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THE EFFECTS OF MANIPULATION.
DISTENTION AND PHYSOSTIGMINE 0N
COMPARTMENTAL 81.009 FLOW IN

THE CAN IKE G E “EMU!

“Inst: for flu Degree of M. S.
MICHiGAN STATE UNEVERSITY

Bradford K. Grassmick
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f.’ ABSTRACT
*jffl\‘ \ THE EFFECTS OF MANIPULATION, DISTENTION
AND PHYSOSTIGMINE 0N COMPARTMENTAL BLOOD FLOW
IN THE CANINE GI TRACT
By

Bradford K. Grassmick

Many studies have shown that the motor activity of the intestinal
smooth muscle can affect local intestinal blood flow. These studies how-
ever, have only measured total blood flow to the intestinal wall, and thus
it is not known whether the distribution of blood flow within the gut wall
is altered by the motor activity. The present study was designed to study
this possibility. The motor activity was altered by 1) manipulation of the
intestinal wall; 2) distention of a segment of the intestine; and 3) intra-
venous infusion of physostigmine while blood flow through the mucosa, sub-
mucosa and muscularis layers of the canine gastrointestinal tract was mea-
sured by the microsphere method. Two types of radioactive microspheres
(15 i 5) labeled with 8SSr and 1“Ge were used. One type of the sphere was
injected before an experimental procedure as a control and the other type
after. A reference arterial sample was withdrawn at 3.88 ml/min for three
minutes from a femoral artery at each injection for the calculation of
blood flow.

In the first series of experiments, two segments of the small intes—
tine were exteriorized. One segment was distended to an intraluminal pres-
sure of 20-30 mm Hg with saline while the other segment was manipulated for
one minute by gently squeezing the wall. In the second series, physostig-

mine salicylate was infused intravenously at a rate between 0.2 and 0.9

Bradford K. Grassmick

ugm/kg/min to increase the lumen pressure of the intestine to 32 i 6
mm Hg.

Tissue samples, in duplicate, were taken from the gastric body,
duodenum, jejunum, ileum and descending colon in both series as well as
from the exteriorized segments in the first series. All samples were
separated into mucosal, submocosal and muscularis layers and weighed.

The radioactivity of the isotopes in each sample was measured and the
blood flow calculated from the radioactivity of the tissue, the reference
sample. Tissue samples were also taken from gall bladder, liver, spleen,
pancreas and adrenal gland in both series.

Manipulation and distention of the exteriorized segments increased
the total wall blood flow, submucosal flow and muscularis flow of the seg-
ments subjected to manipulation and distention. The mucosal flOW'WaS not
significantly altered. The percentage of the total wall blood flow per-
fusing the submucosa and muscularis was significantly increased. Blood
flow and its distribution in the wall of the gastric body, duodenum, je-
junum, ileum and descending colon which were left intact within the abdom-
inal organs studied was also not significantly altered.

Infusion of physostigmine significantly decreased the total blood
flow and mucosal blood flow of the gastric body, duodenum, jejunum and
descending colon. Muscularis and submucosal blood flows were not signifi-
cantly altered. The percentage of total wall blood flow perfusing the
submucosa and muscularis layers was significantly increased. Blood flow
to the spleen was significantly decreased, but flow to the other abdominal
organs studied was not altered by physostigmine.

In order to verify the results obtained by the microsphere method
in the first two series of experiments, the effects of manipulation, dis-

tention and physostigmine on intestinal blood flow were studied by directly

Bradford K. Grassmick

measuring the venous outflow in the third series of experiments.

A loop of the small intestine was exteriorized and divided into
two segments such that each was drained by a single vein. The veins were
then cannulated for the measurement of venous outflow. Manipulation and
distention both increased the venous outflow. The venous outflow, however,
was significantly decreased after the infusion of physostigmine. The magni-
tude of the changes in venous outflow produced by the three experimental
procedures were similar to the changes in total flow observed in the first
two series of experiments which utilized microspheres to measure blood flow.

These studies indicate that although total blood flow to the gut
wall is increased by manipulation and distention and decreased by the ad-
ministration of physostigmine, the changes are not shared equally by the
three tissue layers of the gut wall. The increased flow is not shared by
the mucosa and the decreased flow occurs only in the mucosa and not in the
submucosa or muscularis layers. All three experimental procedures redis-
tribute the flow among the three tissue layers in favor of the muscularis
and submucosa. The redistribution probably results from increases in the

motor activity of the muscularis.

THE EFFECTS OF MANIPULATION, DISTENTION
AND PHYSOSTIGMINE 0N COMPARTMENTAL BLOOD FLOW

IN THE CANINE GI TRACT

By

Bradford K. Grassmick

A THESIS

Submitted to
Michigan State University
in partial fulfillment of the requirements

for the degree of

MASTER OF SCIENCE

Department of Physiology

1975

ACKNOWLEDGMENTS

The author would like to extend his appreciation to Dr. C. C.
Chou for his diligence and patience in advising the preparation of this
thesis. The author would also like to express his thanks to Dr. Emerson

and Dr. Grega for their service on the examination committee.

ii

TABLE OF CONTENTS

List of Tables

List of Figures

Chapter
I. Introduction
II. Review of Literature

III.

IV.

VI.

Materials and Methods

1. Series A

2. Series B

3. Calculation of Blood Flow
4. Series C

5. Statistical Analysis
Results

1. Series A

2. Series B

3. Series C

Discussion

Summary

List of References

iii

Page
iv

24
25
27
28
29
31
32
32
33
40
48

59

61

LIST OF TABLES

TABLE

1.

The weight distribution of the three layers of the gut
wall in the five sections of the canine gastrointestinal

tract

Compartmental blood flow (ml/min/gm) in the manipulated

and distended segments (mean i-S.E.M.) N=10

Blood flow (ml/min/ gm) of the various abdominal organs
before (control) and after (experimental) manipulation
and distention of the exteriorized segments (mean i S.E.M.)

N310

Effects of physostigmine on the compartmental blood flows
(ml/min/gm) in five sections of the gastrointestinal tract

(mean t S.E.M.) N=7

Blood flow (ml/min/gm) of the various abdominal organs
before (control) and after (experimental) infusion of

physostigmine (mean f S.E.M.) N=7

iv

PAGE

30

36

39

43

44

LIST OF FIGURES

FIGURE

1. Total wall blood flow and its percentage distribution
to the mucosa, submucosa and muscularis-serosa in the

manipulated and distended segments N=10

2. Total wall blood flow and the percentage distribution
to the mucosa, submucosa and muscularis-serosa of the
remainder of the gastrointestinal tract within the

abdominal cavity N=7

3. Total wall blood flow and its percentage distribution
to the mucosa, submucosa and muscularis-serosa in the
wall of the gastrointestinal tract before and after

infusion of physostigmine N=7

4. Percentage change from control in venous outflow after
manipulation (N=7), distention (N=5), and the infusion

of physostigmine (N=lO) (mean i S.E.M.)

PAGE

35

38

42

47

CHAPTER I

Introduction

The motor activity of the intestinal smooth muscle has been
shown to affect local intestinal blood flow. Rhythmic contractions of
the intestine can produce a corresponding rhythmicity in intestinal
blood flow (56) while tonic contractions may decrease total blood flow
(58). Distention of the intestine may alter its motor activity and has
been shown to initially attenuate blood flow in proportion to the intra—
luminal pressure (44). These previous studies have only measured total
blood flow to the intestinal wall and thus it is not known whether
changes in blood flow due to distention or motoractivity are shared
equally by all three layers of the tract wall, i.e., the mucosa, sub-
mucosa and muscularis—serosa. It has been shown that luminal placement
of glucose or food increased blood flow only to the mucosal layer of the
gut wall. It is therefore possible that an increase in intestinal motor
activity may only affect blood flow to the muscularis layer. The pres-
ent study was designed to investigate this possibility. Intestinal
motor activity was stimulated by manipulation and distention of the gut

wall and by intravenous infusion of physostigmine.

CHAPTER II

Literature Review

A. Effects of Distention on Intestinal Blood Flow

The effect of intestinal obstruction and distention on blood
flow to the intestine has long been of physiological interest due to
its clinical consequences. By the turn of the century there were
nearly 5000 reports in the literature dealing with the effects of intes-
tinal obstruction or distention.

Mall in 1896, as cited by Lawson and Chumley (45), based on his
work with intestinal motor activity and distension, postulated that while
an increase in intraluminal pressure of the gut produced by distention
clearly decreased intestinal blood flow, such an increase might be expec-
ted to interfere with blood flow less than an increase in gut pressure
produced solely by increased muscle activity. It was thought that the
mucosal and inner muscle layers would buffer the distending pressure so
that the outer muscle layers would be little affected. Increased motor
activity, however, would directly compress the vasculature of the entire
wall.

Von Zwalenburg (63) in 1907, placed a cystoscopic light in the
lumen of an exteriorized canine intestinal segment and observed the effect
of increasing intraluminal pressure on blood flow in the intestinal wall.

