ANTAGONISM 0F HISTAMINE EOEMA FORMATION BY NOREPINEPHRINE IN THE 'cANINE FORELIMB f Thesis for the Degree Of M. ‘S. ' . 2 MICHIGAN STATE UNIVERSITY _ 7 . - DOUGLAS LEE MARCINIAK 1975 21:. \ IIIUIHI Q n '* v. . . .u. ‘ . v¢¢ I I - I . , , §:l I .- r I - . .. .. . . . .1 . .. - - . . , .. , Ira .- - . . ' ' ‘ r . ' . .. . . v- .- - up. a. '= t . ’ ‘ ' . . u .1. “a ' , , . . . - - o , -. . .pgx ' - ' o - v . - - ‘ . cu ~ - “ ‘ . , I. . I _ . '. , " ‘ . ‘ v . '. . 'TD ' - - - . .- - - .~I-~ . . -. fl ' I ' ' us ' n ' - . ' . I - O . -a . . . . .. . db . . _ . . . ‘ ' . . 33%?! _ S W... ‘ I ‘..'~ . ABSTRACT ANTAGONISM 0F HISTAMINE EDEMA FORMATION BY NOREPINEPHRINE IN THE CANINE FORELDWB BY Douglas Lee Marciniak There is clearly a route-dependent differential action of histamine on forelimb transvascular fluid flux. Local (intra-arterial) infusions of histamine (4 to 64ug base/min) into the brachial artery of the forelimb causes marked increases in transvascular fluid flux and forelimb weight owing to an increased capillary hydrostatic pressure gradient and an increased permeability to plasma proteins. Massive visible edema is seen with dosages of 15 to 60mg base/minute. In contrast, systemically (intravenously) infused histamine at rates that produce blood concentrations calculated to equal or exceed those achieved during local infusions, results in sustained net extravascular fluid reabsorption. Possible explanations for this route-dependent differential action of histamine are: the lungs may actively destroy histamine since it must first pass through the pulmonary circuit before reaching the systemic microvessels during intravenous infusion; that factors within the blood may destroy or inactivate histamine before it reaches the systemic microvasculature during systemic infusions; that substances Douglas Lee Marciniak released (e.g., catecholamines) during a sympathoadrenal discharge, subsequent to a marked and immediate hypotension produced only with the systemic infusions (20 to 800ug base/min), may effectively antagonize the edemogenic actions of histamine by a direct blockade of histaminels permeability increasing effects on the microvessels, a shunting of blood to non-nutritional channels or a combination of both. Forelimb weight and hemodynamic studies were conducted during systemic histamine infusions (400 to 800ug base/min) into the vena cava, the left ventricle of the heart and during local infusion into the brachial artery of the forelimb (4 and 64ug base/min). This allowed for comparisons of systemic and local infusions and evaluation of the possible role the lungs may have upon the route-dependent differential actions of histamine. To test the hypothesis that substances liberated during a hypotensive induced sympathoadrenal discharge antagonize histamine's action on the microvasculature, systemic histamine, acetylcholine, and hemorrhagic hypotension were produced for 60 minutes. This was followed by a 30 or 60 minute infusion of histamine (4 and 64ug '. base/min,I.A.) into forelimbs perfused at constant inflow during the systemic hypotension. To further test the hypothesis that substances released during a sympathoadrenal discharge antagonize histamine at the size of the microvasculature, weight and hemodynamic studies were conducted in which norepinephrine (4ug base/min) was simultaneously infused with histamine (4ug base/min) into the forelimb brachial artery. Histamine (400 and 800ug base/min) infused into the vena cava and left ventricle of the heart for 90 minutes produced marked Douglas Lee Marciniak hypotension and only very slight increases in forelimb skin lymph flow rate and total protein concentration. Forelimb weight continually decreased during these infusions. There was no discernable differences in any of the hemodynamic parameters during intravenous or left heart infusions. Thus, the route-dependent differential actions of histamine cannot be attributed to uptake or destruction of histamine by the lungs. The greater transit time required for histamine to reach the microvasculature during systemic infusions than during local infusions, which would allow more time for factors within the blood to destroy or inactivate histaime, cannot account for the route—dependent differ- ential actions of histamine either. This is demonstrated by the fact that large increases in lymph flow rate and total protein concentration are witnessed during local histamine infusions into forelimbs perfused at constant inflow. In these preparations, histamine must travel through about 3 feet of polyethylene tubing before reaching the fore- limb. Since this distance is equal to or greater than the distance histamine must travel during systemic infusions, if factors within the blood were destroying histamine than such profound histamine effects (increased lymph flow and total protein concentration) would not be expected during the local infusions into forelimbs perfused at con- stant inflow. Histamine infused locally (4 and 64ug base/min) into forelimbs perfused at constant inflow completely prevented the increase in total protein contentration of lymph following 60 minutes of systemic histamine, acetylcholine or hemorrhagic hypotension. This indicates that hypotension results in the release of substances (e.g., Douglas Lee Marciniak catecholamines) that effectively antagonizes histamine‘s actions on the microvasculature. The stimultaneous infusion of norepinephrine (4ug base/min) with histamine (4ug base/min) into forelimbs perfused at constant inflow prevented the usual large increases in lymph flow and total protein concentration seen during histamine infusions alone. This convincingly demonstrates the antagonistic action of norepinephrine on histamine at the site of the microvasculature. In conclusion, the failure of systemically infused histamine to exert effects on the microvasculature similar to those produced by locally infused histamine is related to hypotension pg: §g_and to antagonism of the microvascular actions of histamine subsequent to the release of substances (e.g., catecholamines) in response to the hypotension produced by systemically infused histamine. This antagonism of histamine by norepinephrine and/or other substances released during a sympathoadrenal discharge could be due to a direct blockage of histamine's permeability increasing effect or to a shunting of blood flow to non-nutritional channels or a combination of both. ANTAGONISM OF HISTAMINE EDEMA FORMATION BY NOREPINEPHRINE IN THE CANINE FORELIMB BY Douglas Lee Marciniak A THESIS Submitted to Michigan State University in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Physiology 1976 DEDICATION This thesis is dedicated to my parents and grandmother. Without their love, understanding, encouragement and willing support, my education and this thesis would never have been possible. ii ACKNOWLEDGMENTS The author wishes to express his deep appreciation to the following peOple for their endless contributions in this endeavor: Dr. Joe M. Dabney, Dr. Francis J. Haddy, Dr. Jerry B. Scott, Mr. Edward Gersabeck, Mr. James Maciejko, and Mrs. Jodi Johnston. A very Special thanks is extended to Dr. George J. Grega, my advisor. It has been a wonderful experience working with him. iii TABLE OF CONTENTS LIST OF TABLES . . . . . . . LIST OF SYMBOLS AND ABBREVIATIONS. . . . . . . . INTRODUCTION. . . . . . . . . . . . . LITERATURE REVIEW . . . . . . . . STATEMENT OF THE PROBLEM. . . . . . . . . . . METHODS . . . . . . . . . . . . . . RESULTS . . . . . . . . . . . . . . . . DISCUSSION . . . . . . . . . . . . SUMMARY AND CONCLUSIONS . . . . . . . APPENDICES Appendix A. Sample Calculations for Blood Concentrations of Histamine . . . . . . . . . . . . . B. Tables . . . . . . . . . . . . . . BIBLIOGRAPHY. . . . . . . . . . . . . iv Page ix '21 23 29 50 60 62 64 137 Table LIST OF TABLES Effects of histamine base infused intravenously for 90 minutes plus histamine infused intra-arterially into the forelimb during the last 30 minutes on forelimb weight, vascular pressures, blood flow and vascular resistances . . . . . . . . . . . . . . Effects of histamine base infused systemically for 90 minutes plus histamine infused intra-arterially into the forelimb during the last 30 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit. . . . . . . Effects of histamine base infused into the left ventricle of the heart and intra-arterially (after a 3 minute delay) into the forelimb for 60 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit. . . . . . . . . . . Effects of locally infused histamine (4 or 64mg base/min, I.A.) for 60 minutes following 60 minutes of hypoten- sion produced by either acetylcholine base infused intravenously at a concentrationanecessary to lower and maintain aortic pressure near 60 mm Hg for the first 60 minutes or following 60 minutes of hemorrhagic hypotension with aortic pressure maintained near 40 mm Hg . . . . . . . . . . . . . . . Effects of histamine (4 ug base/min, I.A.) and histamine plus norepinephrine (4 ug base/min, I.A. of each) infused locally into naturally perfused forelimbs on weight, vascular pressures, blood flows and vascular resistances . . . . . . . . . . . . . Effects of histamine plus norepinephrine (4 ug base/min, I.A. of each) infused locally into forelimbs perfused at constant inflow on forelimb weight, vascular pressures, blood flows and vascular resistances . . Page 37 39 41 42 43 4S Table Page 7. Effects of histamine base and/or norepinephrine base infused intra-arterially into the forelimb for 60 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . . . 46 A-l. Effects of histamine base infused intravenously for 90 minutes plus histamine infused intra-arterially into naturally perfused forelimbs during the last 30 minutes on forelimb weight, vascular pressures, blood flow and vascular resistances . . . . . . . . . . . . 65 A-2. Effects of histamine base infused intravenously for 90 minutes plus histamine infused intra-arterially into forelimbs perfused at constant inflow during the last 30 minutes on forelimb weight, vascular pressures, blood flow and vascular resistances . . . . . . . 69 A—3. Effects of histamine base infused into the left ventricle of the heart for 90 minutes plus histamine infused intra- arterially into naturally perfused forelimbs during the last 30 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . 73 A-4. Effects of histamine base infused intravenously for 90 minutes plus histamine infused intra-arterially into naturally perfused forelimbs during the last 30 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . . . . 76 A-s. Effects-of histamine base infused into the left ventricle of the heart for 90 minutes plus histamine infused intra-arterially into forelimbs perfused at constant inflow during the last 30 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . . . . . . . . . . . . . 79 A-6. Effects of histamine base infused intravenously for 90 minutes plus histamine infused intra-arterially into forelimbs perfused at constant inflow during the last 30 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . 82 A-7. Effects of histamine base infused into the left ventricle of the heart and intra-arterially (after a 3 minute delay) into naturally perfused forelimbs for 60 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . . . . 85 vi Table Page A-8. Effects of histamine base infused into the left ventricle of the heart and intra-arterially (after a 3 minute A delay) into forelimbs perfused at constant inflow for 60 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . . . 88 A—9. Effects of 4 ug histamine base/minute infused intra- arterially for 60 minutes into forelimbs perfused at constant inflow following 60 minutes of hypotension produced by acetylcholine base infused intravenously at a concentration necessary to lower and maintain aortic pressure near 60 mm Hg for the first 60 minutes . . . 91 A-lO. Effects of 4 ug histamine base/minute infused intra- arterially for 60 minutes into forelimbs perfused at constant inflow following 60 minutes of hemorrhagic hypotension with aortic pressure maintained near 40 mm Hg . . . . . . . . . . . . . . . . _93 A-ll. Effects of 64 ug histamine base/minute infused intra- arterially for 60 minutes into forelimbs perfused at constant inflow following 60 minutes of hemorrhagic hypotension with aortic pressure maintained near 40 mm Hg . . . . . . . . . . . . . . . . 95 A-lZ. Effects of 4 ug histamine base/minute infused intra- arterially into naturally perfused forelimbs on weight, vascular pressures, blood flows and vascular resistances . . . . . . . . . . . . . . . 97 A-13. Effects of histamine plus norepinephrine (4 ug base/minute, of each) infused simultaneously intra-arterially into naturally perfused forelimbs on weight, vascular pressures, blood flows and vascular resistances . . . 101 A-l4. Effects of histamine plus norepinephrine (4 ug base/minute, of each) infused simultaneously intra-arterially into forelimbs perfused at constant inflow on weight, vascular pressures, blood flows, and vascular resistances . . . . . . . . . . . . . . . 105 A-lS. Effects of 4 ug histamine base/minute infused intra- arterially into naturally perfused forelimbs for 60 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . . . 109 vii Table Page A-l6. Effects of simultaneous intra-arterial infusions of histamine and norepinephrine, 4 pg base/minute of each, into naturally perfused forelimbs for 60 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . . . . . . . lll A-l7. Effects of 4 pg histamine base/minute infused intra- arterially into forelimbs perfused at constant inflow for 60 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . 114 A-18. Effects of simultaneous intra-arterial infusions of histamine and norepinephrine, 4 pg base/minute of each, into forelimbs perfused at constant inflow for 60 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . . . ll7 A-19. Effects of 4 pg norepinephrine base/minute infused intra- arterially into forelimbs perfused at constant inflow for 60 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . 120 A-20. Effects of 64 pg histamine base/minute infused intra- arterially into naturally perfused forelimbs for 60 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . . . 123 A-21. Effects of 64 pg histamine base/minute and 4 pg norepin- ephrine base/minute infused simultaneously intra- arterially into naturally perfused forelimbs for 60 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . . . 125 A-22. Effects of 64 pg histamine base/minute infused intra- arterially into forelimbs perfused at constant inflow for 60 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . 128 A-23. Effects of 64 pg histamine base/minute and 4 pg norepin- ephrine base/minute infused simultaneously intra- arterially into forelimbs perfused at constant inflow for 60 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrits. . 131 A-24. Effects of 64 pg histamine base/minute and 16 pg norepin- ephrine base/minute infused simultaneously intra- arterially into forelimbs perfused at constant inflow or 60 minutes on lymph flow, protein concentration of lymph and plasma, vascular pressures and hematocrit . . 