuvers would decrease arterial resistance, these results suggest myogenic mechanisms are operative in the intestine and liver. The venous-arteriolar response has not been consistent- ly observed in skeletal muscle vascular beds. Nagle g£_al. (88) and Folkow and Oberg (36) were able to consistently observe vasoconstriction with venous pressure elevation in denervated canine gracilis muscles and in the hindlegs of reserpinized cats. However, Jones and Berne (74) and Hanson (49) only observed a venous-arteriolar response in a small 25 portion of their isolated canine hindlimb preparations. It has been shown that the capillary filtration coeffi- cient (CFC) of limbs is sensitive to intravascular pressure, an increase in pressure causing the CFC to decrease. In the hindquarters of cats, Mellander gt_gl. (84) observed that simultaneous 50 mm Hg elevations of both venous and arterial pressure elicits an 18% increase in vascular resistance and a 59% reduction in CFC. Since both these responses were abolished by the smooth muscle poison chloral hydrate, they were attributed to a myogenic constriction of precapillary sphincters triggered fur elevated intraluminal pressure. There is also evidence to suggest that skeletal muscle vessels respond to the transmural pressure changes associ- ated with pulsatile perfusion. Rovick and Robertson (99) examined the effects of pulse pressure distension upon vascular resistance in the isolated dog tongue. At mean arterial perfusion pressures of from 80 to 160 mm Hg, vascu- lar resistance increased when pulse amplitude was increased from 0 to 60 mm Hg, with the effect being most pronounced at pulse pressures of 30-40 mm Hg. Mellander and Arvidsson (85) have performed similar experiments in the sympathectom- ized lower leg muscles of the cat. A sudden shift from non- pulsatile to pulsatile perfusion (mean perfusion pressure kept constant) elicited a 6% increase in muscle vascular resistance. When the vasculature was poisoned by papaverine, this response was abolished and replaced by a 10% decrease 26 in vascular resistance. These data indicate that the pulse pressure induced stretch of the vascular smooth muscle ini— tiates a myogenic constrictor response which overrides the effect of passive distension. Mellander and Arvidsson also examined the contribution of pulse pressure distension to the increased vascular resistance observed with elevated transmural pressure. Simultaneous 20 mm Hg elevations of venous and arterial pressure elicited a 7% increase in vascu- lar resistance during non-pulsatile and a 14% increase during pulsatile perfusion. In the papaverine treated, passive vasculature, this same elevation of transmural pres- sure decreased resistance by 20%. Thus, the distending effect of increased transmural pressure observed in the passive vasculature was abolished and replaced by an active, presumably myogenic, constrictor response in the normal vascular bed, but more effectively so during pulsatile than during non-pulsatile perfusion. METHODS Adult mongrel dogs of either sex, weighing 27—32 kg, were anesthetized with sodium pentobarbital (30 mg/kg, i.v.) and a cuffed endotracheal tube was inserted to provide a patent airway. Supplements of sodium pentobarbital were given as necessary during the surgical and experimental procedures. The right gracilis muscle was exposed via a cutaneous incision and the overlying fascia was stripped away. All branches of the gracilis artery and vein which did not go directly to or come directly from the muscle were ligated, as were other vascular connections to the muscle not arising from the gracilis artery and vein. The tendons of origin and insertion were tied to prevent collateral blood flow. After surgery was completed, sodium heparin (Wolins Pharmacal Corp., Mellville, New York) was administered in an initial dose of 700 USP units/kg body weight with hourly supplements of 300 USP units/kg body weight. Gracilis artery and vein pressures were measured from small bore (outside diameter (O.D.) 0.038"-0.048") polyethylene (P.E.) cannulae (Intramedic Tubing, Clay Adams, Parsippany, New Jersey) inserted into side branches of the gracilis artery 27 28 and vein. Gracilis muscle blood flow was determined by a different method for each experimental series (see below), and gracilis muscle vascular resistance was calculated by dividing the pressure drop across the vasculature by gracilis muscle blood flow. In all experiments mean systemic arterial pressure was measured from a P.E. 240 catheter (O.D. — 0.095“) inserted into the thoracic aorta via the left carotid artery. A11 cannulae used for pressure measurements were filled with saline and connected to low volume displacement transducers (No. P23 Gb, Statham Laboratories, Hato Rey, Puerto Rico) coupled to a direct writing oscillograph (No. 7784A, Hewlett Packard Co., Waltham, Massachusetts). The animals were warmed with a heating pad (Walker Co., Middleboro, Massachusetts) and with radiant heat supplied from a lamp (No. 1755, Burton Manufacturing Co., Van Nuys, California) directed at the legs and abdomen, so that rectal and muscle surface temperatures were maintained between 36.5 and 38° C. Rectal and gracilis muscle surface temperatures were monitored via thermister probes coupled to a direct reading tele-thermometer (No. 44TD, Yellow Springs Instru- ment Co., Yellow Springs, Ohio). The gracilis muscle was covered with saline saturated gauze which was coated with an inert silicone spray (Antifoam A, Dow Corning, Midland, Michigan) to prevent drying. 29 Series I: Naturally perfused, innervated gracilis muscles exposed to local hypotension during normovolemic and hypovolemic periods. In 10 spontaneously breathing dogs, the right gracilis muscle was prepared as described above with the gracilis nerve left intact. The gracilis vein was cannulated with a 6-8" section of P.E. 240 tubing downstream from the site of venous pressure measurement. Flow from the vein was directed into an Open reservoir maintained at a constant volume by a variable speed, roller pump (Lange Model RE 161, Extracorporeal Medical Specialties, Inc., Mt. Laurel Town- ship, New Jersey) which returned blood to the animal via a large vein. Gracilis muscle blood flow was determined by timed collection of venous outflow. An illustration of the experimental protocol is pro- vided in Figure 1. After a 30-40 minute control period dur- ing which intravascular pressures and muscle blood flow were allowed to stabilize, mean gracilis artery pressure was reduced approximately 20 mm Hg (minute zero on abscissa) by compression of the gracilis artery with a screw clamp. Intravascular pressures were recorded continuously and flow was determined 2 and 4 minutes after the reduction in gra— cilis artery pressure. This procedure for pressure reduc— tion and data collection was repeated 3 more times, with mean gracilis artery pressure being lowered an additional 10-25 mm Hg each time. The clamp was then released so that 30 A.c0HumHHommo ooaflmumo How uxou memo .ucoEHummxm 30am amusum: omuomamm m Scum mum sumo .cowmcmuomwc Hmflumuum UHEmummm mo mHm>mH 03D mcfiuso can MHEoHo>OEHoc and sync coflmcmpomxn HmooH on mmcommmu :fl AGHE\00V 30Hm cflm> mflaflomum one .Amm EEV wusmmmum humunm mflaflomnm .Amm EEV musmmmum amaumuum OHEoumhm coma CH momcmco .H madman H musmwm mm...32_s_ 03 ON. 00— 8 8 O¢ ON 0 0.: 40 (083.064... {It 3.903008%... 0000.741 m a... «lo... . J m M!» .6. (in... m «It... .... 39.. 25> 2.848 18 n I rfi ~61 - ..l_ r) 3. r1 2. w.t r...w .IJ _ III {to r) I 0.: .0 O 00:.ii I 9%fimwkijhtkdogmtflfig 32 perfusion pressure and blood flow returned to pre-clamp con- trol levels. After a recovery period which was terminated when pres- sures and flow had stabilized (minute 37 on abscissa), the animals were rapidly hemorrhaged from a carotid artery into a pressurized reservoir until mean systemic arterial pres— sure was reduced by approximately 10%. When the hemorrhage induced constriction had developed, the procedure described above for graded gracilis artery pressure reduction and data collection was repeated while the animals were maintained hypovolemic and hypotensive. After the last flow determina- tion, the clamp was released to allow pressures and flow to return to pre—clamp control levels. In 4 of the 10 experiments, the animals were rapidly hemorrhaged again (minute 89 on abscissa) to a systemic arterial pressure approximately 20% below normal. When pressures and flow had stabilized following the second hemorrhage, the procedure described above for local, graded hypotension and data col- lection was again repeated exactly as before. Series II: Pump perfused, innervated gracilis muscles exposed to alterations in mean arterial pressure during normovolemic and hypovole- mic periods. The right gracilis muscles of 8 dogs were prepared as described in Series I and a variable speed, roller pump (Lange Model RE 161, Extracorporeal Medical Specialties, Inc., Mt. Laurel Township, New Jersey) was interposed 33 between the femoral and gracilis arteries. The pump was fitted with a 4—5" section of silicone treated (Siliclad, Clay Adams, Parsippany, New Jersey) latex rubber rubing so that pump flow rate was independent of outflow resist- ance. Latex rubber tubing with an internal diameter (i.d.) of 0.094" was used so that pulse frequency was approximately 9/min/cc of flow delivered. A t-tube connected to an air- filled 12" section of P.E. 360 tubing was inserted into the gracilis arterial perfusion line to serve as a compliance chamber for damping the pulse delivered by the pump. Pulse pressure was maintained approximately 1/3 of mean gracilis artery pressure by adjusting the volume of air within the compliance chamber. The d.c. output of the pump was coupled to a direct writing oscillograph (No. 7784A, Hewlett Packard Co., Waltham, Massachusetts) and pump flow was calibrated before each experiment so that each 1 cc increment of flow produced a 4 mm pen deflection. The relationship between pump flow and galvanometer pen deflection was linear through- out the flow ranges studied and the calibration was checked periodically during each experiment by comparing a timed collection of gracilis vein flow with the oscillographic pen deflection. A heat exchanger was placed around the arterial inflow tubing between the pump and the muscle. Gracilis arterial blood temperature was measured downstream from the heat exchanger by a thermister probe (No. PC-130, Gormann- Rupp Industries, Belleville, Ohio) coupled to a direct In (I! 34 reading temperature controller (No. EC-250, Gormann-Rupp Industries, Belleville, Ohio) which maintained gracilis arterial blood temperatures between 37 and 38° C by con- tinuously adjusting fluid temperature in the heat exchanger. A servosystem (Figure 2) continuously adjusted the flow rate of the perfusion pump to maintain mean gracilis artery pressure constant at 60, 80, 100, 120, 140, 180, or 200 mm Hg. After the muscles had been prepared and pump perfusion established, 10-20 minutes were allowed to: A) determine 7 optimal combinations of the servosystem's setpoint, pro— portional gain, and integral control action required to main- tain mean gracilis artery pressure constant at 60, 80, 100, 120, 140, 180, and 200 mm Hg; and B) determine the 6 optimal combinations of the servosystem's proportional gain and integral control action required to change mean gracilis artery pressure from 140 to 60, 80, 100, 120, 180, and 200 mm Hg in 1—2 seconds with little or no overshoot (step pres- sure change). The servosystem parameters determined in B were used to return gracilus artery pressure from the pres— sures mentioned in B back to 140 mm Hg. After a 30 minute control period in which the muscle was perfused at a mean gracilis artery pressure of 140 mm Hg, the servosystem control parameters were adjusted to that mean gracilis artery pressure was rapidly reduced to and maintained at 80 mm Hg for 4-5 minutes. The servosystem control parameters were then adjusted so that mean gracilis 35 .oecHeucHeE mu consmuem 30am uceumcoo .eooE HenceE cuu3 “uceumcoo encemenm c0umsmuem cweucueE ou oeaaouucoo mu oeemm dado c0umsmued .eooE ouueEouse cuuz .eooa fiancee ecu cu coHuHmom uceEeHe Houucoo Hosea ecu mo uceEumsfloe Hencee muHE them one .meooE HenceE one owueEouse ceeSuec hammeadadc Eeumxm Houucoo ecu muemmcemullmuusouuo mmucouuzm Azlcv Hescmzrouuefiousm .Hmcmum usmuso HeumquEe ecu wo eosuwamae cam muuueaom ou ocuouoooe ucen nude udduso ecu mewue> “HeumuadEe ecu Scum emeuao> e we eocemce ecu cfl He>eH mauueuedo uceumcoo e um usmuso ucenuso e mafieucmea oce meofl>oudllaouucoo usmuso Aemwe mcuumSHoc uceuusuv .B.¢.O .emeuao> eoce nuemmuo ecu ou Hecowuuomond emeuao> usmuso cm meou>oum one memeu IHo> coecoeew oce uouue oeuwuooa ecu meuemEooulueuwHHmEe Houucou .Heaaouucoo ecu mo ucen nude unduso ecu Eoum oe>uneo mu emeuao> coecoeem ecB .memeuao> concoeem one Houue ecu ou ecowuoc5m meH no .eueu one uemeH .cuem Hmc0uuu0dond eaceumsfloe mooeilmucefiumshoe Hecem,aouucou .oemoHe>eo mu emeuao> Houue cm .ucHOQ uem Eoum meueu>eo eaceuue> oeaaonu ncoo ecu cecz .ueHMHHdEe Dc ecu Eoum emeuao> ecu ou oeuemeoo mu couc3 emeuao> eoceuemeu e mecmuaceumennuusouuo ucuom uem .H .eusmmeud mueuue muauoeum Houucoo ou pee: Seummmo>uem ecu mo ouueaecom .m eusmflm 36 N ehsmum mo<.50> xogomwn. wkzwsthoao< 4wz :8 £288 4.: 38:: 93.64% H $638245. .424 .mmwmwu.+...m.m. in? 4.3.648 4: 4 .I IIIIIIIIIIIIIIIIIIIIIIIIII I. 94¢ . _ 304m: _ SE8 :85 42:... . 5.8.5....“ _ _ _ _ _ _ _ 422. 3586 40:50 9586 8459 _ .22. 62.18.26 5&8 5.28 mzimtzm _ $59.. :4 H40 _ _ _ _ _ PI amnjombzoo Edd ON¢ aDmIkmoz oz< mowmc I_ 37 artery pressure was rapidly restored to and maintained at 140 mm Hg for 4 minutes. The procedure just described for a square wave change in mean gracilis artery pressure from 140 to 80 and back to 140 mm Hg was then repeated for mean gracilis artery pressures of 60, 100, 120, 180, and 200 mm Hg in a random sequence. Finally, the protocol described above for pressure reduction to 80 mm Hg and pressure restoration to 140 mm Hg was repeated. While mean gracilis artery pressure was maintained constant at 140 mm Hg by the action of the servosystem on pump flow rate, the animals were rapidly hemorrhaged from a carotid artery into a pressurized reservoir until mean systemic arterial pressure fell to 100 mm Hg. After a con- trol period of 20—40 minutes during which the hemorrhage induced constriction was allowed to develop fully, the pro— tocol described for alterations in mean gracilis artery pressure was repeated while the animals were maintained hypovolemic and hypotensive. During hypovolemia the se- quence of gracilis artery pressure changes and the times at a given pressure were matched to those obtained during normovolemia. After the last gracilis artery pressure manipulation, the gracilis nerve was coated with a local' anesthetic (Cetacaine, Cetylite Industries, Long Island City, New York), severed, and the responses to denervation were followed for 30-40 minutes. 38 As described above, mean gracilis artery pressure was altered throughout each experiment by changing the servo- system's control parameters so as to alter pressure within 1-2 seconds (Figure 3). The most immediate blood flow re- sponses to sudden changes in perfusion pressure are flow minimums or maximums occurring within 5-10 seconds after the pressure change. Similar transient responses have also been described by other investigators (116, 117), and are thought to result from passive collapse or expansion of the vasculature subsequent to the change in distending pressure (117) (see Discussion, Section I-B). Since the initial, rapid changes in pressure reported here are the result of sudden alterations in pump flow rate, it is possible that a significant portion of the recorded changes in blood flow represent artifacts of the servocontrolled pump flow rate and do not accurately reflect passively mediated changes in vascular resistance. To resolve this question, a manually operated pump (Lange Model RE 161, Extracorporeal Medical Specialties, Inc., Mt. Laurel Township, New Jersey) fitted with latex rubber tubing, was connected in series with the servocontrolled pump to provide a downstream resistance. When flow rate in the manually operated, downstream pump was halved or doubled within 0.2 seconds, the servocon— trolled pressure and flow transients lasted approximately 1.0 seconds (Figure 4, panel B). Since all recorded blood flow transients to square wave changes in gracilis artery 39 .mEm m.mm mcucmfles eaomse muauoenm oemsmuem QEDQ oeuoeaem e Eoum ewe eueo .emGOQmeH eueum moeeum ou eEuu mu0flmeo me eHHcB .muceumcenu 30Hm Edfiuxea no ESEHGHE xeed ou eEHu meueoflocu H9 .cHE\HE Cu oeucemeumen mu 30Hm mueuue mflauoenm “mm 85 CH oemmeudxe eue meusmmenm Heuueuue OHEeumhm one muauoenm ceez .ensmmeum coumsmnem CH memceco meum ou memcommen 3oam oooac mueuue mHHHoeuw .m eusmum 40 00. 09 FFLc- ON. 00_ m museum mozoomm F—. cm a cc J O n 3.3... o_ >mmum4 m. 4.364% ON MM. mmammmmn. 8. 