The following is an excerpt from his study:

3

1. At 30 mm Hg pressure we found that some capillary streams
were arrested.

2. At 60 mm, many small veins had their currents arrested and
in most of them the stream.was so slow that individual
corpuscles could be seen.

3. At 90 mm a most interesting study presented itself. A11
blood streams were moving slowly and many were not moving
at all. One was impressed with the difficulties encount-
ered by the circulation in its attempt to find some point
of least resistance through which the stream might insin-
uate itself. Currents would at one time go in one direc-
tion and the next moment in the opposite. At the point of
branching of vessels, streams were seen going in all sorts
of direction.

4. At 130 mm all circulation had ceased--the corpuscles stan-
ding still in the blood vessels. Many vessels, however,
still carried the impulse of the heart beat--the corpuscles
moving to and fro, but retaining their relative positions
in the field. The pulsation was seen in some of the larg—
est and the most superficial arteries after the pressure
was carried up to 250 mm.

As the intraluminal pressure increased, blood flow became more
attenuated. Von Zwalenburg concluded that distention interferes with
the circulation in the intestine and allows infiltration of fluid into
the intestinal lumen and walls.

Van Buren (62) in 1920 studied the relation of intestinal dis-
tention and vascular resistance in the canine small intestine. The
following excerpts summarizes the results of this work:

"The greater the distention of the intestine, the less the

residual elasticity of its wall and the vessels in it. As

for the vessels themselves, their elongation results in a

narrowing of their lumen and a thinning of their walls. The

thinning of the walls results in greater compressibility.

These conditions are maximum at the anti-mesenteric border of

the intestine where the terminal vessels anastomose and, fail-

ing any interference with the mesenteric vessels, tissue dam-
age is earliest and most apparent on the antimesenteric
surface".

Van Buren found that necrosis of the intestine occured with dis-

tention of the gut. The degree of necrosis correlated with the duration

of distention indicating the cessation of blood flow was due to distention.

Catch, Trurler and Ayers, in 1927 (25, 26) found that gaseous
distention of a loop of canine small intestine decreased blood flow which
was prOportional to the distending intraluminal pressure. Blood flow was
measured by collecting the venous outflow of an exteriorized intestinal
segment. A minimal blood fIOW'WaS maintained in the presence of high
distending pressure. They postulated this flow to be from anastomotic
vessels in the gut wall. When the distending pressure approximated the
arterial pressure, there was a complete stasis of blood flow. Upon rel-
ease of the distention, a period of hyperemia was observed. Gatch pos-
tulated that distention producing hypoxia, could produce gangrene accou-
ting for the clinical manifestations of intestinal obstruction.

Dragstedt, Lang and Millet, in 1929 (19) produced gaseous disten-
tion in various portions of the canine gastrointestinal tract. In a
series of 10 observations on 4 animals, intraluminal pressures from 20 to
100 mm Hg were produced in the duodenum, jejumun, ileum and colon. Blood
flow measurements were made by cannulating the veins of an intestinal
segment and collecting the outflow. They found that blood flow through
the small and large intestine is inversely proportional to the intra-
luminal pressure in acute distention. The attenuation of blood flow was
most profound in the duodenum and distention had the least effect in the
more muscular colon. Dragstedt and coworkers reported a residual blood
flow that could not be further attenuated by increased pressure. Like
Catch, they considered this residual flow to represent anastomotic blood
flow.‘ The results of the venous outflow studies were confirmed by obser-
ving mucosal blood flow with a cystoscope placed in the lumen of the dis-
tended segment. It was observed that blood flow in the vessels of the

duodenum ceased at a much lower intraluminal pressure than in the colon.

With time, blood flow returned to control levels in all the distended
segments. .

Culbertson and Catch (24) in 1935 confirmed earlier work which
indicated that intestinal blood flow decreased with increased intralu-
minal pressure. It was observed that the venous outflow from a can-
nulated intestinal segment decreased from a control value of 18
mllminute during control to 3 mllminute at an intraluminal pressure of
200 mm Hg. Through microscopic studies they also found that the mucosal
layers were the most affected by distention. The mechanism by which
blood flow was attenuated was thought to involve both compression of the
capillary beds as well as occlusion of the veins by the increased intral-
uminal pressure. It was also observed that intestinal motility appeared
to decrease with intraluminal pressures above 10 mm Hg and that anemia
of the mucosa could be induced with intraluminal pressures as low as 20
mm Hg. Both secretion and absorption of the distended intestine appeared
to be impaired as well. The results are consistent with the earlier fin-
dings of Catch as well as those of Von Zwalenburg and Dragstedt. The duo-
denum, which has the largest percentage of mucosa per unit of wall weight,
was the most affected by increased intraluminal pressure.

Using an in §i£g_intestinal preparation, Lawson and Chumley (44,45)
in 1940 studied the effects of intestinal distention on both arterial in-
flow and venous outflow. A canine intestinal segment was pump perfused
with blood from a donor animal and the segment was distended by inflating
an intraluminal balloon. Blood flow changed in three phases during and
after distention. Upon inflation of the intraluminal balloon, blood flow
promptly decreased reaching a minimum level in 8 to 10 seconds (phase one).

In spite of maintaining the inflated balloon volume at the same level, the

blood flow returned towards control levels and remained steady at this
level within one to three minutes (phase two). The new fIOW‘WaS near con—
trol levels if the distending pressure was below 30 mm Hg. If the disten—
ding pressure was more than 30 mm Hg, the new flow was below control values.
However, in some animals blood flow near control values were obtained even
with distending pressures as high as 60 mm Hg. Upon release of the dis-
tention, blood flow increased above control.levels, reaching a‘maximum
level when the ballon was completely deflated (phase three). If the dis-
tention was maintained longer than six minutes, two additional phases of
blood were observed after deflation. The hyperemia following deflation
was often interrupted by periods of intense phasic motor activity during
which blood flow became phasic corresponding to rhythmic contraction of
the intestine (phase four). With cessation of the motor activity a period
of increased blood flow was again observed (phase five).

These responses were not observed in all animals during the exper-
iments. Upon distention to 50 mm Hg blood flow in four of five animals
during phase two increased above control levels and follOwing the release
of distention blood'flow in phases three, four, and five was normal.

When the intestine was prevented from enlarging in response to dis-
tention by encasing it in plaster, no recovery of blood flow during phase
two was observed (45). Upon deflation the encased segment showed recovery
of blood flow to control levels, but the hyperemia observed in the unen-
cased segments with release of distention was not observed. The encased
segments were shown to be capable of reactive hyperemia by clamping the
artery for 1 1/2 minutes or more, indicating that reactive hyperemia is
not a major factor in the recovery of blood flow. Reactive hyperemia was

elicited with intraluminal pressures as high as 80 mm Hg.

 

 

Lawson and Chumley (45) also studied the response of the intes-
tine to stretch without distention to see if stretch alone were the mech-
anism by which blood flow was altered. Intestinal segments were laid open
along the antimesenteric border and stretched transversely by hand. In
four of five animals tested flow increased with stretch and the flow res-
ponse was blocked by the local application of 0.1% atropine. The results
indicate that flow recovery may involve mechanisms which are independent
of increases in extravascular pressure and may involve intrinsic nerves
in the wall. Local anesthetics were also found to decrease the ability
of the distended segment to recover its blood flow during phase two (45).
When 1% cocaine hydrochloride was topically applied to the mucosa of the
distended segment the maximum distending pressure at which blood flow
could recover was reduced. The period of hyperemia, phase 3, following
the release of distention was markedly reduced as well. Intrinsic nerves
may then operate in a resistance lowering mechanism to compensate for the
effects of distention on blood flow.

The results of Lawson's and Chumley's study indicate that the
hyperemia observed after release of distention is not identical with the
phenomenon of reactive hyperemia. It also indicates that recovery of
blood flow is dependent upon the ability of the intestine to stretch in
response to increased intraluminal pressure and upon the integrity of
intrinsic nerves within the intestinal wall.

Noer and Derr, in 1949 (48) used India ink to visualize changes
in the blood vessels of the human intestine. By inflating an intralu-
minal balhxxxin isolated segments of the gut with gas, they raised the
intraluminal pressure to 70 mm Hg and released the distention stepwise
in 10 mm Hg decrements. Larger arteries began filling with ink when the

pressure was lowered to 50-—60 mm Hg, but capillary beds were not filled

until complete deflation of the segment (less than 10 mm Hg). Because

the segments were not perfused the results may indicate passive mechanical
effects of distention and not what would happen in a perfused an in_§itg_
intestine.