134 viii NF CF IA IV U8 min ml H4 H64 H400 H800 N4 LIST OF SYMBOLS AND ABBREVIATIONS Naturally perfused forelimb. Forelimb perfused at constant inflow. Drug infusion into the left ventricle of the heart. Drug infusion intra-arterially into the forelimb. Drug infusion into the vena cava. Microgram. Minute. Milliliter. litter. grams. Kilograms Millimeters of mercury pressure. Acetylcholine infused intravenously along with 4pg/min of histamine base. Hemorrhage along with 4pg/min of histamine base. Hemorrhage along with 64pg/min of histamine base. 4pg/min of histamine base. 64pg/min of histamine base. 400pg/min of histamine base. 800pg/min of histamine base 4pg/min of norepinephrine base. ix N16 l6Ug/min piqm PEODS P:Ob1 pimm of norepinephrine base. Relative to 0 minute Relative to 0 minute Relative to 60 minute Relative to 60 minute control. control. value. value. INTRODUCTION Histamine (B-imidazolylethylamine) was first subjected to scientific investigation in 1910 by Dale and Laidlow (8). They found that it stimulated a variety of smooth muscle and markedly depressed blood pressure when injected intravenously. In 1927 Best gt 21° (4) determined that histamine was actually an autocoid and at the same time Lewis (37) showed that it was liberated in response to tissue injury and antigen-antibody reactions. The work by these investigators led to histamine being one of the most studied compounds in the history of biomedical sciences. Over the past 50 years histamine has been implicated in many pathophysiological conditions such as anaphylaxis, allergy, tissue injury and shock. When histamine release is confined to localized areas such conditions may be associated with tissue edema. It is now becoming apparent that histamine may be involved in various normal physiological mechanisms such as microcirculatory regulation, tissue growth and repair, central nervous system function and possibly gastric secretory responses (33). Of primary interest in this study are those actions of histamine relevant to pathological conditions which are manifested by abnormal transvascular fluid fluxes. Local (intra-arterial) infusions of histamine in large doses (15-60 pg base/min) increases transvascular fluid filtration leading to massive edema formation. This edema is attributable to both a rise in the transmural capillary hydrostatic pressure gradient subsequent to arteriolar vasodilation and a fall in the transmural colloid osmotic pressure gradient owing to an increased microvascular per- meability to plasma proteins. Even if capillary hydrostatic pressure is mechanically prevented from increasing, edema still occurs indi- cating that the fall in the transmural colloid osmotic pressure gradient exerts the dominant effect, at least with the higher doses (15-60 pg base/min). In contrast, histamine administered systemically (intravenously), at rates that produce concentrations estimated to equal or exceed those achieved locally, not only fails to promote edema formation but rather leads to sustained net reabsorption of extra- vascular fluid (19). The mechanism of these little noted route- dependent differential actions of histamine on transvascular fluid fluxes is unknown. Although the estimated blood concentrations of histamine may be equal during both local and systemic administration of this agent, there are other obvious differences that may account for the observed route-dependent differential actions of histamine on fluid filtration. First of all, during local infusions of histamine capillary hydro- static pressure (Pc) increases whereas it decreases during systemic infusions of this agent. Also during intravenous infusions, histamine passes first through the pulmonary circuit before reaching the systemic microvessels. Thus it is possible that passage through the pulmonary circuit may result in uptake, metabolism, or biotransformation of histamine by the lungs. During local administration histamine first enters the microcirculation and then, upon recirculation, enters the lung. The transit time from the point of introduction to the micro- circulation is also greater during systemic than during local infusions, possibly resulting in destruction by factors within the blood. Additionally, systemic pressure is little affected during local administration of histamine (5 to 60pg base/min) or falls only slightly after edema has already developed (20). In contrast, during systemic administration there is an immediate and sustained fall in systemic pressure which, at least initially, appears to be dose related within a range of 20 to 800pg of histamine base/min (19). The fall in arterial blood pressure is undoubtedly associated with a marked sympathoadrenal discharge. It is conceivable that substances liberated subsequent to the hypotension effectively antagonize the usual edemogenic action of histamine. It is, therefore, the aim of this study to elucidate the mechanism(s) of the route-dependent differential actions of histamine on transvascular fluid fluxes. These findings may lead to a more complete understanding of the microcirculatory actions of histamine, and may also have relevance to the pathophysiology of certain anaphy- lactic and circulatory shock states which are associated with extremely high blood levels of histamine (up to lug/ml whole blood [28]) but persistent net extravascular fluid reabsorption occurs. LITERATURE REVIEW The movement of fluid across the capillaries was first proposed in 1653 by Bartholinus (3) and again in 1753 by Hales (25). The first definitive theory concerning capillary filtration was proposed in 1861 by C. F. Ludwig (38) and it was not until 1896, when Starling published an article entitled "On the absorption of fluids from the connective tissue spaces" (62), that the physical determinants of fluid exchange across the capillaries began to be understood. The physical basis of fluid filtration is expressed by the following equation: F.M. = K (Pc - Pi - up + Di) where F.M. represents fluid movement across the capillary wall. When the equation is positive, filtration occurs (movement of fluid into the interstitium from the vasculature) and when negative, fluid reabsorption (movement of fluid from the interstitium into the vasculature) occurs. Pc represents capillary hydrostatic pressure and Pi represents the hydro- static pressure of the interstitial fluid. Colloid osmotic pressure (oncotic pressure) is symbolized by Np for plasma and Ni for inter- stitial fluid. The symbol K is a proportionality constant representing the capillary filtration coefficient. It is a measure of the permea- bility of the capillary wall to isotonic fluids (relative to plasma) and is determined by the product of capillary permeability and surface area available for diffusion. Capillary hydrostatic pressure (Pc) is the force acting to move fluid outward into the interstitium across the capillary wall. Its immediate determinants are capillary blood volume and capillary compliance. Since the capillaries are relatively stiff, compliance is usually not considered to significantly change. Clough 33 31' (7) reported changes in radius of only about 0.1 micrometer during systole in capillaries of cat mesentery. This rigidity may be due to elastic properties of the capillary wall (5) or to the incompressible nature of the surrounding gel preventing capillary distention (15). Hence, capillary hydrostatic pressure is determined by capillary blood volume which is regulated by several physical factors affecting both inflow ’ and outflow. These factors are related by the equation: “U 0 ll - Rv (Pa - PV) m 4' PV where: Pc mean capillary hydrostatic pressure Pa = mean arterial blood pressure Pv = venous or outflow pressure Rv = venous resistance to outflow Ra = arterial resistance to inflow Thus, capillary hydrostatic pressure is directly related to changes in mean arterial blood pressure, venous outflow pressure and venous resistance; and is indirectly related to arterial resistance. Arterial and venous resistances are related to vessel caliber which is deter- mined by active changes (vascular smooth muscle activity), passive changes (transmural pressure) and blood viscosity. The hydrostatic pressure of interstitial fluid (Pi) is analogous to the capillary hydrostatic pressure but is that pressure found in the interstitial spaces. The classical view is that inter- stitial tissue pressure is positive and consequently Opposes fluid movement out of the capillaries. Recently, however, new information has come forth challenging this viewpoint suggesting that interstitial fluid hydrostatic pressure is slightly negative in a variety of tissues (21). If this is the case then fluid movement out of the capillaries would be enhanced by interstitial hydrostatic pressure since there would be no opposing force to even the minimal capillary hydrostatic pressure. This issue remains to be resolved. Plasma colloid osmotic pressure (Np) is the pressure resulting from the concentration of dissolved proteins in the plasma and other’ physical—chemical factors not completely understood. The plasma proteins are a mixture containing albumins, globulins and fibrinogen. Albumin has an average molecular weight of 69,000 and its concentration 0 is normally found to be about 4.5 grams 6. The globulins average molecular weight is 140,000 with a concentration of about 2.5 grams %. Fibrinogen is the largest with a molecular weight of about 400,000 and is found in concentrations of approximately 0.3 grams %. This yields a total plasma protein concentration of 7.3 grams % and a normal plasma colloid osmotic pressure of around 28 mm Hg of'which 19 mm Hg is attributable to the proteins themselves and 9 mm Hg to charged ions being held by electronegative forces from the plasma proteins. (The values stated are human plasma values; the values for a dog tend to be slightly less.) Since albumin is only about half the size of the globulins and its concentration is almost twice as much, the osmotic effect of albumin is about 70% of the total colloid osmotic pressure. Colloid osmotic pressure of interstitial fluid depends upon the amount of protein dissolved within the interstitium. Small amounts of plasma proteins normally cross the microvascular wall usually at the immediate postcapillary venule (41). Albumin, since it is smaller, crosses at about 1.5 times the rate of the globulins. The total quantity (grams) of the interstitial proteins is about equal to that of plasma but since interstitial fluid water volume is about 4 times greater than plasma, the effective protein concentration (grams %) and hence, osmotic pressure, is much less. Normal concentration of total inter- stitial proteins is about 2.0 grams % which gives an interstitial colloid osmotic pressure of about 4.5 mm Hg in tissues such as skin and skeletal muscle. This pressure is disproportionately less compared to plasma because at low concentrations of protein the colloid osmotic pressure becomes mostly a function of concentration alone. The electronegative forces become minimal. More recent findings suggest that the total protein concentration of interstitial fluid is about 3 grams % and the colloid osmotic pressure about 10 mm Hg (66). Protein concentration and colloid osmotic pressure will vary from one vascular bed to another depending upon the capillary permeability. An example is in the liver where the heptic sinuses are extremely permeable to proteins thereby allowing for a very rapid and an effective exchange of nutrients between blood and liver cells. When discussing tissue colloid osmotic pressure certain reservations must be kept in mind especially concerning absolute values. Measurements using implantable devices such as perforated capsules that theoretically equilibrate with interstitial fluid, may be inaccurate because of the possiblity of contamination by plasma or that sampled fluid may not contain all the various osmotically active particles. Lymph fluid analysis, the most common method, makes the assumption that it is a true reflection of interstitial fluid. However, there could be drastic changes in lymph composition as it flows from terminal lymphatics upwards to larger vessels and/or that gradients of protein concentration exist within the interstitial spaces. Mayerson and associates (45, 49) and Garlick and Renkin (17) have performed studies showing exchange occurs only at lymph nodes, not in the lymphatic trunks. Therefore, if lymph is sampled before it reaches a node it should be a true reflection of what is at the terminal lymphatic vessel. Furthermore, Renkin and Garlick (53) using dextran molecules of known molecular weight and size found that the concentra- tions were equal between lymph and interstitial fluid for the dextran molecules and presumably for albumins. Their conclusion was that there is no substantial protein concentration gradients within the intersti- tial spaces beyond the capillaries. So at least under normal con- ditions, it appears that lymph protein concentration is a satisfactory reflection of interstitial fluid protein concentration. However, there is the possibility that under abnormal conditions the lymph proteins may not remain in equilibrium with the interstitium. The capillary filtration coefficient is actually a composite of several variables affecting transcapillary fluid movement. These variables are the surface area of the capillary wall available for filtration (Am), thickness of the capillary wall (Ax), fluid viscosity (n) and the specific filtration constant of the membrane (Km). Looking at these factors in terms of an equation we see: Capillary filtration coefficient (K) = fimAfim Since viscosity of the fluid and capillary wall thickness remains fairly constant, the primary determinants for the capillary filtration coefficient is the actual permeability of a given vascular bed and the surface area available for diffusion. Furthermore, within a given capillary bed, permeability remains fairly constant under normal conditions. Hence in normal physiological conditions it is usually changes in surface area available for diffusion that alters the capillary filtration coefficient and consequently affects net fluid movement. In many pathOphysiological states, such as anaphalaxis, allergy, tissue injury and shock, all of the parameters affecting fluid movement may be markedly altered and subsequently cause drastic changes in transcapillary fluid movement. Histamine, released in such conditions is one agent that could be involved in at least some of the cardiovascular alterations associated with these pathophysiological states. Hinshaw found that mean blood levels of histamine increase linearly from a control of 0.1 pg/ml to about 1.0 pg/ml over a time span of 180 minutes after intravenous administration of a lethal dose of endotoxin to anesthetized dogs (28). Similar changes were observed in monkeys under the same experimental conditions, however mean values were much lower. Histamine increased from control values of nearly zero to about 0.4 pg/ml of blood within one hour after endotoxin administration (29). Schayer (57, 58) demonstrated that in response to not only endotoxin shock but to stress induced shock (achieved by intramuscular injections of 20 or 40 pg epinephrine in 10 mice) that there was a threefold increase in skin and lung tissue histidine decarboxylase activity. Norepinephrine produced similar results in skin at the same dosages but with 3 separate subcutaneous injections. Histidine decarboxylase is the enzyme involved with the conversion of histidine to histamine. An increase in its activity presumably reflects an increase in the rate of histamine synthesis. Dekanski observed in mice that the total body histamine content almost doubled from a control of about 8.6 pg/gram of tissue within 10 minutes after being plunged into 60°C water fur 30 seconds (10). Schayer and Ganley repeated these experiments and found a threefold increase irnhistidine decarboxylase (59). When the stimulus for histamine release is a localized condition such as a burn, bee sting, etc., the amount of histamine released is extremely variable being dependent not only upon the nature and severity of the insult but upon the sensitivity of the individual. Consequently, to study the local effects of histamine, investigators usually administer doses of histamine that produce effects similar to those seen clinically. In most vascular beds the local administration