34.554 08 6.2396 Mm: mmammmma 8. >554 08 2.545 41 .mmEnd ecu neezuec eunmmeum mnuaaouunoo mu Eeummmo>uem .mfinm oeaaouunooo>uem ecu ou eoneumumeu Eeeuuenzoo e oeou>oum dang oeueuemo maaennee ecu uecu 0m mend oeaaouunooo>uem ecu Eouu Eeeuuensoo meuuee nu oeuoennoo mend oeueuemo haaennee .m Henem .mmEnm ecu neezuec eunmmeum mnuaaouunoo mu Eeummm |o>uem .dEnm oeaaouunooo>uem ecu ou eounow 30Hu Eeeuummn nm oeou>oum mend oeueuemo maaennea ecu uecu om mfinm oeaaouu Inooo>uem ecu ou Eeeuummn meuuem nu oeuoennoo mend oeueuemo maaennea .d Henem .oeoa nu memneco meum ou AnuE\ooo euen 30am mesa one com EEV eunmmeum oeHHouunooo>uem mo memnommem .4 eunmum 42 mozoomm s N_Ae m e .. N o ! ul 1 a # q d E q d d 4 a d 1 L a v. N. O. m w v. N no 1 d d 4 E d 1 q u - d a d 4 d .v_ N_ n: w my v qldliqddd-ddd ‘1 #1 4 4 l 0. ON On 0_ ON On On 00. tom. 4 eunmum 8200mm 4. N. o_ m o 4 m o - d M d 1 d 4 4 d u q u d u N L a v_ N. O. m w v N O - d a d a q 1 q q q u d d d 1 ¢_ N. 0_ MW mw ¢ N no ! q d d d d + d J. H d d d d 1 221351211? < 1 l L O o . 30...... owccomkzoo 0N >44mmm On 0 mmnmmmma n omsuomuzoo m -o>mmm 43 pressure took at least 6 seconds to develop (see Results, section II—A) , the oscillographic blood flow recordings were assumed to reflect actual changes in vascular resistance. After each step change in gracilis artery pressure, transient muscle blood flow responses similar to those depicted in Figure 3 were observed in all 8 experiments. Data collected for subsequent analysis included: a) pres- sure and flow values prior to a change in perfusion pres- sure; b) intravascular pressures and peak minimum or maximum flow transient values at time T1 following a step change in perfusion pressure; c) the time to reach these initial flow transients (Tl); d) the time to reach the new steady-state level of flow (T2); e) intravascular pressures and muscle blood flow at time T2; f) the total time at each Pressure; and g) the planimetered area of the oscillographic flow recording above or below a horizontal line projected to the left from the steady state flow at time T2. Series III: Naturally perfused, denervated gracilis muscles exposed to elevation of mean vascular distending pressure and to local hypotension during pulsatile or non-pulsatile perfusion. The right gracilis muscles of 10 dogs were prepared as described above and acutely denervated. In all experiments, the animals were ventilated with room air through a cuffed endOtracheal tube with a positive pressure respirator (No. 607 , Harvard Apparatus Co., Dover, Massachusetts). 44 The muscle was perfused through a 6-8" section of sili- cone treated (Siliclad, Clay Adams, Parsippany, New Jersey) latex rubber tubing (i.d. = 0.25") inserted between the femoral and gracilis arteries. Within the length of rubber tubing a t-tube was inserted to which was attached a 25" section of P.E. 360 tubing. The distal end of the P.E. 360 tubing was fitted with a rubber bulb containing an air inlet valve to control pressure within the air-filled length of P.E. 360 tubing (compliance chamber). When pressure within the compliance chamber was equal to mean gracilis artery pressure, perfusion could be suddenly converted from pulsa- tile to nearly non-pulsatile without changing mean pressure by removing a clamp placed at the blood-air interface in the compliance chamber. In addition to mean gracilis artery and venous outflow pressures, gracilis artery pulse pressure was also measured from a side branch of the gracilis artery. The gracilis vein was cannulated with P.E. 240 tubing downstream from the site of pressure measurement and the venous outflow was directed into the right femoral vein via a servocontrolled pump (Lange Model RE 161, Extracorporeal Medical Specialties, Inc., Mt. Laurel Township, New Jersey) which.maintained a constant gracilis vein pressure. The servosystem depicted in Figure 2 was used except that the direction of control was reversed so that, for example, elevations in gracilis vein pressure caused by an increased muscleeblood flow resulted in an increased pump flow. 45 That is, the pump functioned as part of a closed-loop, nega- tive feedback, system which operated to keep gracilis vein pressure nearly constant. As described in Series II, the pump was fitted with latex rubber tubing so that pump flow rate was independent of outflow resistance. An air-filled compliance chamber was inserted on the inflow side of the pump to remove pulse pressure oscillations originating from the pump. The d.c. output of the pump was coupled to a direct writing oscillograph (No. 778A, Hewlett Packard Co., Waltham, Massachusetts) and pump flow rate was calibrated before each experiment so that a 1 cc increment of flow corresponded to a 4 mm pen deflection. The relationship between pump flow and galvanometer pen deflection was linear throughout the flow ranges studied and the calibration was checked several times during each experiment by comparing timed collections of pump flow with oscillographic pen deflections. To determine that the oscillographic recording of pump flow accurately reflected changes in gracilis vein flow, a manually operated pump (Lange Model RE 161, Extracorporeal Medical Specialties, Inc., Mt. Laurel Township, New Jersey) fitted with latex rubber rubing, was connected in series upstream to the servocontrolled pump, thereby providing a flow source to the servocontrolled pump. When the manually operated, upstream pump flow was halved or doubled within 0.2 seconds, the servocontrolled pressure and flow transients 46 lasted approximately 0.8 seconds (Figure 4, Panel A). Since all recorded flow transients in this series of experi- ments took at least 4 seconds to develop (see Results, section III-A) the oscillographic blood flow recordings were assumed to accurately reflect gracilis muscle blood flow. After the muscles had been prepared and perfusion established, the servosystem parameters required to maintain gracilis vein pressure constant at 4 or 24 mm Hg were determined and the preparation was allowed to stabilize for a 30-40 minute period during non-pulsatile perfusion with gracilis vein pressure held constant at 4 mm Hg. As illus- trated in Figure 5, when intravascular pressures and muscle blood flow had stabilized, a screw clamp on the gracilis artery was tightened so that mean gracilis artery pressure fell 20 mm Hg. After the transient flow responses had sub- sided, the arterial screw clamp and the setpoint of the out- flow pump servocontroller were adjusted so that mean gracilis arterial and venous pressures were simultaneously increased 20 mm Hg, producing an elevation in mean vascular distending pressure with no change in effective pressure gradient. After the transient flow responses had subsided, vein pressure was reduced to the control level by adjusting the outflow pump servocontroller parameters and pulsatile perfusion was restored by closing off the arterial compli- ance chamber. Control pressures and flow during pulsatile perfusion were determined and the procedure just described 47 .noumnmuem eHHuemHnm mnuuno mm ES ow one .om .ooa .oma ou mmeum nH oeonoeu mes eunmmeum hueuue mflawoeum neeE uXez .nOHmnmued eHHuemHnm mnauno necu one eHHuemHnmlnon mnuuno on E8 om oeue>eae umuwm me3 eunmmeum mnuoneumuo ueHnome> neez .uneEuuemxe HHH meuuem oeuoeaem e mo nouuuom Heuuunu ecu Eoum eEuu mo enouuonnm we chem ooa\nuE\ooo 30am nue> mflauoeum one can 85o meunmmeum mnone> one Heuueuue mHHuoeuw .m eunmum 48 On O¢ On m eunmum mmhnzi ON O. J O. n. ON ON O¢ Om ON 00. ON. OS 00. 30...”. Z_m> 94.055 mmommmmd 49 for elevation of mean vascular distending pressure was re- peated while the perfusion was pulsatile. When pressures and flow had stabilized after the manuever to increase mean vascular distending pressure, the screw clamp on the gra- cilis artery was tightened so that mean gracilis artery pressure was reduced in steps to 120, 100, 80, and 60 mm Hg. After each pressure reduction, time was allowed for the transient responses to subside. The clamp was then released so that pressures and flow returned to control levels and the entire protocol sequence depicted in Figure 5 (minutes 0-50) was repeated except that the pulsatile mode of per- fusion was reversed. To determine that the observed resistance responses resulted from active changes in vascular radius, a constant infusion of papaverine hydrochloride (a smooth muscle poi- son) was administered into the gracilis arterial supply. In all experiments a papaverine concentration was used that increased muscle blood flow 5-8 fold without reducing mean gracilis artery pressure more than 5-10 mm Hg. In 4 separate preparations the concentration range of papaverine used in these experiments was found to be effective in blocking the normal gracilis vasculature responses to test doses of norepinephrine and acetylcholine (Table 1). Because it was not possible to produce step changes in mean gracilis artery pressure by adjustment of the arterial screw clamp, an analysis of blood flow transients 50 Table 1. Effects of papaverine infusions on gracilis muscle vascular responses to graded infusions of norepinephrine and acetyl— choline. Values are means from 4 experiments. Gracilis Gracilis Gracilis % of % of Artery Vei Vascular Control Control Pressure Flow Resistance Flow Resistance Before Papaverine Infusion Control 142.2 13.4 10.6 100 100 Norepinephrine 0.1 Ug/min 142.3 7.9 17.9 58.9 169.2 0.5 pg/min 141.7 3.2 44.0 23.7 416.1 1.0 ug/min 141.9 1.4 101.2 10.6 955.2 Acetylcholine 0.1 ug/min 141.9 34.3 4.2 261.5 38.7 0.5 ug/min 142.3 52.1 2.7 400.2 24.9 1.0 ug/min 141.8 65.1 2.2 493.1 21.0 Constant Papaverine Infusion 1.9 mg/min Control 138.1 64.9 2.16 100 100 Norepinephrine 0.1 ug/min 138.1 61.8 2.21 95.4 104.8 0.5 Ug/min 138.3 59.4 2.34 92.5 109.4 1.0 ug/min 138.2 57.3 2.42 86.9 114.2 Acetylcholine 0.1 ug/min 138.1 65.1 2.12 100.4 99.8 0.5 ug/min 138.1 65.5 2.10 101.1 96.4 1.0 ug/min 138.3 66.6 2.07 103.3 95.2 Constant Papaverine Infusion 3.8 mg/min Control 133.2 93.9 1.41 100 100 Norepinephrine 0.1 ug/min 133.4 91.2 1.45 97.2 102.8 0.5 ug/min 133.4 90.4 1.47 96.2 104.3 1.0 ug/min 133.3 89.6 1.49 95.4 105.7 Acetylcholine 0.1 ug/min 133.1 94.2 1.41 100.3 99.8 0.5 ug/min 133.2 95.0 1.40 101.2 99.4 1.0 ug/min 133.1 95.3 1.39 101.5 99.2 amm Hg bcc/min/lOO gms C mm Hg/cc/min/lOO gms 51 subsequent to reductions in perfusion pressure or elevations of mean vascular distending pressure was not performed. However, since it was possible to instantaneously switch the mode of perfusion from pulsatile to non-pulsatile or vice- versa, analysis of the transient blood flow patterns result— ing from a change in the mode of perfusion was accomplished. After each mode change of gracilis artery perfusion, muscle blood flow responses similar to those depicted in Figure 6 were observed throughout all 8 experiments. Data collected from subsequent analysis were: a) mean and pulsatile intra- vascular pressures and venous flow prior to a change in the mode of perfusion; b) intravascular pressures and peak maxi- mum of minimum transient flow values resulting from a change in the mode of perfusion; c) the time required to reach these initial flow transients; d) the time required to reach the new steady-state level of flow; and e) intra- vascular pressures and muscle blood flow at the new steady state. Figure 6. 52 Gracilis arterial and venous pressure (mm Hg) and venous flow (cc/min) responses to changing the mode of perfusion from non-pulsatile to pulsatile and vice versa. Data are from a selected naturally perfused gracilis muscle weighing 103.5 gms. GRACILIS VEIN PRESSURE NEAN GRACILIS ARTERY PRESSURE GRACILIS ARTERY PRESSURE GRACILIS VEIN FLOW I50 53 0 4O 80 l20 ISO 200 240 . 280 I SECONDS Figure 6 DATA ANALY S I S Differences among means (muscle blood flow and vascu- lar resistance) at the different levels of local hypoten- sion obtained during normovolemia, hypovolemia, pulsatile perfusion, or non-pulsatile perfusion were compared by using the Student-Newman-Kuels procedure, a stepwise method in which the range is the test statistic. A Students t test for paired observations was used to analyze whether hemorrhage (Series II) or non-pulsatile perfusion (Series III) produced significantly different values from their respective controls with regard to a given parameter. A least square linear regression analysis was performed on each set of % of control flow and resistance data obtained over the autoregulatory range of perfusion pressures. The slopes obtained from the linear regression of control data were compared with the slopes of experimental data by using an analysis of covariance. In all three series of experi- :ments, blood flow autoregulation was determined to be present if a given percent change in local perfusion pres— sure elicited a smaller percent change in flow and a direc- tionally similar change in vascular resistance. For all comparisons, differences between means or regression coefficients were considered significant only if the 54 55 probability of making a type I error (a) was less than 0.05. A more detailed description of the statistical methods used is presented in Appendix B. RESULTS I. Series I: Naturally Perfused, Innervated Gracilis Muscle; Effects of Local Hypotension During Normovolemic and Hypovolemic Periods A. Muscle Blood Flow Figure 7 reports gracilis vein flow as a function of gracilis artery pressure during graded, local hypotension in normovolemic and hypovolemic dogs. When the animals were normovolemic, muscle blood flow was significantly re- duced from control with each step reduction in perfusion pressure. The gracilis vasculature of these normovolemic dogs displayed blood flow autoregulation in response to graded local hypotension as evidenced by the fact that a 45 i 3.1% decrease in arterial pressure from 130 :_4.2 to 72 i 2.6 mm Hg elicited only a 39 :_2.6% decrease in flow from 12.6 i 2.0 to 7.7 i 1.1 cc/min/lOO gms. When gracilis artery pressure was reduced from the normovolemic control value of 130 i 4'2 to 115 :_4.1 mm Hg by hemorrhage, muscle blood flow decreased significantly from 12.6 i 2.0 to 6.8 :_0.7 cc/min/lOO gms. This hemor- rhage induced reduction of gracilis artery pressure is quantitativelysfinfilar'to the first local reduction of per- fusion pressure obtained during normovolemia. The fall in blood flow accompanying hemorrhage is however, significantly 56 57 .muneE luuemxe 0H Eouu muouue oueoneum.H mneeE unemeumeu euec .mm 88 nu eunmmeum mueuue mwauoeum muuomeu emmuomce one mam ooa\nue\oo nfl 30am nwe> mwauoeum muuomeu euenflouo .onm nwe> mwauoeum no moofluem .meHOHHoV OHEeHo>om>c one .muoo ouaomo ouEeHo>oEuon mnfluno noumneuommc HeooH mo muoemwm .n eunmflm 58 h eunmam mmommmmn. >mwkm< 94.0410 Om. mN_ OO. 0% 00 em .. >>On_n. Z.w> m...:o muawoeum eunemeumeu euenuouo .moOHHem cmeuenvm one meaoufloo ouEeHo>om>c oBu one cmuooo UHEeHo>oEuon mnwuno noumneuommc HeooH mo muoemmm .m eunmfim 61 00. m eunefim wmowmwmn. >mwhm< m..._o £4.30 Lg 3 62 those accompanying equivalent local pressure reductions produced during normovolemia. During moderate hypovolemia (line connecting circles), step reductions of gracilis artery pressure from a control value of 120.8 : 5.6 to 94.3 i 7.2 mm Hg and below signifi— cantly decreased gracilis muscle blood flow. The ability to autoregulate blood flow was not compromised during the first hemorrhage, since a 46 i 2.5% local reduction in gracilis artery pressure from 120 i 5.6 to 65 :_2.4 mm Hg produced only a 28 i 2.3% fall in muscle blood flow from 7.9 :_1.4 to 5.7 :_0.9 cc/min/100 gms. During the second, more severe hemorrhage (line connecting squares), step reductions of gracilis artery pressure from a control value of 102.5 : 4.8 to 81 i 2.9 mm Hg and below signifi- cantly decreased gracilis muscle blood flow. The ability to autoregulate blood flow was again maintained since a 43 :_3.1% reduction in gracilis artery pressure from 103 i 4.8 to 59 i.2°5 mm Hg elicited only a 24 i 2.4% fall in .muscle blood flow from 5.1 :_l.6 to 3.9 i 0.5 cc/min/lOO gms. To more accurately describe and compare the relation- ships between muscle blood flow and perfusion pressure during normovolemia and hypovolemia, the data reported in Figure 7 over the gracilis artery pressure range of 70-120 Imn Hg were normalized to percent of control and subjected to linear regression analysis (Figure 9). The data obtained during normovolemia demonstrated a significant regression 63 I enm.o u mm .om.4 .>oase + He.mm + ..mmmna I o no w. mmo.o + mam.o u .umumem I o no m I mam.o u mm .mm.4 .>oEnoz + es.4m + ..mmmnn I o no w. aso.o + Nae.o u .umummm I o no w I mauve u an as u . . .mmm I. O . " .>0 mm m + an mm + . mm o e w. 4mo o + Hon 0 esoun I o no w I mambo u we .