Boley and coworkers in 1969 (8) studied the effects of distention
of the canine intestine with several methods. Intraluminal pressures
were raised from O to 210 mm Hg in 30 mm increments and the effects on
total wall blood flow were studied with non-cannulating electromagnetic
flow probes. No consistent changes were observed in blood flow with intra-
luminal pressures of under 30 mm Hg. With further increases in lumen
pressure blood flow showed a reciprocal decrease until a pressure of 90
to 120 mm Hg was reached. Above this pressure further increases could not
reduce blood flow further. A residual blood flow, accounting for between
20 to 35% of the control blood flow remained at this level. Upon the

release of distention blood flow increased above control levels and syste-

matic pressure transiently decreased.

It was also found that with increased distention, the_p02 of the
venous effluent increased. This was thought to indicate an increase in
the percentage of blood flow passing through non-nutritional or.anastome
otic blood vessels.

Boley also used the Kr-85 washout technique to determine the dis-
tribution of blood flow within the intestinal wall. The washout curve
was resolved into three components accounting for 26%, 53%, and 21% of
the total flow and represent mucosal, submocosal, and muscle-serosal flows,
respectively. With distention only the muscle-serosal and submucosal comr
ponents were observed indicating a shift of blood flow away from the

mucosa. This observation is consistent with the views of Catch who pos-

tulated that the mucosa was the most sensitive to the effects of distention-

Measurements of venous outflow done by Hanson and Moore (33, 34)
in 1969 indicate that the canine colon has some ability to autoregulate
its blood flow. This ability to autoregulate was lost when either the
venous pressure or the intraluminal pressure was increased. When the
intraluminal pressure in the colon was increased to 50 mm Hg blood flow
was found to decrease 65%. As intraluminal pressure was further increa-
sed the venous outflow from the distended colon was further diminished.
They also found that when venous pressure was raised to 25 mm Hg, blood
flow decreased 44% in the colon.

-More recently Hanson (32) observed that while intestinal blood
flow initially decreased with distention to an intraluminal pressure of
50 mm Hg in the canine ileum, flow tended to show recovery within a few
minutes. The recovery of blood flow in the distended gut was attributed
to the attenuation of lumen pressure by stress relaxation of the gut wall.
Part of the recovery was also attributed to an active autoregulatory mech-
anism. It was found that infusion of papaverine abolished the ability of
the distended intestine to autoregulate. Before distention, blood flow
in the papaverinized animal was higher than in the untreated animal. With
distention blood flow in the papaverine treated animal fell to below the
level seen with distention in the untreated animal.

In summary, all these studies on distention and blood flow indicate
that acute distention which raises intraluminal pressure above 30 mm Hg
decreases blood flow to the distended segment. Distention has the grea-
test effect on the mucosal layer of the gut wall. Blood flow tends to
return to control levels within a few minutes if the distending pressure
is below 30 mm Hg and sometimes with intraluminal pressures of up to 60

mm Hg. The mechanism of blood flow recovery seems to involve both stress

lO

relaxation of the wall as well as intrinsic nerves which can be affec-
ted by local anesthetics. Hanson (32) postulated that an active auto-
regulatory mechanism was also responsible for some of the recovery of
blood flow.

B. Effects of Acetylcholine and Intestinal Motility_on Intestinal
Blood Flow.

 

Mott and Haliburtion (46) in 1897 studied the effects of choline
in the dog. Intravenous injections of choline, isolated from the cere—
brospinal fluid, produced a marked vasodilation in the intestinal vas—
culature of the dog as well as a decrease in both heart rate and sys-
temic pressure.

Hunt (36) in 1918 studied the effects of acetylcholine in the
vasculature of the cat. Acetylcholine, in a dose of 2.4 x 10-9 mgm/kg,
produced a negative inotropic effect on the heart. It was also observed
that acetylcholine produced a vasodilation of the limbs as measured with
a plethysmograph. Dale as cited by Hunt (36), working about the same
time, found that acetylcholine can produce an increase in the intestine
as indicated by increased volume on a plethysmograph. Larger doses of
acetylcholine increased intestinal motor activity which decreased blood
flow in the intestine as measured by collection of venous outflow. It
was also found that stimulation of intestinal motor activity by acetyl-
choline could be blocked by the administration of atropine.

Anrep,Cergua and Samaan, (l) in 1934 studied the effects of motor
activity on blood flow in the canine intestine. Ileal segments, 4-6 cm
long, were perfused with blood from a donor animal at constant pressure.
It was observed that both spontaneously occurring intestinal contractions
and those elicited by vagal stimulation caused blanching of the intestinal

segment. Stronger contractions were accompanied by a greater decrease in

11

intestinal blood flow. These observations were confirmed by determining .
the hemoglobin content of the contracting and quiescent segments. Con-
tracting segments were found to have 40-60% less hemoglobin than the quiet
segments indicating a lower blood flow in the contracting segments.

Sidky and Bean (57) in the 19503 studied the effects of spontan-
eous motor activity on blood flow in the canine gut. Isolated intestinal
segments were perfused with heparinized dog blood and kept at constant
temperature. Blood flow was measured by drop recorders and a thermopile.
It was found that rhythmic motor activity produced a rhythmic alteration
in venous outflow. Venous outflow was transiently augmented with contrac-
tion of the wall while arterial inflow decreased. Upon relaxation the
reverse was observed,_arterial inflow was increased and venous outflow
diminished. 0n the basis of these results, Sidky and Bean postulated that
rhythmic contractions of the intestine acted as a muscle pump to increase
blood flow. In support of this hypothesis was the observation that during
contraction venous pressure often was elevated above the perfusion pres-
sure. Attempts to perfuse the segment backwards through veins indicated
considerable resistance possibly due to venous valves which would be func-
tionally consistent with the action of a muscle pump.

Later studies of Sidky and Bean (4, 58) indicate the importance of
wall tension in intestinal blood flow. It was observed that decreased
wall tension will augment blood flow while increased tension will restrict
blood flow. Consistent with this observation is the effect of tonic intes-
tinal contractions on intestinal blood flow. Tonic contractions were ob-
served to increase venous outflow transiently as blood was forced from the
vessels of the wall, after which flow decreased. Following the tonic con-

traction, flow was observed to increase above control levels. The hyperemia

12

following tonic contraction was postulated to be the result of the release
of vasoactive metabolites from the muscle.

Acetylcholine, used by Sidky and Bean (5), was found to increase
blood flow in the intestine in the absence of any motor activity. Larger
doses of acetylcholine were found to cause tonic contractions and decrease
intestinal blood flow. Dilation of the intestinal vasculature invariably
preceded the onset of motor activity.

Boatman and Brody (7) in 1963 also studied the effects of acetyl-
choline on intestinal blood flow in the dog. It was found that infusion
of 1-20 ugm bolus injections of acetylcholine produced an initial decrease
in ileal blood flow. The increase in vascular resistance was hypothesized
to be the result of norepinephrine release from tissue stores. A sympa-
thetic depleting agent, reserpine, was applied, but the effect remained
indicating that it was not a sympathomimetic effect. A ganglionic bloc-
king agent, hexamethonium, was also applied, but the increase in resis-
tance was still elicited. AtrOpine blocked the effect indicating that
acetylcholine acted directly upon the intestinal muscle. Following the
initial vascular constriction, resistance decreased and the motility of
the-gut increased. During the increased motility, blood flow was found
to be at control levels.

Scott and Dabney (54) studied the relationship of but motility
and blood flow in the canine intestine. A segment of the ileum was exter-
iorized and the veins cannulated for the measurement of outflow. It was
foundthatintravenous infusion of acetylcholine at 5 ug per minute produced
an increase in lumen pressure from 15 mm Hg to 35 mm Hg. Ileal blood flow
also increased from 0.3 ml/min/gm of tissue weight to 0.5 ml/min/gm with

acetylcholine infusion. At higher infusion rates ileal lumen pressures

13

were further increased, but blood flow decreased somewhat though it
remained above control levels. When infusion was stopped, lumen pressure
dropped to control levels and a period of hyperemia was observed. It was
concluded that intestinal motility could markedly effect vascular resist-
ance in the intestine. It was also observed that the vasodilatory effects
of acetylcholine could be masked by extravascular compression produced

by increased motor activity.

The dual effect of acetylcholine on blood flow was also reported by
Chou,Dabney, Scott and Haddy in 1967 (11, 12, 13). Using pump perfused
canine ileal segments, acetylcholine was infused and the effect on intes-
tinal compliance and resistance were determined. Acetylcholine, infused
at l ug/min, decreased vascular resistance with little effect on ileal wall
tension. Infusion at 4 ug/min decreased vascular resistance still further,
but increased ileal wall tension. At 10 ug/min acetylcholine produced a
marked increase in ileal wall tension and lumen pressure; vascular resist-
ance showed a concomitant increase as well. It was concluded that at the
higher infusion rate the increased wall tension masked the active vaso-
dilation produced by the acetylcholine. It was postulated that the increase

in vascular resistance was the result of vascular compression by the vis-

- ceral smooth muscle.