of histamine in doses ranging from 3-64 pg base/min intra-arterially or 1.0 to 28.0 pg injected subcutaneously greatly increases net transvascular fluid filtration (6, 9, 20, 24, 32, 35). Massive visible edema, similar to that observed clinically, is produced with the medium to high infusion rates (16-64 pg base/min). This increased net fluid filtration is attributable to both a rise in the transmural capillary hydrostatic pressure gradient and a fall in the transmural colloid osmotic pressure gradient. The rise in the transmural hydrostatic pressure gradient is due to an increased capillary hydrostatic 11 pressure subsequent to arteriolar vasodilation. The fall in the transmural colloid osmotic pressure gradient is attributable to an increased microvascular permeability to plasma proteins and exerts the dominant edemogenic effect with the higher doses of histamine. The alterations of these two determinants of fluid filtration is witnessed by marked increases in organ weight, volume and circumference, and both flow rate and total protein concentration of lymph draining the vascular bed. The latter approaches plasma values with high doses of histamine (6, 20, 23, 32, 52). The increased protein efflux is usually attributable to an increased pore size subsequent to a direct action of histamine on the postcapillary microvascular membrane (39, 40, 41, 42, 67). These microscopic studies offer evidence that gaps appear between the endothelial cells of the microvasculature, primarily in venules of 20 to 30 micra diameter. It has been postulated that histamine may actually cause contraction of actomyosin-like fibrils within the endothelial cells resulting in their "rounding up," thereby effectively creating a larger than normal intercellular cleft, i.e., pore (42). The concept of histamine causing an increased pore size has recently been challenged by Renkin and co-workers (6, 32, 51, 52). Rather than an increased microvascular pore size, allowing for an increased outward diffusion of proteins, they postulate that histamine is a stimulus for an increased vesicular transport of proteins across the capillary membrane thereby accounting for the increased protein efflux witnessed with histamine administration. Vesicular transport, or pinocytosis, is a process whereby the capillary endothelial cell invaginates.and subsequently engulfs a small portion of plasma. An intracellular, or pinocytotic vesicle is then formed which diffuses 12 to the other side of the cell, where its contents are released into the interstitium. Definitive support for this hypothesis being the major protein transport mechanism stimulated by histamine is still lacking. Controversy surrounds the possibility of capillary hydrostatic pressure contributing to an increased microvascular permeability and, if so, how important a role it may play. Shirley gt a}, (60) have postulated the concept of the "stretched pore phenomenon" based on the fact that investigators have found that increasing capillary hydrostatic pressure leads to an increase in the size of the macromolecules appearing in lymph (36, 44, 60). Haddy §t_al, (23) have shown that mechanical increases in blood flow and small vein pressures for 30 minutes to levels identical with those seen during local histamine infusions results in a slight increase in albumin concentration in lymph while globulin concentration was unchanged. When venous pressure alone was increased for 90 minutes they found slight increases in total con- centration of lymph protein within the first 10 minutes after ele- vation (12). This was attributable to an increased albumin concentra- tion and no change in globulin concentration. The initial increase in lymph total protein may be a result of a washout of protein from the free fluid phase of the interstitial space. Apparently passive stretching of microvasculature pores occurs but its importance seems to be relatively minor based on the available data, especially in the presence of histamine (20). Of the many publications of the effects of histamine on fluid filtration there is surprisingly only a small number concerning the effects of systemically administered (intravenous) histamine on 13 transcapillary fluid exchange (9, ll, 19, 22, 50). In general these studies show that, in contrast to locally administered histamine, systemic administration promotes sustained net reabsorption of extra- vascular fluid. Daugherty gt_§l. (9) noted that in comparing intra-arterial and intravenous infusions of histamine that large increases in hindlimb weight occurred during intra-arterial administration whereas during intravenous infusions, weight was increased only very slightly with the lowest dose employed (20.6 pg base/min) and was reduced with higher doses, falling below control with the highest dose used (82.4 pg base/min). Haddy (22) offered evidence indicating that differences in capillary hydrostatic pressure contributed, at least partially, to the failure of intravenous histamine to increase transcapillary fluid filtration. During local administration of histamine, skin small vein pressure (which represents a minimum for capillary hydrostatic pressure) increases to levels just slightly lower or equal to normal colloid osmotic pressure (22). Hence, net fluid filtration is favored across the vascular bed in question. In contrast, during intravenous infusion of histamine small vein pressures increased slightly with a small dose of 5pg base/min. This may explain the increase in forelimb weight that Daugherty gt_al, observed with their lowest dose. With a higher dose (34pg base/min) small vein pressures decreased well below control suggesting a reduction in microvascular pressure which would favor net extravascular fluid reabsorption. Remington and Baker (50), using changes in specific gravity of blood and plasma as indicators found no evidence for leakage of plasma proteins (indicating an increased capillary permeability) during intravenous infusions of histamine. 14 Deyrup (ll), injecting histamine subcutaneously in the thigh of the dog hindlimb, used changes in plasma volume as an indicator of histamine’s effects on transcapillary fluid flux. She found during the first 30 to 90 minutes after histamine injection (3 to 12 mg base/kg), when blood pressure was markedly reduced, that plasma volume was unchanged or moderately increased and in a few cases slightly reduced. There was no evidence for increased capillary permeability since the escape of the albumin-bound dye T-l824 from the vasculature did not increase significantly. In a study performed by Grega gt a1, (19) using isolated, naturally perfused, innervated canine forelimbs, it was found that‘ instead of the typical large increases in forelimb weight seen with the medium to high doses of locally infused histamine (20-60 pg base/ min) (20), there was a persistent dose-dependent (20-800 pg base/min) weight loss of a magnitude that could not be accounted for solely by decreased intravascular volume when infused intravenously. This was best demonstrated by the fact that during the time period of the greatest fbrelimb weight loss, the total forelimb small vessel and large vein (capacitance vessels) resistances were not changing. This implies that vessel caliber and consequently intravascular blood volume was constant during this time. Hence, extravascular fluid reabsorption must have occurred to account for the observed weight loses. Most likely this was a result of a decreased transmural capillary hydrosta- tic pressure gradient. This is suggested because all vascular pressures, including small vein pressures, and forelimb blood flow fell markedly below control. Unfortunately flow rate and total protein concentration of lymph were not measured in any of the systemic studies. 15 Thus only speculations can be made concerning any permeability changes and consequently the transmural capillary colloid osmotic pressure gradient. If systemically administered histamine does increase micro- vascular permeability and hence decrease transmural colloid osmotic pressure gradients, it must be effectively counteracted by an even greater fall in the transmural capillary hydrostatic pressure gradient. Local histamine increases permeability sufficiently to increase fore- limb weight significantly even when capillary hydrostatic pressure (Pc) is mechanically prevented from increasing above normal plasma colloid osmotic pressure (20). In this situation, fluid filtration still occurs when perfusion pressure is kept at or below normal plasma colloid osmotic pressure. Capillary hydrostatic pressure then must certainly have been well below normal plasma colloid osmotic pressure. Since locally administered histamine increases microvascular permeability and subsequently causes net fluid filtration even when perfusion pressure and capillary hydrostatic pressure are exceedingly low, it is perplexing why fluid filtration does not occur during systemic administration of histamine when perfusion pressure, blood flow and calculated blood concentrations (assuming no degradation) are at least equal or exceed the minimal dosage that produces fluid filtration when infused locally (see Appendix A for calculations). There are, however, several differences that may account for the route- dependent differential actions of histamine on transvascular fluid flux. First of all, during local infusions of histamine, capillary hydrosta- tic pressure (Pc) increases, whereas Pc decreases during systemic infusions of histamine (19, 20). Also during systemic infusions, histamine first passes through the pulmonary circuit before reaching 16 the systemic capillary beds. This passage through the pulmonary circuit may result in the uptake, metabolism, or biotransformation of histamine by the lungs. During local administration, histamine first passes through the microvasculature and then, upon recirculation, enters the lung. The transit time from the point of administration to the microvessels is also greater during systemic administration than during local intra-arterial infusions. This added time may possibly allow destruction of histamine by factors within the blood. Additionally, systemic pressure is little affected during local administration of histamine or falls only slightly after edema has already developed. In contrast, during systemic administration there is an immediate precipitous and sustained fall in systemic arterial blood pressure. Hence, the latter response is most likely associated with a marked sympathoadrenal discharge subsequent to the hypotension. It is con- ceivable that substances liberated from the sympathoadrenal discharge such as the glucocorticoids and catecholamines, effectively antagonize the usual edemogenic action of histamine seen with local histamine administration. Schayer proposed that there exists a balance between catecholamines and histamine which subsequently forms a component of circulatory homeostasis in stressful conditions (57). Hence, the catecholamines are very likely candidates. Evidence for adrenal gland discharge subsequent to histamine administration is offered by Weiss gt_§l, (65). While studying the effects on blood pressure in normal human subjects they observed that upon termination of a prolonged histamine infusion (several hours) of 25 to 50 pg base/min that there was a significant increase in systemic blood pressure above normal control levels. Also, skin (such as of 17 the face) that was previously flushed appeared very pale. They con— tributed these opposite responses to "antagonistic substances" and/or "vasomotor reflexes" having been developed which are antagonistic to histamine. Their hypothetical substance and vasomotor reflex had actions very similar to the catecholamines, norepinephrine and epine— phrine which are released from sympathetic nerve fibers and the adrenal gland, respectively. Further evidence for an adrenal gland discharge subsequent to histamine administration is offered by Roth and Kvale who used histamine as a test for pheochromocytoma (56). The intravenous injections of histamine into patients suspected of having the disease resulted in marked increases in blood pressure which was identical to the typical paroxysmal hypertension (a result of hyper- secretion of adrenal catecholamines) often seen in such patients. Furthermore, Robinson and Jochim (54) found that during histamine induced hypotension from intravenous administration that there was marked increases in blood levels of catecholamines (from a control of 0.035 pg/kg/min up to 0.25 pg/kg/min), leaving the adrenal glands via the lumboadrenal vein in dogs. Hakfelt, using rats, found that the suprarenal content of'norepinephrine was significantly reduced following subcutaneous injections of histamine (30). It has also been shown that adrenalectomized rats and mice (26, 27, 43) are rendered hypersensitive to histamine. This hypersensitivity (measured by comparisons of mortality rates to dose levels between normal and adrenalectomized animals) is at least partially reversible when epinephrine is given prior to histamine administration (27). The primary stimulus for catecholamine release from the adrenal gland appears to be the hypotension produced regardless of the 18 causative mechanism. Rosenberg §£_al, (55) produced hypotension by two different methods in dogs. Within 5 minutes after an intravenous injection of endotoxin (7.5 mg/kg) they found venous plasma epinephrine concentration had increased to 20pg/l. A 30—fold increase above control. Norepinephrine increased to about 17 pg/l representing a 9-fold increase above control. Hemorrhagic hypotension, produced by an acute loss of 36% of total blood volume, within 5 minutes resulted in epinephrine increasing to 35 pg/l and norepinephrine to 14 pg/l, a 70 and 5 fold increase above control, respectively. The discrepancy between concentrations achieved by these two different mechanisms are most likely a result of the arterial blood pressure values. Within 5 minutes after endotoxin administration, arterial blood pressure fell to about 80 mm Hg whereas during hemorrhage it fell to about 50 mm Hg. Furthermore, this hypotension induced release of catecholamines appears to function via a neurogenic mechanism as demonstrated by Nykiel and Glaviano (48) and Egdahl (13) who observed no change in adrenal cate- cholamine output following intravenous administration of sublethal doses of endotoxin in dogs with their splanchnic nerves sectioned or spinal cord transected between C-7 and T—l, respectively. Without question, the catecholamines are released during hypotension, however, much disagreement exists within the literature concerning the catecholamines effects upon the various vasculature sites and subsequently transvascular fluid flux. In general, the cateCholamines are believed to be vasoconstrictors of both resistance and capacitance vessels except for epinephrine which in low concentra- tions is said to dilate the precapillary vessels of skeletal muscle (18). In studies of naturally perfused forelimbs and ileum segments 19 (14, 24) small vein pressures are sometimes seen to rise, sometimes fall, or remain unchanged in response to norepinephrine. The organ weight (forelimbs or ileum segment) was found to vary directly with small vein pressure in these studies. This association between small vein pressure and consequently organ weight can be best explained in terms of varied effects upon the pre/post capillary resistance ratio. If the venous segment of the vascular bed constricts more than the arterial segment, capillary outflow would be impeded and hence capillary hydrostatic pressure will increase, thereby favoring an increase in filtration and consequently an increased organ weight. If the arterial segment was constricted proportionately more than the venous segment, then capillary