>oEnoz umH.m + m>.mm + A.mmeum I U m0 we Nmo.o + mmm.o BOHM U m0 w .mumwaene nOummenmeu ueenua euenom umeea mc oenueuco uum umec mo menua ecu eue eueo mo uem coee cmnoucu menuq .uneEuuemxe coee you oecuuo Imeo emneu ecu nucufl3 eunwmeum hueuue muauoeum umecmuc ecu ue oenHeu Ico emocu mmesae eues eueo ecu erHeEuon ou oemn menae> Houunoo ece .oeunooeu eue mooHuem UHEeHo>0d>c one ouEeHo>OEuon cuoc uou emneu ennm Imenm eEem ecu nucufl3 oenueuco eueo emocu mano .uneEuuemxe ne>Hm e mom .mm SE omflIoh mo emneu eunmmeum mueuue mflafloeum ecu ue>o muneEHHemxe 0H Eouu eneeE Henou>flonu eue eueo .eunmmeum mueuue muawoeum Houunoo mo w unemeudeu eemmuomce «eoneumumeu neanome> eHomnE muafloenm Houunoo mo w one 30am nue> muawoeum Houunoo mo w unemeumeu meuenuouo .cHened neBoHV eoneumumeu neanome> eaomnE mHHHoeum one AHenem uedmnv 30am nHe> muauoeum no moOMHem .menwa oecmeo one meaonuo. ouEeHo>ommc one .menHH mnonnuunoo one muoo. oHEeHo>0Euon mnuuno nOumneuomwc Heooa mo muoeumm .m eunmflm m eunmflm wmommmmn. >mwhm< 94.04.10 40m....ZO0 ....O .x. 8. om om 04. om. om it 1 ON m02<...m.mmm . om 40m....200 “.0 o\o M . 09 OO_ Om Om O... OO 1 . . . I i . . J 4 ow I On . Om .504... . om 40mFZOO u.O o\o I 00. 65 of flow as a function of perfusion pressure with a slope of 0.62, indicating that the gracilis vasculature autoregulated its blood flow. The data obtained during hypovolemia also showed a significant regression of flow as a function of perfusion pressure with a slope of 0.70. Because this regression coefficient is not significantly different from that observed during normovolemia, the hemorrhage induced vasoconstriction did not impair the ability of the gracilis vasculature to autoregulate flow in response to local hypo- tension. B. Muscle Vascular Resistance Figure 10 reports gracilis muscle vascular resistance as a function of gracilis artery pressure during graded, local hypotension in normovolemic and hypovolemic dogs. When the animals were normovolemic, reduction of gracilis artery pressure from 112.1 :_3.6 to 72.4 :_2.6 mm Hg elicit- ed a significant reduction in muscle vascular resistance from 12.8 i 1.9 to 10.4 :_1.6 mm Hg/cc/min/lOO gms. When gracilis artery pressure was reduced below the normovolemic control of 130 i 4.2 to 114 i 4.1 mm Hg by hemorrhage, muscle vascular resistance increased significantly from 11.7 i 1.8 to 17.8 i 1.9 mm Hg/cc/min/lOO gms. During hypo- volemia, local step reductions of gracilis artery pressure from a control value of 115 i 4'1 to 77.9 i 3.0 and 66.4 i 3.0 mm Hg significantly reduced gracilis muscle vascular 66 .muneEuuemxe oa Eonm muouue oneoneum H mneeE ene eueo .mm as nu eunmmeum mueuue mHHHoeum munemeumeu emmuomce one mam ooa\nHE\00\mm EE nH eoneumumeu ueHnome> eHomnE mHHHoenm munemeumeu euenHouo .eoneumflmeu Heanome> eHomnE mflauoenm no wooeuem .meaouwo. ouEeHo>om>c one .muoo. aneHo>oEuon mnfluno nOHmneuommc HeooH mo muoeuwm .oa eunmflm 67 cu museum wmommwmd >mm....m.< m.4_0omhc eueueooz 0.4 m e.mn m.H m o.m m.m m m.omu m.4 m m.NmH Honucoouumon 0.4 + a.mn 4.H + a.» e.m + m.omH s.4 + m.N~H Homecoo .4 u n. eHEeHo>ommc oHHz o.m m m.mn e.n m m.HH o.m m m.ema m.4_m m.ama Houpcoouumom o.m + m.mu e.u + m.uu o.m + m.emn m.m + o.N4H Honueoo Av H CV GHEOHO>OEHOZ m.m m 4.5a m.o m o.» 4.4 m m.mna m.4 m e.mHH Honucoonumom m.u + m.uu m.o + m.e H.4 + m.mHH m.m + H.mHH Houuaoo .oa n n. euEeHo>om>c m.u m m.HH o.m m e.mu m.4 m m.HmH m.4 m m.mma Honucoqumon o.m + m.nu o.m + e.NH N.4 + e.omu m.4 + H.5ma Honucoo Aoa n no eHEeHO>OEuOZ em on on mm .Ame eone Iumumeu Heunome> eHomnE euafloeum one ..mm. 30Hm nHe> muafloeum .com. eunm Imeum aneune muauoeum neeE .cmm. eunmmeum Hewueuue ouEeumMm neeE now muouue oueoneum one menae> neeE ene eueo .euEeHo>0m>c one eHEeHo>OEuon cuoc onu Iuno noumneuomwc oeoeum .aeooH .Houunoqumom. Heume one .Houunoo. ou uoflum oe>nemco .mEm ooa\nHE\00\mm ES. meoneumwmeu ueanome> eaomnE one .cmEm ooa \nHE\oo. mBOHu oooHc euomne ..mc ES. meunmmeum Heanome>euunu mo nowanemeoo .m eucee 70 II. Series II. Pump Perfused, Innervated Gracilis Muscle; Effects of Alterations in Mean Arterial Pressure During Normovolemic and Hypovolemic Periods A. Muscle Blood Flow Figure 11 reports changes in gracilis muscle blood flow from a selected experiment in which perfusion pressure was altered locally during both normovolemia and hypo— volemia. During normovolemia, reducing gracilis artery pressure from 140 to 80 mm Hg by adjustment of the perfu- sion pump servocontroller parameters caused an initial rapid decrease in flow to a minimum value at time T Flow 1’ then rose slowly to a new steady state value at time T2. During hypovolemia, a step change in gracilis artery pres- sure from 140 to 80 mm Hg elicited a similar pattern of blood flow responses. When the animals were normovolemic, restoration of gracilis artery pressure to 140 mm Hg re- sulted in an initial, rapid increase in flow to a maximum value at time T1, which was followed by a slower return of blood flow to a new steady state at time T2. During hypo- volemia, restoration of gracilis artery pressure to 140 mm Hg elicited muscle blood flow responses similar in form to those observed during normovolemia. The time to reach the minimum or maximum transient flow values (T1) and the time to reach the new steady state levels of flow (T2) were always longer during hypovolemic than during a correspond- ing normovolemic period. Figure 11. 71 Gracilis artery blood flow responses to step changes in perfusion pressure during normo- volemic and hypovolemic periods. Mean gra- cilis and systemic arterial pressures are expressed in mm Hg; gracilis artery flow is expressed in ml/min. T1 indicates time to peak minimum or maximum flow transient values, while T2 depicts time to steady state responses. Data are from a seleCted, pump perfused gracilis muscle weighing 93.5 gms. GRACI LIS ARTERY PRESSURE SYSTEMIC ARTERIAL PRESSURE GRACILIS ARTERY F LOW GRACILIS ARTERY PRESSURE SYSTEMIC ARTERIAL PRESSURE GRACILIS ARTERY FLOW 72 HYPOVOLEMIC ‘ .__-——. A ‘ A -A_. A. AA. A. . f .v— ‘4— w .v w h M A— - .---n __ v v OI' L 4 4 A A A L 4 L L L 1 O 20 4O 60 80 IOO I20 I40 I60 I80 200 SECONDS 200 NORMOVOLEMIC I50 - .. - ---... loot-‘1 ._ .. I 50 200 I50 WWW I00 50 20 OZO406DBOIOOI20I40I60I80 secouos Figure 11 73 Figure 12 reports gracilis muscle blood flow responses to step changes in gracilis artery pressure.from 140 to 80 and back to 140 mm Hg. Data are mean values and standard errors for responses obtained during normovolemia (upper panels) and during hypovolemia (lower panels). When the animals were normovolemic (upper left panel), reducing gra- cilis artery pressure from 140.1 : 0.7 to 80.1 :_0.6 mm Hg in 1.7 i 0.3 seconds elicited a fall in muscle blood flow from 10.0 i,1°2 cc/min/lOO gms to 2.6 i 0.3 cc/min/lOO gms. This minimum flow was reached in 13.8 i 0.5 seconds and corresponded to a 74.5% reduction from the control value of 10.0 cc/min/100 gms observed at a perfusion pressure of 140 mm Hg. The initial period of declining flow was brief and while gracilis artery pressure remained at 80 mm Hg, muscle blood flow rose to a steady state value of 6.6 i 1.0 cc/min/lOO gms. This steady state level of flow was at- tained at 40.6 :_l.l seconds and was 36.3% less than the control flow rate of 10.0 cc/min/100 gms. The gracilis vasculature of these normovolemic dogs displayed blood flow autoregulation, since steady state flow fell only 36.3% when perfusion pressure was reduced 42.9% (from 140 to 80 mm Hg). The area enclosed within the oscillographic flow recordings below horizontal lines projected to the left from the steady state flows averaged 63.6 :_7.7 ml x seconds. 74 .muneEunemxe m Eonm muouue oueoneum one menae> neeE eue eueo Hae one monooem x no nfl oemmeumxe eue meeue emece .mzoam eueum moeeum ecu Eoum umea ecu ou oeuoehoum menHH HeunoNuuoc .mHenem ucmuuo e>oce no .maenem umeu. soaec mmneouooeu 30am oooHc oacmeumoHHfiomo ecu mo eeue oeneueEuneHm emene>e ecu munemeumen Henem coee nucuflz HecEnn necuo ece .3oHu EnEuer no Ennunufi xeem ecu ue oenueuco 3oam Houunoo mo uneenem ecu unemeumeu mHenem onec ucmwu ecu nH m3oHu EnEuer ceem ecu e>oce one maenem onec umeH ecu nu mzoam EnEwnuE ceem ecu Boaec menHe> uneouem ece .eEHu one 30am you muouue oueoneum N muneeeumeu munuom eueo ecu cmnoucu uec HeunoNHuoc one Heofluue> coee one 3oam mom muouue oueoneum unemeumeu munfiom eueo mnfluoennoo menua Boaec one e>oce meene oeoecm ece .monooem nu eEHu unemeumeu eemmwomce one mEm ooa\nuE\oo nfi 30H“ nwe> muawoeum unemeumeu meuenuouo .mm as 04H ou om Eoum mnOHue>eae eunmmeum noumnmuem ou memnommeu 3oam 30cm ucmuu ecu no mHenem .mm ES om ou 04H Eoum mnouuonoeu eunmmeum nOumnmuem ou memnommeu 30Hm uoameo umea ecu no mHenem .moOHHem .muenem oBu ueBoHv ouEeHo>om>c one .maenem OBu Hemmno UHEeHo> IOEuon cuoc mnunno oenueuco mm ES o4a ou om ou 04H Eouu mnOAueHeuHe eunmmeum aneuue mflauoeum HeHunenUem ou memnommeu.onm oooac eaomnz .m. museum 75 NH eunmum N o N MOI-I NIBA SI'IIOVHS mozoomm |o.m . o... . Io . mozoomm . 8n . 8» 08 o 8 8 . o I smorwmoe \ III E c :. «.036. § \ m m mem o. o. s...\ m. 9 8.93.9... 828mm 8 Om O¢ O . . . . . 828mm new . ImmmI . m8 0 on . 04 . lo 0 a $33.3 S +. 5.- . m .\He+. m ‘\ .\ §+ -9 ._ .9 new“ m.__~ .49 .0. 4+ .8 - 4.4.4.3.» .8 -8 M013 NI3A SI'IIOVHS 76 When the animals were hypovolemic, reducing gracilis artery pressure from 140.2 : 0.6 to 79.9 :_0.5 mm Hg in 1.7 H- 0.4 seconds elicited a fall in muscle blood flow from 6.1 i 0'7 cc/min/lOO gms to 2.5 i 0.3 cc/min/lOO gms (Figure 12, lower left panel). This minimum flow level was reached in 19.8 i,1°l seconds and represented a 59.1% reduction from the control flow rate. This 59.1 i 1.0% fall in blood flow observed in response to perfusion pressure reduction from 140 to 80 mm Hg during hypovolemia was significantly less than the 74.5 i 0.9% reduction observed in response to a similar maneuver performed during normovolemia. In addition, the time required to reach this peak minimum flow was sig- nificantly longer during hypovolemia than during normo- volemia. As observed during normovolemia, the initial period of declining flow associated with a decrease in perfusion pres— sure was brief and flow subsequently rose to a steady state value of 4.3 :_0.6 cc/min/lOO gms. This new steady state rate was 31.1':_1.3% below the control rate of 6.1 cc/min/ 100 gms. The percent decrease in steady state flow accom- panying a perfusion pressure reduction from 140 to 80 mm Hg was significantly less during hypovolemia (31.1 i 1.3% versus 36.3 i_l.5% during normovolemia), indicating that blood flow autoregulation may actually be enhanced by hemor— rhage. However, the 74.4 :_3.0 seconds required to reach steady state flow during hypovolemia was significantly 77 greater than the 40.6 i 1.1 seconds required during normo— volemia, indicating that the autoregulatory response to a step reduction in perfusion pressure develops more slowly during systemic arterial hypotension. The area enclosed within the oscillographic blood flow recordings below horizontal lines projected to the left from the steady state flows averaged 61.8 :_8.2 ml x seconds during hypovolemia and was not significantly different from that observed during normovolemia. These areas were not significantly different even though the amplitude of the transient and steady state flow responses to perfusion pres- sure reduction during hypovolemia were significantly smaller than those observed during normovolemia. This discrepancy results from the longer time required to reach steady state flow responses during hypovolemia. After gracilis artery pressure had been maintained at 80 mm Hg for 295.3 :_9.4 seconds during normovolemia (Figure 12, upper right panel), elevation of perfusion pressure to 140 :_0.8 mm Hg in 1.8 :_0.5 seconds elicited an increase in muscle blood flow from 6.6 i 1.0 cc/min/lOO gms to a maximum of 19.7 :_2.2 cc/min/100 gms. This peak flow took 8.8 i 0.5 seconds to develop and represented an elevation to 318 i_14.8% of the control value of 6.6 cc/ min/100 gms observed at a perfusion pressure of 80 mm Hg. This initial period of rising flow was brief and muscle blood flow subsequently fell to a steady state value of 78 10.0 i 1.3 cc/min/lOO gms. This steady state level of flow required 53 :_1.6 seconds to develop and corresponded to a 150.1 :_2.7% increase from the control rate observed at 80 mm Hg. The gracilis vasculature displays blood flow auto- regulation in response to perfusion pressure elevation from 80 to 140 mm Hg since the 175% elevation in perfusion pres- sure elicited only a 150% increase in flow. The area en- closed within the oscillographic blood flow recordings above horizontal lines projected to the left from the steady state flows averaged 211.9 : 26.3 ml x seconds. After perfusion pressure had been maintained at 80 mm Hg for 281.1 i.13-6 seconds during hypovolemia (Figure 12, lower right panel), elevation of gracilis artery pressure to 140.1 1 0.5 mm Hg in 1.8 :_0.3 seconds elicited an in- crease in muscle blood flow from 4.3 : 0.6 cc/min/lOO gms to a maximum of 14.4 :_l.9 cc/min/lOO gms. This peak flow took 22.5 i 1.8 seconds to develop and represented a flow elevation to 346.2 :_13.8% of the control rate observed at a perfusion pressure of 80 mm Hg. This 346% increase in blood flow observed in response to perfusion pressure ele- vation from 80 to 140 mm Hg during hypovolemia was signifi— cantly greater than the 318% increase in flow observed in response to a similar maneuver performed during normo- volemia. Furthermore, time required to reach peak flow was significantly greater during hypovolemia than during normo— volemia. 79 As observed during normovolemia, the initial rising phase of blood flow accompanying increased perfusion pres— sure was brief and flow subsequently fell to a steady state level of 6.1 :_0.8 cc/min/lOO gms which corresponded to a flow elevation to 145.2 1 1.8% of the control level of 4.3 cc/min/lOO gms observed at 80 mm Hg. Since this 145% flow increase observed in response to a 175% elevation of perfu- sion pressure from 80 to 140 mm Hg during hypovolemia was significantly less than the 150% elevation of flow accom- panying a similar maneuver performed during normovolemia, blood flow autoregulation again appeared to be more pro- nounced during systemic arterial hypotension. However, the 85.8 :_4.1 seconds required to reach this steady state level of flow during hypovolemia was significantly greater than the 53.0 i_l.6 seconds required during normovolemia, indicating that the autoregulatory response to a step ele- vation of perfusion pressure develops more slowly during systemic arterial hypotension. The area enclosed within the oscillographic blood flow recordings above horizontal lines projected to the left from the steady state flows averaged 213.1 1 29.6 ml x seconds during hypovolemia and was not significantly differ- ent from that observed during normovolemia. These areas were not significantly different in normovolemic versus hypovolemic periods even though the amplitude of the trans- ient and steady state flow responses to perfusion pressure 80 elevation during hypovolemia were significantly smaller than those observed during normovolemia. This discrepancy results from the longer time required to reach the steady state flow responses during hypovolemia. During both normovolemic and hypovolemic periods, the time required to reach peak minimum flow when perfusion pressure was reduced to 80 mm Hg was significantly greater than that time required to reach peak maximum flow when perfusion pressure was restored to 140 mm Hg. In addition, the time required to reach a new steady state flow after perfusion pressure reduction to 80 mm Hg was always sig- nificantly less than that time required to reach a new steady state flow after perfusion pressure was restored to 140 mm Hg. Table 3 reports all data described above in Figure 12 as well as the flow responses to step changes in perfusion pressure to and from 200, 180, 120, 100, and 60 mm Hg. The data depicted in Figure 12 are contained in the first four rows of Table 3 and represent responses to the first set of gracilis artery pressure alterations performed in each normovolemic and hypovolemic period. The last four rows of Table 3 report the flow responses to this same set of gracilis artery pressure manipulations performed at the end of each normovolemic and hypovolemic period. Because all responses to step changes in gracilis artery pressure from 140 to 80 and back to 140 mm Hg were statistically 81 concaucoo 04+ 0.0+ 3.4+ 0.0.“ 0.3M 0.3..” T? 04+ TOM Woo... 0.2.... 0.0.,“ m.o+ NAM «.mva H.0 0.00 o.ova H.mam m.0vm v.0a m.mm m.H H.ova H.me m.v 0.00 H.ooa Tm.“ 0.0M 0......“ m6“ Nd...“ mam Ton H4“ You m.oH m0.“ To.“ 0.0M HAM 0.00 m.v 0.05 0.00 m.a0t m.ov m.m m.ma n.H m.mn n.0mm H.0 m.ova H.ooa urn...“ mg.“ 04.“ 0.0.... 80%“ HEN N.N.+. m6“ To.“ m.oH man 04...“ man m.mH H.oma o.oa o.mm H.00H m.HHm 0.0Hm n.ma 0.0 m.a o.ovH m.mmm 0.0 H.00 m.ava 5% 04H 34“ To.“ HKH To“. TOM 90H To“. 0.0M m.m.+. «AH Ton 06H n.m0 0.0 0.00 H.om 0.m0n 0.0m 0.m m.ma h.H H.om v.mvm o.oa H.00H m.ava Uwom 0.0 m9 Um fl Uwom 0.0 mm. as um 08 um mm mm mmmZOmmmm medem woémem mumZOmmmm BzmHmz .mocoomm x 00 ca mmum 0cm “mccoomm CH mafia “mam ooa\CwE\oo CH mBOHm «mm 55 ca commoumxm mum mausmmmum .mmcfiouoomu 3on ucoflmcmuu onmn no m>onm mmumud “30Hm cwo> mwaflomum Houucoo mo usmoquHUmwm “moam> 30H0 zoom» 0» omuflsqou mEHuuhB «madmmmum 30c nommu on powwowmu oEwuumB «coaumm Houucoo mo coflumu50nua “30Hm cfio> mflafiomumuom «musmmoum ammuum maawomum cmofinum «madmmoum Hmwuouuw Uflfimpmxm :mmEumm .oHSmmmum coflmswumm CH mmmcmso down on momcommou 30am vooHQ maum5E mwaaomuo .m magma 005000000 82 0.0+ 0.0+ 0.0+ 0.0+ 0.0+ 0.0+ 0.0M 0.0+ 0.0“ 0.0“ 0.0+ 0.0M 0.0+ 0.0+ 0.00 0.0 0.00 0.000 0.0 0.00 0.0 0.00 0.0 0.000 0.000 0.0 0.000 0.000 0.0“ 0.0“ 0.0“ 0.00 0.00“ 0.0“ 0.0“ 0.0“ 0.0“. 