Price, Shedaheh, Thompson, Underwood and Johnson, (51) in 1969
studied the effects of motility on mesenteric blood flow. Blood flow in
the mesenteric artery of the canine gut was measured with an electromag-
netic flow probe and motility determined with an intraluminal ballon at-
tached to a pressure transducer. Acetylcholine was infused at 14 Ugm/min
for 7 minutes. An increase in peristaltic waves was Observed, accompanied

by an increase in lumen pressure from 6 mm Hg before infusion to 26 mm Hg.

14

Blood flow was found to increase to 68% above control levels. The increase
in blood flow was found to precede the increase in motility by approximately
30 seconds. When the lumen of the intestine was filled with saline to in-
crease wall tension mesenteric blood flow did not decrease. Price con-
cludes that a large increase in wall tension need not increase vascular re-
sistance. These views are in contrast to those of Chou, Scott, and Dabney
(11,12, 13, 54).

Brobman, Jacobson and Brachner (9) in 1970 studied the responses of
canine intestinal segments to increased intraluminal pressure and infusion
of acetylcholine. Isolated ileal sugments were perfused with blood from
a donor animal and the venous outflow was measured. It was found that in-
flation of an intestinal segment to an intraluminal pressure of 40 mm Hg
Caused blood flow to decrease from 21 ml/min before inflation to 9 ml/min
in that segment. A 10 pg injection of acetylcholine increased blood flow
from 27 ml/min during control to 39 ml/min without increasing intraluminal
pressure. In one series of experiments the ileal segments were filled with
saline. When sufficient acetylcholine was infused to elicit motor activity
and increase intraluminal pressure, blood flow decreased. Saline was then
withdrawn to keep intraluminal pressure constant and an increase in intes-
tinal blood flow was observed. Rhythmic changes in lumen pressure resul-
ted in rhythmic blood flow though average flow was not altered.

Semba,Kazumoto and Fuiji, in 1971 (56) measured the effects of
rhythmic and tonic contractions on blood flow in the canine small intestine.
Intestinal pressure was measured in an exteriorized loop and vagostigmine
(0.05 mg/kg), eserine sulfate (0.1-0.2 mg/kg) or hypertonic salt were ad-
ministered to alter intestinal activity. Arterial and venous blood flow
were measured with electromagnetic flow probes. Application of hypertonic

salt to the serosa side of the intestine transiently inhibited intestinal

15

motility and concurrently increased blood flow indicating an inverse
relationship between intestinal motility and blood flow. They also found
that intestinal contractions caused a periodicity in arterial and venous
blood flow which he divided into three patterns. The patterns are the
contraction phase type, the relaxation phase type, and the combination
types. Intestinal contractions were then elicited by eserine or vagos-
tigmine and it was observed that venous flow increased but arterial flow
showed a simultaneous decrease. The increased venous flOW'WaS attributed
to active expiration of blood from the wall by the contraction of the wall.
As arterial flow began to recover there was a corresponding decrease in
venous flow which was attributed to refilling of blood vessels in the wall.
Semba referred to this as the contraction phase pattern.

The relaxation phase pattern occurred when both arterial and venous
flow were decreased with intestinal contraction. Minimal blood flow cor-
responded to the peak of contraction. With relaxation of contraction,
blood flow was augmented.

A combination of the contraction and relaxation types was charac-
terized by an initial increase in both arterial and venous blood flow with
the beginning of contraction. With maximal contraction flow decreased in
both arteries and veins and was increased with relaxation. Rhythmic con-
traction resulted in an increase in venous flow with each contraction and
a corresponding decrease in arterial flow. During relaxation the reverse
was observed, arterial flow increased and venous flow decreased. The
results indicate that intestinal motor activity can actively influence
intestinal blood flow.

Kenwater (42) in 1971 infused acetylcholine at varied rates and

measured the effects on intestinal motility and blood flow in the cat.

16

Blood flow was measured in the superior mesenteric artery with a closed
drop recorder and intraluminal pressure was determined from an exter—
iorized segment of the proximal jejunum. It was found that spontane—
ously occurring contractions did not alter intestinal blood flow. In-
fusion ofacetylcholine between 0.1 and 100 ugm/min produced increases in
motility roughly proportional to the dose. As is consistent with the res-
ults of other investigators, it was found that strong contractions of the
intestine decreased blood flow. Following intestinal contraction, a per-
iod of hyperemia was observed. Rhythmic contractions of the intestine
were not observed to exhibit the positive pumping action described by
Sidky and Bean.

In 1973 Zeigler, Barton, and Swan (67) studied the effects of mo—
tility on intestinal blood flow in the in gigg_canine gut. Both acetyl-
choline and metacholine (which is not vasoactive) were administered to
stimulate intestinal motility. It was found that acetylcholine infusion
at 1.5 ugm/kg/min increased superior mesenteric blood flow as measured by
an electromagnetic flow probe. Motility, which was measured by an index
of magnitude and duration of activity was found not to be significantly
greater than spontaneously occurring activity. Intraluminal pressure was
significantly increased from 20 mm Hg during control to 40 mm Hg. Meta-
choline, infused at 0.3 ugm/kg/min produced a similar increase in intra—
luminal pressure, but superior mesenteric blood flow was not significantly
altered. Infusion of metacholine at 0.7 ugm/kg/min produced a signifi-
cant increase in the index of motility and intraluminal pressure reached
peaks of 100 mm Hg. A typical intraluminal pressure tracing of all three
infusions showed rhythmic contractions with each. It was found that mes—
enteric blood flow was not significantly altered by the increased motility

stimulated by metacholine. Zeigler and coworkers conclude that intestinal

17

motor activity and blood flow are not necessarily related in canine gut.
They postulate that results obtained by other workers in the field may be
the result of the vasoactive properties of the stimulating agents such as
acetylcholine.

The majority of evidence in the literature supports the following
conclusions offered by Jacobson (38). Lumen pressure and intestinal blood
flow seem to be inversely related. Acetylcholine tends to increase blood
flow if vigorous motor activity is not elicited. Vigorous contractions
decrease intestinal blood flow due to increased wall tension. Contrary to
Sidky and Bean, contractions of the intestine do not seem to exert a posi—
tive pumping action on the blood.

C. Measurement of Compartmental Blood Flow

Several methods have been utilized by researchers for determining
compartmental blood flow in the gastrointestinal tract. In 1949 Kety as
cited by wagnerémd Ledingham (64), first described the use of radioactive
gas to determine blood flow distribution. An intra-arterial injection of
either 85Kr or 133)(e is made and the washout of the gas is measured by a
scintillation counter placed near the organ. The radioactivity is plotted
with time on semi-log paper and the resulting washout curve is resolved
into its various components. Assuming equal distribution of the gas init-
ially, the radioactivity of the gas will be proportional to the blood flow
in that compartment.

Kampp and Lundgreen (40) in 1966 used the method to determine comr
partmental flow in the cat intestine. They resolved the multi-exponential
washout curve into 4 components. The first component consisted of 35-50%
of the initial activity and also had the shortest half time value. The
half time value is the time required for the initial activity to be re-

duced by 50% and is an indicator of the rate of blood flow. A short half

18

time value indicates a high rate of washout and thus, a high rate of
blood flow. Kamp and Lundgreen suggest the first component represents
counter current exchange in the villi of the mucosal layers. The second
component had a longer half time value and accounted for less than 30
to 40% of the initial activity and was thought to represent blood flow
within the mucosal layer. Blood flow in the mucosal layer was estimated
to be 30-70 ml/min/lOO grams of tissue. The third component of the wash-
out curve was thought to represent muscularis flow and accounted for ap-
proximately 25% of the initial activity. Muscularis blood flow was esti-
mated as 10-20 m1/min/100 gm. The last component was thought to be the
result of blood flow through the perivascular fat rather than a component
within the intestinal wall. To verify the results of the 85Kr washout
technique, Kampp and Lundgreen autoradiographed sections of the intestinal
wall and the weights of the various wall components were compared and
found in agreement with the results of the washout technique.

Selkurt,Scibetta and Cull (55) in 1967, used the xenon washout
method to study flow distribution in the canine gut. Intra-arterial in-
jections of 133Xe were made and the resulting washout curve was resolved
into three components. In order of the magnitude of blood flow, these
components were thought to represent epithelial gland tissue (148.8 ml/
min/100 gm), smooth muscle (35.7 ml/min/lOO gm) and supportive and con-
nective tissue (3.4 ml/min/lOO gm). .Blood flow to the whole wall was
calculated to be 64.2 m1/min/100 gm from the various compartmental values.
Using electromagnetic flow probes flow to the whole wall was simultane-
ously measured. By this method, the flow was found to be 63.1 m1/min/100
gm which is in close agreement with the 133Xe method.

Another method of determhfing compartmental blood flow is the use

uz 86
of an arterial injection of radioactive potassium ( K) or rubidium ( Rb).