inflow would be impeded, thus capillary hydrostatic pressure would decrease, thereby favoring net fluid re- absorption which would account for the observed weight decreases. Kaiser and Diana (34) found no significant alteration in isogravimetric capillary pressure nor the capillary filtration coeffi- cient during infusions of norepinephrine (0.9 to 2.9 pg base/min/kg tissue) under both constant flow and constant perfusion pressure con- ditions in isolated dog hindlimbs. Mellander and Nordenfelt (46) studying the human hand (skin) and calf (skeletal muscle) also found no significant effects upon the capillary filtration coefficient by norepinephrine administered intravenously at dosages of 0.05 to 0.3 pg base/kg/min for 30 minutes. The results of these two studies indicate that both capillary surface area available for diffusion (determined by the precapillary sphincters) and capillary permeability were uneffected by norepinephrine. However, Jfirhult (31) has reported that under constant perfusion pressure conditions, in denervated 20 skeletal muscle of the lower leg of cat hindlimbs, that norepinephrine (mean dosage of 2.7 pg base/kg/min) increases the capillary filtration coefficient indicating an increased capillary surface area available for diffusion. In contrast, Appelgren and Lewis (1) reported a decreased capillary permeability-surface area product (PS) in naturally perfused human skeletal muscle when solutions of 0.04 mg/ml norepine- phrine were injected locally. Uptake of 86Rb, used as an index of capillary surface area available for diffusion, has been shown to increase (indicative of an increased surface area) during constant inflow conditions in isolated dog forelimbs during intra-arterial infusion of 0.015 to 0.03 pg epinephrine/min (2) and in isolated dog hindlimbs following an intra- arterial injection of 2 pg epinephrine or norepinephrine (16). However, in an isolated, pump perfused, canine gracilis muscle pre- 86Rb paration, Szwed and Freedman (63) have reported a decreased in uptake reflecting a decreased capillary surface area available for diffusion during intra-arterial infusion of 0.0012 to 0.06 pg/min/gram of tissue of epinephrine or norepinephrine. STATEMENT OF THE PROBLEM The objective of this study was to attempt to determine the mechanism(s) of the route-dependent differential actions of histamine on forelimb transvascular fluid flux. Local (intra-arterial) infusions of histamine (4 to 64pg base/min) into the forelimb increase transvascular fluid flux and forelimb weight producing massive visible edema with the medium to high infusion rates (l6-64pg base/min). The increased fluid flux and consequently edema is attributable to both a rise in the transmural capillary hydrostatic pressure gradient (due to arteriolar vasodilation) and a fall in the transmural colloid osmotic pressure gradient (due to an increased microvascular permeability to plasma proteins). In contrast, systemically (intravenously) infused histamine at rates that produce histamine blood concentrations cal- culated to exceed those achieved by local infusion, results in sus- tained net extravascular fluid reabsorption. Since during intravenous infusions, histamine must first pass through the pulmonary circuit before reaching the systemic micro- vasculature, the possibility exists that the lungs may remove, destroy or inactivate histamine. It is also possible that since the transit time during systemic administration is much greater than during local infusions, that factors within the blood may destroy or inactivate histamine before it reaches the forelimb microvasculature. 21 22 During local infusions of histamine (3 to 64pg base/min) systemic arterial blood pressure is minimally affected or falls only slightly after edema has already deve10ped. In contrast, there is an immediate and sustained fall in aortic pressure during systemic administration of histamine which is, at least initially, dose related (20-800pg base/min). This marked reduction in systemic blood pressure is undoubtedly associated with a sympathoadrenal discharge. Hence, it is conceivable that substances released subsequent to the hypotension (e.g., catecholamines) effectively antagonize the usual edemogenic actions of histamine. This antagonism may be a result of a direct blockade of histamine's permeability increasing effects upon the microvasculature, or a shunting of blood flow to non-nutritional channels or a combination of both. These possibilities will be systematically evaluated to deter— mine if and if so, to what extent, they may affect the route- dependent differential actions of histamine. METHODS Mongrel dogs weighing approximately 18 kilograms were anesthetized with sodium pentobarbital (30mg/kg). Normal saline was administered at 15ml/kg to assure good hydration and ample time was allowed for equilibration. Blood clotting was prevented by adminis- tering 10,000 U.S.P. units of Sodium Heparin. Small incisions were made in the right forelimb over the brachial artery, cephalic vein (above the elbow), second superficial dorsal metacarpal vein in the paw and also over the femoral triangle. In all experiments a lymph vessel in the area of the cephalic vein abbve the elbow was isolated and cannulated. The lymph vessels in this area drain primarily the forelimb skin and paw (47). As many vessels as could be found (usually 2 or 3) were tied off and one of them was then cannulated distally with PE-lO polyethylene tubing about 10cm. in length and beveled at the cannulating end. Initial puncture of the lymph vessel was accomplished by using a 23 gauge hypodermic needle. Lymph was collected over 10 minute periods in miniature graduated cylinders (about 0.3m1 capacity) constructed from narrow plastic pipettes to minimize evaporation. Lymph total protein concentration was measured by the spectrOphotometric method of Waddel (64) on a Beckman DB spectrophotometer (Model 24, Beckman Instruments, Inc., Fullerton, California). 23 24 In all experiments the femoral vein was cannulated to allow administration of saline, heparin and nembutal as needed. Pressures in skin small veins were obtained by cannulating upstream one of the small surface veins on the dorsal side of the paw with PE-60 poly- ethylene tubing. All pressures were measured with Statham pressure transducers (Model P23Gb, Statham Instruments, Inc., Oknard, California). connected to a direct writing Sanborn oscillograph (7700 series, Hewlett Packard Co., Palo Alto, California). Drugs utilized were histamine diphosphate, levarterenol- bitartrate (norepinephrine) and acetycholine hydrochloride in solutions of isotonic saline which were administered with a Harvard Apparatus‘ infusion/withdrawal pump (Harvard Apparatus Co., Inc., Millis, Maryland). The volume delivery rate of the infusate was either 0.2cc/min or 0.4 cc/min depending on the dose being used. In those experiments utilizing a naturally perfused forelimb, a small side branch of the brachial artery above the level of the elbow was isolated and cannulated upstream with PE-SO polyethylene tubing for local (intra-arterial) infusion of drugs. In cases where both hista- mine and norepinephrine were simultaneously infused, two small side branches were isolated and cannulated. Systemic arterial blood pressure was measured by a cannula inserted upstream through the femoral artery into the descending aorta. In experiments utilizing a forelimb perfused at constant inflow, the brachial artery was isolated, tied off and transected about 2 inches above the level of the elbow. Blood was obtained from a cannula in the femoral artery and pumped at.constant but controllable flow into the transected brachial artery by a Sigmamotor pump 25 (Model T68H, Sigmamotor Inc., Middleport, New York). Perfusion pressure was monitored by cannulating a small side branch of the brachial artery distal to the site of transection. Systemic arterial blood pressure was measured by inserting a cannula upstream through the brachial artery, proximal to the transection, to approximately the subclavian artery. Local (intra—arterial) administration of drugs in these experiments was achieved by infusion into the pump circuit behind the Sigmamotor pump. In four series (the;number of animals per series is reported in the tables) of experiments, a PE-240 polyethylene tubing was inserted down the right common carotid artery into the left ventricle of the heart. The catheter was initially connected to a pressure transducer and successful placement was confirmed by a typical left ventricular pressure tracing. Drug administration was then achieved by infusion into this catheter. In three series of experiments, systemic drug administration was accomplished by infusion into the femoral vein. In these experi- ments the saphenous vein of one of the hindlimbs was isolated and cannulated in case supplemental nembutal was necessary after beginning intravenous drug infusion into the femoral vein. In all experiments, arterial blood samples of about 3ml were withdrawn from the cannula used to obtain aortic pressure. A sample was taken during the control period and at 30 minute intervals during the experiments. Hematocrits and total plasma protein concentration 0 in grams 6 were determined from these samples. In three series of experiments, histamine (4 and 64pg base/min) was infused locally for 60 minutes into forelimbs perfused at constant 26 inflow during or following acetylcholine or hemorrhagic induced hypo- tension. Acetylcholine hypotension was produced by intravenous infusions of dosages necessary to lower and maintain aortic pressure at or near 60 mmHg for 60 minutes. The dosage at 60 minutes was then continued for the remainder of the experiment (total duration 120 minutes). Hemorrhagic hypotension was produced by removing the necessary quantity of blood (through a PE-360 polyethylene catheter inserted into the femoral artery) to lower and maintain aortic pressure near 40 mmHg for 60 minutes. Vascular pressures, lymph flow rate, plasma and lymph total protein concentrations and hematocrits were obtained as described above during the control period, hypotensive period, and the drug infusion period during the last hour of the 120 minute experimental period . In five series weight and hemodynamic measurements were made in collateral-free, innervated canine forelimbs. The surgical pro- cedure consisted of sectioning the skin circumferentially about 1-2 inches above the elbow of the right forelimb. The forelimb nerves (median, ulnar, radial, and musculocutaneous), brachial artery, and brachial and cephalic veins were isolated and coated with an inert silicone spray to prevent drying. The muscles and connective tissue in this area were then sectioned by electrocautery. The humerus was cut with a bone saw and the ends of the marrow cavity were packed tightly with bone wax to prevent bleeding. At this point blood entered the forelimb only through the brachial artery and.exited only via the brachial and cephalic veins. The brachial and cephalic veins were partially transected at about the level of the elbow. They were then cannulated with short sections of PE-320 polyethylene 27 tubing. The sections of tubing were about 6 inches in total length with a 90° bend at about the 5 1/2 inch mark. The short 1/2 inch section of the bent tubing was then inserted into the veins. The cannulas were secured such that the outflow tips were at the same level as the veins. The venous outflow was directed into a reservoir and returned the dog via the jugular vein by a variable speed Holter pump (Model RE161, Extracorporeal Medical Specialties, King of Prussia, Pennsylvania) at a rate that kept reservoir volume constant. Blood flow was determined by timed collections of the venous outflows into graduated cylinders and calculated as ml of flow/min/lOO grams of forelimb tissue. The median cubital vein (the major anastomosis between the brachial and cephalic veins) was tied off in these experiments. This provided a more accurate mean for comparing resistance changes between muscle and skin vascular beds since the cephalic vein drains primarily skin and the brachial vein primarily muscle (47). The brachial and cephalic venous pressures were measured by inserting PE-60 polyethylene tubing through side branches found at a level of about 1-2 inches below the elbow. Aortic, perfusion (constant inflow conditions) and skin small vein pressures were all obtained by the methods described above. When surgery was completed, the forelimb was suspended on a wire mesh tray that was attached to a sensitive strain gauge I-beam balance. The balance was wired to the Sanborn oscillograph thereby allowing changes in forelimb weight to be continuously monitored throughout the experiment. This system was calibrated by the addition of a 2 gram weight to the center of the isolated forelimb which produced a pen deflection of about 10-22 mm. Total skin and muscle vascular 28 resistances were calculated by dividing cephalic or brachial blood flows into their respective pressure gradients. The data was analyzed by Analysis of Variance utilizing a Randomized Complete Block Design (61). All means were compared to the 0 minute control mean and significant difference was determined for P equal to or less than both 0.05 and 0.01 by the Least Significant Difference Test (61). When applicable, means were compared back to minute 60 and control. RESULTS Table 1 In naturally perfused forelimbs intravenously infused histamine (400 + 800pg base/min) produced a continuous decrease in forelimb weight. Aortic pressure and forelimb cephalic and brachial vein pressures and outflows were all markedly reduced. Total skin resistance was only minimally affected being statistically greater than control' only at minutes 30, 35, and 90. Total muscle resistance was signifi- cantly increased throughout the infusion period. The local infusion of histamine (64pg base/min) produced no further alterations in any of the measured parameters. In forelimbs perfused at constant inflow there was a rapid marked increase in forelimb weight within 5 minutes. The peak of the weight increase was reached within 15 minutes and remained elevated throughout the infusion period. Aortic pressure fell markedly whereas perfusion and cephalic vein pressures were only modestly reduced, the latter declining slowly with time. Brachial vein pressure was un- effected whereas skin small vein pressure was moderately increased throughout the experiment. No effect was observed upon cephalic venous outflow but a slight reduction was seen in brachial venous outflow during the first 35 minutes. There was no appreciable effect on total skin resistance. Total muscle resistance increased slightly 29 30 during the first 15 minutes, returned to control, and was then signifi- cantly reduced from 60 minutes on. Local histamine, as in naturally perfused forelimbs, produced no further changes in any of the para- meters measured. Table 2 In naturally perfused forelimbs histamine (400 + 800pg base/min) infused either intravenously or into the left ventricle markedly reduced aortic pressure and either failed to alter or only slightly decreased skin small vein pressure. Lymph flow rate and lymph total protein concentration were moderately increased. Plasma protein con: centration was not changed and hematocrits were greatly elevated. The simultaneous local infusion of histamine (64pg base/min,I.A.) during the last 30 minutes of systemic infusion produced no further alterations in any of the measured parameters relative to minute 60. In forelimbs perfused at constant inflow the systemic infusions of histamine either intravenously or into the left ventricle resulted in a moderate reduction of perfusion pressure. All other responses were similar to those seen in naturally perfused forelimbs