0.0“ 0.0“ 0.0“ 0.0“ 0.0“ 0.000 0.0 0.000 0.000 0.00 0.000 0.00 0.00 0.0 0.000 0.000 0.0 0.000 0.000 0.0M 0.00 0.0“. 0.0M 0.0M 0.0M 0.00 0.00 0.0M 0.0M 0.0H 0.0M 0.0M 0.0“ 0.00 0.00 0.00 0.000 0.00- 0.00 0.0 0.00 0.0 0.000 0.000 0.00 0.000 0.000 0.00 0.00 0.0“ 0.00 0.0“ 0.0“ 0.0“ 0.0“ 0.00 0.0“ 0.0“ 0.0M 0.0M 0.00 0.000 0.00 0.00 0.000 0.00 0.000 0.00 0.00 0.0 0.000 0.000 0.00 0.000 0.000 0.0“ 0.00. 0.0“ 0.00 0.00 0.0“ 0.0“ 0.0“ 0.0“ 0.0“ 0.00“ 0.0“ 0.0“. 0.00 0.00 0.0 0.00 0.000 0.00 0.00 0.0 0.00 0.0 0.000 0.000 0.00 0.000 0.000 0.0M 0.0M 0.0M 0.0M 0.00M 0.0“ 0.0“ 0.0M 0.0“ 0.0“ 0.0M 0.0“ 0.0“ 0.0“ 0.000 0.00 0.000 0.000 0.000 0.000 0.00 0.00 0.0 0.000 0.000 0.0 0.000 0.000 0.0M 0.00 0.00. 0.00 0.0“ 0.0“ 0.0“ 0.0“ 0.0“ 0.0“ 0.0“ 0.0“ 0.0“ 0.0H 0.00 0.00 0.00 0.000 0.00: 0.00 0.0 0.00 0.0 0.000 0.000 0.00 0.000 0.000 0.0“ 0.00 0.00 0.00 0.00M 0.0“ 0.0“ 0.0“ 0.0“ 0.0“ 0.0M 0.0M 0.0M 0.0“ 0.000 0.00 0.00 0.000 0.000 0.000 0.00 0.00 0.0 0.000 0.000 0.00 0.000 0.000 0 0000 00 a 00 0 0000 00 00 00 00 00 00 00 00 000200000 00000 000000 000200000 020002000 0000200 000:0ucoonum 00009 83 Umscflucoo 0.0+ 0.0+ 0.0+ 0.0M. 0.0+ 0.0+ 0.0M 0.0+ 0.00 0.0“ 0.0+ 0.0+ 0.00 0.0+ 0.000 0.0 0.00 0.000 0.00 0.000 0.00 0.00 0.0 0.000 0.000 0.0 0.000 0.000 0.0“ 0.00. 0.0“ 0.0“ 0.0M 0.00 0.0“ 0.0“ 0.0M 0.0“ 0.0“ 0.00 0.00 0.0“ 0.00 0.0 0.00 0.000 0.00- 0.00 0.0 0.00 0.0 0.000 0.000 0.0 0.000 0.000 0.0M. 0.00 0.0“ 0.00 0.00 0.0“ 0.0“. 0.0“. 0.00 0.0“ 0.0H 0.0M 0.0M 0.00 0.000 0.00 0.00 0.000 0.00 0.000 0.00 0.0 0.0 0.000 0.000 0.0 0.000 0.000 0.00 0.0“ 0.00 0.00. 0.00 0.00 0.0“ 0.0“ 0.00 0.00 0.00 0.00 0.00 0.00 0.00 0.0 0.00 0.000 0.00- 0.00 0.0 0.00 0.0 0.000 0.000 0.00 0.000 0.000 0.0M 0.00 0.00 0.00 0.00 0.00 0.0“ 0.00 0.0H 0.00 0.00 0.00 0.00 0.00 0.000 0.0 0.00 0.000 0.00 0.000 0.0 0.00 0.0 0.000 0.000 0.0 0.000 0.000 0.0H 0.0M 0.0M 0.0...“ 0.0.“ 0.0.“ 0.0M 0.0...H 0.0H 0.00.0 0.000 0.0|+. 0.0.“ 0.0M 0.00 0.0 0.00 0.000 0.00- 0.00 0.0 0.00 0.0 0.000 0.000 0.0 0.000 0.000 0.0“ 0.00 0.00 0.00 0.0“ 0.0“ 0.00 0.00 0.0M 0.00 0.0“ 0.0“ 0.0“ 0.00 0.000 0.00 0.00 0.000 0.00 0.000 0.00 0.00 0.0 0.000 0.000 0.0 0.000 0.000 0.0“ 0.00 0.00 0.00 0.00 0.0“ 0.0“ 0.0M 0.00 0.00 0.0“ 0.00 0.00 0.0“ 0.00 0.0 0.00 0.000 0.00- 0.00 0.0 0.00 0.0 0.000 0.000 0.00 0.000 0.000 0000 00 00 00 0 0000 00 00 00 00 00 00 00 00 000200000 00000 000000 000200000 020002000 0000200 vmscflucoouum 00309 84 N.nu 5.0+ o.m+ 5.QH m.mmu o.mau m.mfl 5.H+ m.o+ o.QH m.mu. m.o+ m.o+ m.mH m.¢va 0.0 v.0m o.ova «.mflm H.mmm m.va m.am m.H H.0vH m.mm~ m.v 0.0m >.ooa H.nu m.qu m.mH m.qu m.mfl o.dH m.qu m.¢u v.qH m.qH m.mH m.qu n.QH m.mu m.vo m.q m.h> 0.0m >.mou v.Hv m.~ 0.0m n.H 0.0m m.vv~ 0.0 m.ova n.ooa o.¢H v.&H m.du n.qu “.mmH m.m&H H.mu m.qu m.qu o.qH ~.aH m.mH m.oH_ o.afl v.mmH H.0H m.vm o.ova «.mmm «.mmm m.mH m.m m.H o.ova v.am~ 0.0 0.0m «.mva m.mu m.&H s.dH v.QH m.mfl m.qu m.QH m.QH ¢.QH m.QH H.mH v.&H B.QH m.mH m.ao o.m m.mv 0.0m v.mo- v.mm m.m o.ma 5.H 0.0m n.mv~ H.0H m.ova «.mva m.mH m.qu “.mu m.qu v.0mfl m.HaH m.mH m.flH v.qu m.QH H.nH v.QH m.QH H.mfl m.ama H.o o.moa m.ova m.vom n.0vm 0.5H m.mm m.H m.ova H.mm~ m.m v.00 n.00H m.fiH v.QH o.mfl m.qH o.@fl m.QH H.QH o.flH v.qu m.qu «.mu m.QH m.qu H.mu n.mm N.m H.voa v.00 m.vm- m.oH o.H o.mm m.H v.00 ~.Hv~ H.o N.ova m.ooa m.¢H m.¢H m.dfl m.QH o.mmu m.mH N.nH m.QH m.qn m.qu «.mu 0.0“. m.oH_ N.QH m.mom 0.0H H.Nn H.0qH m.mam m.mvm m.om m.HH m.H H.0VH m.ov~ m.q m.om o.mva o.&n o.qu H.dH m.oH. m.mH m.qu ~.qu m.qH m.qu m.qu m.nH m.&u m.ou. m.oH m.mv m.q m.hn m.om m.vo- m.mH m.a m.NH m.a m.oo «.mvm H.0H H.owa o.mva owom um ma om ¢ owom mm me me mm 09 mm wm mm mmmzommmm mamem unamem mmmzommmm azmHmz¢ms qomezou vmscwucooanm magma 85 similar before and after each experimental period, the prep- arations were assumed to have been stable throughout the normovolemic and hypovolemic periods. Transient and steady state flow responses to gracilis artery pressure alterations to and from 120, 100 and 60 mm Hg during normovolemia and hypovolemia are similar to those responses described above for perfusion pressure altera— tions to and from 80 mm Hg. For example, after each change in perfusion pressure to and from 120, 100, 80, and 60 mm Hg, more time was required to reach the peak transient and steady state flows during hypovolemia than during normo— volemia. In addition, the areas above the minimum trans— ient flows and below the maximum transient flows were the same in normovolemic versus hypovolemic periods. As ob- served for pressure alterations from 80 to 140 mm Hg, changes in gracilis artery pressure from 120, 100, or 60 to 140 mm Hg elicited greater percent changes in initial transient flow in hypovolemic versus normovolemic periods. Furthermore, a given change in gracilis artery pressure always produced a proportionately smaller change in steady state flow, indicating that the vasculature autoregulated its blood flow in response to each of the step changes in perfusion pressure. Changing gracilis artery pressure to 180 or 200 mm Hg and back to 140 mm Hg elicited a somewhat different set of flow responses than those described above for perfusion 86 pressure alterations below 140 mm Hg. As described previous- ly, more time was required to reach transient and steady state flows when the dogs were hypovolemic. However, when gracilis artery pressure was elevated from 140 to 180 or 200 mm Hg, the percent increases in flow occurring at T1 and T2 were not significantly different during normovolemic versus hypovolemic periods. Further, the gracilis vascula- ture appeared less able to autoregulate its blood flow in response to elevations in perfusion pressure above 140 mm Hg. This conclusion is based on the observation that 29 and 43% increases in gracilis artery pressure (140 to 180 and 140 to 200 mm Hg, respectively) elicited 45 and 92% increases in steady state flow. Figure 13 reports steady state gracilis vein flow as a function of gracilis artery pressure when perfusion pres- sure was altered from 140 mm Hg by changing the perfusion pump servosystem parameters during normovolemic (dots) and hypovolemic (circles) periods. During normovolemia, reduc- tions of gracilis artery pressure from 140 to 100 mm Hg and below, or elevations of perfusion pressure from 140 to 180 and 200 mm Hg significantly altered muscle blood flow. During hypovolemia, muscle blood flow was significantly reduced from the normovolemic control values observed at each level of perfusion pressure. Except for the local pressure reduction from 140 to 120 mm Hg, all changes of gracilis artery pressure from 140 mm Hg performed during 87 .mucmeflnmmxo m Scum mnouum cumccmum _H mmsHm> some ucmmmummu mama .mm ES CH madmmmum wuwunm mHHHomum manommn mmmflomnm cam mam ooa\cHE\oo ca 30am cwo> mflaflomum muHOQoH mumcflvuo .3oam cwm> mflawomum co mpownmm Ammaouwov oHEmHo>ommn can Amuoo UAHOmV owEmHo>oenoc mCHHsU macapmuwuam whammmum mumuum mHHwomum mo muommmm .ma musmflm 88 ma 353m mmammmmn. >mu...m< 94.0416 emu om. om. ow. ow. cm. 0.0 mm . o . o . o. .. n. l 8 30d z_m> 9.533 l mu 89 hypovolemia significantly altered gracilis muscle blood flow. To more accurately compare the relationships between muscle blood flow and perfusion pressure during normovolemia and hypovolemia, the flow data reported in Figure 13 were normalized to percent of control (Figure 14). During normo- volemia the vasculature autoregulated its flow at perfusion pressures of 140 mm Hg and below since a 57% local reduc- tion in gracilis artery pressure from 140 to 60 mm Hg eli- cited only a 51.1 11.0% decrease in blood flow. However, when pressure was altered from 140 to 180 or 200 mm Hg (29 and 43% increases respectively), blood flow increased 45.4j;5.l and 92.3 i 9.0%. The fact that steady state flow increased proportionately more than pressure indicates that the gracilis vasculature is less able to autoregulate flow at high perfusion pressures. The ability to autoregulate flow is not completely lost at these higher pressures since flow always declined after the initial passive dila- tion associated with a given pressure elevation (Table 3). The ability to autoregulate flow in response to per- fusion pressure alterations below 140 mm Hg was evident during systemic hypotension since a 57% local reduction in perfusion pressure from 140 to 60 mm Hg elicited only a 47.3 i 1.2% decrease in steady state blood flow. In fact, hemorrhage appeared to potentiate autoregulation slightly since step reductions in gracilis artery pressure from 140 9O .mucmEHummxm m Eoum muouum mummcmum + mmDHm> cmme unommnmmu mumo .ucmEHHmmxm comm Eoum mumm on» muflamfinoc on com: mums mm 88 ova mo monsmmmum ammuum mfiaflomum um Umcfimuno mumo .mm 85 CH whammmum mumuum mflaflomum muuommu mmmflomnm mam 30Hu cfl0> mflaflomum Houucoo mo w muuommu mumcflmuo .Honucoo «o w on mmuwameuoc 30am cfim> mwawomum co mmoflumm Ammaouflov UflEmHo>om>n mam Ampom mwaomv UHEmHo> noEHoc mcflusm macaumumuam musmmmum mumuum mHHHomum mo muommmm .VH musmflm 91 CON «a magmas umawmmma >mw._.m< 94.036 om. om. cc. on. 00. xx" oo 1 I dl .0? 1 ON. OS .09 ow. 304n— Jomkzoo no $ cow 92 to 120, 100, 80, and 60 mm Hg each elicited a significantly smaller percent change in flow during hypovolemia. As dur- ing normovolemia, blood flow autoregulation during hypo- volemia was diminished at pressures above 140 mm Hg since 29 and 43% elevations of perfusion pressure from 140 to 180 and 200 mm Hg elicited 38.5 i 5.4 and 89.5 :9.5 percent increases in blood flow. These flow changes in response to increasing perfusion pressure from 140 to 180 and 200 mm Hg during hypovolemia were not significantly different from those observed in response to identical pressure manipula- tions performed during normovolemia. B. Muscle Vascular Resistance Figure 15 reports steady state gracilis muscle vascu- lar resistance as a function of gracilis artery pressure when perfusion pressure was altered from 140 mm Hg by chang- ing the perfusion pump servosystem parameters during normo- volemic (dots) and hypovolemic (circles) periods. During normovolemia, elevation of gracilis artery pressure from 140 to 180 or 200 mm Hg significantly reduced muscle vascu- 1ar resistance from the control value observed at 140 mm Hg; all other perfusion pressure manipulations failed to significantly alter vascular resistance. During hypovolemia, muscle vascular resistance was Significantly elevated from the normovolemic control values observed at each level of perfusion pressure. As was 93 .mucmEHummxm m Eon“ muonnm mummcmum.H mmsHm> cmmE ucmmmummu mumo .mm EE ca musmmmum mumuum mflHHomum manommu mmmflombm mam mam ooa\cHE\oo\mm as :a mocmumfimmn Hmasomm> maomsfi mHHHomHm muuommn mumcwmuo .mocmumflmmu umasomm> mHOmDE mwaaomum so mmoflumm Ammaouflov owEwHo>ommn mam Amuom UHHOmV UHEmHo> noauoc mafiuso macaumnmuam magmmmum mumuum mHHHomum mo muomwmm .mH musmflm 94 me masons mmnmmwmm >mwhm< m...:o cmmE ucmmmummu mumo .ucmEHummxm comm scum mumm mcu meHmEHo: on mmms mumz mm EE ova mo musmmmum mumuum mwawomum um mmcflmuno mmocmumflmmm .mm as cw mummmmum humunm mflafiomum muuommu mmmwombm mam mocmumflm 1mm Hmasomm> mHOmDE mHHfiomum Houpcoo mo unmonmm manommu mumCHUHo .Houucoo mo usmoumm ou mmufiHmEHoc mocmumwmmu umasomm> mHomSE mflaflomum co wmoflumm Ammaouflov anmHo>om>£ mam Amuom meOmv owEmHo> Iosuoc mceusm mcowumumuam musmmmum mumuum mHHHomum mo muoomwm .ma musmflm 97 .OON we magmas mmammmma >mmkm< m_n=oom>m H.mfl H.NH «.mfl h.QH h.QH m.mH v.nH vmum>umccH m.mma v.mm m.mm 0.0 H.0va n.ooa v.~oH memHo>om>m o.mfl v.&H n.QH_ w.nH m.mH pmum>umccH ooa OOH m.mH H.0H o.ova v.mva H.mo Houucou mocmumammm 30Hm mocmumamom 30am musmmmum musmmmum usmfiaummxm Houucou Houucoo “manomm> cam> >umuu< Hmwumuuc mo mafia «o w mo w odomsz mwawomuo mwaaomuw UHEmummm . mHHHomuo cmmz cmmz sown muouum mummcmum mam mosam> :mms mum mumo mafinz mmuSCAE ca Ummmmumxm ma mega .mucmsaummxm m .mm as ca mommmMQXm mum mmusmmmum .Amsm ooa\cfiEK00\mm EEV mocmumwmmu uma:omm> mam Amsm 00H\cwa\oov 30am vocab maomse mwawomum co mmmzuuoEms meansm coflum>umcom mo muommwm .w magma 100 their normovolemic control values. Vascular resistance decreased from 163.2 :_3.1 to 122.5 i.2-9% of control resistance when the muscles were acutely denervating during hypovolemia, suggesting that approximately 60% of the resistance elevation observed with arterial hemorrhage to 100 mm Hg could be attributed to neurogenically medi- ated vasoconstriction. However, the interpretation of these data is complicated by at least two factors which make it difficult to assess what portion of the hemorrhage induced vasoconstriction is neurally mediated: l) a por- tion of the decrease in resistance observed in response to acute denervation during hypovolemia could be due to a removal of the neural component of basal vascular tone that is present during normovolemia since acute denervation dur- ing normovolemia also increases muscle blood flow; 2) a portion of the decrease in resistance observed in response to acute denervation during hypovolemia could be due to increased sympathetic vasodilator fiber activity secondary to the trauma the nerve receives with section. 101 III. Series III: Naturally Perfused, Denervated Gracilis Muscles; Vascular ResponseS‘to Increased Venous and Mean Distendingv Pressures and to Local Hypotension During Pulsatile or Non— Pulsatile Perfusion A. Vascular Response to Alterations in Perfusion Mode Figure 17 reports data from a selected experiment in which the mode of perfusion was altered from non-pulsatile to pulsatile and back to non-pulsatile. Panels on the left show responses to perfusion mode alterations in the normal vasculature, those on the right depict vascular responses to similar alterations performed during papaverine infusion. In the normal vasculature, switching from non—pulsatile to pulsatile perfusion (second 0 on abscissa) elicited a rapid increase in muscle blood flow followed by a slower decline to a new steady state flow rate lower than that observed during non-pulsatile perfusion. When non—pulsatile perfu- sion was restored (second 160 on abscissa), there occurred an initial, rapid decrease in flow.followed by a gradual return to a new steady state value higher than that observed with pulsatile perfusion. The effect of papaverine infusion on the blood flow responses to changes in perfusion mode are reported in the right hand panels of Figure 17. During papaverine infusion, switching from non-pulsatile perfusion (second 0 on abscis- sa) resulted in a rapid, sustained increase in muscle blood 102 .25 mimoa 9:. Inmflm3 maomsfi mHHHomHm mmmamumm maamusumc m cufl3 unmaaummxm pmuooamm m Eoum mum mumo .mmcoomm cw mfiflu vowmmp mmmmwomnd .cHE\oo ca 30am cflm> meHomum mam “mm 85 ca whammmum humuum mHHHomum Umumnmmucfi loos mom mummmmnm mumuum mflawomum Ammumummucfi MHHmUACOHuomHmV cmmE .mHSmmmud cflm> mflaflomum vowmmp mmumcwpuo .mcwum>mmmm mo comeMCH HmooH m mcflusp mmmcommmu 305m “amen may :0 mmosu .musumasommb HmEHo: may ca mcofiumumuam once scamsmumm ou mmmcommmu uoflmmp puma on» so mamcmm .mHHummHsmlcoc ou xomn mam wawummasm on mHHummHsmlcoc Scum cowmsmnmm mo mpoE map mcwmcmco cu noncommmu “manomm> maomss mHHHomnw .na musmflm 103 mozoowm O ow. .9... . 0.. .10... a 1 an #1 Low 00 on. CON l__l—l—1 On 00. on. CON 5 g 0. ON WW 304.... z_m> 94.055 8. mmammmma 55.54 94.055 wmammmma >mwkm< 24w: 0 mmammuma z.w> 94.0410 ha musmem mozoomm 0mm O¢N CON 8. ON. q d d d d a i O. n. ON 00 on. on on _ CON 0 0. ON 30...“. 