19

If the extraction of the isotopes is equal in all tissues then the radio-
activity in any tissue will be proportional to the blood flow through
that tissue. This method was used by Delaney and Grim (17) in 1964 to
determine the distribution of flow in the gastric body of both anesth-
etized dogs. In the anesthetized animals mucosal flow accounted for 72%
of the total flow through the wall, submucosal flow accounted for 13% and
muscularis flow was 15% of the total. In the unanesthetized animal, the
flow in the mucosa accounted for 74% of the total while the submucosa and
muscularis accounted for 14% and 12%, respectively.

The most recent means of estimating compartmental blood flow is
the radioactive microsphere technique. Glass microspheres were first used
by Prinzmetal et al. (52) to demonstrate functional arteriovenous shunting
in the heart and other organs. Ryan as cited by Wagner and Ledingham (64)
first produced radioactive microspheres by bombarding the glass micro-
spheres with neutrons and converting some of the sodium in the glass to
2“Na. The microspheres which would become trapped in the tissues would
then be an indicator of blood flow. The glass microspheres had the dis-
advantage of being much more dense than blood and tended to settle out.
Although the glass microspheres did not exhibit the same rheology as blood,
Grim and Lindseth (30, 31) used them in 1958 to study the canine small
intestine. The microspheres (12 u diameter) were injected into a local
artery of an isolated loop of the intestine. Blood flow from the 100p
was also measured by collecting the venous outflow. The wall of the seg-
ment was separated into mucosal, submucosal, muscularis and serosal layers
for measurement of compartmental radioactivity. Blood flow in all four
layers was calculated to be approximately the same averaging 40-50 ml/min/

100 gm of tissue in the jejunum. In the ileum, muscularis blood flow was

20

found to be slightly higher. Anastomotic blood flow although low, 2-4%
of the total, was also demonstrated in the intestine with the use of mic-
rospheres. The finding of anastomotic flow through the intestine tends
to support the hypothesis of Dragstedt and others who postulated that the
irreducible flow found with distention was, at least in part, through
anastomotic vessels.

Gastric blood flow in the dog was studied with the microsphere
method by Delaney and Grim (17) in 1964. Glass microspheres labeled with

24
Na, 16-20 u in diameter,were injected in the celiac artery along with

“2K and tissue samples were taken from the gastric body. Blood flow was
found to be distributed primarily to the mucosa, 68%, with the submucosa
and muscularis accounting for 12% and 20%, respectively. These values are
in close agreement with those obtained from the ”2K.method reported earlier.
Total gastric blood flow was calculated to be 0.54 ml/min/gm which is con-
sistent with the findings of Grim and Lindseth.

By 1966 both ceramic and plastic microspheres were available in
a variety of diameters and labeled with any one of a number of radionuclides.
Many studies have been done to determine the accuracy of the microsphere
method. According to Wagner et al., the validity of the method as an ac-
curate indicator of bloOd flow rests upon four assumptions (64, 65).

The first assumption, that the microspheres are adequately mixed
with the blood and exhibit the same rheology as red blood cells, has been
adequately verified by Phibbs, et al., (49, 50), Nuetz et al., (47) and
Kiahara et al., (43). Kiahara,Vanheerden, Migata and Wagner, (43) found
that the spheres were adequately mixed with the blood (for the systemic
circulation) if they were injected either in the left atrium, left ven-

tricle or in the origin of the aorta. The left ventricle and aorta did

not provide adequate mixing time for the coronary circulation, however.

21
The results of Phibbs studies indicate that within the arteries, the
smaller diameter spheres, 7-10 and 15 u, distrflnna themselves in the
axial stream in approximately the same proportion as red blood cells.
Larger diameter spheres showed more deviation from the natural distri-
bution.

The second assumption, that the microspheres themselves do not
alter the circulation, has been demonstrated by Kaihara et al., (43) as
well as Hoffbrand and Forseyth (35). It was found that injections of up
to four different types of microspheres did not alter the blood flow
distribution. In one study a small, but statistically significant change
in resistance was found with the third injection.

Another assumption of Wagner and Ledingham (64) is that the radio-
active label remains bound to the microspheres and that the microspheres
remain within the capillary beds. Since the nuclide is an integral part
of the sphere rather than a coating on the surface it is not possible for
the two to become separated. Kaihara and coworkers (43) have shown that
the spheres are neither moved nor metabolized for up to two weeks. An
intracardial injection of 50 u microspheres was made and the activity of
the various organs was monitored for two weeks with an external counter.
Urine and feces were also checked for the first five days following the
injection. No change in the radioactivity of the organs was detected nor
was any significant radiation found in the urine or feces.

The final assumption is that the spheres are removed from the blood
in the first pass through a capillary bed. Kaihara et al., (43) found
that 5-10% of the 15 u diameter spheres could pass through a capillary bed,
but that virtually all the spheres were removed by the pulmonary capil-
lary beds. This indicates that the microsphere method may not measure

total blood flow through an organ, but only flow through nutritional channels.

22

It is also indicated that recirculation of the spheres is not a problem.
Greenway and Murthy (29) utilized the microsphere method to study
blood flow in the intestinal wall of the cat. Carbonized plastic micro-
spheres (15 i 5 u) labeled with either Ce-14l or Cr-Sl were injected in
a local intestinal artery. Although both the spheres were obtained as
15 i’S u, it was found that the spheres differed in size. The 1“Ce
labeled spheres were significantly smaller in diameter (12 l 0.15 0)
than the 51Cr labeled spheres (17 i 0.16 u). When the smaller micro-
spheres were used, the distribution of microspheres in the wall was pri-
marily to the mucosa which received 47.4% of the injected activity. The
submucosa received 27.5% and the muscularis accounted for 7.8% of the total.
The remainder of the activity was found in the mesentery. When the larger
ler labeled spheres were used, the distribution of the spheres was altered.
The mucosal layer received only 26.3% of the total activity while the sub-
mucosa accounted for 50.0% of the total. The muscularis activity remained
relatively constant at 7.8% of the total. Since the smaller spheres ap-
peared to penetrate further into the mucosal layer than the larger spheres,
it was postulated that the mucosal and submucosal vascular beds were in
series. If the beds were in parallel the size of the microspheres would
. not alter the distribution. Since the distribution of the spheres to the
muscularis was not affected by the size of the spheres, the muscularis
vascular bed was though to be parallel with the mucosa-submucosa bed.
Injections of dye into the intestine tend to support this hypothesis.
The dye studies indicate an almost complete lack. of capillary sized ves-
sels in the submucosa except near the mucosal border. The submucosal ves-

sels seemed to be larger than 30 0 except near the mucosarnuino arterio—

venous anastomoses were observed.

23

Greenway and Murthy (29) performed another series of experiments
to validate this theory. The intestinal vasculature was maximally constric—
ted by intravenous infusion of vasopressin and the two types of micro-
spheres were injected. A segment of the intestine was then excised for
determining the distribution of the microspheres. The vasculature was
then dilated with isoprenaline (isoproterenol) and another intestinal tis-
sue sample was removed. When the vasculature was constricted, the larger
ll”Ce labeled spheres were found primarily in the submucosa (79.1%) with
the remainder in the mucosa. After dilation the distribution of spheres
between the mucosa and submucosa was altered. The spheres had redistri-
buted themselves such that 44.4% of the activity was found in the mucosa
and 55.6% of the total was in the submucosa. The smaller Ce labeled
spheres had initially been distributed nearly equally between the mucosa
(51%) and the submucosa (49%). Upon dilation the microspheres showed a
shift towards the mucosal layer (63.1%) away from the submucosa (36.9%).

As a result of this experimental evidence, Greenway and Murthy (29)
conclude that the microsphere method is not a valid technique for use in
vascular beds which lie in series with one another. In this case, Green-
way and Murthy indicate that the microsphere method will not be able to
differentiate mucosal and submucosal blood flow accurately. The relative
distribution of microspheres being determined by the diameter of the sub-
mucosal vessels. The muscularis vascular bed is in parallel with the

mucosa and submucosa and thus, may be distinguished from them.

CHAPTER III
Materials and Methods

Three series of experiments were performed to study the effects
of manipulation of the intestine, distention of the intestine and the
intravenous infusion of an anticholinesterase, physostigmine salicylate,
on total wall blood flow and flow distribution within the wall of the
gastric body, duodenum, jejunum, ileum and descending colon. Mongrel
dogs, weighing between 10 and 15 kilograms and fasted for 24 hours were
anesthetized with sodium pentobarbital (30 mglkg) and ventilated with a
positive pressure respirator (Harvard, Model 607, Dover, Mass.). Sys-
temic arterial pressure was monitored from a catheter in a femoral artery
which was connected to a pressure transducer (Statham, Model P23Gb) and
recorded on a direct writing oscillograph (Sanborn, MOdel 296).