except skin small vein pressure was increased and the rise in lymph flow rate and lymph total protein concentration was markedly greater. The simultaneous local histamine infusion produced no further alterations as in naturally perfused forelimbs. Table 3 In forelimbs perfused either at natural or constant inflow the simultaneous infusion of histamine (400pg base/min) into the left ventricle and locally (64p base/min,I.At initiated 3 minutes after 31 beginning systemic infusion) markedly reduced aortic pressure, produced no significant change in perfusion pressure and plasma protein concen- tration and increased hematocrits. Skin small vein pressure was signi- ficantly reduced in naturally perfused forelimbs and was markedly increased in forelimbs perfused at constant inflow. In naturally perfused forelimbs, lymph flow and lymph total protein concentration were unaltered whereas in forelimbs perfused at constant inflow they were significantly increased. Table 4 Hypotension induced by hemorrhage resulted in lower aortic pressures than that produced by acetylcholine infusion. Acetylcholine hypotension produced no effect upon perfusion pressure, skin small vein pressure, lymph flow or lymph and plasma total protein. The simultane- ous infusion of local histamine (4pg base/min,I.A.) initiated at minute 60 failed to produce any further alterations of aortic pressure, skin small vein pressure, plasma protein and hematocrits. It did, however, produce a significant reduction in perfusion pressure relative to control and minute 60. Lymph flow rate and lymph protein concen- tration were slightly increased; however, the latter was not signifi- cantly elevated until the last 20 minutes of histamine infusion. Hemorrhagic hypotension produced a marked increase in perfusion pressure, either a slight reduction or no change in skin small vein pressure and a slight but not significant increase in lymph flow rate. Lymph total protein was either unchanged or modestly increased whereas plasma protein concentration was either unchanged or slighly reduced. Local histamine (4pg base/min) produced no further change in aortic 32 pressure, a moderate reduction in perfusion pressure and a slight increase in skin small vein pressure relative to minute 60. Although skin small vein pressure was increased by local histamine it still remained below control. No further alterations were observed in lymph flow rate or lymph and plasma protein concentration. The high dose of histamine (64pg base/min) produced a significant reduction relative to minute 60 in aortic and perfusion pressure. Skin small vein pressure was increased relative to minute 60 and was also significantly greater than control. Lymph flow rate was moderately increased being significantly greater than both control and minute 60. No further changes were observed in lymph or plasma total protein concentration.- In all three series the hematocrits were significantly elevated with no further change produced by either dose of histamine. Table 5 Table 5 shows the effects of histamine alone (4pg base/min, I.A.) and in combination with norepinephrine (4pg base/min, I.A. of each) in naturally perfused forelimbs on forelimb weight, vascular pressures and resistances and blood flows. The effects of histamine alone and in combination with norepinephrine produced no effect upon aortic pressure and markedly increased skin small vein pressure. Histamine alone produced a continuous increase in forelimb weight throughout the infusion period. Venous pressures and outflows were all increased by histamine alone whereas both skin and muscle total resistances were reduced. The combination of histamine and norepinephrine in contrast, produced an initial fall in forelimb weight during the first 5 minutes 33 which then slowly increased to a value slightly above control at the end of the infusion period. Cephalic vein pressure and outflow and brachial venous outflow were all markedly reduced whereas brachial venous pressure was significantly increased. Both skin and muscle total resistances were greatly elevated throughout the infusion period. Table 6 Table 6 shows the effects of histamine and norepinephrine (4pg base/min,I.A. of each) in forelimbs perfused at constant inflow on forelimb weight, vascular pressures, resistances and blood flows. Aortic pressure was significantly increased during the first 30 minutes whereas perfusion and skin small vein pressures remained markedly ele- vated throughout the entire 60 minute infusion period. Forelimb weight was significantly increased by 15 minutes and then continued to rise slowly throughout the remainder of the infusion period. Cephalic and brachial venous pressures were both significantly increased initially but then returned towards control with cephalic pressure returning slowly with time and brachial pressure returning abruptly. Both cephalic and brachial venous outflows were only minimally affected being slightly reduced when statistically significant. Total skin and muscle resistances were increased and fairly well maintained throughout the infusion.period. Table 7 In naturally perfused forelimbs, local histamine (4pg base/ min,I.A.) alone or in combination with norepinephrine (4pg base/min, I.A.) produced no effect upon aortic Pressure and markedly increased skin small vein pressure. Histamine alone caused marked increases in 34 lymph flow and lymph protein concentration but with the simultaneous infusion of norepinephrine no change was observed in either of these parameters. During the simultaneous infusion of histamine and nore- pinephrine no change in plasma protein concentration and a slight increase in arterial hematocrit was observed. These parameters were not monitored for histamine infusion along. In forelimbs perfused at constant inflow, histamine (4pg base/min,I.A.) failed to alter aortic or skin small vein pressure and significantly reduced perfusion pressure. Histamine infused simultane- ously with norepinephrine (4pg base/min,I.A. of each) or norepinephrine alone (4pg base/min) produced marked increases in these vascular pressures. Histamine alone greatly increased lymph flow and lymph total protein concentration but failed to alter plasma protein concen- tration or arterial hematocrit. Only a slight increase in lymph flow and no change in lymph or plasma total protein concentration was observed during the combination infusion. Norepinephrine alone did not alter lymph flow rate or plasma total protein concentration but produced a slight decrease in lymph total protein concentration during the last 30 minutes of infusion. Arterial hematocrits were slightly elevated during the infusion of norepinephrine. In naturally perfused forelimbs the local infusion of the high dose of histamine (64pg base/min,I.A.) alone or in a combination with norepinephrine (4pg base/min,I.A.) resulted in decreased aortic pressures and increased skin small vein pressures. The magnitude of the increased small vein pressure was much lower during the combina- tion infusion. Histamine alone produced marked increases in both flow rate and total protein concentration of lymph. Plasma proteins and 35 arterial hematocrits were not monitored in this series. The simultane- ous infusion of norepinephrine with histamine prevented the large increases in flow and protein concentration of lymph. Only slight increases were seen relative to control. The combination infusion produced no effect upon plasma protein concentration but significantly increased arterial hematocrits. In forelimbs perfused at constant inflow the high dose of histamine (64pg base/min, I.A.) significantly reduced both aortic and perfusion pressures but failed to alter skin small vein pressure. Lymph flow and lymph total protein concentration were markedly increased throughout the infusion period. The simultaneous infusion of nore- pinephrine (4pg base/min,I.A.) with the high dose of histamine failed to prevent the fall in aortic pressure but caused significant increases in both perfusion and skin small vein pressure. There was a large increase in lymph flow rate throughout the infusion period being much more pronounced than with histamine alone. Lymph total protein con- centration was increased throughout the infusion period with values being similar to those seen during histamine infusion alone. However, the change from control was much less. The simultaneous infusion of norepinephrine (16pg base/min,I.A.) with the high dose of histamine produced significant increases in aortic, perfusion and skin small vein pressures, the latter two being profound. Lymph flow rate was markedly increased but the magnitude was not nearly as great as that seen with the low dose of norepinephrine infused simultaneously with the high dose of histamine. The values were similar to those seen during infusion of the high dose histamine alone but the rate of increase was much greater with the combination infusion. The combination of the 36 high dose of norepinephrine with histamine attenuated the increase in lymph total protein concentration seen with both the combination of the low dose of norepinephrine with histamine and histamine alone. *In all three series no change was observed in plasma total protein concentration except for histamine alone which caused a slight decrease at minute 60. Arterial hematocrits were significantly increased in all cases. In comparing the high to low dose of histamine alone in both natural and constant flow conditions it is apparent that the magnitude of the increased lymph flow is dose dependent (being directly related) and is markedly greater in naturally perfused forelimbs. 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Systemic administration of histamine (400 to 800pg base/min, I.V.) in naturally perfused fore- limbs produces a continuous decine in forelimb weight throughout the infusion period (Table 1). In contrast, during the local infusion of histamine (4pg base/min, I.A.) in naturally perfused forelimbs, there is a continuous increase in forelimb weight (Table 5). Furthermore, systemic histamine (infused either intravenously or into the left heart) produced only minimal increases in lymph flow and total protein concentration (Table 2) when compared to local histamine (Table 7). These differences occur despite the fact that the calculated blood concentration of histamine during systemic infusion (assuming no degradation) greatly exceeds or is at least equal to that which produces profound effects on the above parameters when infused locally. There is the possibility that the lungs destroy or inactivate histamine, thereby accounting for the route-dependent differential action of histamine by drastically reducing the effective concentration at the site of forelimb microvasculature. Such a function being served by the lungs is disproved by comparison of systemic histamine infusions into the vena cava which first must pass through the 50 51 pulmonary circuit before entering the general circulation, and into the left ventricle of the heart whereby histamine bypasses the pulmonary circuit (Table 2). There is no discernable difference between either route of systemic histamine administration upon any of the measured parameters. There still existed the possibility that histamine may be inactivated by factors within the arterial blood before reaching the studied vascular bed. This possibility was ruled out since, in fore— limbs perfused at constant inflow the local infusion of histamine (4 and 64pg base/min, I.A.) produced marked increases in flow rate and total protein concentration of lymph. The increases in these two parameters is attributable to an increased microvascular permeability since they occurred in the face of unaltered skin small vein pressure and slightly reduced perfusion pressure (a minimum and maximum for capillary hydrostatic pressure respectively). During constant inflow the locally infused histamine must travel through about three feet of polyethylene tubing before reaching the forelimb. This distance is as much or more than histamine would travel during left ventricular infusion. Consequently, if factors within the blood were destroying histamine then one would not expect to see such large increases in flow and total protein concentration of lymph during local infusions at constant inflow. This should not imply that histamine destruction does not occur to some extent. It does however, imply that whatever the degradation rate may be, it is not sufficient to account for the route- dependent differential actions of histamine. In naturally perfused forelimbs the systemic administration of histamine caused marked reductions in forelimb blood flow, either no effect or slight reductions in skin small vein pressure and increased 52 total resistances in the skin and muscle vascular beds. Consequently, the observed forelimb weight loss can be attributed to both a decreased intravascular blood volume (inferred from increased resistances) and to a reduced capillary hydrostatic pressure (inferred from flow and skin small vein pressure decreases) which would favor tissue fluid re- absorption. However, these hemodynamic changes alone cannot solely account for the failure of histamine to produce marked increases in forelimb weight and flow rate and total protein concentration of lymph as seen during local administration. This conclusion is drawn from three different observations. First of all, Grega et_al, (20) demonstrated that in forelimbs perfused at constant inflow at a pressure at or below normal colloid osmotic pressure (thereby greatly favoring tissue fluid reabsorption) that histamine (60pg base/min, I.A.) produced a weight gain of 19 grams within 10 minutes. This indicated that histamine acts, at least partially via a pressure inde- pendent mechanism, i.e., an increase in capillary permeability. Secondly, the fact that histamine is present in the naturally perfused forelimbs during systemic administration is evidenced by the slight increases in lymph flow rate and total protein concentration seen. This occurs in spite of a reduced capillary hydrostatic pressure inferred from the marked fall in skin small vein pressure. Hence, these observed increases must be due to a slight increase in micro— vascular permeability. In contrast to the local infusion of histamine in naturally perfused forelimbs, the increased flow and total protein concentration of lymph are quite small during systemic infusion com- pared to the lowest dose of histamine (4pg base/min, I.A.) infused locally. This attenuation is not likely due to a proportionally 53 decreased histamine blood concentration in the forelimb, since during systemic administration it is calculated to be higher or at least equal to the concentration during local histamine infusion (assuming even distribution of the infused histamine) even though forelimb blood flow is reduced. Thirdly, during systemic infusions of histamine (400 to 800pg base/min) into forelimbs perfused at constant inflow (Table l and 2) the increase in forelimb weight, lymph flow and lymph total protein concentration is actually less than that produced by equal concentrations of histamine infused locally (64pg gase/min) in fore- limbs perfused at constant flow. These changes during systemic administration occur in the presence of an increased microvascular pressure (inferred from the increased skin small vein pressure) whereas microvascular pressure is unchanged relative to control during the local infusion of histamine. Based on these observations, one would expect substantially greater increases in these three parameters during systemic administration. The above observed differences between local and systemic histamine infhsions on weight and lymph hint to the possibility that the effects of histamine during systemic infusion are being antagonized