25> $354.10 wmammmmm >mmkm< m_.:omm.._.m< 94.055 24% wmzmmwmm z.w> «3.055 104 flow. Restoration of non-pulsatile perfusion (second 80 on abscissa) resulted in a rapid, sustained decrease of blood flow. Table 5 reports mean values and standard errors from 10 experiments for gracilis vascular responses to perfu- sion mode changes in normal and papverine treated muscles. In the normal vasculature, rapid elevation of gracilis artery pulse pressure from 2.6 i 0.8 to 40.9 i 5.5 mm Hg elicited a 43.2 i 3.7% increase in blood flow from 14.7 i 2.1 cc/min/lOO gms to 21.8 :_2.7 cc/min/lOO gms. This peak maximum flow was reached in 5.3 :_0.4 seconds and occurred because vascular resistance fell 22% from 11.5 i.1-8 to 8.4 :_1.7 mm Hg/cc/min/lOO gms. Following the initial, rapid elevation, muscle blood flow declined to a steady state value of 10.2 :_l.3 cc/min/lOO gms. This steady state level of flow required 78.5 i 4.3 seconds to deve10p and represented a 25.7% reduction from the flow rate ob- served during non-pulsatile perfusion. Thus, switching from non-pulsatile to pulsatile perfusion elicited an ini— 'tial, rapid fall in muscle vascular resistance followed by a slower rise in resistance to a new steady state value 30.6 i 3.2% above the level observed during non-pulsatile perfusion. During papverine infusion, the secondary phase of in- creasing resistance was not observed; switching from non- Pulsatile to pulsatile perfusion elicited only a rapid 105 Table 5. Gracilis muscle vascular responses to perfusion mode changes in normal and papaverine treated muscles. Pressures are expressed in mm Hg; flow in cc/min/lOO gms; resistance in mm Hg/cc/min/lOO gms; and time in seconds. values : standard errors from 10 experiments. Data are mean Steady Transient State Control Responses Responses Normal Vasculature Mean Perfusion Pressure 141.4 :_2.8 141.5 :_l.8 141.5 :_l.9 Pulse Pressure 2.6 :_0.8 40.9 :_5.5 40.9 :_5.4 Muscle Blood Flow 14.7 :_2.1 21.8 i 2.7 10.2 :_1.3 Muscle Vascular Resistance 11.5 i 1.8 8.4 :_1.7 15.6 :_2.0 Time to Response - 5.3 :_0.4 78.5 :_4.3 % of Control Flow 100 143.2 :_3.7 74.2 i 2.2 t of Control Resistance 100 78.0 :_l.9 130.6 : 3.2 Mean Perfusion Pressure 142.2 :_2.4 142.2 :_2.3 142.2 : 2.3 Pulse Pressure 40.4 :_5.1 2.7 i 0.8 2.7 :_0.8 Muscle Blood Flow 10.1 :_1.2 8.7 :_1.0 14.6 :_2.6 Muscle Vascular Resistance 15.7 :_2.1 17.9 i 2.6 11.6 :_l.7 Time to Response - 5.7 :_O.3 81.5 :_4.6 % of Control Flow 100 88.1 :_2.1 137.4 :_3.4 % or Control Resistance 100 113.2 i 2.4 77.8 :_1.9 Papaverine Treated Vasculature Mean Perfusion Pressure 133.9 :_2.3 - 133.8 :_2.2 Pulse Pressure 1.7 :_0.6 - 32.8 i_3.4 Muscle Blood Flow 64.3 i 7.4 - 78.7 :_8.8 Muscle Vascular Resistance 2.0 i 0.6 - 1.7 i 0.5 Time to Response - - 5.8 :_0.5 % of Control Flow 100 - 125.5 3 4.1 % of Control Resistance 100 - 83.7 i_2.1 Mean Perfusion Pressure 132.6 :_2.4 - 132.7 :_2.4 Pulse Pressure 32.4 :_3.1 - 1.8 :_0.6 Muscle Blood Flow 78.9 :_8.6 - 63.7 :_7.2 Muscle Vascular Resistance 1.7 :_O.5 - 2.0 :_0.6 Time to Response - - 6.3 i_0.3 % of Control Flow 100 - 81.2 :_1.9 % of Control Resistance 100 - 127.1 :_4.7 106 16.3% reduction in resistance from 2.0 i 0.6 mm Hg/cc/min/ 100 gms to a steady state value of 1.7 i 0.5 mm Hg/cc/min/ 100 gms. This response was very similar to the initial transient response observed prior to papaverine treatment, suggesting that the normal vasculature's transient re- sponses to switching the mode of perfusion from non- pulsatile to pulsatile probably results from a passive expansion of vascular radius due to pulse pressure disten- sion. Since papaverine removed the secondary slow phase of increasing resistance, this response probably represents an active, myogenic reduction in vascular radius triggered by the increase in pulse pressure. Restoration of non-pulsatile perfusion in the normal vasculature elicited a pattern of responses that were an approximate mirror image of those described above to switching from non-pulsatile to pulsatile perfusion. Blood flow fell rapidly and then slowly rose to a new steady state value 37.4% above the level observed during pulsatile perfusion. During papaverine infusion, switching to non- pulsatile perfusion resulted in a rapid fall in flow to a new steady state value 18.8% below the level observed with pulsatile perfusion. As was observed for the opposite perfusion mode change, papaverine removed only the slow phase of changing resistance, suggesting that the initial, rapid responses to perfusion mode changes are passive, while the more slowly developing responses represent 107 actively mediated alterations in vascular radius. B. Vascglar Responses to Increasing Mean Distending Pressure Table 6 and Figure 18 report gracilis muscle vascular responses to 20 mm Hg elevations in mean vascular distend- ing pressure performed during pulsatile and non-pulsatile perfusion in normal and papaverine treated muscles. When gracilis arterial and venous pressures were simultaneously elevated 20 mm Hg during non-pulsatile perfusion, muscle blood flow was reduced 25.3% below control as the result of a 15.6 :_1.7% elevation of vascular resistance from 12.2 i 1.6 to 14.0 :_l.8 mm Hg/cc/min/lOO gms. However, during pulsatile perfusion, increasing mean vascular distending pressure 20 mm Hg elicited a significantly greater percent reduction in muscle blood flow as the result of a 26.6 i 1.8% elevation of vascular resistance from 14.9 :_l.9 to 20.1 i 2.2 mm Hg/cc/min/lOO gms. When the vasculature was treated with papaverine, increasing mean distending pressure produced significant elevations in muscle blood flow due to an 18.2 i 1.5% reduction in resistance during pulsatile perfusion and an 18.8 :_1.6% resistance decrease during non-pulsatile perfu- sion. Therefore, elevations of mean vascular distending pressure in the face of a constant pressure head results in a passive dilation when the vascular smooth muscle cells are paralyzed with papaverine. In the normal vasculature 108 m.m H 6.8 in H 999 90 + B.~ m6 H 23 To H mam Wm H mag Ho H 54 may: 03 OS de mm.m méH 99 HoH m.m m.mH ma: NdH ~.o muzo TNH mém o.~H mama 20H 34 a.mH m6... Honam YNH 6A2 ~.mH m.~m .85 03 03 Ho H mo..~ Wm H ~13 6.0 H m.m m.~ H :3 TN H 6.3 mo QHDUMHDOWMNV UUUMQHH. 0CHHO>MQMQ RHH Hm: m.mH man méHmJA mAH Tm m.on.mm o.~Hh.mmH 80H HN me: I I l I l :2 OOH 00H m.H + H.NH n.H + H.HH m.o + m.m m.H + m.mHH «.0 + m.o m o a; H mfifi Hm H 4.8 HNH 5.3 m.oH m.m m.oH 8mm TNH 463 im H ~44 82+ 03 03 m; H 93 H; H o.m «.6 H m.m m4 H Hm: Hm H 8: mo .. . HmHuB psoomm m4 H ~63 Hm H 98 «to. H Tom m6 H m.m To H 82.. m4 H 6.03 2m H 0.3 82+ 03 03 m4 H 93 a; H Tm «.6 H m.m m; H Home in H 93 mo 54 H 8m: m.m H Hi m4 H 0.3 HA H Hm «.0 H m.m~ o.m H To: m6 H m.~ 32+ ooa 03 m; H HS 64 H 2: m.o H m5 m4 H ma: To H «.0 .726 Hague umuem musumasomm> Hmsuoz QOCMUWHmmm 30.7mm mugumfimmm 30Hh whammflum QHSmmwhm 0H§mm0Hm Houusou Houucoo umHsomm> sHm> sHm> >umuu¢ mmasm no a mo a meaeomuo meawomuo meaeomuo mwaeomuo sumuum smmz mHHHomuU lfiU mflfld—mmmhm .musmEHummxm 0H Eouu muouum oummsmum.H mosam> smmE mum mama Imam cmmE :H soHum>mHm mm as om a ma2+ mam .Houusoo oHHummHsmlsos u mu .mnsmmmum mcwmcmu U “Houusoo mHHummHsm .mEm ooa\:Hs\oo\mm as :H mosmumwmou «mam ooa\sHa\uu sH 30am no: as sH consummummu mum .moaomss mmummuu mcwuw>mmmm mam Hmsuo: cH sowmsuuom «HHummHsmnso: mam mHHummHsm msHHsp musmmmum mswmcmumwm amasomm> smma cw msOHum>me on noncommmu umHsomm> odomsa mHHHomuU .0 mafia. 109 .mquEHummxm 0H scum mnouum pummCmum.H mCmmE mum mumo .moCmumHmmH umHComm> mHomCE mHHHomum CH mmCmCo quoumm on» mHOHmmp wumCHpHo .mmHomCE Amumn pCmC uanHV Umummuu mCHum>mmmm pCm Amumn UCmC ummHv HmEHOC CH ConCmqu mHHummHCQICOC no mHHummHCm mCHuCp ousmmmum mCHmCmumHm umasomm> CmmE CH mCOHum>mHm ou mmmCommmH moCmumHmmu mHomCE mHHHomuU .ma musmem 110 we mnsmem F-i 401F200 m2_mw>+ I. I. I. I. .I I2 OOH OOH H.O + mO.N O.m + H.Hm H.O + H.m H.H + H.HHH O.O + H.H m o I. .I .I I. I. I. .I m H.O + m.NOH O.H + m.mm m.O + mO.H O.O + m.OO H.O + O.mm H.H + m.mmH H.H + m.mm >+ I I I I I m OOH OOH m.O + mm.H m.m + O.me m.O + O.m H.H + O.mmH H.H + m.mm o mHsumHComm> pwummue mCHHm>mmmm O.N.H O.OHH H.H.H O.NO O.H.H O.mH H.H.H H.m m.O.H O.mm O.N.H e.mmH 0.0.H H.H m>+ OOH OOH H.H.H m.HH H.H.H O.HH m.O.H O.m O.~.H H.OmH O.O.H H.H mIzo II II .I .II II II I m H.H + H.ONH O.H + H.Hm H.H + H.OH m.O + O.m m. O + O.mm H.H + H.OHH H.m + H.Hv >+ I I I I I m OOH OOH H.H + «.mH H.H + m.OH O. O + O.m H.H + H.OvH H.m + H.HH o HmHHB pCoomm .I .I I. .I I. I. I. m H.H + m.mmH O.H + O.Om H.H + H.ON 0.0 + m.m m.O + O.mm m.H + O.OHH m.m + O.Hv >+ I I I I I m OOH OOH O.m + O.mH m.H + ~.OH m.O + O.mm O.H + m.OHH m.m + O.Hv o I I I I I I m H.H + m.HmH H.H + H.HO O.m + H.HH H.H + m.m H.O + O.mm O.H + m.HvH m.O + H.H >+ I. I. I. I. .I mIz OOH OOH O.H + m.HH H.~ + H.HH m.O + O.m O.H + m.HvH O.O + H.H o - HmHue pmHHm mHCpmHComm> Hmeuoz mocmumHmmm onm moCmumHmmm onm wusmmmum musmmmum musmmmum I HouuCou HouuCoo HmHComm> CHm> CHo> wumuum mmHCm mo w mo w mHHHomuw mHHHomuw mHHHomHU mHHHomHO mumuHC 2.002 mHHHUMHU .mquEHHmmxm 0H Scum muouum pumpCmum.H mmsHm> CmmE mum mumo .musmmmum CHm> mumo CoHum>mHm mm 88 ON u m>+ pCm .mumc HonuCoo mHHummHsm u no .mump HOHUCOU oHHummHsm ICOC u .mEm 00H\CHE\oo\mm BE CH moCmumHmmu UCm «mam 00H\CHE\oo CH 30Hm .mm BE CH pmusmm Imummu mum mmusmmmum .mmHomsE pmummuu mCHHm>mmmm UCm HmEHOC CH Consmuwm mHHummHsmICOC mCm mHHummHCm OCHHCU whammmum msocm> mo mCoHum>mHm mm BE ON on mmmCommmu umHComm> mHomsE mHHHomHU .5 anme 113 .mpCmEHHmmxw 0H Scum muouum UHmUCmum H mCmmE mum mumo .QOCmumHmmH HmHComm> mHomCE mHHHomHm CH mmCmCo quoumm mCu mHOHmmm mumCHUHO .mmHomCE Hmumn pCmC quHHV mmummnu mCHHm>mmmm mCm Amumn pCmC ummHv HmEHOC CH ConCmHmm mHHummHleCOC Ho mHHummHCm mCHHCU mummmmum mCOCm> mo mCOHum>mHm mm as om on mmeommmH mOCmumHmmu mHomCE mHHHomH0 .OH mHsmHm .1114 OH onsmHm mzzhtéidm Jomhzoo I.| ll .202 3.532.. _ _ I-H wozflrwawm z. moz HOHUCoo mCB .mm 55 owHIom mo mmCmu musmmmum mumuum mHHHomHm mCu Hw>o mquEHHmmxm oH Comm mmCHm> HmCmH>HmCH mum mumo .musmmmnm humunm mHHHomum HouuCoo mo quoumm mqummHmmu mmmHombm «moCm IumHmmH HmHComm> wHomCE mHHHomHm HonuCoo mo quouwm mqummHmmH mumCHpuo .moCmumHmmu CmHsomm> mHomCE mHHHomHm Co COHmCmumm AmmHoHHov mHHummHCm ICOC mCm Hmuoov mHHummHCm mCHHCp ConCmuommn HmOOH .mmpmum mo muomwmm .HN musmHm 1.21 00_ Hm musmHm wmammmma 5.5.52 m_I:o<¢@ Jomkzoo to .x. om om Ob Co on d. 1 o I 00. D 00 o o I ON. 0 wozS—hawm . on. Jomhzoo no $ 122 pressure. However, the slope for the resistance data ob- tained during non-pulsatile perfusion was -0.381 i 0.089; a value significantly different from that observed during pulsatile perfusion. Thus, blood flow autoregulation was significantly attenuated during non—pulsatile perfusion. Table 8 reports the vascular responses to 20 mm Hg reductions in gracilis artery pressure during non-pulsatile perfusion in normal and papaverine treated muscles. When gracilis artery pressure was reduced 14.2% below the con- trol value of 139.7 1 2.0 to 119.9 1 1.9 mm Hg in the normal vasculature, gracilis muscle blood flow fell 21.7% below the control level of 14.9 i 2.2 to 11.1 i 1.7 cc/min/ 100 gms due to the fall in perfusion pressure and to a 9.3 i 3.9% increase in muscle vascular resistance. However, during papaverine infusion, a 14.8% reduction in gracilis artery pressure from 133.3 :_2.7 to 113.6 : 2.3 mm Hg elicited a significantly greater percent reduction in flow as a result of a 23.7 i 4.3% increase in gracilis muscle vascular resistance. These data indicate that although muscle blood flow autoregulation was attenuated with non- pulsatile perfusion, the vasculature was still capable of some degree of flow autoregulation since the resistance increase observed in re3ponse to gracilis artery pressure reduction was greater in the papaverine treated versus the normal vasculature. 123 l i l m.v.H n.MmH m.m.H m.mo m.o + mm.m m.v H m.mv 5.0 + m.m m.m + o.MHH ~.o H «.0 m+ I I .I I I mIz 00H 00H v.0 + mo.~ m.m + H.vm 5.0 + m.m h.N + m.MMH 0.0 + n.H U mHCumHsomm> mmumwua mCHHm>mmmm I. I. II II II I. I. 0 m.m + m.m0H v.m + m.mn m.H + H.NH n.H + H.HH m.o + m.m m.H + m.mHH m.o + m.o m+ I I I I I I2 OOH 00H m.H + m.HH m.m + m.vH 0.0 + m.m o.m + h.mMH 0.0 + n.m m U mHCumHComm> Hmsuoz moCmumHmmm BOHm moCmumHmmm onm musmmmum musmmmum musmmmum Houucou HouuCou HmHComm> CHm> CHm> mumunfl mmHsm mo w Ho w mHHHomHo mHHHomHo mHHHomuw mHHHomHU humuud Cmmz mHHHomHo .mquEHummxm 0H Eon muouum pummCmum.H mmsHm> CmmE mum mumo .mHCMmmum aumuum mHHHomum mo COHuoCmmu mm 65 on u m+ pCm «mump HOHuCoo mHHummHCm ICOC u m U .msu 00H\CHE\00\mm BE CH moCmumHmmH pCm «mam OOH\CHE\oo CH onu «mm ES CH pmqummHmou mum mmusmmmum .mmHomCE pmummuu mCHum>mmmm UCm HmEHOC CH COHmCmumm mHHummHCmICOC mCHHCm musmmmnm humuum mHHHomum mo mCOHuosmmu mm BE ON on mmmCommmu HmHComm> mHowCE mHHHomuo .m mHnt DISCUSSION 1. Series I and II: Naturally Perfused and Pump Perfused, Innervated Gracilis Muscles; V Gracilis Artery Pressure Alterations During Normovolemic and Hypovolemic Periods A. Steady State Autoregulatory Responses to Local Alterations in Perfusion Pressure For naturally perfused muscles, the autoregulatory adjustments in vascular tone accompanying step reductions in gracilis artery pressure were not significantly altered by hypovolemia (Figure 9). However, when gracilis artery pressure was lowered from or increased to 140 mm Hg in pump perfused muscles, a small improvement of blood flow auto- regulation was detected when vascular tone was elevated by hemorrhage (i.e., a given percent change in pressure produced a smaller percent change in flow during hypovolemia) (Table 3; Figures 14, 16). Bond and Green (13, 43) also reported improved autoregulatory adjustment of canine hind- limb blood flow in response to local perfusion pressure alterations performed during hypovolemia. Both metabolic and myogenic mechanisms may contribute to the maintenance or improvement of blood flow autoregula- tion when vascular tone is elevated by hemorrhage. 124 125 The reduced blood flow levels accompanying hemorrhage could lower the flow to metabolism ratio and increase the tissue concentration of vasodilator metabolites thereby enhancing blood flow autoregulation via metabolic mechanisms. Some support for this view is provided by the studies of Stainsby (115) in which blood flow autoregulation was found to be more effective when the flow to metabolism ratio was lowered by skeletal muscle exercise. Furthermore, Jones and Berne (70) report that muscle blood flow autoregulation is more pronounced in preparations displaying low venous blood oxygen saturation. It is also possible that myogenic mechan- isms, augmented by a reflex increase in vascular smooth muscle tension, account for the improved autoregulation seen during hemorrhage. It has been shown for example that isolated arterial strips exhibit increased myogenic re- sponsiveness to passive stretch when resting tension is high (112, 113). B. Transient Vascular Responses to Altera- tions in Perfusion Pressure In the pump perfused muscles, each elevation of arteri- al pressure elicited a rapid, disprOportionate increase in flow which was followed by a slower decline of flow toward control levels (Table 3; Figures 11, 12). Other workers have suggested that this initial, brief period of declining resistance is due to a passive vascular distention (43, 116, 117). Since the vascular wall should be less compliant 126 during elevated levels of vasomotor tone accompanying hemor- rhage, it might be expected that the peak transient flow would be proportionately less during hemorrhage if this initial period of rising flow is due to purely passive vascular behavior. However, just the opposite occurred: when perfusion pressure was elevated to 140 mm Hg, the ini- tial peak flows were proportionately greater during hypo- volemia than during normovolemia (Table 3). Assuming that the vasculature is less compliant during hemorrhage, these results suggest that the initial period of declining re— sistance observed in response to perfusion pressure eleva- tion is not due solely to a passive distension of the resistance vessels. The metabolic hypothesis provides one explanation for the relatively greater increase in peak transient flow observed during hemorrhage. Since the initial level of tissue vasodilator metabolites should be higher at the low flow rates accompanying hemorrhage, elevation of perfusion pressure could via metabolic mechanisms, elicit a propor- tionately greater initial decrease in resistance during hemorrhage. Some support for this interpretation is obtained from an analysis of Jones' and Bernes' (73) data, where perfusion pressure elevations in muscles having low flows and low venous oxygen content elicited initial vasodilations which were relatively greater than those 127 observed in preparations having higher resting flows and venous oxygen contents. When perfusion pressure was lowered during normOF volemia, muscle blood flow at first rapidly decreased to minimum values and then slowly rose toward control levels (Table 3; Figures 11, 12). When vascular tone was elevated during hypovolemia, perfusion pressure reductions elicited proportionately smaller reductions in initial transient flows (Table 3; Figure 12). For example, a 57% reduction in gracilis artery pressure from 140 to 80 mm Hg elicited a transient 75% reduction in flow during normovolemia, whereas flow fell only 59% below control when local pressure was similarly reduced in hypovolemia. Other investigators have proposed that this initial rise in resistance accom- panying perfusion pressure reduction is due to passive vascular collapse (116, 117). If it is assumed that the vasculature is less distensible when vascular smooth muscle tone is elevated by hemorrhage, the observation that per- fusion pressure reductions elicit proportionately smaller initial reductions in flow during hemorrhage supports this hypothesis. When perfusion pressure was altered to or from 140 mm Hg, more time was required to reach the minimum and maxi- mum transient flow values when vascular tone was elevated by hemorrhage. These data possibly reflect a decreased 128 distensibility of the vascular smooth muscle cells when active tension is elevated by hemorrhage. During hemorrhage, more time was also required to reach the steady state flows after gracilis artery pressure alter— ations (Table 3), indicating that the autoregulatory responses to step changes in perfusion pressure develop more slowly during systemic arterial hypotension. This central, vasoconstrictor