In the first two series of experiments, series A and B, compart-
mental blood flows in the five sections of the gastrointestinal tract
were determined by the radioactive microsphere method (14). For this
purpose, an indwelling cardiac catheter was inserted through the chest
wall into the left ventricle of the heart for the injection of the mic-
rospheres. Presence of the catheter was confirmed by recording the left
ventricular pressure. Another femoral artery was cannulated with a poly-
ethylene tube (PE 280) filled with heparinized saline for the withdrawal

of a reference arterial blood sample.

24

25

Carbonized plastic microspheres, 15 i 5 u in diameter, labeled
with either strontium-85 (BSSr) or cerium-141 (1“ICe) were used to deter-
mine tissue blood flow. The microspheres lodge in the systemic capillary
beds in proportion to the blood flow through the bed. Blood flow through
a tissue was determined by comparing the radioactivity of the tissue to
that of the reference arterial sample withdrawn at a known rate as the
microspheres were injected (14). One type of microsphere was injected
before an experimental procedure as a control and the other types after.
The order of injection of the two types of microspheres was randomized
in the experiments. The spheres were obtained in 1.0 millicurie amounts
in 10 ml of a 10% Dextran suspension (0.1 millicurie/ml) from the Nuclear
Products Division of the Minnesota Mining and Manufacturing Company (3M
Center, St. Paul, Minn.). A drop of Tween 80 was added initially to the
stock suspension to prevent aggregation of the microspheres. Before use
in an experiment, the stock suspension was thoroughly mixed on a vortex
mixer and an aliquot of 0.4 ml, containing approximately 1.7 x 106 spheres,
was withdrawn and added to 2.0 m1 of 20% Dextran solution. This mixture
was then mixed with an ultrasonic sonifier cell dispersion of the spheres
just prior to injection. As each injection was made a reference arterial
blood sample was withdrawn at 3.88 ml/min for 3 minutes with a Harvard in-
fusion/withdrawal pump (Model 999) for the calculation of blood flow.

Series A--Manipulation and Distention

 

Ten to fifteen minutes following the cardiac and femoral cathe-

terization, one type of microsphere was injected and simultaneously a

reference arterial sample was withdrawn to estimate control blood flow in

26

the intact intestine.1 The abdominal cavity was then opened via a midline
incision and the most accessible loop of the small intestine was carefully
exteriorized. The loop happened to be the jejunum in 7 of the 10 dogs and
the ileum in the remaining three. The loop was then divided into two seg-
ments such that each segment was perfused by a single artery. A rubber
tube, 0.5 cm o.d., was placed in the lumen of one segment for the intro-
duction of saline. The mesentery was cut and both ends of the segments
were tied to prevent collateral blood flow; The segments were kept moist
and covered with a plastic sheet. One of the exteriorized segments was
manipulated by gently and thoroughly squeezing the wall between thumb and
fore finger for one minute. The other segment was distended by introdu-
cing 15 to 30 m1 of saline to the lumen to raise intraluminal pressure to
about 20 mm Hg. The second type of microsphere was then injected and a
second reference arterial sample withdrawn. The animal was then sacrificed
by an intraventricular injection of a saturated potassium chloride solu-
tion.

The two exteriorized segments and a segment of the gastric body,
mid-duodenum, jejunum, ileum and descending colon were excised. These seg-
ments were then separated into three portions, i.e., mucosa, submucosa,
and muscularis-serosa with a blunt instrument. Tissue samples were also
taken from the liver, spleen, the body of the pancreas, gall bladder and
adrenal gland. The tissue samples, in duplicate, were placed in tubes of
known weight and.‘reweighed for determining the net wet weight of the tis-

sue o

 

1The reason for measuring control flow before laparotomy and ex-
teriorization of the intestinal segments was that these procedures per se
redistribute the blood flow within the gut wall in favor of the muscularis
(unpublished observation). Since blood flow in an intact state was used
as a control, the changes in flow observed following one minute manipula-
tion and distention actually resulted from these maneuvers as well as sur-
gical manipulation performed during preparation of the experimental segments.

27

The radioactivity of each sample was measured in a two channel gamma
scintillation spectrometer (Packard Instrument Co., Model 3002, Tricarb scin-
tillation spectrometer). The channels of the spectrometer were set so that
the counting window of channel A included the primary gamma energy peak of
1“Ce and the window of channel B included that of 85Sr. Because of the
overlap of the emission energies of 85Sr, and ll”Ce, the Ce counting chan-
nel also measured some of the Sr emissions. The degree of overlap was de-
termined from the reference arterial blood samples and was corrected for in
each experiment. Each sample was counted for 10 minutes by which time be-
tween 4000 and 10,000 counts were accumulated. The radioactivity of each
sample was expressed as counts per minute per gram of wet tissue weight
(cpm/gm).

Series B

In this series, the effects of physostigmine on gastrointestinal
blood flow and its distribution within the wall were studied. After car-
diac and femoral catheterization, the abdomen was Opened via a small mid-
line incision and a portion of the small intestine was exteriorized.
Through an incision in the wall of the intestine a ballon was introduced
into the lumen and directed to stay in the area 20 cm from the incision.
The incisions were closed, the balLux1filled with water and connected to
a Statham pressure transducer through a rubber tube to monitor intestinal
luminal pressure. Following closure of the abdominal cavity, one type of
microsphere was injected and a reference arterial sample withdrawn. Phys-
ostigmine salicylate was then infused intravenously at a rate between 0.15
and 0.9 ug/kg/min until an increase in intraluminal pressure was observed.
When the intraluminal pressure showed a significant increase, between 20

to 50 mm Hg, infusion was stOpped and the second type of microsphere was

28

injected during the period of increased intraluminal pressure. A ref-
erence arterial blood sample was withdrawn after which the animal was
sacrificed. A segment of the gastric body, mid-duodenum, jejunum, ileum
and descending colon were excised. Each segment was then divided into
three portions, i.e., mucosa, submucosa, and muscularis. Tissue samples
were also taken from the liver, spleen, body of the pancreas, gall bladder
and adrenal gland. The radioactivity of the tissue samples was measured
as described for series A.

Calculation of Blood Flow

 

The results of series A and B are expressed as 1) total wall
blood flow, 2) compartmental, i.e., mucosa, submucosa, and muscularis
blood flow, and 3) the percentage of total wall blood flow to each com-
partment. Since it is technically difficult to cut small samples of the
gut wall which contained representative weights of each of the three
layers, the radioactivity of the whole wall was calculated from the radio-
activity of each of the layers and the weight distribution among the layers.
The weight distribution among the three tissue layers of the gastric body,
duodenum, jejunum, ileum and descending colon were determined previously
by Luke and Ya Mei Yu (66) in our laboratory (Table 1). With this mean
weight distribution and the radioactivity (cpm/gm) of each tissue layer,
the total radioactivity of the whole wall was calculated as follows:

Wall Activity = (cpm/gm of mucosa x % wt. mucosa) + (cpm/gm of

(cpm/ gm)
submucosa x % wt. submucosa) + (cpm/gm of mus-
cularis x % wt. muscularis)

From the radioactivity of the whole wall (RV), and the withdrawal

rate (3.88 mllmin) and the radioactivity of the reference arterial sample

(RA), the blood flow to the wall was calculated as follows (14):

29

3.88 mllmin
RA (Cpm)

 

Wall Blood Flow = Rw (cpm/gm) x
(ml/min/gm)

Similarly, blood flow in each tissue layer was calculated:

 

Tissue Blood Flow = Radioactivity in the tissue (cpm/gm)x 3&88(:1;?1n
(ml/min/ gm) A P

The blood flow to each layer in % of total wall blood flow was calculated
in the following manner:
Blood flow in one layer = radioactivity of the layer (cpm/gm) x
(Z of total blood flow)

weight percentage of the layer (Z)
radioactivity of the whole wall (Rw, cpm/gm)

 

Series C

The purpose of this series of experiments was to confirm the values
of total blood flow of the first two series of experiments in which micro-
spheres were used to estimate intestinal blood flow. The blood flow of two
adjacent igL§itg_jejunal segments was directly measured by timed collections
of their venous outflows under the same experimental conditions performed
in the first two series of experiments, i.e., manipulation, distention, and
the infusion of physostigmine.

A loop of the jejunum was exteriorized via a midline incision and
divided into two segments such that each was drained by a single vein.

After an administration of sodium heparin (6 mg/kg) the veins were can-
nulated for the collection of the venous outflow. The venous outflow was
drained into a reservoir and pumped back to the dog via a femoral vein at

a rate equal to the venous outflow. A rubber tube was placed in the lumen of
each segment for the introduction of saline and for monitoring intraluminal
pressure. The mesentery was then cut and both ends of the segments tied

to prevent collateral blood flow. The venous outflow from the two segments
were continuously or periodically collected in one-minute samples in grad-

uated cylinders.