at the site of the microvessels by some endogenous factor. Support of this possibility is offered by the fact that after 60 minutes of systemic histamine infusion (400 to 800pg base/min) the initiation of a 30 minute local infusion of histamine (64pg base/min, I.A.) produces no further alterations in any of the observed parameters (Table 2). One possible explanation for this is that the responsiveness of the microvasculature to histamine diminished with time, thereby making the local infusion of histamine ineffective. This possibility is ruled out 54 though since systemic infusion of histamine (400ug base/min) into the left heart simultaneously with local histamine (64pg base/min, I.A.) in which, after a delay of only three minutes, the local histamine infusion failed to produce any major differences in results when com- pared to the other systemic studies (Table 3). From this it is obvious that some factor(s) are present that effectively antagonize the local actions of histamine upon the microvascular. One readily observable difference between systemic and local infusions of histamine is the effect upon systemic arterial blood pressure. During local histamine infusions (4 or 64pg base/min, I.A.) there is either no effect or only a moderate decrease in aortic pressure. The latter occurring with the high dose of histamine. In contrast, during systemic infusions of histamine (400 to 800pg base/min) there is a large precipitous fall in aortic pressure. This hypotension would elicit a generalized sympathoadrenal discharge which may effectively antagonize the effects of histamine via catecholamines and/or other substances released in response to hypotension. To test the hypothesis that endogenous catecholamine release, subsequent to hypotension, may antagonize the microvasculature actions of histamine, three series of experiments were performed in which hypotension was produced by means other than histamine (Table 4). Hypotension was produced by both hemorrhage and acetylcholine for 60 minutes prior to initiation of a local infusion of histamine (4 or 64pg base/min, I.A.) into forelimbs perfused at constant inflow. Following 60 minutes of hemorrhagic induced hypotension, the local infusion of histamine (4pg base/min, I.A.) for 60 minutes failed to produce any further alterations in lymph flow or total protein concentration. The increased perfusion SS pressure prior to initiating the local histamine infusion is indicative of a sympathoadrenal discharge which would tend to compensate for the loss of blood and blood pressure. In contrast, the high dose of histamine (64pg base/min, I.A.) produces moderate increases in lymph flow rate, but even in the face of an increased microvasculature pressure (inferred from the increased skin small vein pressure) this increase was greatly attenuated compared to local infusion of the same dose of histamine alone in forelimbs perfused at constant inflow. No further change was observed in lymph total protein concentration in spite of the increased lymph flow rate thereby indicating that the local action of histamine on the microvasculature was effectively antagonized. Acetylcholine hypotension did not produce as severe a reduction in aortic pressure as hemorrhage did. The local infusion of histamine (4pg base/min, I.A.) following 60 minutes of acetylcholine hypotension produced only a very slight increase in lymph flow rate and essentially no change in lymph total protein concentration. Hence, by comparison of all three types of hypotension and both doses of local histamine, it appears that the same antagonist is present (i.e., the catecholamines) regardless of how hypotension is produced and that it antagonizes histamines microvascular actions. To test the hypothesis that the catecholamines are antagonistic to histamine's action on the microvasculature, experiments were per— formed to compare differences in forelimb weight changes, hemodynamics, lymph flow and lymph total protein concentration between local histamine infusions alone and in combination with norepinephrine. The doses of histamine employed were 4 and 64pg base/min, the former being a more physiological dose. The dosage of norepinephrine employed S6 was 4ug base/min except in one series where 16ug base/min was infused. In naturally perfused forelimbs, histamine (4pg base/min, I.A.) infused locally produced a continuous increase in forelimb weight, reaching 44 grams by minute 60 (Table 5). Forelimb vascular pressures and blood flows were all significantly increased and forelimb resis- tances decreased throughout the infusion period. Lymph flow rate and lymph total protein concentration were both markedly increased within the first 10 minutes of infusion and continued to rise modestly or remained fairly constant throughout the infusion period (Table 7). In one series of experiments norepinephinre (4pg base/min, I.A.) was infused into forelimbs perfused at constant inflow. There was no observable effect upon lymph flow rate and a slight decrease in lymph total protein concentration during the last 30 minutes of infusion (Table 7). The simultaneous infusion of norepinephrine with histamine (4pg base/min, I.A. of each) into naturally perfused forelimbs, in contrast to histamine alone, produced an initial decline in forelimb weight which then slowly increased reaching +5 grams by minute 60. There was no change in lymph flow or lymph protein concentration. Thus, the effects of local histamine on forelimb weight, lymph flow and lymph total protein concentration were effectively antagonized by norepinephrine. To determine possible contributions of reduced forelimb blood flow per se during the simultaneous infusions of histamine and nore- pinephrine into naturally perfused forelimbs, these infusions were repeated in ferelimbs perfused at constant inflow (Tables 6 and 7). 57 An increase in forelimb weight was observed, reaching 15 grams by minute 60, and only slight increases in lymph flow rate which was significantly less than that seen during histamine infusion alone. Lymph total protein concentration did not change. Hence, the increase in forelimb weight and lymph flow must have been a result of an increased fluid filtration due to an increased microvascular hydrosta- tic pressure (inferred from the increased skin small vein pressure). Since forelimb inflow is held constant, and there was no significant shunting of blood flow between skin and muscle, the increased micro— vascular pressure can be attributed to a venoconstriction (skin large vein resistance markedly increases) produced by norepinephrine. This data illustrates that the simultaneous infusion of norepinephrine with histamine (4pg base/min, I.A. of each) completely prevents the normal histamine effect upon the microvessels, i.e., histamine failed to produce an increase in microvascular permeability (inferred from the failure of lymph protein concentration to change). The local infusion of the high dose of histamine (64pg base/ min, I.A.) produces drastic increases in both lymph flow rate and lymph total protein concentration. The latter approaching plasma protein values. These effects are largely pressure independent since at constant inflow microvascular pressure remains constant (inferred from the unaltered skin small vein pressure) but lymph total protein concentration still increases (relative to control) to about the same values as seen during natural flow. The simultaneous infusion of norepinephrine (4pg base/min, I.A.) with histamine (64pg base/min, I.A.) in naturally perfused forelimbs appeared to produce a slight increase in lymph flow rate 58 (although not statistically significant) and lymph total protein con— centration. These dosages in forelimbs perfused at constant inflow produced an increased lymph flow rate greater than that produced by histamine alone at constant inflow and similar to that seen at natural inflow. This large increase occurred however, in the face of an increased microvascular pressure (inferred from the increased skin small vein pressure) substantially greater than that seen in natural flow. Hence, much of this increase can be attributed to fluid filtra- tion across the capillary membrane due to an increased capillary hydrostatic pressure. The fact that lymph total protein concentration failed to increase relative to control as much as with histamine alone, in the constant inflow experiments indicates at least some antagonistic action of norepinephrine even though such a disproportionate concentra- tion ratio exists. The higher dose of norepinephrine (l6pg base/min, I.A.) infused simultaneously with histamine in forelimbs perfused at constant inflow, prevented as marked an increase in lymph total protein concentration, relative to control, as seen during histamine infusion alone. Lymph flow rate increased substantially more than during histamine infusion alone but was less than that seen during simultaneous infusion with the low dose of norepinephrine even though microvascular pressure is inferentially greater. Thus, it appears that the antagonism of histamine's microvascular effects by norepinephrine is dose dependent. This conclusion is in line with Schayer's proposed balance of histamine and catecholamines producing a degree of circulatory homeostasis during shock or stressful states (57). 59 This antagonism of the microvascular actions of histamine by norepinephrine could be due to a complete shunting of blood from nutritional to non-nutritional channels and/or a direct antagonism of the action of histamine on the microvascular membrane by norepinephrine. Additional experimentation is needed to resolve this point. SUMMARY AND CONCLUSIONS It is clearly demonstrated that a route-dependent response exists concerning histamine's microvascular effects. In naturally perfused forelimbs during systemic histamine administration (400 to 800pg base/min) there is a continuous decline in forelimb weight. In contrast, local histamine (4pg base/min, I.A.) produces a continuous increase in forelimb weight throughout the infusion period. This differential action cannot be explained on the basis of destruction or inactivation of histamine by the lungs or factors within the blood since infusions upstream and downstream to the lung produce essentially identical effects on the forelimb microvasculature. Prior hypotension for 60 minutes produced by either systemic histamine, acetylcholine or hemorrhage completely prevented the marked rise in lymph total protein concentration during local infusions of histamine (4 and 64pg base/min, I.A.). These data suggest that sub- stances liberated subsequent to hypotension (e.g., catecholamines) effectively antagonize the local microvascular actions of histamine. At constant inflow, the combined infusion of histamine and norepinephrine (4pg base/min, I.A. of each) prevented the increase in lymph flow and total protein concentration seen during infusion of histamine alone (4pg base/min, I.A.) and did not produce any significant 60 61 shunting of blood flow between skin and skeletal muscle. These data demonstrate that norepinephrine effectively antagonizes the action of histamine on the microvasculature. This antagonism of histamine by norepinephrine could either be due to a direct blockade of histamine's effect on the microvascular membrane or a drastic shunting of blood flow to non—nutritional channels. Further experimentation is necessary to completely resolve this matter. In conclusion, it seems likely that the severe hypotension produced by systemic infusions of histamine functions as a stimulus for a sympathoadrenal discharge, and that the released catecholamines (and perhaps other substances) effectively antagonize the microvascular actions of histamine. This would account, at least in part, for the route-dependent differential actions of histamine on the microvascula- ture . APPENDICES APPENDIX A SAMPLE CALCULATIONS FOR BLOOD CONCENTRATIONS OF HISTAMINE APPENDIX A SAMPLE CALCULATIONS FOR BLOOD CONCENTRATIONS OF HISTAMINE Preliminary Information: 1. About 8% of total body weight in grams equals the total blood volume of the dog in milliliters. Thus a 20 Kg dog has a total blood volume of 1600 ml. 2. Forelimb blood flow (control) is about 25 ml/min/lOO grams of forelimb. The average forelimb weight for the dogs used in these studies is about 600 grams. Thus, the control blood flow through a 600 gram forelimb would be 150 ml/min. Calculations: A. Systemic Administration general formula: infusion rate % total blood volume = blood concentration after one minute 1. Histamine infusion = 400 ug base/min. Blood concentration = 0.25 ug/ml after one minute. 2. Histamine infusion rate = 800 pg base/min. Blood concentration = 0.50 ug/ml after one minute. 62 63 B. Local Administration general formula: infusion rate % ml/min forelimb blood fIOW'z forelimb blood concentration. 1. Histamine infusion rate = 4 ug base/min. Blood concentration = 0.03 ug/ml. 2. Histamine infusion rate = 16 pg base/min. Blood concentration = 0.11 ug/ml. 3. Histamine infusion rate = 64 ug base/min. Blood concentration = 0.43 ug/ml. These calculations are for comparative purposes only. It is not intended that these calculations represent actual blood concentrations since degradation rate and distribution within the blood volume will obviously affect the concentration of histamine at any given site. APPENDIX B TABLES APPENDIX B TABLES Appendix B lists, in the form of tables, all the individual observations for the experiments performed in this study. Also listed are the means, standard error of the mean, and statistical signifi- cance. The data in the appendix tables corresponds to the mean values in Tables 1-7 as follows: Table Number Appendix Table Number 1 Al, A2 2 A3, A4, A5, A6 3 A7, A8 4 A9, A10, A11 5 A12, A13 6 A14 7 A15, A16, A17, A18, A19, A20, A21, A22, A23, A24 64 6S N N N N N N N N N NH NH N N N N N N H N N N N N N N N N N N H N N N NNN NNN H N N N N H H N N NH NH NHNNNNHN NHN> NNHNNNNN NH NH NH NH NH NH NH NH NH NH NH HNHHN NHNNNNHN INN INN INN INN INN INN INN INN INN NNN NNN NNNNa NN NN NN NN NN NN NN NN NN NNN NNN NN NN NN NN NN NN NN NN NN NNN NNN NN NN NN NN NN NN NN NN NN NNN NNN NN NN NN NN NN NN NN NN NN NNN NNN NN NN NN NN NN NN NN NN NN NNN NNN AN: NNN NHNNNNHN NN NN NN NN NN NN NN NN NN NNN NNN NooHN NNHHNHHN NHNNHNNN NH NH NH NH NH NH NH NH NH N.H NH HoHHN NHNNNNHN «wan Nod: «mHI ymHa «NH! «mu *5: NV: «H: m. o mcmma NN- NH- NH- NH- NH- N- N- N- N H N NN- NN- NN- NN- NN- NN- NH- NH- NH- N N N- N- N- N- N N N N N N N N- N N N N N N N N N N NN- NN- NN- NN- NN- NH- NN- NN- N- N N NNNNHNN NN- NN- NN- NN- NN- NH- N- N- N- H N HNNHNz NH NNNNNN NN NN NN NN NN NN NN NH N N N- NNNHNNNNN NNHN NN: I NNN: NNNN NNN: Uofihom CmeS-«cm HOHHCOU .Noocmumwmon NNHSUNN> paw 30am pecan .NoHSNNopm HNHsomm> Ingmaoz pawfiouom co Newscfla om «NNN egg magnau mnsfiaouom wemsmuom xaamusuan ous“ xHHNNHouum -muucw womsmcfi ocfismumwn mafia Newscws ca pom Aflmsoco>wuucw comamcw omen ocwswumwn mo muoommm-.H< oHan Table Al.