influence on blood flow autoregula- tion could be mediated by several different mechanisms. Folkow (35) has proposed that the smooth muscle of resist- ance vessels is arranged in two functionally different sheaths; one inner, myogenically active layer surrounded by an outer sheath where the smooth muscle cells essentially lack myogenic activity but are subordinated to adrenergic vasoconstrictor fiber control. According to this hypothe- sis, the myogenically active inner cells would become unloaded by contraction of an outer, well innervated layer during hemorrhage, possibly leading to a prolonged develop- ment of autoregulatory adjustments in vascular caliber during hemorrhage. Of course, this hypothesis could also be extended to encompass delayed local metabolic influences on the inner smooth muscle cells. However, it is not necessary to propose a differential neuroeffector organization within the vascular wall to explain the prolonged develOpment of autoregulatory adjust- ments in vascular tone during hemorrhage. Vasodilator 129 metabolites could exert their effects more slowly on the same population of smooth muscle cells which are driven by neurally released or blood borne vasoconstrictors during hemorrhage. Some support for this view is provided from the studies of Chalmers gt_§1. (20) who observed that the rate of development of gracilis exercise hyperemia is progressively slower at increasing levels of vasomotor tone elicited by systemic hypoxemia. Alternatively, during hemorrhage the myogenically active cells may be driven to such a large extent by neurally released or blood-borne vasoconstrictors that they are unable to respond quickly to changes in transmural pressure. When gracilis artery pressure was altered from or increased to 140 mm Hg, the areas under or above the trans- ient flow peaks were the same in normovolemic versus hypo- volemic periods (Table 3). The fact that these areas were the same in normovolemic and hypovolemic periods provide further indication that the overall autoregulatory response is not appreciably altered by hemorrhage induced elevations in vascular tone. II. Series III: Naturally Perfused, Denervated Gracilis Muscles; Vascular Responses During Pulsatile and Non-pulsatile Perfusion A. Vascular Responses to Alterations in Perfusion Mode Switching the mode of perfusion from non-pulsatile to pulsatile elicited a rapid increase in blood flow followed 130 by a slower decline to a new steady state flow rate lower than that observed during non-pulsatile perfusion (Figure 17; Table 5). Conversely, restoration of non-pulsatile perfusion elicited an initial, rapid decrease in flow followed by a gradual return to a new steady state level higher than that observed with pulsatile perfusion (Figure 17; Table 5). The initial, rapid changes in resistance seen with perfusion mode alterations are attributed to the addition or removal of pulse pressure induced vascular dis- tension since qualitatively similar passive responses were observed when perfusion mode alterations were performed in vascular networks poisoned with papaverine (Figure 17, Table 5). The secondary phases of increasing resistance observed after switching to pulsatile perfusion or decreas- ing resistance after switching to non-pulsatile perfusion probably represent active responses since papaverine abol- ished these changes in vascular resistance (Figure 17; Table 5). When pulse pressure was increased from 2.6 to 41 mm Hg in these experiments, steady state gracilis vascular resistance increased 31% above control levels (Table 5). Rovick and Robertson (99) reported quantitatively similar increases in dog tongue vascular resistance in response to similar increases in pulse pressure. These investigators observed a progressive increase in muscle vascular resist- ance in response to step elevations in pulse pressure, 131 with a maximum 37% increase in resistance occurring when pulse pressure was increased to 40 mm Hg. With further elevations in pulse pressure to 64 mm Hg, tongue vascular resistance was observed to return to control levels. However, when Mellander and Arvidsson (85) elevated arterial pulse pressure from 8 to 56 mm Hg in sympathectomized cat hindlimbs, they observed only a 6% increase in muscle vascular resistance. It is possible that these relatively small resistance responses observed by Mellander and Arvidsson are due to species and/or preparation differences. However, an analysis of Rovick and Robertson's data suggests that only small increases in resistance would be expected with pulse pressure elevation from 8 to 56 mm Hg, since passive vascular distension apparently overrides the active vasoconstriction when pulse pressure is increased beyond 40 mm Hg. The increased vascular tone accompanying pulse pressure distension could be either metabolically or myogenically mediated. When perfusion is altered from non-pulsatile to pulsatile, it is conceivable that flow could be redistrib- uted within the muscle microvasculature in such a manner as to produce transvascular exchange conditions that would result in a lower tissue concentration of vasodilator metabo- lites leading to an increased total muscle vascular resist— ance via metabolic mechanisms. However, because the venous-arteriolar response is more pronounced during 132 pulsatile versus non-pulsatile perfusion (Figure 19; Table 7), it is more probable that the increased resistance seen with pulsatile perfusion represents an active myogenic reduction in vascular radius triggered by the increase in pulse pressure. Transmural pressure changes associated with pulse pres- sure distension could elicit increased myogenic activity by at least two mechanisms. The increased vascular resistance seen with pulsatile perfusion could result from an increased myogenic activity elicited by the higher peak systolic dis- tending pressures. Alternatively, myogenically active pace- maker cells could also be sensitive to the dynamic stretch stimulus afforded by repetitive pulsatile distensions. Both mechanisms seem plausible since active tension development and spike frequency in isolated artery and vein strips is directly related to the rate and increment of stretch as well as to resting tension (61, 112, 113). B. Vascular Responses to Increased Venous Pressure A 20 mm Hg increase in vein pressure elicited a 21.5% increase in gracilis vascular resistance during non-pulsatile perfusion and a 29.3% increase in resistance during pulsatile perfusion (Table 7; Figure 19). These responses were almost completely abolished when the vasculature was poisoned with papaverine (Table 7; Figure 19). Since the muscles were acutely denervated, these venous-arteriolar responses do not 133 appear to result from neural reflexes. Because the meta- bolic hypothesis predicts a decreased resistance with venous pressure elevation and because the venous—arteriolar response should persist during papaverine administration if tissue pressure effects mediate the response, these data indicate that myogenic mechanisms of local blood flow con— trol are present within the gracilis muscle vasculature. While it is possible that local neural networks mediate these venous-arteriolar responses, this seems unlikely since the response is present in the hindlimbs of reserpinized cats (36). Also, Johnson (62) has demonstrated that pro- caine administration does not effect the venous—arteriolar response in the intestine. The venous-arteriolar response has not been consistent— ly observed in skeletal muscle vascular beds. Jones and Berne (73, 74) and Hanson (49) observed small venous- arteriolar responses in only a few of their isolated canine hindlimb preparations. Since in these experiments, the muscles being studied were sectioned, it is possible that the preparations were less myogenically active. When venous pressure was elevated in muscle preparations that required little surgical disruption (canine gracilis and cat hind— quarters), Nagle et_al. (88) and Folkow and Oberg (36) con— sistently observed large venous—arteriolar responses. The presence and magnitude of the venous—arteriolar response also appears to depend on the resting blood flow 134 level. In experiments reported here, gracilis muscle blood flow averaged 10 cc/min/100 gms and all preparations dis- played prominent venous-arteriolar responses. The gracilis muscles used by Nagle et 31. (88) and the cat hindlimbs studied by Folkow and Oberg (36) displayed similar resting blood flows per unit tissue weight and quantitatively simi— lar venous-arteriolar responses. Resting blood flow averaged only 3.4 cc/min/lOO gms in the canine hindlimb prep- arations used by Jones and Berne (73, 74), and these prep- arations usually did not exhibit venous-arteriolar responses. In the few preparations which displayed relatively high control blood flows and venous oxygen content, Jones and Berne did observe an appreciable increase in vascular resistance with elevated venous pressure. These results suggest that at low resting blood flow rates, increased tissue concentrations of vasodilator metabolites may over- ride myogenic mechanisms so that the venous-arteriolar response is abolished. In the experiments reported in this thesis, venous pressure elevation produced larger elevations in gracilis vascular resistance during pulsatile compared to non- pulsatile perfusion (Table 7, Figure 19). This potentia- tion of the venous-arteriolar response by arterial pulse pressure distension implies that the gain of the myogenical- ly active smooth muscle cells is increased with repetitive 135 pulse pressure distension. Pulsatile changes in transmural pressure could elicit increased myogenic activity in response to elevation of venous pressure by at least two mechanisms. Myogenic pacemaker cells could be more active at the higher peak systolic distending pressures since it is known that isolated artery strips develop more active tension in response to a given increment of passive stretch when resting tension is high (112, 113). Alternately, myo— genic pacemaker cells could become more sensitive to a given increment of stretch when they are exposed to the dynamic changes in length associated with repetitive pulse pressure distension, since active tension development and spike fre- quency in isolated artery and vein strips are increased with elevated rates of stretch (61, 112, 113). C. Vascular Responses to Increasing Mean Distending Pressure When mean vascular distending pressure was elevated by increasing mean gracilis arterial and venous pressure 20 mm Hg, muscle vascular resistance increased 15.6% during non-pulsatile and 26.6% during pulsatile perfusion (Table 6, Figure 18). When the vasculature was treated with papa- verine, this same increase in mean distending pressure elicited an 18.2% reduction in resistance during pulsatile perfusion and an 18.8% resistance decrease during non- pulsatile perfusion (Table 6, Figure 18). Mellander and Arvidsson (85) have obtained similar results in the 136 sympathectomized lower leg muscles of the cat. When these investigators increased mean distending pressure 20 mm Hg in the normal vasculature, they observed a 7% increase in vascular resistance during non-pulsatile and a 14% increase during pulsatile perfusion, whereas the same elevation in transmural pressure in the papaverine treated vasculature decreased resistance by 20%. These data indicate that the distending effect of in- creased transmural pressure observed in the passive muscle vasculature is abolished and replaced by an active, presum- ably myogenic, constrictor response in the normal vascular bed. Further, since the constriction is more pronounced with pulsatile perfusion, it would appear that myogenically active smooth muscle cells are also sensitive to the trans- musal pressure changes accompanying pulse pressure disten- sion. These data do not conclusively demonstrate a vascu- lar sensitivity to pulse pressure distensiOn because, in the same experiments, simultaneous elevations of mean arterial and venous pressures are accompanied by greater increases in peak systolic pressures during pulsatile com- pared to non-pulsatile perfusion (Table 6). It could be argued that the greater constriction observed in response to elevation of mean distending pressure during pulsatile perfusion is instead related to the larger increase in systolic pressure during pulsatile compared to non-pulsatile perfusion. However, this seems unlikely since, when venous 137 pressure alone is elevated, the venous-arteriolar response is also more pronounced during pulsatile perfusion (Table 7, Figure 19). Thus, it appears that transmural pressure changes associated with pulse pressure distension potentiate the myogenically mediated vasoconstriction observed in response to elevations of mean distending pressure. Since it has been shown that isolated artery strips develop more active "tension to a given increment of stretch when resting tension is high (112, 113), myogenically active pacemaker cells could be operating more efficiently at the higher systolic distending pressures thereby eliciting more pronounced con- striction in response to mean distending pressure elevation during pulsatile perfusion. Because isolated vessels do show more active tension development and elevated spike frequency at increasing rates of stretch (61, 112, 113), these pacemaker cells could also be sensitive to the rate of transmural pressure change and operate at higher gain during pulsatile perfusion. D. Vascular Responses to Graded, Local HypotenSion When mean gracilis artery pressure was progressively lowered from 140 to 60 mm Hg during pulsatile perfusion, both pulse pressure and muscle vascular resistance decreased significantly (Figures 20, 21). However, during non-pulsa- tile perfusion, the same maneuver elicited a progressive 138 rise in muscle vascular resistance (Figures 20, 21). Even though resistance increased in response to graded pressure reductions during non-pulsatile perfusion, some degree of blood flow autoregulation remained because the papaverine treated, passive vasculature displayed significantly larger increases in resistance when perfusion pressure was reduced from 140 to 120 mm Hg (Table 8). Vascular resistance was observed to decrease only when both mean arterial and pulse pressure were reduced, indi— cating that muscle blood flow autoregulation is mediated to a large extent by alterations in pulse pressure rather than simply by changes in mean transmural pressure. The fact that the venous-arteriolar response and the constriction in response to mean distending pressure elevation are both larger during pulsatile perfusion suggests that alterations in pulse pressure mediate blood flow autoregulation by myo- genic rather than metabolic mechanisms. Myogenically active pacemaker cells could elicit autoregulatory decreases in resistance when pulse pressure is reduced by relaxing in response to lower peak systolic pressures. This interpreta- tion is consistent with the reports of other investigators that isolated artery strips develop less active tension to a given increment of stretch when resting tension is low (112, 113). SUMMARY AND CONCLUS IONS 1. A given percent change in gracilis artery pressure elicited either similarly less proportionate (naturally perfused muscles) or slightly smaller (pump perfused muscles) percent changes in flow during hypovolemia, indicating that steady state blood flow autoregulation was either maintained or slightly improved when vascular tone was elevated by hemorrhage. Both metabolic and myogenic mechanisms may have contributed to the maintenance or improvement of steady state autoregulatory adjustments in vascular tone. 2. The rate of development of autoregulatory responses to perfusion pressure alterations were prolonged during hemorrhage induced elevation of vascular tone due to some interaction or competition between local metabolic and/or myogenic mechanisms and remote vasoconstrictor influences. 3. Altering the mode of perfusion from non-pulsatile to pulsatile or vice versa elicited initial passive changes in muscle resistance due to the addition or removal of pulse pressure induced vascular distension. These brief, passive alterations in resistance were followed by the addition or removal of pulse pressure induced myogenic vascular tone. 139 140 4. Increased venous or mean vascular distending pres— sures produced more vasoconstriction during pulsatile versus non-pulsatile perfusion, indicating that myogenic mechanisms are present within the muscle vasculature and are sensitive to pulse pressure induced changes in trans- mural pressure. 