30

Table 1. The weight distribution of the three layers
of the gut wall in the five sections of the
canine gastrointestinal tract

 

 

Organ Mucosa 7° Submucosa 7o Muscular-is Z
Gastric Body 49.9 20.3 29.8
Duodenum 66.6 ' 10.8 22.6
Jejunum 63.1 . 11.9 25.0
Ileum 55.5 13.4 31.1

Colon 36.1 ' 18.6 45.3

 

31

When systemic arterial pressure and venous outflow became stable,
one of the exteriorized segments was manipulated for one minute while the
other segment was distended with saline to an intraluminal pressure of
about 20 mm Hg. Measurement of venous outflow was continued until flow
became stable which usually occurred within 15 minutes. Physostigmine
was then infused via a femoral vein until the intraluminal pressure in-
creased to between 20 and 50 mm Hg. Venous outflow was measured until
blood flow became stable.

Statistical Analysis

 

In both series A and B the experimental and control flows were
compared using the Student's t test modified for paired comparisons. In
series C blood flows over time were compared with the control blood flow
using a Randomized Complete Block design analysis of variance and the
Student-Newman-Keuls test for the comparison of means (60). The results
of series A and B were compared with those of series C with a Completely
Randomized Design analysis of variance and the Student—Newman—Keuls test.

P values less than or equal to 0.05 were accepted as significant.

CHAPTER IV

Results

The systemic arterial pressure which ranged 85 to 135 mm Hg,was
not altered by any of the experimental procedures in all three series of
experiments.

Series A--Manipulation and Distention

 

Systemic arterial pressure ranged between 87 and 130 mm Hg,
(120 1'16, mean I S.E.M) prior to manipulation and distention. The
pressure was not significantly altered (P> 0.05) following manipulation
and distention, averaging 110 I 17 mm Hg.

Figure 1 shows the effects of manipulation and distention on
total wall flow and its distribution in the wall of the distended and
manipulated segments. Prior to manipulation and distention, the total
blood flows to the walls of the two segments were not significantly dif-
ferent (P> 0.05), averaging about 0.6 mllmin/gm in each. The distribution
of wall blood flow to the three layers of the gut wall were also similar
in the two segments; 82-85% of the wall blood flow perfused the mucosal
layer, 4-6% perfused the submucosa and 10.8% perfused the muscularis-serosa
layer.

Manipulation and distention produced similar changes in total blood
flow and compartmental blood flow distribution. Total wall flow was sig-
nificantly increased (P< 0.05), as the result of increases in muscularis

and submucosal flow. As shown in Table 2, muscularis blood flow increased

32

33

from 0.30 to 1.93 ml/min/gm following manipulation and increased from 0.28
to 1.24 ml/min/gm following distention. Submucosal flow was also increased
significantly, but mucosal blood flow was not significantly altered (Table 2).
These changes in flow caused changes in percentage distribution of the total
wall flow to the three tissue layers. The mucosal share of the total wall
flow decreased from 82-85% to approximately 45% while the muscularis share
increased from 10.8% to 37-49% following either manipulation or distention.
An increase in motor activity was visually observed in the exteriorized
segments following either manipulation or distention.

Figure 2 shows the total wall flow and its distribution within the
wall of the remainder of the gastrointestinal tract which was left undis-
turbed inside the abdominal cavity. Manipulation and distention of the ex—
teriorized segments did not significantly alter the wall blood flow nor its
distribution in the remainder of the tract. Blood flow was not altered to
the liver, pancreas, spleen, gall bladder, or adrenal gland (Table 3).

The results indicate that manipulation or distention of a segment of the
small intestine affects only the submucosal and muscular blood flow of the
segment and does not affect the mucosal flow of the segment or flows in the
undisturbed gastrointestinal tract and various abdominal organs.

Series B--Infusion of Physostigmine

 

Systemic arterial pressure was not significantly altered (P> 0.05)
by the infusion of physostigmine. The arterial pressure was 110 I 25 mm Hg
(mean I S.E.M.) prior to infusion and 113 i 11 mm Hg following infusion.

Intravenous infusions of physostigmine increased the lumen pres-
sure of the small intestine to 32 i 6 mm Hg. The tracings of intestinal
intraluminal pressure showed that the elevated lumen pressure was usually
at a constant level with few fluctuations. This indicated that the intes-

tinal motility response was a tonic contraction. The vascular effects of

Figure l.

34

Total wall blood flow (bars) and its percentage
distribution to the mucosa (bottom), submucosa
(middle), and muscularis-serosa (top of the bars)
in the manipulated and distended segments. C =
control, E = following manipulation or disten—

tion, d (mean i'S.E.M.) = difference between con-

trol and experimental total wall blood flows.

~The values within the bars represent the percen-

tage of total wall blood flow to that compart-

ment. N=10

 

L2-

L0-

Blood Flow (mllminlgm)
8 8

O
L.

02

 

0.0

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Figure l

"r , .
\ Musculans S
Submucosa EJ
Mucosa :3
* p<.05
d=.581;1&
48.5% __
\\ \
P 5
\ 6s.45:.|7\
*
\ 6.8%
g *
I'\°'° ’° 7.7% \
{4% l0.0%
o *
8437.43.99. 6-4" 161%
821%
t
45.7%
C E C E
Manipulated Distended

Table 2.

36

Compartmental blood flow (ml/min/gm)

in the manipulated and distended seg-
ments (mean i S.E.M.). N=10

 

 

Manipulated Segment Distended Segment
Control Exp. Control Exp.
Mucosa
0.83 i .12 0.93 i .28 0.86 i .17 0.76 i .17
Submucosa
0.25 4_- .0 0.84 i .33* 0.28 i .07 1.18 i- .47"c
Muscularis
0.30 i .07 1.93 i .50* 0.28 t .08 1.24 + .22"

 

Exp. = following manipulation or distention.

denotes values which are significantly different

from control (P(0.05)

Figure 2.

37

Total wall blood flow (bars) and its percentage
distribution to the mucosa (bottom), submucosa
(middle), and muscularis-serosa (top of the bars)
in the remainder of the gastrointestinal tract
within the abdominal cavity. C = control, E =
following manipulation and distention, d (mean t
S.E.M.) = difference between control and experi-
mental total wall blood flows. The values within
the bars represent the percentage of total wall

blood flow to that compartment. N=7

 

38

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Table 3.

39

Blood flow (ml/min/gm) of the various abdominal organs
before (control) and after (experimental) manipulation and

distention of the exteriorized segments
(mean i S.E.M.) N=10

 

 

Organ Control Experimental
Spleen 0.92 t .17 1.27 i .04
Liver 0.46 i .09 0.57 f .35
Pancreas 0.31 i .06 0.27 :-.08
Gall Bladder 0.36 i .08 0.17 i .06
Adrenal Gland 3.03 i .81 3.11 f 1.10

 

40

infusion of physostigmine are shown in Figure 3. Total wall blood flow
was significantly decreased in the stomach, duodenum, jejunum and colon.
Blood flow decreased in the ileum as well though the change was not sta-
tistically significant (Figure 3). The decrease in flow was primarily
the result of decreased mucosal flow in these organs (Table 4). Muscu-
laris and submucosal blood flows were not significantly altered. These
changes in compartmental blood flow produced a significant decrease in
the percentage of total wall flow perfusing the mucosal layer and a cor-
responding increase in the percentage of total wall flow perfusing the
muscularis of the small intestine (Figure 3). Although the distribution
of flows within the gastric and colonic wall was n0t significantly altered,
the same tendency of decreasing mucosal and increasing muscular shares of
the total wall flow was observed. As shown in Table 5, infusion of phy-
sostigmine decreased blood flow to the spleen, but did not alter flow to

the liver, pancreas, gall bladder or adrenal gland.

Series C--Direct Measurement of Venous Outflow of Jejunal Segments

 

The purpose of this series of experiments was to provide an in-
depenent verification of the microsphere method. The experimental pro-
cedures of the first two series, i.e., manipulation, distention and the
. administration of physostigmine, were repeated and the magnitude and di-
rection of changes in venous ourflow were compared with the results of
series A and B. All three experimental procedures, manipulation, disten-
tion and the administration of physostigmine, produced an increase in in-
testinal motor activity.

Manipulation and Distention

 

The systemic arterial pressure was not significantly altered,

(P> 0.05) by any of the procedures, averaging 120 I 5 mm Hg (mean I S.E.M.)

Figure 3.

41

Total wall blood flow (bars) and its percentage
distribution to the mucosa (bottom), submucosa
(middle), and muscularis-serosa (top of the bars)
in the wall of the gastrointestinal tract. C =
control, E = after infusion of physostigmine,

d = difference (mean t S.E.M.) between control
and experimental total wall flows. The values
within the bars represent the percentage of total

wall blood flow to that compartment. N=7

 

42

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44

Table 5. Blood flow (ml/min/gm) of the various
abdominal organs before (control) and after
(experimental infusion of physostigmine
(mean f S.E.M.).