-Continued. Infusion Period Control H800 H800 + H64 H400 30 35 45 6O 65 75 90 15 Time (minutes) 2* 1* 1* 2* 2* 2* 2* 2* 3* means standard error :1 11 ll Brachial Vein Pressure (mm Hg) 66 3 10 7 12 7 12 3* 3* 3* 4* 4* 4* 4* 4* 4* means standard error i1 Cephalic Flow 12 13 12 13 (ml/min/lOO grams) 29 14 29 14 14 7* 5* 3* 3* 3* 4* 4* 4* 4* 14 :3 means standard error 67 NN NN NN NN NN NN NN NN NH HH HH NH NH NH NN NN NN NN NH NH NH NH H -N NNH NH HH NH NH NN NH NN HN NH NN NN x H-Ha x NHs x N: NNN NH HH HH NH NN NN NN NH N NH NH NocmHNHNNN NHomsz HNHNN NH NH NH NH NH NH NH NH HH HH HH HNHHN NHNNNNHN INH NH NH NH NH +NH +NH N N NH HH Names NH NH NH NH NH NH NH NH N N N NN NH NH NH NH NH N N N N N NN NN NN HN NH NH NN N NH NH NH N N N N N N N N N NH NH HH-N NNH N N N N NH N N N N NH HH H -Hs x :Ha x N: NNN HN NH NH NH NH NN NN NH N NH NH NNNNHNHNNN :HHN HNHNN N.H N.H N.H N.H NH N.H NH N.H HH HH HH HoHHN NHNNNNHN INN N N H NIH NIH «H «N «N m m mamma— N N H H H H H H H HH HH H H H H H H H H H N N H H H H H H H H N NH NH N N N H H H H N N NH NH N N N N H N H N N N N HNNNHN NNHNNHNNHNN N N N N H H H N N N N onN HNHHNNHN NN NN NN NN NN NN NN NH N N N- HNNHNNHNN NEHN NNN I NNN: NNN: NNN: flowhom fimeSmcH HOHHCOU .NNNNHHNNN-.H< NHHNN 68 .NosHN> ouscHE co anm ucohommHv xHucmonchHm one: Newscfle om Ho mm .mo mom mosHN> cues oz .Houucoo op o>HuNHoH mo.o w.m u + .Houucou on o>HHNHoH Ho.o w.m u I NH NH NH NH NH NH NH NH NH NH NH HoHHo NHNNNNHN 0N +NN *Hm Nmm Now NHM «mm tom +wN OH ma mfimOE 0N 0N vm mm NV cw om Nm nv NH NH om om mm Hm av NV on vv v0 VN VN NN NN NN NN NN NN NN NH N N N- HNNHNNHNN NEHN v0: + com: com: oov: onHeN :onnmcH Houpcoo .NNNNHHNoN-.H< NHNNN NNH NNH NNH NNH NNH NNH. NNH NHN NNH NNH NNH NN NN NN NN NNH NN NHH NNH NNH HNH NHH NN NN NN NN NN NN NN NNH NN NHH NNH NN NN NN NN NN NN NN NN NN NHH NHH Hm: NNN NN NN NN NN NN NN NN NN NN NHH NHH NHNNNNHN NNHNNNHNN NH NH NH NH NH NH NH NH NH NH NH HNHHN NHNNNNHN INN INN INN INN INN INN INN INN INN NNH NHH memos NN NN NN NN NN NN NN NN NN NNH NNH NN NN NN NN NN NN HN NN NN NN NN NN NN NN NN NN NN NN NN NN NNH NNH NN NN NN NN NN NN NN NN NN NNH NNH NH NH NH NN NN NN NN NN NN HNH NNH HNN NNN NHNNNNHN NN NN NN NN NN NN NN NN NN NNH NNH NooHN HNHHNHHN oHeNHNNN NHH NH NH NH NH NH NH NH NH N.H NH HNHHN NHNNNNHN «OH «ma NHN NHN NNN NNN NNN NNN *hH N. O mcwos NN HN NN NN NN NN NN NN NN N. N NN NN NN NN NN NN NN NN HN N N N N N N N N N N N N. N NH- NH- NH- N- N- N NH NH NH H N N HH NH NH NN NH NH NH N H- N HNNNHNN NN NN NN NN NN NN NN NN NN N. N HHNHNN NH NNNNNN NN NN NN NN NN NN NN NH N N N- HNNHNNHNN NNHN NN: I NNN: NNN: NNNN UOMHOQ fiOHmSMGH HOHHfiOU .NoocmumHNoH HNHnoNN> van 3on wooHn .mousmmoum HNHSUNN> .unwwoz peHHoHom co Nouasfle on HNNH on» wcfinsv onm:H uzmumcoo um vomsmnom mneHHeHom ouaw NHHNNHNHHN -NHH:N womamcfi ocHENuNHn msHm Newscwe om Hem NHmsoco>Nuuafi Newswaw ommn ocHsNuNH: mo Nuoommm-.m< NHANH 7O HH HH HH HH HH HH HH HH NH NH NH HNHHN NHNNNNHN IN IN IN IN IN N N N N N N Names N N N N H H H H N H H H H H H N N N N N H H N N N N N N N NH NH HH HH N H N N N N N N N N N H H H H N N N N N N N HNN NNN N N N N N N N N N N N NHNNNNHN NHN> NHHNNNNN NH NH NH NH NH NH NH NH NH NH NH HNHHN NHNNNNHN INH INH INH INH INH INH INH INH IHN NH N NNNNE NN NN NN NN NN NN NN NN NN NH NH NN NN NN NN NN NN NN NN NN NH NH N N N N N N N N N N N NH HH NH NN NN NH NH NH NN NH HH NH NH HH N N N N NH NH N N HN: NNN N N N N N N N N N N N NHNNNNHN NHN> HHNNN NHNN NH NH NH HHH NH NH NHH HNH NHH NH NH HNHHN NHNNNNHN INN INN INN INN INN INN IHN NHH NNH NHH NHH NNNNe NN NN NN NN NN NN NN NN NN NN NN NN NN NN NN NN NN NN NH N N N- HNNHNNHNN NNHN NNN I NNN: NNN: NNN: @Owhom fiofimsmcm Houucou .NNNNHucoN-.N< NHNNN 71 N N N N N N N N N N N NH NH NH NH NH NH NH NH NH NH NH NH NH NH NH NH NH NH NH NH NH NH HNNNHN NNHNNHNNHEN NH NH HH N N N N N N NH NH onN HNHHNNHN NH NH NH NH NH NH NH NH NH NH NH HNHHN NHNNNNHN NH NH NH NH NH NH NH NH NH NH NH Names NH NH NH NH NH NH NH NH NH NH NH N N N N N N N N N N N NN NN NN NN NN NN NN NN NN NN NN NH NH NH NH NH NH NH NH NH NH NH N N N N N N N N N N N HNENHN NNHNNHENHNN NN NN NN NN NH NN NN NN HN NN NN onN NHHNNNNN HH HH HH NH HH HH NH NH NH HH HH HNHHN NHNNNNHN NH N N N N N N N N N N NNNNN HH NH HH NH NH N N N N N N N N N N N N N N N N N N N N N N N N N N N N NH NH NH NH N NH N N NH HH NH N N N N N N N N N N N HNN NNN NH NH NH NH NH NH NH NH NH N NH NHNNNNHN NHN> HNHHNNHN NN NN NN NN NN NN NN NH N N N- HNNHNNHNN NEHN NN2 I NNN: NNN: NNN: VOMHOQ fiOmmefiH HOHHGOU .NNNNHHNoN-.N< NHNNN 72 .NesHN> ouscHE oo Eopm unoHeMMHv NHucmonHcme one: NewscHe om Ho mN .mo New Neus> :moe oz .HOHucoo op o>wpmHou mo.o.w m u + .Houuaoo on m>HHNHoH Ho.o.w.m u I HH .3.- HH NH NH. H“ M“ NH mu. HH 3.. INC-Hum whmvcmum IN IN IN IN N N N IHH IHH N N NNNNN N N N N N N N N N N N N N NH NH N NH HH NH NH HH NH HH N NH NH NH NH NH NH NN NH NH N N N N N N N NH N N N H -N NNH N N N N N N N N N N N x H-He x NHN H N: NNN N N N N N N N NH N N N NNNNHNHNNN NHost HNHNN NH NH NH NH NH NH NH NH NH NH NH HNHHN NHNNNNHN N N N NH N N N NH N NH NH NNNNN N N N N N N N N N N N NH NH HN NN NH NH NN NN NN NN NN N N N N N N N N N N N N N N N N N N N N N N H -N NNH NH NH NH HN NH NH N NH NH NH NH x H-He x NHN H N: NNN N N N N N N N N N N NH NNNNHNHNNN NHHN HNHNN HH HH HH NH NH HH HH NH HH HH HH HNHHN NHNNNNHN NH NH NH NH NH IHH IHH INH IHH NH NH NNNNN NH NH NH NH NH NH N N NH NH NH NH NH NH NH NH NH NH NH NH NH NH NN NN NN NN NN NN NN NH N N N- HNNHNNHNN NNHN NNN I NNN: NNN: NNN: UOHHOQ :OHMMSMCH HOHHCOU .NNNNHHcoN-.N< NHNNN 73 HH HH HH HH HH NH HH HH HH HH HH HoHHo NHNNNNHN NH HH HH HH HH NH HH HH N HH HH NNNNN NH NH NH NH NH NH NH HH NH NH NH N N N N N N N N N N N NH NH NH HH HH HH NH NH N NH NH HH NH NH N NH N N HH N HH HH NH NH NH HH NH NH NH NH HH HH HH NH NH HH NH NH NN NH NH N NH N HN: NNN HH HH NH NH NH NH NH HH HH NH NH NHNNNNHN NHN> HHNEN NHHN NHH NH NH NH NH NH NH NH NH NH NH HNHHN NHNNNNHN IHN INN INN INN INN INN INN INN INN NNH NNH NNNNN NN NN NN NN NN NN NN NN NN NNH NNH NN NN NN NN NN NN NN NN NN NNH NN NH NH NH NH NN NN NN NN NN NNH NNH NN NN NN NN NN NN NN NN NN NNH NNH NN NN NN NN NN NN NN NN NN NNH NNH NN NN NN NN NN NN NN NN NN NNH NNH HN: NNN NHNNNNHN NN NN NN NN NN NN NN NN NN NNH NNH NNNHN HNHHNHHN NHENHNNN NN NN NN NN NN NN NN NN NH N NH- HNNHNNHEN NEHN NNN I NNN: NNN; NNN: flowhmm cofimnmcH HOHHCOU .uHHUOHNEo: use Nopsmmoum HNHsoNN> ImENNHm cam :aexH mo :oHHNHucoocoo :Hououm I3on :QENH co NouscHE om HNNH ecu wcHusv NnEHHoHom vomsmuom NHHNHSHN: oucfl NHHNNHouHm-muuzw vomnmcfi ocwsmumwn NsHm NewscHE om How Hume; can we oHoHHuco> umoH esp ou:« pomsmcw omen ocHENuNH: mo Nuoemmm-.n< oHnmh 74 N.N N.N N.N N.N N.N N.N N.N N.N N.N H.N H.N N.N HN NNNHNN N.N N.N N.N N.N NHNHNHN NENNHN N.H N.H N.H N.H N.H NIH N.H H.N N.H N.H N.H .HOIHHo FEE—mum IN.N IH.N IN.N IN.N N.N IN.N IN.N IN.N IN.N N.N N.H NNNNN N.N N.N N.N N.N H.N N.N H.N N.N N.N N.H N.H N.N N.N H.N N.N N.N N.N N.N N.N N.N N.N H.N N.N N.N N.N N.N N.N N.N N.N N.N H.N N.N N.N N.N N.N N.N N.N N.N N.N N.N N.N N.N N.N N.N N.N H.N N.N N.N N.N N.N N.N N.N H.N N.H N.H HN NNNHNN N.N N.N N.N N.N N.N N.N N.N N.N N.N N.H N.H NHNHNHN HNHNN NNENN NN. HN.H HN.H NN.H NN. NN.H NN.H NN.H NN.H HN.H NNN.H HNHHN NHNNNNHN NN. NN. NN. INN. INN. INN. IHH. INH. INN. NN. NN. NNNNN NN. NN. NN. NN. NN. NN. NN. NH. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. HN. NN. NH. NN. NN. HH. NH. NH. NN. NN. NN. HN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NH. NH. NN. NN. HN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. HNHE NHNHsN NN. NN. NN. NH. HN. NN. NN. NN. NN. NN. NN. NHNN onN HNNNH NN NN NN NN NN NN NN NN NH N NH- HNNHNNHNN NEHN NNN I NNNN NNN: NNN: wowhmm fimechH HOHHGOU .NNNNHHaou-.N< NHNNN 75 .NosHN> ouncHE oo EOHM unopomva HHucmonwcmHm one: Nouncwe om Ho ow ION pom mesHm> cave oz .Honucoo on o>HuNHeH mo.o v m u + .Houueoo ou o>HuNHoH Ho.o w.m u I NH mH mH NH Hopno vnmvcmum *mm *vm Nom 0? WCNQE Nm mm NN Nm mm om mN mm No No No NN mN HN HN ON ON mm mm mN cm mm mm ON mm NN mm ON HHHooHNEN: N.H N.H N.H N.H Houno vnmvcwum H.m m.m H.m o.m Names N.N o.m N.N N.N N.N N.N N.N m.m NN NN NN NN NN NN NN NN NH N NH- HNNHNNHNN NEHN Nor I com: com: CON: voHHoN :onsmcH Honucou .NNNNHHNNN-.N< NHNNN NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. HN. HN. NN. NN. NH. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NN. NH. NH. NH. NN. NN. HNHE NHNHEN NN. NN. NN. NN. NN. NN. NN. NN. HN. NN. NN. NHNN onN NNNNH HH HH HH HH HH HH HH HH HH HH HH HNHHN NHNNNNHN «0 NB «5 NB NB +w «h +w +w m CH mfiwmfi N N N N N N N N N N N N N N N N N N N N NH HH N N N N N N N N N NH NH N N N N N N N N N N N N N N N N N N N N HH NH HN: NNN N N N N N NH NH NH N N N NHNNNNHN NHN> HHNeN NHHN NH NH NH NH NH NH NH NH HH NH NH HNHHN NHNNNNHN INN IHN INN INN INN IHN INN INN INN NHH NHH NNNNE NN NN NN NN NN NN NN NN NN NNH NNH NN NN NN NN NN NN NN NN NN NHH NHH NN NN NN NN NN NN NN NN NN NNH NNH NN NN NN NN NN NN NN NN NN NNH NNH N NH NN NN NN NN NN NN NN NN NN HN: NNN NHNNNNHN NN NN NN NN NN NN NN NN NN NHH NHH NNNHN HNHHNHHN NHENHNNN NN NN NN NN NN NN NN NN NH N NH- HNNHNNHEN NEHN NN: I NNN: NNN: NNN: UOMHOQ CmeDMCH fichuflou .HHHoouNEo: paw NNHSNNNHQ HNHsomN> INENNHQ can cmst mo mafipmnuzoocoo :HNHOHQ Ionm :QEHH :o NouscHE on HNNH any wcHnsw NDENHoHom vomsmaem NHHmusumc oucfl HHHNHHoHHm -mHucH womsmcw mmHENpNH: msHm Newscfie om How HHmsoce>mHucw vomsmzw ommn ocfiemumfin mo Nuuommm-.N< oHnNH Table A4.-Continued. Infusion Period Control H800 H800 + H64 H400 20 30 40 SO 60 70 80 90 10 -10 Time (minutes) .02 .11 .07 .07 .02 .02 .01 .01 .01 .01 .01 .01 .01 .01 .01 .02 .01 .01 .01 .01 .01 .01 .03 1.01 .02 .06* .06* .06* .05+ .04 .03 .02 .02 $.02 i.03 $.02 i.01 .01 $.01 1.01 i.01 i.001 .02 i.001 means standard error 3.2 3.7 3.0 2.8 3.8 3.1 2.6 3.1 3.5 1.9 3.5 2.0 3.4 2.0 2.7 1.7 2.4 2.2 1.7 2.4 Lymph Total Protein 3.8 3.5 3.8 (grams %) 2.3 2.7 3.5 3.2 3.5 3.5 3.4 3.3 3.4 2.2 2.1 77 2.3 2.2 2.1 2.7 3.0 3.3 3.6 3.6 3.5 3.1 1.4 2.4 2.6 3.0 3.0 3.1 2.9 2.6 3.2 1.4 2.0 2.9 2.1 2.0 2.0 1.6 1.6 1.5 1.5 1.7 1.7 1.6 1.9 2.1 2.3 2.3 2.6* 2.9* 3.0* 3.0* 3.0* 3.2* 3.2* means standard error +| MNQOQP’) VLOVVVLO \DOI—IOON VI!) LOU) VII) hwfi‘fl’l—OV V’VLOOLDLD c>pqc><'05fl' mvmmvm Plasma Protein (grams %) USN V-H 03H V‘H v LO 0 Ln means standard error I") +| N +l 78 .mosam> ouncfis co Scum acouommfiw Afipcmoflwwcwfim «no: mouscwe om 90 cm .on How mosfim> came oz .Houusoo ou o>fium~ou mo.o.w.m u + .~0hu:oo ou o>wumHou Ho.o w.m u 4 NH NH m“ an Honno wamwcmum «Hm «mm «.wv vm mammfi om Hm we mm mm cm mm mm 0v av me mm Hm Nm we mm 0v nv ov Hm 8 me mm mm 9208.26: om cm on oo om ow on ON oH o oH- nmopseflsv mafia com + com: com: 00v: weapon :owmsmcH Hopucou .eoscflucoo--.v< macaw 79 ma ma 0H wH ma ma 0H NH VA MH ma OH NH NH NH OH OH . NH NH NH OH OH OH OH OH OH OH OH OH OH NH OH OH HO: EEO OH ON OH OO OO ON HO ON OO OH NH «HOOOOHO :Ho> HHmeO :HHO NH NH OH OH OH OH OH OH OH OH OH HOHHO OHOOOOHO aflm amm *mm mvm +©m +00 +wm mom «we mofi moH mamme OHH ONH OOH OO OO OOH NO OO OO OHH OHH OOH NO OO OO OO OO OOH NO ON OHH OOH OO OO ON OO OO OO NN ON NO OOH OOH OOH OO OO OO OO OO OHH OHH OO OOH OOH OO OO NO OO OO OO OO OO ON OHH OHH HO: EEO ON OO OOH OHH ONH OHH OHH OO OO OHH ONH. OHOOOOHO OOHOOHHOO OH OH OH OH OH NH OH OH NH OH OH HOHHO OHHOOOHO fiwN amN amN *mN «om «OM «mm *Hm mom wHH Rafi mCdOE ON ON ON ON ON NO NO Ow OO ONH ONH ON ON ON NN NN NN OO NN NN OOH ONH OH OH OH NO OO OO OO OO NN OOH OOH OH OH ON ON ON ON OO ON ON NOH NOH ON ON ON ON NN OO OO OO OO ONH ONH HO: EEO OHOOOOHN NO OH OO OO OO NO OO NO OO ONH ONH OooHO HOHHOHH< OHEOHONO OO OO ON OO OO OH OO ON OH O OH- HOOHOOHEO oaHO «Oz + OOO: OOO: OOO: wofihmm :oflmfimcH HOthOU .uwnooumfio: was mousmmoum ansomm> .wsmmHm ecu :mexH mo coaumuucoocoo :Houonm .3on smexH :o mouscfie om ummH esp wcflusu onmcfi pcmumcoo um womsmnom mnEHHonom oucfl OHHmOnounm-muu:O vomnmafl osmewumfls OsHm mouscfla om uom.uwmo: may we oHoOnuao> pmoH on» ousfi comsmcw ammo unflamumwn mo Ouoommm-u.m< oHnme 80 O.H O.H O.H o.H O.H O.H O.H O.H O.H O.H O.H Houno unmvcmum HO.O ON.O HN.O «0.0 «0.0 HO.O «N.O «o.O OH.O m.H m.H memos 0.0 0.0 0.0 O.c H.O 0.0 0.0 0.0 o.N 0.0 N.O o.O 0.0 0.0 0.0 0.0 o.O N.O o.O O.N H.H O.H o.O m.N m.N o.O 0.0 0.0 0.0 o.O O.N O.H o.H 0.0 m.O o.O N.O 0.0 0.0 0.0 0.0 m.N N.N O.N N.O 0.0 0.0 m.O H.O N.O o.O 0.0 O.H H.H N.H HO OEOHOU 0.0 0.0 N.O 0.0 0.0 0.0 0.0 0.0 o.N O.H N.H :kuopm HOHON :QENH No.H No.H No.H No.H Oo.H Oo.H Oo.H Oo.H Oo.H Ho.H Ho.H noun» wumwcwum co. Oo. +00. OOH. «OH. OOH. *ON. «HO. OOH. No. No. m:moa No. Oo. HH. OH. ON. NO. 0O. OO. OH. HO. Ho. oH. Oo. HH. OH. OH. OH. ON. OO. OO. No. No. Oo. No. O0. O0. Oo. co. No. ON. Oo. No. No. HH. OH. OH. NH. ON. NN. OO. OO. NH. Oo. Oo. Oo. me. me. mo. OH. 0N. NN. ON. co. Ho. Ho. HcHE oH\Hev Oo. NH. co. OH. O0. O0. OH. NN. Oo. Ho. Ho. mama onm :msNH HH HH HH OH OH NH OH OH OH HH HH Hoano Oumwcmum OH OH OH NH OH NH OH NH OH OH OH mcmoa HH OH O O a HH NH NH NH NH NH OH OH OH OH NH NH NH NH NH OH OH om oO ON 00 om OO 0O 0N 0H 0 oH- HmopscHev oEHH O0: + OOO: OOO: OOO: OoHHom consmcH Honucou .Ooschcou--.O< oHOOO 81 .Ooon> ouocHE oo soum acouommHo NHpcmonHcmHm onoz mouscHa om no CO .oN How moon> came oz .Houpcoo op o>HumHoH Oo.o.w m u + .Honunoo op o>HumHon Ho.o.w m u « OH OH OH NH Honuo wnmwcmum OOO HOO *HO OO OOOOE NO OO NO NO OO NO OO OO OO OO OO HO OO NO OO OO OO OO OO OO NO HO OO OO HHHOOHOEOO M.H N .H M.H v.“ .HOHHG flung—mam O.O O.O O.O O.O mamas O.O H.O 0.0 H.N O.O O.O O.O O.O O.O O.O N.O O.O N.O H.O O.O O.O O.O N.O O.O O.O HO OEOHOO O.O O.O N.O O.O :HOHOHO OEOOHN OO OO ON OO OO OO OO ON OH O OH- HOOHOOHEO OEHH OO: + OOO: OOOO OOO: HuOmHmnH conSmcH HOHHHHOU .OosaHuaoo--.O< OHOOH NH NH OH OH HN HN OH NN ON OH OH OH HN OH OH OH ON ON ON HO NH OH ON ‘ON ON ON ON ON ON ON HN OH OH HO: EEO HO OO HO OO NO HO ON HO NN OH OH OHsOOOHO OHO> HHOEO OHHO OH NH OH OH OH OH OH NH OH OH OH HOHHO OHOOOOHO -OO HON OON HON HON HON -OO *ON HNN HOH HOH Osmoa OO OO OO NO OO OO NO OO NO OO OO ON ON ON ON ON ON NO ON ON OOH OOH OO OOH OO OO OO OO OO OO OO OHH OHH OO OO OO NN OO OO OO OO ON OHH OHH HOO EEO OO OO OO OO OO OO OO OO ON OO OO OHOOOOHO OOHOOOHOO NH OH OH OH OH OH OH NH NH OH OH HOHHO OHOOOOHO .NO ONO HNO ONO HOO HHO HOO .ON HHO HHH NHH mamas ON ON ON ON ON ON NN ON ON OOH OOH ON ON ON ON ON NN OO ON ON OO OO ON ON ON ON NN OO OO OO OO ONH ONH OO OO OO OO NO OO OO OO OO ONH OOH OO OO OO OO OO OO OO OO OO ONH ONH HO: EEO OHOOOOHO ON ON ON ON ON ON ON NN OO OOH OOH OOOHO HOHHOHHO OHEOHONO OO OO ON OO OO OO OO ON OH O OH- HOOHOOHOO OsHN OO: + OOO: OOO: OOO: wowhmm Seams-«HHH HOHHCOU .HHHooumEoc wow monommonm HmHsomm> .OEOmHm wow HQENH mo :oHHmHucoocoo :Hououm .3on :QENH :o OopscHs 0O ume on» maHuzv onmaH ucmumcoo um vomomnom OQEHHoHom ouaw NHHOHHQHHO -OHH:H womsmcH ocHEOumH: msHm mouaaHE om How NHmsoco>mhch oomsmcH ommn ocwsmpmws mo muoommm-.o< oHan 83 N H O H N H N H O.H O.H O.H O.H O.H N.H N.H Mon-Ho uumwcmum HN.m «h.m ¥w.m «h.m aw.m *w.m *m.m «H.v w.N m.N M.N mflmoa 0.0 O.N O.N N.N O.N O.N O.N O.N O.N H.N O.N N.O O.O O.O O.O N.O N.O O.O H.O 0.0 H.O H.O O.O O.O H.O N.O 0.0 0.0 0.0 N.O H.N O.H N.H O.O O.O N.O N.O O.O N.O 0.0 N.O N.N O.H H.N O.O O.O O.O O.O O.O H.O 0.0 N.O H.O O.N N.N HO OOOHOO O.O H.O N.O N.O O.O O.O O.O O.O O.N O.N N.N :HOHOHO HOHOO HOENH OO.H NO.H OO.H OO.H OO.H OO.H OO.H OO.H OO.H NOO.H NOO.H HOHHO OHOOOOHO OH. OO. HNH. +HH. OOH. .NH. .NN. .HO. HNH. NO. NO. OOOOE NO. NO. OO. NO. NO. OO. NO. OO. OO. NO. NO. OO. OH. OH. OO. NH. OH. ON. OH. NH. HO. HO. NN. OO. OH. OO. OO. HH. ON. ON. NH. NO. NO. OO. OO. OH. OH. NN. HN. NN. NO. OH. NO. NO. OH. OH. ON. NN. OO. OO. OO. OO. OO. NO. NO. HOHa OH\HaO OO. OO. OO. OO. OH. OO. OO. OO. OO. HO. HO. OHOO onO OOENH OH OH OH OH OH OH OH OH OH HH HH HOHHO OHOOOOHO «HN «HN *HN *NN {ON HON HON «NN HHN MM MH mcon NN OH ON OH HO OH HH NN HH HH HH NH NH OH HH HH HH OH HH OH OH OH OO OO ON OO OO OO OO ON OH O OH- HOOHOOHEO OaHO OO: + OOO= OOO: OOO: flown-Hon— CONmHHmcH HOHHHHOU .OOOOHHOoo-.O< «HOOP 84 .OooHO> ouocHE OO Eonm HconomeO NHuomonHcmHm one: mouscHE om no 0O .ON now OosHO> :Ooe oz .Honucoo oH o>HHOHon Oo.o.w m u + .Honucoo on o>HHOHon Ho.o.w m n O OH OH OH NH nonno Onmvcmum Ham H9O OOO NO mcmoa NO NO OO ON NO OO NO OO NO NO OO OO mm OO NO NO OO OO OO HO OO mO ON NO OHHnoouOEo: O.H O.H N.H H.H nonno Onmvcmnm 0.0 0.0 0.0 0.0 mamas 0.0 N.O 0.0 0.0 N.O 0.0 0.0 0.0 0.0 N.O N.O 0.0 0.0 0.0 N.O 0.0 N.O O.O N.O O.O HO OEOHOO 0.0 0.0 0.0 0.0 :Houonm OEOOHQ om 0O ON OO om 0O 0O ON oH o oH- HmouscHEv OEHN OO: + QOO: OOO: OOO: OoHnom :onomcH Honucou .OOOOHHaoo-.O< OHOOn 85 HO. NO. NO. OO. OO. NO. NO. NO. NO. HO. NO. HO. HO. NO. NO. NO. OO. OO. OO. NO. NO. NO. NO. NO. HOHE OHNHsO OO. OO. OO. OO. OO. OO. NO. NO. OHOO :oHO OOEHH HH HH HH HH HH HH HH HH nonno vnmwcmum «h *5 «h *0 an *0 NH HH mfimmfi N O N O O N OH NH N N N O O O OH OH O O O N O O HH HH O O O O O O O O N N O O O N OH NH HO: EEO N N N O O O O O OHOOOOHO OHO> HHOEO OHHO OH OH OH OH OH NH OH OH nonno wnmvcmum Hmv «vv HHV «mm *mm «QM MNH mNH mfimoa NN OO OO ON OO OO OOH OOH OO OO OO NO OO OO ONH ONH OO OO OO OO OO OO NOH NOH OO OO NO OO OO OO OOH OOH OO OO OO OO OO OO OHH OHH HO: EEO OHOOOOHO OO OO OO OO ON ON OO OOH OooHO HOHHOHHO OHOOHONO OO OO OO OO ON OH O OH- HOOHOOHEO oEHn OOO + OOOO vowhom COMM—HHH; HOHHHHOU .HHnooHOEo: Ocm monommonm ansomm> .OEOOHQ vow :QENH mo :oHumnucoocoo :Houonm .3on :QENH :o mouocHE 0O nom OOEHHmnom Oomsmnom NHHwnsuO: oucH HNOHOO ouscHe O O noummv NHHOHnoHnO uwnucH Ocm unmo: on» mo oHoHnuco> HmoH on» oucH OomsmcH ommn ocHEOHOH: mo muoommm-.N< oHnwh Table A7.