5. When mean gracilis artery pressure was progressively lowered from 140 to 60 mm Hg during pulsatile perfusion both pulse pressure and muscle vascular resistance decreased, whereas during non—pulsatile perfusion the same maneuver elicited a progressive rise in vascular resistance. These data indicate that muscle blood flow autoregulation is mediated to a large extent by alterations in pulse pressure induced myogenic activity rather than simply by changes in mean transmural pressure. APPENDICES APPENDIX A PRESSURE, FLOW, AND RESISTANCE DATA 141 142 Table A-1. Effects of local hypotension during normovolemic and hypo- volemic periods on mean systemic arterial and gracilis artery and vein pressures (mm Hg), gracilis vein flow (cc/min/lOO gms), and vascular resistance (mm Hg/cc/min/lOO gms). Values are means : standard errors from 10 experi- ments. Systemic Gracilis Gracilis Gracilis Muscle Arterial Artery Vein Vein Vascular Pressure Pressure Pressure Flow Resistance Normovolemia Control 137:4.3 130:4.2 4.8:l.6 12.6:2.0 11.8:2.0 Clamp 137:4.0 112:3.6 3.4:1.4 9.9:}.4 12.8:l.9 136:3.9 98:3.6 2.7:l.3 8.8:}.2 12.1:l.8 137:4.1 86:3.4 2.4:}.2 8.3:}.2 11.5:1.6 137:4.2 72:9.6 2.131 3 7.7:}.1 10.4:l.6 Control 137:4.2 132:4.3 4.8:l.5 12.7:2.0 ll.6:l.5 Hypovolemia Control 118:3.9 115:4.1 l.8:0.8 6.8:0.8 17.8:1.9 Clamp 118:4.3 103:3.6 l.6:0.8 6.1:p.7 l7.8:1.9 119:4.3 91:4.0 1.4:0.7 5.5:0 6 l7.1:l.8 11814.3 78:3.0 1.3:0.6 5.3:p.5 15.2:l.8 118:4.2 66:3.0 l.l:0.6 4.919 5 l3.9:1.7 Control 119:4.3 115:4.4 1.9:0.8 7.Q:0.8 17.4:2.2 143 Table A-2. Effects of local hypotension during normovolemic and two hypovolemic periods on mean systemic arterial and gracilis artery and vein pressures (mm Hg), gracilis vein flow (cc/ min/100 gms), and vascular resistance (mm Hg/cc/min/lOO gms). Values are means 1 standard errors from 4 experiments. Systemic Gracilis Gracilis Gracilis Muscle Arterial Artery vein Vein Vascular Pressure Pressure Pressure Flow Resistance Normovolemia Control 14215.3 13815.0 4.511.5 ll.511.7 12.313.0 Clamp 14114.0 11712.6 3.811.4 10.011.9 13.114.3 14212.9 10115.1 3.011.4 9.211.7 12.113.9 14012.5 9113.0 2.811.3 8.811.6 11.413.6 14113.6 7412.5 2.411.3 7.711.4 10.312.9 Control 14014.3 13715.0 4.411.5 ll.311.7 12.313.0 Mild Hypovolemia Control 12214.7 12015.6 2.811.2 7.911.4 15.914.0 Clamp 12115.1 10814.9 2.111.1 6.911.l l6.514.4 12215.5 9417.2 1.810.9 6.511 0 15.013.9 12115.1 7913.4 l.710.9 6.010 9 13.113.0 12214.9 6512.4 1.610 8 5.71p.9 ll.312.3 Control 12314.9 12015.5 2.91l.2 8.011.3 15.714.0 Moderate Hypovolemia Control 10414.1 10314.8 l.410.9 5.110.6 20.813.9 Clamp 10414.2 9113.9 l.310.9 4.610.? 20.513.9 10314.4 8112.9 1.210 8 4.41p.6 l9.213.8 10314.3 6912.5 1.210.8 4.31p.5 16.813.6 10314.3 5912.5 l.010.7 3.919 5 16.113.8 Control 10414.4 10214.5 1.411.0 5.31p.7 20.213.9 144 Table A—3. Effects of local gracilis artery pressure alterations during normovolemic and hypovolemic periods on gracilis muscle vas- cular resistance. Pressures are expressed in mm Hg; resist- ance in mm Hg/cc/min/lOO gms. Values are means + standard errors from 8 experiments. '— Systemic Gracilis Gracilis % of Arterial Artery Vascular Control Pressure Pressure Resistance Resistance Normovolemic Control 14316.0 14010.7 15.912.6 100 Local pressure change 14316.1 20111.3 11.711.8 74.813.5 14316.2 18010.9 14.112.2 88.413.5 14316.1 12110.5 16.513.1 100.412.4 14316.2 10110.7 15.813.l 95.712.5 14316.8 8010.4 15.012.9 91.212.9 14316.2 6010.9 14.712.6 90.812.l Hypovolemic Control 10012.1 14010.7 25.813.4 100 Local pressure change 10012.1 20111.4 l9.713.3 76.214.1 10012.0 18010.9 23.213.5 89.913.8 10012.2 12110.5 24.110.4 93.911.7 10012.1 10110.8 23.613.5 89.411.7 10012.9 8010.5 22.112.4 84.713.5 10012.1 6010.8 19.613.0 83.212.2 145 figmmam immmév némoafl mamas. m.omm.m 0.0mmm ~.omm.o v.HHm.nm n.flHm.mw m.#Hm.ma m.£Hm.m m.@Hm.m o.fiHmn m.@Hn.m o.flHo.mm o.flHn.mn m.flH®.ma m.oHa.m m.DHm.m m.MHooa o.flHm.n m.a+o.oo m.a+n.mm m.a+m.va H.H+H.m m.o+m.m m.o+mma v.m+m.ma msmao 03 03 o .mflofin m .HHN .3 ~1on .m w 4H2: m .mHm .3 H9528 coflmsmuom mawummaom H.mmo.m3 fimmmam figmeaa odWé m.omm.m mammm o.omo.o m.flHm.vHH o.flHo.mv n.flHm.~H o.HHm.o m.on.m h.chn O.DHo.o m.flHm.moa H.flHm.vm n.flHv.mH v.flHm.m v.o+m.m v.9Hooa N.DHm.o m.m+m.moa v.m+m.mn o.H+m.NH m.a+m.aa m.o+m.m m.o+HmH m.o+a.a memao 03 OS fianm .2 fimnh .3 m .on.m mJHNE m .oHn .m H828 coflmsmuom maflummasmncoz mocmumflmmm 30Hm oocmumfimmm 30Hm whammmum ousmwmum whammmum Houucoo Houucoo uma:omm> cam> cam> mumuum mmasm mo w mo w mHHHomuo mflafiomuo mflafiomuu mflaflomuo mumuum 2mm: mHHHomuo .mucmefinmmxm 0H Bonn muouum oumwcmum.H mcmwfi mum mosam> .mEm ooa\cfl8\oo\mm ES ca mocmumwmmu cam «mam ooa\cHE\oo ca 30am «mm 85 ca omucmmoummu mum monommmum .coflmswuom oHflUMmHDQ Icon cam maflummasm mcfluso coamcouomxs Hmooa .Umomum on momcommwu HmHsomm> oaom55 mflawomuw .vud magma APPENDIX B STATISTICAL METHODS 146 APPENDIX B STATISTICAL METHODS I. Series I Gracilis vein flow and muscle vascular resistance were determined for control periods and during four experimental periods of local hypotension obtained during normovolemia or hypovolemia. For each period individual means (ii) were calculated for each parameter from two values obtained 2 and 4 minutes after gracilis artery pressure stabilized. The individual means were used to calculate a grand mean (i), variance (82), and standard error of the mean (SE?) for each period during normovolemia or hypovolemia as follows: E. x = z —1— i=1 n —2 (2x.) 222 _ 1=11 ._ i n S2 = 1-1 n - 1 SEE =7 82 /‘/ n A. Comparison of Control Means with Four Experimental Means Produced by Local Hypotension During Normovolemia or Hypovolemia An analysis of variance was performed to determine if the population means for each parameter at the five different 147 148 perfusion pressures were identical. Prior to the analysis of variance, the sample mean variances among periods (82) were determined to be homogeneous (a prerequisite for analy- sis of variance) by using the F max test. In this procedure 2 the maximum variance ratio (8 max/ 82 .n) is computed mi from the greatest (S2 ) and the smallest (S2 . ) sample max min variances. This test statistic is compared with critical values (Fmax’ 0.05, k, U) from a tabled cumulative probabil- ity distribution of maximum F for 2 sided comparisons based on k experimental periods and U degrees of freedom (n - 1). In all vases the test statistic was smaller than the tabled 2 . 2 _ F max value, so the null hypotheSis (S m — S min) was ax accepted, indicating that the sample mean variances among periods were homogeneous. An analysis of variance partitions the total variation of items into two distinct sources of variation: 1) that due to variation among groups (treatment effects); 2) that variation within groups (inter-dog variation). The varia- tion among groups (mean square error among groups, MMamong) is expressed as the sum of squares among groups divided by the n - 1 degrees of freedom (SSamong/n ~ 1). The sum of squares among groups is computed from the following expres- sion: a n a n 2 Z ( Z Y ) - -—- ( Z Z Yi) _ 1 SS ‘ H i=1 i=1 among ‘Mhere a is the number of treatments, n the number of 149 experiments, and Yi are the individual parameter values. The mean square error among groups (MSamong) is based on the variance of group means, which describes the dispersion of the group means around the grand mean. The variation within groups (mean square error within groups, MS ) is ex- within pressed as the sum of squares within groups divided by a(n - 1) degrees of freedom (SS /a(n - 1). The sum of within squares within is computed from the following expression: 2 a n 1 a n 2 SS . . = - - Z ( Z Y.) + Z Z Y where: Within n = '=1 1 i=1 i=1 a is the number of treatments, n is the number of experi- ments, and Yi are the individual parameter values. The mean square error within groups (MS ) gives the average dis- within persion of the n items in each group around the group means. The test statistic for the analysis of variance is: MS /MS among within' The test statistic was compared With critical values (F, 0.05, Ul’ 02) from an F distribution table for 01 degrees of freedom of numerator mean square and 02 degrees of freedom of denominator mean square. If the test statistic exceeded the critical value, the null hypo- thesis of no added effect of treatments was rejected and the alternative hypothesis of a significant effect of local hypotension on vascular resistance or muscle blood flow was accepted. 150 If the analysis of variance demonstrated that local hypotension produced a significant change in a parameter value within the range of treatments, it was of interest to determine which treatments were significantly different from each other. If c independent comparisons among means are made, the probability of finding at least one signifi- cant comparison by chance is l - (1 - 0:)C (where m = error rate for each comparison) (78). Therefore, as the number of comparisons increases, the probability of finding at least one spuriously significant result also increases. The Student-Newman-Kuels procedure is a method in which comparisons are made in a stepwise fashion so that the probability of making a type I error (a) for the collection of comparisons can be set at a desired level. The Student- Newman-Kuels procedure was used in the present study to determine what levels of local hypotension produced signifi— cant changes in parameter values. The procedure requires the calculation of the standard error of a group (5;) using the error mean square (MS ) from the analysis of vari- within ance as a pooled estimate of the variance among items within a group: MS . . S—-= // Within y a(n - 1) The least significant ranges (LSR) for 2, 3, 4, and 5 means were then calculated using a table of studentized ranges (Q) at a significance level of 0.05 for groups 5, 4, 3, and 151 2 means with a(n - 1) degrees of freedom: LSR (for k groups) = [Q, 0.05, k, a(n - 1)] x S; The means were then arranged in order of magnitude and the ) total range from the largest to the smallest mean (yg - §i was compared with the LSR for k = 5. If this range was significant, the range from the smallest to the next to largest mean (y; - §i) was compared with the LSR for k = 4. The testing was continued in a similar manner until a range was encountered that was not significant and the testing was then discontinued. If the range enclosed by any two means was greater than the critical LSR the null hypothesis (u = ub) was rejected and the alternative hypothesis a (pa # ub) was accepted. B. Comparison of Gracilis Muscle Vascular Responses to Local Hypotension Produced During Normovolemia with Responses Obtained During Hypovolemia Gracilis artery pressures, muscle blood flows and vascular resistances obtained at the animals' prevailing systemic arterial pressure either during normovolemia or hypovolemia were used as control values to normalize the data obtained in response to graded local hypotension to percent of control. To determine whether hemorrhage in- fluenced gracilis muscle vascular responses to local hypo- tension, separate least square linear regressions were performed on the normovolemic and hypovolemic percent of 152 control flow and resistance data, as functions of percent of control gracilis artery pressure. Only data obtained over the autoregulatory range of perfusion pressures (70-120 mm Hg) were used for the regressions and the analysis was per- formed on the Control Data Corporation 6500 Digital Computer using the MSU STAT LS program. The slopes of the regression equations for flow and resistance obtained during normo- volemia or hypovolemia were compared by an analysis of co- variance. In an analysis of covariance the features of regression and analysis variance are combined to provide a test statistic. Briefly, the procedure requires that a regression of pooled normovolemic and hypovolemic data be performed and the slope of this pooled regression compared to the normovolemic or hypovolemic regression line slope using the F statistic. If the calculated F value exceeds the critical F value (F, 0.05, u 02) from an F distribu- 1! tion table, the null hypotheSis (Slopenormovolemic = SloPehypovolemic) was rejected and the slopes were conSid- ered to be significantly different. II. Series II and III The statistical methods described for Series I (F max test, analysis of variance, Student-Newman-Kuels procedure) were used to determine whether alterations in gracilis artery perfusion pressure during either normovolemia, 153 hypovolemia (Series II), pulsatile or non-pulsatile perfu- sion (Series III) produced significant changes in gracilis muscle blood flow and vascular resistance. As described for Series I experiments, separate least square linear regressions were performed on the percent of control flow and resistance data obtained in response to graded local hypotension as functions of percent of control gracilis artery pressure during either normovolemia, hypovolemia, pulsatile, or non-pulsatile perfusion. Only data obtained below 140 mm Hg gracilis artery pressure were used for the regressions and as described for Series I, analysis of co- variance was used to determine whether hypovolemia or non- pulsatile perfusion significantly altered gracilis muscle vascular responses to local hypotension compared to their respective controls. A standard paired difference test was used to deter- mine whether blood flows and vascular resistances during local hypotension were significantly different in normo- volemic vs. hypovolemic periods (Series II) and in pulsatile vs. non-pulsatile perfusion (Series III) at corresponding gracilis artery pressures. The test statistic (ts) for each comparison was: where: 154 d = mean difference between control and experimentsl values (i.e., mean pulsatile control resistance minus mean non—pulsatile resistance at gracilis artery pressure = 100 mm Hg). Sd = standard deviation of the difference between the control and experimental mean. n = number of observations. The test statistic was compared with critical values (t0.05, 0) obtained from a Students t distribution table. If tS exceeded t0.05, u, the null hypothesis (ua-= 0) was rejected and the alternative hypothesis (ua-# 0) was accepted. The paired difference test was also used to evaluate: 1) whether hemorrhage produced significantly dif— ferent transient vascular responses to a step change in perfusion pressure compared to control (Series II); and 2) whether elevation of mean vascular distending pressure produced significantly different vascular responses during pulsatile vs. non-pulsatile perfusion (Series III). In these studies a significance level of 0.05 was used for all comparisons. BIBLIOGRAPHY 10. ll. BIBLIOGRAPHY Anderson, D. K., S. A. Roth, R. A. Brace, D. Radawski, F. J. Haddy, and J. B. Scott: Effects of hypokalemia and hypomagnesemia produced by hemodialysis on skeletal muscle vascular resistance: Role of potassium in active hyperemia. Circ. Res. 31:165-173, 1972. Anrep, G.: On local vascular reactions and their inter- pretation. J. Physiol. 45:318-326, 1912. Axelsson, J., B. Wahlstrom, B. Johansson, and O. Jonsson: Influence of the ionic environment on spontaneous electrical and mechanical activity of the rat portal vein. Circ. Res. 21:609-618, 1967. Bachofen, M., A. Gage, and H. Bachofen: Vascular respon- ses to changes in blood oxygen tension under various blood flow rates. Am. J. Physiol. 220:1787-1789, 1971. Baez, S.: Bayliss response in the microcirculation. Fed. Proc. 27:1410-1418, 1968. Baez, 8.: Supporting tissue tension and microvascular reactions. Microvas. Res. 4:95-103, 1972. Baez, S., H. Lamport, and A. Baez: Pressure effects in living microscopic vessels. 1g: Flow Properties of Blood. Eds. A. L. Copley and W. Stainsby, Pergamon Press, London, p. 122, 1960. Bayliss, W. M.: On the local reactions of the arterial wall to changes in internal pressure. J. Physiol. (London) 28:220-231, 1902. Berne, R. M.: Cardiac nucleotides in hypoxia: possible role in regulation of coronary blood flow. Am. J. Physiol. 204:317-322, 1963. Berne, R. M.: Regulation of coronary blood flow. Physiol. Rev. 44:1-205, 1964. Berne, R. M.: Metabolic regulation of blood flow. Circ. Res. 15:Suppl. 1, 261-268, 1964. 155 12. 13. 14. 15. 16. 17. 18. 19. 20. 21. 22. 23. 24. 156 Bevan, J. A. and B. Ljung: Longitudinal propagation of myogenic activity jg; rabbit arteries and in the rat portal vein. Acta Physiol. Scand. 90:703-715, 1974. Bond, R. F. and H. D. Green: Skeletal muscle autoregu— lation during changes in vascular tone induced by hemorrhage. Fed. Proc. 26:662, 1967. Bond, R. F., R. F. Blackard, and J. A. Taxis: Evidence against oxygen being the primary factor governing autoregulation. Am. J. Physiol. 216:788-793, 1969. Bozler, E.: Conduction, automaticity and tonus of visceral muscles. Experientia 4:213-218, 1948. Britton, K. E.: Renin and renal autoregulation. Lancet 2:329-333, 1968. Burgi, S.: Zur physiologie und pharmakologie der uber- lebenden arterie. Helv. Physiol. Pharmacol. Acta 2:345-353, 1944. Burnstock, G. and C. L. Prosser: Conduction in smooth muscle: Comparative electrical properties. Am. J. Physiol. 199:553-559, 1960. Carrier, 0., J. R. Walker, and A. C. Guyton: Role of oxygen in autoregulation of blood flow in isolated vessels. Am. J. Physiol. 206:951-954, 1964. Chalmers, J. P., P. I. Horner, and S. W. White: The control of the circulation in skeletal muscle during arterial hypoxia in the rabbit. J. Physiol. (London) 184:698-716, 1966. Daugherty, R. M., J. B. Scott, J. M. Dabney, and F. J. Haddy: Local effects of O and CO on limb, renal and coronary vascular resi tances. Am. J. Physiol. 213:1102-1109, 1967. Davignon, J., R. R. Lorenz, and J. T. Shepherd: Response of human umbilical artery to changes in transmural pressure. Amer. J. Physiol. 209:51-59, 1965. Deal, C. P. and H. D. Green: Effects of pH on blood flow and peripheral resistance in muscular and cutaneous beds in the hindlimb of pentobarbitalized dogs. Circ. Res. 2:148-156, 1954. Detar, R. and D. F. Bohr: Oxygen and vascular smooth muscle contraction. Am. J. Physiol. 214:241-244, 1968. 25. 26. 27. 28. 29. 30. 31. 32. 33. 34. 35. 157 DeWardener, H. E. and B. E. Miles: Autoregulation of the kidney blood flow. Clin. Sci. 11:267, 1952. Dobson, J. G., R. Rubio, and R. M. Berne: Role of adenine nucleotides, adenosine, and inorganic phos- phate in the regulation of skeletal muscle blood flow. Circ. Res. 29:375-384, 1971. Driscol, T. E., T. W. Moir, and R. W. Eckstein: Vascu- lar effects of changes in perfusion pressure in nonischemic and ischemic heart. Circ. Res. 15: Suppl 1, 94-102, 1964. Duling, B. R. and R. M. Berne: Possible mechanisms for the participation of oxygen in local regulation of blood flow. Circ. Res. 27:669-678, 1970. Duling, B. R. and R. M. Berne: Oxygen and the local regulation of blood flow: Possible significance of longitudinal gradients in arterial blood oxygen saturation. Circ. Res. 29:Supp1. 1, 65-69, 1971. Ekstrom-Jodal, B.: Effect of increased venous pressure on cerebral blood flow in dogs. Acta Physiol. Scand. Suppl. 350:51-93, 1970. Emanuel, D. A., J. B. Scott, and F. J. Haddy: Effects of potassium on small and large vessels of the dog forelimb. Am. J. Physiol. 197:637-644, 1959. Emerson, T. E., J. L. Parker, and G. W. Jelks: Effects of local acidosis on vascular resistance in dog skeletal muscle. Proc. Soc. Exptl. Biol. Med. 145:273-276, 1974. Fog, M.: Reactions of the pial arteries to a fall in blood pressure. Arch. Neurol. Psychiat. 37:351—359, 1937. Folkow, B.: Intravascular pressure as a factor regulat- ing the tone of small vessels. Acta Physiol. Scand. 17:289-310, 1949. Folkow, B., B. Oberg, and E. H. Rubenstein: A proposed differentiated neuro-effector organization in muscle resistance vessels. Angiologica 1:197-208, 1964. 36. 37. 38. 39. 40. 41. 42. 43. 44. 45. 158 Folkow, B. and B. Oberg: Autoregulation and basal tone in consecutive vascular sections of the skeletal muscles in reserpine treated cats. Acta Physiol. Scand. 53:105-113, 1961. Forrester, T. and A. R. Lind: Identification of adeno- sine triphosphate in human plasma and the concen- tration in the venous effluent before, during, and after sustained contractions. J. Physiol. (London) 204:347-364, 1969. Friedman, S. M. and C. L. Friedman: Effects of ions on vascular smooth muscle. 1p; Handbook of Physiol- ogy, Ed. W. F. Hamilton and P. Dow, Sec. 2, Vol. 2, pp. 1135-1166, Williams and Wilkins, Baltimore, 1963. Funaki, 8.: Studies on membrane potential of vascular smooth muscle with intracellular microelectrodes. Proc. Jap. Acad. 34:534-536, 1958. Funaki, S.: Spontaneous spike discharge of vascular smooth muscle. Nature (London) 191:1102-1109, 1961. Gellai, M., J. M. Norton, and R. Detar: Evidence for direct control of coronary vascular tone by oxygen. Circ. Res. 32:279-284, 1973. Glover, W. E., I. C. Roddie, and R. G. Shanks: Effect of intraarterial potassium chloride infusions on vascular reactivity in the human forearm. J. Physiol. (London) 163:22P, 1963. Green, H. D., C. E. Rapela, and R. F. Bond: Cerebral vs. cutaneous and skeletal muscle vascular responses to localized hypotension and to systemic hypotension induced by hemorrhage and shock. 12; Reversibility of Cellular Injury Due to Inadequate Perfusion, ed. by T. I. Malinin, Thomas, Springfield, Illinois (1972), p. 135. Guyton, A. C., J. B. Langstone, and G. Navar: Theory for renal autoregulation by feedback of the juxta- glomerular apparatus. Circ. Res. 15:Suppl. 1, 187-196, 1964. Haddy, F. J., J. B. Scott, M. A. Florio, R. M. Daugherty, and J. N. Huizenga: Local vascular effects of hypo- kalemia, alkalosis, hypercalcemia, and hypomag- nesemia. Am. J. Physiol. 204:202-210, 1963. 46. 47. 48. 49. 50. 51. 52. 53. 54. 55. 56. 57. 159 Haddy, F. J. and J. B. Scott: Effects of flow rate, venous pressure, metabolites, and oxygen upon resistance to blood flow through the dog forelimb. Circ. Res. 15:Suppl. 1, 49-54, 1964. Haddy, F. J. and J. B. Scott: Role of transmural pres- sure in local regulation of blood flow through kidney. Am. J. Physiol. 208:825-831, 1965. Haglund, U. and O. Lundgren: Reactions within consecu- I tive vascular sections of the small intestine of the cat during prolonged hypotension. Acta Physiol. Scand. 84:151—163, 1972. Hanson, K. M.: Autoregulation in the hindlimb. Circ. Res. 15:Supp1. 1, 25-29, 1964. Hanson, K. M.: Experiments on autoregulation of hepatic blood flow in the dog. Circ. Res. 15:Suppl. Hanson, K. M. and P. C. Johnson: Vascular resistance and arterial pressure in autoperfused dog hindlimb. Am. J. Physiol. 203:615-620, 1962. Hanson, K. M. and P. C. Johnson: Evidence for local arteriovenous reflex in intestine. J. Appl. Physiol. 17:509-513, 1962. Hanson, K. M. and P. C. Johnson: Local control of hepatic arterial and portal venous outflow in the dog. Am. J. Physiol. 211:712-720, 1966. Hinshaw, L. B.: Mechanism of renal autoregulation: Role of tissue pressure and description of a multi- factor hypothesis. Circ. Res. 15:Suppl. 1, 120-129, 1964. Holman, M. E., C. B. Kasby, M. B. Suthers, and J. A. F. Wilson: Some properties of the smooth muscle of rabbit portal vein. J. Physiol. (London) 196:111- 132, 1968. Honig, C. R.: Control of smooth muscle actomyosin by phosphate and 5' AMP: Possible role in metabolic autoregulation. Microvas. Res. 1:133-146, 1968. Hutchins, P. M., R. F. Bond, and H. D. Green: Partici- pation of oxygen in the local control of skeletal muscle microvasculature. Circ. Res. 34:85-93, 1974. 58. 59. 60. 61. 62. 63. 64. 65. 66. 67. 68. 69. 160 Jarhult, J. and S. Mellander: Autoregulation of capil- lary hydrostatic pressure in skeletal muscle during regional arterial hypo- and hypertension. Acta Physiol. Scand. 91:32-41, 1974. Johansson, B. and D. F. Bohr: Rhythmic activity in smooth muscle from small subcutaneous arteries. Amer. J. Physiol. 210:801-806, 1966. Johansson, B. and B. Ljung: Spread of excitation in the smooth muscle of the rat portal vein. Acta Physiol. Scand. 70:312-322, 1967. Johansson, B. and S. Mellander: Static and dynamic components in the vascular myogenic response to passive changes in length as revealed by electrical and mechanical recordings from the rat portal vein. Circ. Res. 36:76-83, 1975. Johnson, P. C.: Myogenic nature of increase in intes- tinal vascular resistance with venous pressure elevation. Circ. Res. 7:992-999, 1959. Johnson, P. C.: Autoregulation of intestinal blood flow. Am. J. Physiol. 199:311-318, 1960. Johnson, P. C.: Review of previous studies and current theories of autoregulation. Circ. Res. 15:Suppl. 1, 2—9, 1964. Johnson, P. C.: Origin, localization, and homeostatic significance of autoregulation in the intestine. Circ. Res. 15:Suppl. 1, 225-232, 1964. Johnson, P. C.: Effect of venous pressure on mean capil- lary pressure and vascular resistance in the intes- tine. Circ. Res. 16:294-300, 1965. Johnson, P. C.: Autoregulatory responses of cat mesen- teric arterioles measured in vivo. Circ. Res. 22: 199-212, 1968. Johnson, P. C.: The microcirculation and local and humoral control of the circulation. 1p; MTP Inter- national Review of Science, Series One, Vol. 1, pp. 163-196, ed. A. C. Guyton and C. E. Jones, Univer- sity Park Press, Baltimore, 1974. Johnson, P. C. and K. M. Hanson: Effect of arterial pressure on arterial and venous resistance of intes- tine. J. Appl. Physiol. 17:503-508, 1962. 70. 71. 72. 73. 74. 75. 76. 77. 78. 79. 80. 81. 161 Johnson, P. C., K. M. Hanson, and O. Thulesius: Pre— and post-capillary resistance in the dog forelimb. Am. J. Physiol. 210:873—876, 1966. Johnson, P. C. and H. Wayland: Regulation of blood flow in single capillaries. Am. J. Physiol. 212:1405- 1415, 1967. Jones, R. D. and R. M. Berne: Skeletal muscle blood flow regulation. Fed. Proc. 20:104, 1961. Jones, R. D. and R. M. Berne: Intrinsic regulation of skeletal muscle blood flow. Circ. Res. 14:126-138, 1964. Jones, R. D. and R. M. Berne: Local regulation of blood flow in skeletal muscle. Circ. Res. 15:Suppl. 1, 30-35, 1964. Jones, R. D. and R. M. Berne: Evidence for a metabolic mechanism in autoregulation of blood flow in skeletal muscle. Circ. Res. 17:540-547, 1965. Katori, M. and R. M. Berne: Release of adenosine from anoxic hearts. Circ. Res. 19:420-427, 1966. Kjellmer, I.: The potassium ion as a vasodilator during musclar exercise. Acta Physiol. Scand. 63:460-471, 1965. Kirk, R. E.: Experimental design: Procedures for the behavioral sciences. Wadsworth Publishing Co., Belmont, Calif., 1968. Kontos, H. A., D. W. Richardson, and J. L. Patterson: Blood flow and metabolism of forearm muscle in man at rest and during sustained contraction. Am. J. Physiol. 211:869-875, 1966. Langston, J. B., A. C. Guyton, and W. J. Gillespie, Jr.: Autoregulation absent in normal kidney but present after renal damage. Am. J. Physiol. 199:495-498, 1960. Lassen, N. A.: Autoregulation of cerebral blood flow. Circ. Res. 15:Supp1. 1, 201-204, 1964. 82. Ljung, B. and L. Stage: Adrenergic excitatory influ- ences on initiation and conduction of electrical activity in the rat portal vein. Acta Physiol. Scand. 80:131-141, 1970. 83. 84. 85. 86. 87. 88. 89. 90. 91. 92. 162 Lundvall, J., S. Mellander, and H. V. Sparks: Myogenic response of resistance vessels and precapillary sphincters in skeletal muscle during exercise. Acta Physiol. Scand. 70:257-268, 1967. Mellander, S., B. Oberg, and H. Odelram: Vascular ad- justments to increased transmural pressure in cat and man with special reference to shifts in capil- lary fluid transfer. Acta Physiol. Scand. 61:34-48, 1964. Mellander, S. and S. Arvidsson: Possible "dynamic" com— ponent in the myogenic vascular response related to pulse pressure distension. Acta Physiol. Scand. 90:283-285, 1974. Molnar, J. I., H. W. Overbeck, and F. J. Haddy: Local effects of oxygen and carbon dioxide upon dog fore— limb vascular resistance. In: Proc. 22nd Intern. Congr. Physiol. Sci. LeydenT_Vol. II, Excerpta Med. Found., Amsterdam (1962), p. 175. Molnar, J. I., J. B. Scott, E. D. Frohlich, and F. J. Haddy: Local effects of various anions and hydrogen ion on dog limb and coronary vascular resistances. Am. J. Physiol. 203:125-129, 1962. Nagle, F. J., J. B. Scott, B. T. Swindall, and F. J. Haddy: Venous resistances in skeletal muscle and skin during local blood flow regulation. Am. J. Physiol. 214:885-891, 1968. Nahmod, V. E. and A. Lanari: Abolition of autoregula- tion of renal blood flow by acetylcholine. Am. J. Physiol. 207:123-127, 1964. Navar, L. G., A. C. Guyton, and J. B. Langston: Effects of alterations in plasma osmolality on renal blood flow autoregulation. Am. J. Physiol. 211:1387-1392, 1966. Nicoll, P. A. and R. L. Webb: Vascular patterns and active vasomotion as determiners of flow through minute vessels. Angiology 6:291-299, 1955. Radawski, D., J. M. Dabney, R. M. Daugherty, F. J. Haddy, and J. B. Scott: Local effects of CO on vascular resistances and weight of the dog forelimb. Am. J. Physiol. 222:439-443, 1972. 93. 94. 95. 96. 97. 98. 99. 100. 101. 102. 103. 104. 163 Radawski, D., W. Hoppe, and F. J. Haddy: Role of vaso- active substances in active hyperemia in skeletal muscle. Proc. Soc. Exptl. Biol. Med. 148:270-276, 1975. Rapela, C. E. and H. D. Green: Autoregulation of canine cerebral blood flow. Circ. Res. 15:Supp1. 1, 205-211, 1964. Richman, H. G. and L. Wyborny: Adenine nucleotide degredation in the rabbit heart. Am. J. Physiol. 207:1139-1146, 1964. Ross, J. M., H. M. Fairchild, J. Weldy, and A. C. Guyton: Autoregulation of blood flow by oxygen lack. Am. J. Physiol. 202:21-26, 1962. Ross, J., G. A. Kaiser, and F. J. Klocke: Observations on the role of diminished oxygen tension in func- tional hyperemia of skeletal muscle. Circ. Res. 15:473-479, 1964. Rovick, A. A.: Active vascular capacity responses in skeletal muscle. Amer. J. Physiol. 210:121-127, 1966. Rovick, A. A. and P. A. Robertson: Interaction of mean and pulse pressures in the circulation of the iso- lated dog tongue. Circ. Res. 15:208-215, 1964. Rubio, R. and R. M. Berne: Release of adenosine by the normal myocardium in dogs and its relationship to the regulation of coronary resistance. Circ. Res. 25:207—216, 1969. Rubio, R. and R. M. Berne: Release of adenosine by the normal myocardium in dogs and its relationship to the regulation of coronary resistance. Circ. Res. 25:407-416, 1969. Rudko, M. and F. J. Haddy: Venous plasma [K+J during reactive hyperemia in skeletal muscle. Physiologist 8:264, 1965. Scott, J. B., R. A. Hardin, and F. J. Haddy: Pressure- flow relationships in the coronary vascular bed of the dog. Am. J. Physiol. 199:765-769, 1960. Scott, J. and F. J. Haddy: Local effects of Cd++ and Hg on dog forelimb vascular bed. Fed. Proc. 22:179, 1963. 105. 106. 107. 108. 109. 110. 111. 112. 113. 114. 115. 164 Scott, J. B., R. M. Daugherty, J. M. Dabney, and F. J. Haddy: Role of chemical factors in regulation of flow through kidney, hindlimb, and heart. Am. J. Physiol. 208:813-819, 1965. Scott, J. B., M. Rudko, D. Radawski, and F. J. Haddy: Role of osmolarity, K+, H+, Mg++, and O2 in local blood flow regulation. Am. J. Physiol. 218:338-345, 1970. Scott, J. B. and D. Radawski: Role of hyperosmolarity in the genesis of active and reactive hyperemia. Circ. Res. 29:Supp1. 1, 26-32, 1971. Scott, J. B., W. Chen, D. Radawski, D. K. Anderson, and F. J. Haddy: Bioassay evidence for participa- tion of adenosine and the adenine nucleotides in local blood flow regulation. Proc. Intern. Union. Physiol. Sci. 9:504, 1971. Selkurt, E. E. and P. C. Johnson: Effect of acute ele- vation of portal venous pressure on mesenteric blood volume, interstitial fluid volume and hemodynamics. Circ. Res. 6:592-599, 1958. Skinner, N. 8., Jr., and W. J. Powell, Jr.: Action of oxygen and potassium on vascular resistance in dog skeletal muscle. Am. J. Physiol. 212:533-538, 1967. Smiesko, V.: Unidirectional rate sensitivity component in local control of vascular tone. Pflugers Arch. 327:324-336, 1971. Sparks, H. V.: Effect of quick stretch on isolated vascular smooth muscle. Circ. Res. 15:Supp1. l, Sparks, H. V. and D. F. Bohr: Effect of stretch on passive tension and contractility of isolated vascu— lar smooth muscle. Am. J. Physiol. 202:835-840, 1962. Speden, R. N.: Electrical activity of single smooth muscle cells of the mesenteric artery produced by splanchnic nerve stimulation in the guinea pig. Nature (London) 202:193-194, 1964. Stainsby, W. N.: Autoregulation of blood flow in skeletal muscle during increased metabolic activity. Am. J. Physiol. 202:273-276, 1962. 116. 117. 118. 119. 120. 121. 122. 123. 124. 125. 126. 127. 165 Stainsby, W. N.: Autoregulation in skeletal muscle. Circ. Res. 15:Supp1. 1, 39-47, 1964. Stainsby, W. N. and E. M. Renkin: Autoregulation of blood flow in resting skeletal muscle. Am. J. Physiol. 201:117-122, 1961. Stainsby, W. N. and A. B. Otis: Blood flow, blood oxygen tension, oxygen uptake, and oxygen transport in skeletal muscle. Am. J. Physiol. 206:858-866, 1964. Steedman, W. M.: Microelectrode studies on mammalian vascular muscle. J. Physiol. (London) 186:382-400, 1966. Stowe, D. F., T. L. Owen, W. T. Chen, D. K. Anderson, and J. B. Scott: Effects of O and CO changes on the vasoactivity of effluent bIood from exercising skeletal muscle. Fed. Proc. 32:436, 1973. Thulesius, O. and P. C. Johnson: Pre« and postcapillary resistance in skeletal muscle. Am. J. Physiol. 210:869—872, 1966. Thurau, K.: The nature of autoregulation of renal blood flow. Proc. III Int. Congr. Nephrol., Washington, Ed. J. S. Handler. Basel, New York, Vol. I, pp. 162-173, 1966. Torrance, H. B.: The control of the hepatic arterial circulation. J. Physiol. 158:39—49, 1961. Wachholder, K.: Haben die rhythmischen spontankontrak— tionen der gefass einen nachweisbaren einfluss auf den blutstrom? Pfluger Arch. Ges. Physiol. 190: 222-231, 1921. Walker, J. R. and A. C. Guyton: Influence of blood oxygen saturation on pressure-flow curve of dog hindleg. Am. J. Physiol. 212:506-509, 1967. Waugh, W. H. and R. G. Shanks: Cause of genuine auto- regulation of the renal circulation. Circ. Res. 8:871-888, 1960. Weideman, M. P.: Effect of venous flow on frequency of venous vasomotion in the bat wing. Circ. Res. 5:641-644, 1957. 166 128. Weideman, M. P.: Enhanced arteriolar activity during elevation of intraluminal pressure. 12: The Microcirculation, Ed. W. L. Winters and A. Brest; Springfield, Illinois, 1969, p. 121. 129. Weideman, M. P.: Contractile activity of arterioles in the bat wing during intraluminal pressure changes. Circ. Res. 19:559-563, 1966. 130. Zsoter, T., L. Banderman, and C. I. Chappel: The effect of '1oca1' pH changes in blood flow in the dog. Am. Heart J. 61:777-782, 1961.