-Continued. Infusion Period Control H400 + H64 60 50 40 30 20 10 -10 Time (minutes) .01 .01 .01 .01 .09 .01 .01 .02 .02 .09 .07 .09 .09 .02 .02 .02 .04 i.01 .03 .03 .04 .04 i.01 .01 i.01 1.01 .02 i.003 .02 .02 means standard error i 2.4 3.1 3.4 3.7 4.0 2.8 2.6 2.3 0.9 2.6 2.4 0.9 Lymph Total Protein 2.6 2.7 0.8 2.1 2.2 0.9 2.7 2.6 (grams %) 1.0 3.3 0.9 1.1 0.9 86 3.8 3.6 3.0 5.9 2.6 3.5 3.1 2.7 3.0 3.8 2.3 3.8 4.3 3.2 3.2 3.1 3.7 5.3 6.3 6.0 5.9 5.5 5.5 2.9 3.1 3.2 3.4 3.4 3.0 2.7 3.0 means standard error +l +| +| (\Ofi'mOO LOOLOOOOO {\O'SVOLOHH mmmxoooo QNOOOV \DOLOOOQ Plasma Protein (grams %) M OH I") OH I!) \0 means standard error V V q +| 87 .Honucoo on o>HumHon Oo.o V D.- II 4. .Honuooo on o>HHOHon Ho.o v a ll 1: N H N H N H HOHHO whmwcmum HOO HHO HO OOOoa OO OO OO OO OO NO OO OO OO NO OO NO OO OO OO NO OO OO HHHUOHOEOO OO OO OO OO ON OH O OH- HOOHOOHEO OeHn OO: + OOOO UOHHQQ SONODMCH Ho-HHHHOU .OOOOHHcoo-.N< OHOOn 88 OH OH OH NH OH NN O HH OH O O HH OH NH OH O OH OH NH OH OH HN OH OH HO: EEO HH NH OH OH HO OO O O «HOOOOHO OHo> HHOOO OHHO OH OH OH OH oHH OH NH OH nonno find—Emu».- OO OO OO OO HOH HO OOH OOH OOOOe ON OO NO OHH OOH OOH OOH OOH OO OO ON OO OO OOH OOH OHH NO ON ON OO NO OO NOH NHH OOH OO OO OO OO ON OO OO NN OO OO NO NO NN NO OO HOO EEO ONH ONH ONH OOH NOH OOH OO OO OHOOOOHO :oHOOOHOO OH OH OH OH NH NH NH HH nonno Honmwcmpm *OM «mm Hem «QM Hmm *VM NOH OOH mammfi OO OO OO OO OO OO OHH OOH NN ON NH NN NN OO OOH NOH OO OO OO OO NO NO NOH NOH OO NO OO NO OO OO NHH OHH NO NO OO NO NO NO NOH OOH HO: EEO OHOOOOHO OO OO NO OO OO OO OOH OOH OooHO HOHHOHHO OHEOHONO OO OO OO OO ON OH O OH- HOOHOOHEO oeHn OO: + OOO: UOMHOQ COHOSMcH Ho-HHCOU .HHnooHOEo: van monommonm ansomO> .OEOOHQ Ocm :QENH mo :oHHOnucoucoo :Houonm .3on :mexH :o mousaHe 0O nom 3oncH Hcmnmcoo HO Oomsmnom OOEHHonom och HNOHOO ouscHa O a noummv NHHOHnounO umnnaH Ocm unmo: ecu mo oHoHnuco> HmoH osn ouaH OomomcH Oman oaHEOnOH: mo Ouuommm-.O< oHnOH 89 N H O.H N.H H.H O.H O.H O.H O.H Henna OHOOOOHO OO.? HN.? «w.m «w.m «m.m +N.m m.N ¢.N mcmmfi O.O O.O N.O N.O N.N O.N N.N O.N O.O O.O O.O O.O O.O 0.0 N.N O.N O.O O.O N.O H.O 0.0 H.O O.N O.N N.O O.O O.O H.O N.O O.O 0.0 H.O 0.0 0.0 N.O 0.0 O.O O.N H.N O.H HO OEOHOO N.O O.O O.O O.O O.N O.N O.H O.H OHOHOHO HOHon OOENH NO.H OO.H OO.H OO.H OO.H OO.H HO. HO.H Honno OHOOOOHO OO. ONH. ONH. ONN. «OO. OON. NO. NO. OOOOa OO. OO. NO. OH. HH. HN. NO. NO. OO. OH. NN. OO. NO. NO. OO. NO. OH. OH. ON. NO. OO. NO. HO. HO. OO. NH. OH. OH. NN. OH. HO. HO. OH. OH. NN. HO. OO. NO. OO. OO. HOHO OHNHEO OO. OO. OH. OH. ON. OH. NO. NO. 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OHOO OOHO OOHNH H H H H OH HH HH HH Honnm vumwcmum HH HH HH HH HH OH OH OH OOOOs O OH OH OH OH O N N OH OH OH OH NH NH OH OH OO OO OO OO ON OH O OH- HOOHOHHOO OeHN OON-Mom fiOm mamO—H HOHHSOU .OOOHHHcou-.NH< OHOOO 116 .Hopucoo op o>HumHmH OO.O.W m u + .Honucoo on o>HumHoH Ho.o.w m u H voHHmm :onzmcH NH NH NH noun» Onmccmpm mm mm NO mamas 0O on NO ON ON ON em on mm ON ON on mm mm NO om NO on HHHooumam: N.H O.H O.H Houuo vuavcmum 0.0 0.0 0.0 mamas N.O 0.0 0.0 N.O 0.0 w.O 0.0 N.O H.O 0.0 0.0 0.0 0.0 H.O c.o Aw memumv H.O 0.0 0.0 :Hououm OEOOHN OO OO OO OO ON OH O OH- HOOHOHHOO OOHH Honucou .OOOHHHcou-.NH< OHOOH 117 ON ON OO ON NO HO O O OO OO OO OO OO OO OH OH NO OO NO OO NO OO HH HH HO: EEO ON NN NN NN ON NN OH HH OHOOOOHO HHo> HHOHO OHHO NH OH OH OHH OHH OHH OH OH HOHHO OHOOHOHO OOOH OOOH OONH OONH OOOH OOOH NHH OHH OOOOO OOH OOH OOH ONH ONH OOH OHH OHH ONH OOH OOH ONH OOH ONH OOH OOH NOH NOH OOH OOH NOH NOH OHH NHH ONH ONH ONH NNH OOH OOH OOH OOH OHN OHN OHN OON OON OON OHH OHH HO: EEO OOH OOH ONH ONH OOH OOH OO OO OHOOOOHO :oHOOHHOO NH OH OH OH OH OH OH OH HOHHO OHOOHOHO OOOH ONOH OOOH OOOH OOOH OOOH OHH OHH Ocmoa OOH NOH NOH NNH ONH NOH ONH ONH NHH NHH OHH NNH NNH NNH NOH NOH ONH ONH ONH NNH ONH OOH OHH OHH NOH OOH ONH NNH ONH NNH NOH OOH OOH OOH OOH OOH OOH OOH NHH OHH HO: OOO OHOOOOHO ONH ONH ONH OOH OOH OOH OOH OOH OooHO HOHHOHHO OHEOHOHO OO OO OO OO ON OH O OH- HOOHOHHEO OOHH UOwHOm CONmDMEH HOHHCOU .HHHuoumEo; can OOHSOOon HmHsumw> OOEOOHQ vcm :QENH mo :oHpmHucmocoo :Hououm Ozoflm :QENH :o OopscHE oo How onmcH ucmumcoo Hm vomswumm OQEHHoHom oucH Onomm mo oH=:HE\ommn N: O .ocHnnmochmHo: wam ocHEOpOH: mo OconsmcH HOHHopHO-muucH OsomcmuHSEHO mo Ouoommm-.NH< oHan 118 O.H m.H O.H O.H m.H m.H N.H N.H HOHHQ wumvcmum O.O H.O H.O N.O N.O O.O O.O O.O memos N.O O.O O.O O.O O.O O.O O.O O.O O.O O.O O.O O.O N.O O.O N.O O.O N.O O.O N.O N.O N.O O.N H.O O.O N.O O.O O.O H.O H.O N.O N.O O.O 0.0 N.O O.O O.O O.O O.O O.O N.O HO 223 O.O H.O N.O O.O O.O O.O H.O 0.0 :HOHOHO HOHOO OOHHH HO.H HO.H NO.H NO.H OO.H NO.H OOO.H OOO.H HOHHO OHOOOOHO OO. OO. OO. HOO. ONO. HOO. NO. NO. OOOOH HO. HO. HO. HO. HO. NO. HO. NO. OO. OO. OO. OH. ON. OH. OO. OO. NO. OO. OO. OO. OO. OO. HO. HO. OO. OO. OO. HH. HH. NO. HO. HO. HO. HO. HO. HO. HO. OO. NO. HO. HOHE OHNHHO HO. HO. HO. HO. HO. OO. OO. NO. OHOO onO OOHHH NH NH NH NH NH NH HH NH HOHHO OHOOHOHO OOO OHO ONO OHO OHO ONO O OH OHOoH OO NO OO OO ON OO O N ON ON ON NN ON ON O O OO OO OO OO ON OH O OH- HOOHOHHHO OOHH OOH-Hon COMmSMCH HOHHGOU .OOOOHHHOO-.OH< OHOOH 119 .3350 ca 03.528 mod IV- a u ._. .Honuzoo op c>HumH0H 8.0 W a u H NH NH NH HOHHO whmwcmum OOO OOO OO OOOOe OO NO OO OO OO HO OO OO ON OO OO NO OO OO OO OO OO OO HHHOOHOHOO N . H N . H m . H HOH-Ho undue—mum O.O O.O N.O mamas O.O N.N O.O N.O 0.0 O.O O.O O.O O.O 0.0 O.O N.O 0.0 N.O 0.0 HO OEOHOO O.O N.O H.O HHOHOHO OeOOHO OO OO OO OO ON OH O OH- HOOHOOHHO OOHH vowhom :OHMHHMCH HOHHGOU .OOOOHHOou-.OH< OHOOH 120 ON OO NO NN ON ON OH HH ON ON ON ON ON ON O OH ON ON NN NN ON ON OH OH HO: EEO NH NH ON NN NN ON OH OH OHOOOOHO OHo> HHOHO OHHO mH mH OH mH mH mH NH MH Honk» vhmvcmum OHNN OONN OOHN OOON OOON OOON OHH OHH OOOOe OHN OHN OHN OHN OOH OOH OHH ONH OON ONN ONN OHN OON OOH ONH NHH ONN ONN OHN OHN OHN OON OHH NOH NON NON OON OON OON OON OHH ONH OHN OHN OHN OON OOH OON OOH OOH HOO EEO OON OON OON OHN OHN OHN NHH OHH OHOOOOHO OOHOOHHOO NH wH wH OH N.H mH NH MH HOHHO whats-mum OOOH OOOH OOOH OOOH OOOH OOOH ONH OHH memos OOH OOH OOH NOH OOH OOH OHH OHH OOH OOH OOH OOH OOH OOH ONH NNH ONH ONH ONH NNH OOH NOH OHH OHH OOH OOH OOH OOH OOH OOH ONH ONH OOH NOH NOH OOH OOH NOH NHH OHH HO: EEO OHOOOOHO OOH ONH ONH ONH OOH ONH ONH ONH OOOHO HOHHOHHO OHEOHOHO OO OO OO OO ON OH O OH- HOOHOOHHO OEHO Oofihom COmmSMCH HO.HHCOU .uHHooumEo; Ocm mmHSOOon HOHSUOO> OOEOOHQ wcm :QENH mo coHuOHucoucou :Hououm Oonm :QENH :o OouscHE oo How onmcH ucmumcou um Ommsmumm OnsHHouom oucH NHHOHHouHm-OaucH OomsmcH muscHENmOwn ocHHcmmchoHo: N: O mo Opuommm-.OH< oHnmb 121 AN OEOHNV :Houonm OEOOHN Honno cumucmum mamas RN OEOHNV :HOHOHm Hmpoh :mENH OOO.H HO.H HO.H HO.H OOO.H HO. HO.H HO.H HOHHO OHOOOOHO NO. OO. NO. NO. NO. OO. NO. NO. OOOOe OO. OO. OO. NO. OO. OO. OO. OO. HO. HO. HO. HO. NO. HO. NO. NO. NO. OO. OO. OO. HO. OO. NO. NO. OO. NO. NO. HO. HO. HO. HO. HO. HO. OO. HO. HO. HO. NO. HO. HO. HHHE OHNHHO OO. OO. HO. HO. HO. NO. HO. HO. OHOO onO HOHHH NH OH OH NH NH OH HH HH Hop-Ho OHOHOEOHO «.HN OONN OONN OOHN OOHN HOMN HH NH mamma— OH OH OH OH OH ON O O OH OH OH OH OH NH HH NH OO OO OO OO ON OH O OH- HOOHOHHEO OOHH OO.H-Mom CowmzmcH HO.H-:50 .OOsHHHOOO-.OH< OHOOH 122 .HOHHOOO 8. o>HHOHOH OO.O W H .-. + .3580 3 3323 8.0 W H n O NH NH HH HO.H.Hw Uhmflcmum OOO +NO OO OOOOH OO OO OO OO OO HO OO OO OO OO NO OO OO OO OO OO OO HO HHHOOHOEO: H . H N . H N . H .Ho-HHO Ohmficmum O.O O.O N.O OOOOE O.O H.O N.O O.O O.O N.O N.O O.O O.O OO OO OO OO ON OH O OH- HOOHOHHOO OeHH woflkom COHmsmcH HOHHCOU .OOOOHHOOO-.OH< OHOOO 123 OH. NN. ON. NO. OO. HO. OO. OO. ON. HO. OO. OO. NO. OO. NO. NO. NN. OO. OO. NO.H ON.N. HO.H NO. NO. HsHa OHNHHO OO. OO. ON. NO. OO.H OO. NO. NO. OHOO onO OOHHH OH OH OH OH OH OH HH HH HoHHo OHOOHOHO ONN ONN OON OON OON OON OH OH mamas OH NH OH OH OH OH OH OH HO HO NO NO OO OO OH OH OH OH OH OH OH ON HH HH ON ON ON ON HN NH OH OH NH HH HH NH NH HH OH HH OO OO HO NO OO OO OH OH HO: EEO OH NH NH OH OH HN NH NH OHOOOOHO cHo> HHOeO OHHO OH OH OH OH NH NH OH OH HOHHO OHOOOOHO «am How *wh *mn Own *Nw wan mam mfimofi ON ON ON ON ON ON NOH NOH HOH OOH OOH OOH NOH NOH ONH ONH ON HN HN ON OO NO OHH OHH OO OO OO OO OO OO ONH ONH OO OO OO OO OO NO NNH NNH OO ON ON ON OO NO ONH OmH HO: OOO OHOOOOHO ON ON ON OO NO ON OHH OHH OOOHO HOHHOHHO OHEOHOHO OO OO OO OO ON OH O OH- HOOHOHHHO OEHH OON-Hon COHWSWCH HOHHCOU .HHHUOHOEo: Ocm OmHSOOon HmHsoOm> OOEOOHQ Ham :QENH mo :OHumhucoocoo :Hououm Oonm nmsxa co OopscHe o0 How OQEHHmHom Oomsmnom NHHmnsumc oucH NHHOHHmpHm-OHHCH OomsmcH upscHs\mmwn ocstHOH: N: Oo mo Ouommmm-.oN< oHnOH 124 .Houucoo ow o>HuOHoH O0.0 W a u + .Houucou ou o>HHmHoH Ho.o w.m u H I HHOEO HHHO HHH NHH OHH OHH OHH OH OH OH HOHHO OHOOOOHO OOO OOO OHO ONO OOO ONO HHH NHH OOOOe OO OO OO OO OO OO OOH OHH NO OO OO NO OO NO OOH OOH OO OOH OOH OOH OHH OO OHH OHH OO OO OO OO OO OO ONH ONH OO OO OO OO OO OO OHH OHH OHH ONH ONH ONH OHH OOH NOH NOH HO: EEO OO OO OO OO OO OO ONH ONH OHOOOOHO HOHHOHHO OHEOHOHO OO OO OO OO ON OH O OH- HOOHOOHEO QEHH OOH-amn— fiOHmSMCH HOHHCOU .uHHuomem: Ocm OoHSOOmHm HOHsoOm> OOEOOHQ was cmENH mo coHumHHcoocou :proam Oonm :QENH co OmpncHE 00 How OQEHHoHow Ommsmuom NHHOHSHOG ousH NHHOHHQHHm-OHHCH NHmsoocmuHssHm OomsmcH ouscHE\oOmn ocHyzaochoHoc N: O wan muchE\oOmn oCHEOHOH: m: Oo mo Opommmm-.HN< OHLOH Table A21.-Continued. Infusion Period Control -10 60 50 40 30 20 10 Time (minutes) .13 .06 .02 .02 .03 .24 .01 .10 .27 .01 .01 .06 .09 .08 .01 .01 .07 .01 .02 .02 .04 .03 .02 .01 .01 .01 .01 .02 .01 .01 .05 .01 .07 .04 .04 .04 .04 .01 .01 .08 means standard error 002 i.002 i. 5.4 4.8 5.2 5.1 4.9 3.2 5.0 3.3 4.5 3.8 Lymph Total Protein M 3.4 5.7 6.8 3.5 3.3 3.0 3.3 4.7 (grams %) 126 5.8 6.8 5.5 5.6 5.8 6.5 3.1 5.2 5.0 4.7 6.9 6.0 5.3 3.1 4.6 2.7 3.7 4.4 2.4 3.7 4.1 2.1 3.7 3.7 2.5 3.7 2.3 1.9 3.0 4.0 3.7 4.2 3.3 3.9 2.8 4.0 3.8 4.3+ 4.3+ 4.5* 4.6* 4.5* 3.6 3.8 4.2+ means standard error +l +| +| qu\tnInOH'¢IO héthOm NZVNIDOVN [\mBOSOOI-n MVLQMOHW [\U;I\O':\O\OU; Plasma Protein (grams %) I\\O OH «)0 OH w \0 means standazd error m +| 127 .Hopucoo ou mififlou O0.0 W m u + .Honucou on OSHHHOHQH 8.0 w- Q n H mH M.H NH HOHHO fihmwcmum ONO ONO OO OOOOO NO OO OO OO NO NO OO NO NO OO OO OO OO OO OO HO OO NN OO OO OO HHHOOHOEOO OO OO OO OO ON OH O OH- HOOHOOHOO OeHO OOHROQ fiOHmDWH—H HO.H-thou .OOOHHHOOO-.HN< OHOOO 128 O O O OH NH HH OH OH OH O O O O HH OH HH OH OH HH NH NH OH HH HH HO: OH. OH OH OH NH OH OH NH NH OHOOOOHO OHO> HHOOO OHHO OH OH OH OH OHH OHH OH OH HOHHO OHOOOOHO Hmw Hmw va *Ow +Om «mu mOH mOH mcmmfi OO OO NO OO OO OO OHH OHH OO NO OO OHH ONH ONH ONH NHH OHH NOH NOH OO OHH NO OHH OOH NO NO OO ON ON ON NOH OOH OO OO NO NN ON ON OO NO HO: H5O OOH OOH OOH OOH OOH OO OHH OHH OHOOOOHO :oHOOHHOO NHH NHH HHH OHH OHH OH OH OH HOHHO OHOOOOHO «ow Ohm «hm Hem *Ow fiNm mHH wan mCNOE OO OO OO OOH NNH ONH NOH NOH OO OO OO OOH OO NOH ONH ONH OO OO OO OO NO OOH OHH OHH OO OO OO OO OO ON OHH OHH OO OO OO NO OO NN OOH OOH HO: EEO OHOOOOHO OOH OOH OOH OOH NO ON NNH ONH OOOHO HOHHOHHO OHOOHOHO OO OO OO OO ON OH O OH- HOOHOOHOO OeHH vow-Hon :ome-HCH HOHHcOU .HHHooumEo: van monummohm HmHsoOm> .mEOmHm Ocm amexfi mo :oHumuucoocoo :Hououm Oonm :QENN :o mouscHa oo How onmaH acmumcoo um womsmuoa OQEHHOHOM cucH NHHOHHmuHm-OpucH OomzmcH muchE\ommn ocHEmumH: m: Oo mo Opoomwm-.NN< oHnmb 129 N.H N.H N.H O.H N.H O.H O.H O.H HOHHO OHOOHOHO OO.O ON.O OO.O OO.O OO.O OO.O H.N O.N mamas 0.0 N.O O.O 0.0 N.O O.N O.N O.N O.O O.O O.O O.O O.O O.O O.H O.H H.O O.O O.O N.O 0.0 H.O H.N O.N 0.0 N.O O.O O.O O.O O.N H.H O.H N.O N.O O.O O.O H.O O.O O.N O.N HO OHOHOO O.O O.O H.O O.O O.O O.N H.N O.H OHOHOHO HOHOH HOOHH OO.H OO.H OO.H OO.H OO.H OO.H NOO.H NOO.H HOHHO OHOOOOHO OON. OON. ONN. OOO. ONO. OOH. HO. HO. OOOOa HN. ON. HO. OO. OO. OO. NO. NO. NN. NN. NN. OO. OO. HO. HO. HO. OO. HH. ON. ON. OO. NH. HO. HO. ON. NO. ON. OO. NO. NN. NO. NO. ON. NO. NO. OO. OO. ON. HO. HO. HOHO OHNHHO OH. NH. ON. HN. NN. OO. HO. HO. OHOO HOHO OOENH HH HH HH HH HH HH HH HH HOHHO OHOOOOHO OH OH OH HH HH NH OH HH mamas N O N N O O N N HH HH HH OH OH NH HH NH OO OO OO OO ON OH O OH- HOOHOHHOO OEHO vow-Hon Seam—HMO; HO.H-«=00 .vcscaucou-n.NN< manmh 130 .HofiEoo ou o>HumHoH mod Iv. m u ._. .3350 cu min—22 8.0 W a n O MH NH NH Ono-aha “Huntsman OOO OHO OO OOOoa NO NO OO OO OO NO OO NO OO OO OO ON NO OO OO NO OO OO HHHOOHOEO: N . H N . H H . H HOH-Ho whmvcmum OH.O O.O 0.0 OOOOs N.O O.O O.O O.O O.O O.O O.O O.O O.O O.O O.O N.O H.O O.O 0.0 HO OEOHOO O.O O.O N.O OHOHOHO OHOOHO OO OO OO OO ON OH O OH- HOOHOHHEO OeHH OOH-Hon teams-fl; HOHHCOU .OOOOHHcou-.NN< OHOOH 131 OH NN ON ON ON NO N N ON ON ON ON OO HO N N ON ON ON HN ON OO O O HO: HHH OO OO OO NO OO OO OH OH OHOOOOHO cHo> HHOHO OHHO OHH OH OH OH OH OH OH OH HOHHO OHOOOOHO OOOH OOOH OOOH OONH +OHH HOH OO OO OHOOs OOH NOH OOH NOH ONH OOH NO OO OHH OHH NHH NOH OO OO OO OO OHH OOH OOH OOH OOH OOH ONH ONH OOH OOH OOH OOH OOH NHH OO OO NHH OHH NOH OOH NO OO NO NO HO: OOO NOH OOH OOH OOH ONH OHH OO OO OHOOOOHO OOHOOHHOO NH NH OH OH mH OH OH OH HOHHo Humvcmum HOO HOO HOO OOO +NO +OO OOH OOH OOOOE OO OO NO OO NO NO NO OO NOH NOH NOH OO OO NN OOH OOH NO OO NO OO OO OOH OOH OOH OO OO NO NN NO ON OO OO NN ON ON NN ON OO OOH OOH HO: OOO OHOOOOHO OO OO NO OO OOH OOH OOH OOH OooHO HOHHOHHO OHOOHOHO OO OO OO OO ON OH O OH- HOOHOHHEO OOHO UOwHoO— COHmSMCH .HOOHHCOU .OHHHoouOEo; can Omusmmonm HOH3omm> .OEOOHQ cam :mENH mo coHuOHucoocoo :Hmuoum Oonm :QENH so OopscHe oo How onwcH unnumcoo Hm Oomsmuom OQEHHoHom oucH NHHmHuouHm-much NHmsoocmuHssHm vowsmcH ouscHE\mOmn mcwuzmmchoHoc N: O can muscHE\ommn ocHEOHOH: m: Oo mo Ouoommm-.ON< oHnmb 132 O.H O H O H O H O H N H O.H O.H Houno OHOOOOHO +O.O OO.O OO.O OO.O OO.O ON.O O O N.O mamas O.O 0.0 N.O O.O O.O O.O N.O N.O N.N H.O O.O N.O 0.0 O.O N.N N.N O.O O.O O.O O.O H.O O.O O.N O.N O.O O.O H.O O.O O.O N.O O.N N.N 0.0 O.O O.O O.O N.O 0.0 O.O O.O HO OOOHOO O.O O.O 0.0 O.O 0.0 N.O O.O O.O OHOHOHO HOHOO OOENH OO.H OO.H OO.H HH.H OH.H OH.H HO.H HO.H HOHHO OHOOOOHO «mm. *mm. #MV. «Hm. *hh. 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OHOO onO :OEHH OH OH OH OH MH OH HH HH H0990 Uhmvcmum OOO OHO OHO OOO OOO OOO NH NH OOOoa NO OO OO OO NO OO HH HH OO OO OO OO OO OO OH OH OO OO NO OO NO OO NH HH ON OO OO OO OO ON OH NH OO OO OO OO ON OH O OH- HOOHOOHEO oaHH UOMHGQ fioflmawcm HOHuflOU .OOOOHHOOO--.ON< OHOOO 136 .Houpcoo op o>HuOHmH mo.o w.m u + .Houucou on o>mum~ou .c.o w.m 4 H O... mH M.H NH HOHHO vhmvzmum «Ev «.0? mm mcmoa NO NO NO OO OO ON OO OO NO OO OO OO OO OO OO NO OO NN OO NO OO HHHOOHOeoz M.H O.H N.H HOHHO vhmwcmum O.O O.O O.O OOOOE O.O O.O N.O O.O O.O O.O O.O O.O O.O O.O N.N H.O H.O H.O O.O 0.0 H.O O.O HO OOOHOO O.O N.O O.O OHOHOHO OaOOHO OO OO OO OO ON OH O OH- HOOHOOHEO «OHO vofiumm consmcH Houucou .OOOOHHOOO-.ON< OHOOO B IBLIOGRAPHY 10. BIBLIOGRAPHY Appelgren, L. and D. H. Lewis. Capillary permeability-surface area product (PS) of Renkin in human skeletal muscle. Acta. Med. Scand. 184:281-282, 1968. Baker, C. H. Epinephrine-induced vascular volume changes in dog forelimbs with controlled flow. Am. J. 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