 

 

Organ Control , Experimental
Spleen 1.02 t .10 0.54 f .17*
Liver 0.65 i .30 0.30 f .12
Pancreas 0.50 i .23 1.10 1- .77
Gall Bladder 0.30 t .07 0.30 f .13
Adrenal Gland 2.87 f .31 2.71 t .41

 

*
denotes values which are significantly different from control (P<0.05)

N=7

45

before and 126 I 2 mm Hg following manipulation and distention.

The average venous outflow before manipulation was 0.62 i .06
(mean * S.E.M.) mllmin/gm which was statistically similar to the value
found in series A, 0.63 I .10 ml/min/gm (Figure 1). Two and 1/2 minutes
after manipulation the venous outflow had significantly increased to 0.75
I .08 mllmin/gm (125% of control) (Figure 4). Statistically, this value,
0.75 ml/min/gm, was not significantly different from 1.20 i .31 ml/min/gm
obtained with the microsphere method in series A (CRD Analysis of Variance
and SNK test). Blood flow remained above control for 15 minutes following
manipulation, but usually returned to control levels within 25-30 minutes.

Similar results were observed in the distended segment. Before
distention, total blood flow was 0.64 I .14 ml/min/gm as measured by col-
lection of the venous outflow. This value was not significantly different
from that obtained with the microsphere method (0.57 i .09 ml/min/gm)
(CRD Analysis of Variance and SNK test). Distention of the segment to 20
mm Hg increased venous outflow to 0.79 i .16 ml/min/gm (125% of control)
after 2.5 minutes (Figure 4). When compared with the value from the micro-
sphere method, O.9 I .19 mllmin/gm, the two were found not to be signifi-
cantly different. Blood flow usually returned to control levels within
25-30 minutes.

Infusion of Physostigmine

 

Infusion of physostigmine increased intraluminal pressure to
approximately 45 mm Hg and decreased venous outflow 38% from a control
level of 0.74 i .11 to 0.46 i .07 ml/min/gm 5.5 minutes after starting in-
fusion (Figure 4). The values obtained in series B (Figure 3), 0.75 i .08
mllmin/gw iuring control and 0.35 i .10 after infusion of physostigmine,
were not gnificantly different from the Venous outflow values. Blood

flow was usually decreased for more than 30 minutes.

46

Figure 4. Percentage change from control in venous outflow
after manipulation (N=7), distention (N=5), and
the infusion of physostigmine (N=10), (mean t
S.E.M.). Control flow for distention = 0.64 i
0.14 ml/min/gm, manipulation = 0.62 + .06 ml/min/
gm, physostigmine = 0.74 mllmin/gm. * Denotes
values which are significantly different from con-

trol (P < 0.05) (Randomized Complete Block Analysis

of Variance and Student-Newman-Keuls test).

 

Percent change from Control

47

 

 

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Time (minutes)

Figure 4

CHAPTER V
Discussion

Many studies have indicated that motor activity of the gastroin-
testinal tract can affect its blood flow (1, 4, 5, 7, 51, 56). The purpose
of this study was to determine whether increased motor activity altered the
distribution of blood flow within the wall and if so, which tissue layer
of the gut wall is most affected. Intestinal motor activity was increased
by manipulation and idstention of the gut wall and by the administration of
physostigmine. Blood flow in the layers of the wall was measured by the
microsphere method.

Validation of the Microsphere Method

The validity of the microsphere method depends upon four factors:
1) adequate mixing of the microspheres with blood, 2) distribution of the
microspheres to the tissues in the same manner as red bkxnicells, 3) re-
moval of the microspheres from the blood in a single pass through a capil-
lary bed and 4) that the microspheres do not alter the circulation them-
selves (64). Adequate mixing of the microspheres with the blood when in-
jected into the left ventricle has been demonstrated (40, 47). Further-
more Phibbs,Dong, wyler and Neutze have demonstrated that the 15 i 5 0
diameter microspheres distribute themselves in the arteries in a manner
similar,if not identical to red blood cells. It has also been shown that
90-95% of the spheres are removed from the circulation by one pass through

a systemic capillary bed and the remainder are trapped in the pulmonary

48

49

circulation (43). Once in the tissues the microsphere remain inert and
do not alter the circulation (35, 43, 66). In addition, the microsphere
method has been shown to yield quantitatively similar results when com-
pared with more widely accepted methods of measuring blood flow such as
“2meethod and collection of venous outflow (17, 30, 31). On the basis
of these finding the microsphere method is indicated to be a reliable
method for measuring tissue blood flow.

In the present study the effects of manipulation, distention and
the administration of physostigmine on total wall flow were measured with
the microsphere method in the first two series of experiments and with
direct measurements made by collection of venous outflow in the third
series of experiments. The microphere method yielded values that were in
close agreement with the collection of venous outflow. For example, before
manipulation blood flow in the exteriorized jejunal segment was 0.63
ml/min/gm with the microsphere method and 0.62 by the collection of venous
outflow.‘ The values for the blood flow as determined by the two methods
were_not significantly different from one another either before or after
any of the experimental procedures.

The distribution of blood flow among the three layers of the gas-
‘trointestinal tract was also determined with the microsphere method. In
the wall of the gastric body, mucosal flow accounted for 71.7% of the
total wall blood flow during the control period. Submucosal flow accounted
. for 10.7% and muscularis flow for 17.6% of the total wall flow respectively.
These values are in close agreement with those reported by Delaney and
Grim (l7, 18) using the 1‘2K method and 2"Na labeled microspheres. The
distribution of blood flow among the three layers of the wall was found to

be mucosa, 72%, submucosa, 13% and muscularis, 15% as determined by the

50

l'ZK.method and 68%, 11% and 21% respectively with the use of 2“Na mic-
rospheres. Although the distribution of flow was found to be similan to
that reported by Delaney and Grim, the absolute values of both the total
wall flow and compartmental blood flows in the gastric body were found to

be lower in the present study. Total wall blood flow was found to be

0.20 ml/min/gm in contrast to 0.54 mllmin/gm reported by Delaney and Grim.
Mucosa, submucosa and muscularis blood flows were 1.1, 0.5 and 0.25 ml/min/gm
as determined by the 1'2K method and 0.91, 0.42 and 0.26 ml/min/gm respec-
tively by the 2”Na microspheres. In the present study however, mucosal

flow was found to be 0.31, submucosa 0.16 and muscularis flow 0.24 ml/min/gm.
The differences in calculated blood flow may be the result of differences

in protocol, technique and/or population samples. The comparison thus may
not actually reflect absolute differences in blood flow.

Blood flow and its distribution in the small intestine was also
determined in the present study. Total blood flow to the wall of the je-
junum.was between 0.93 and 0.62 mllmin/gm in the determinations made. The
results are consistent with those of Selkurt, Scibetta and Cull (55) who,
using the washout technique of 133Xe in the small intestine, reported total
wall flow to be 64.2 m1/min/100 gm. Grim and Lindseth (30) using 12 u glass
microspheres, found jejunal blood flow to be slightly less, averaging 50
‘ml/min/lOO gm. Blood flow was found to be similar in the three layers of
the wall averaging 40-50 m1/min/100 gm in each layer (30). The results of
this study indicate that the distribution of blood flow within the wall of
the gut is not so uniform as suggested by Grim and Lindseth (30% however,
Mucosal blood flow was found to be between 0.83 and 1.3 ml/min/gm while

submucosal flow was between 0.25 and 0.50 ml/min/gm. Muscularis blood flow

was found to be between 0.10 and 0.50 ml/min/gm which is in close agreement

with Selkurt. Using the washout of 133Xe, blood flow in the three layers

51

was found by Selkurt to be 148.8 ml/min/lOO gm in the mucosa, 35.7
mllmin/lOO gm in the submucosa and 3.4 ml/min/lOO gm in the muscularis (55).
The distribution of blood flow within the wall of the jejunum, as determined
by Yu (66) was primarily to the mucosa (70.1 t 3.5%). The submucosal
layer received 5.3 i 0.8% of the total flow to the wall and the muscu-
laris accounted for 24.6 I 3.3%. These values are in close agreement with
the distribution of wall flow found in this study. The mucosa received
79.1 i 5.3% and the submucosa and muscularis received 4.8 t 1.1% and 16.7
i .6%, respectively.

The measurements of blood flow and its distribution within the
wall of the gastroindstinal tract obtained by the microsphere method in
the present study are consistent with the measurements obtained with the
“2K, 133Xe methods, the measurement of venous outflow as well as those of
other workers using the microsphere method.

Response to Manipulation and Distention

The effect of intestinal distention has long been of physiolog-
ical interest due to the clinical consequences in humans. As a result,
the effect of distention on intestinal blood flow has been studied by
many investigators. The results of the present study indicate that dis-
tention of an intestinal segment to an intraluminal pressure of 20-30 mm
Hg was accompanied by a significant increase in total wall blood flow of
the distended segment from 0.63 to 0.90 ml/min/gm, within three minutes
after distention. However, the increase in the wall blood flow was not
evenly shared by all three layers of the wall, but was primarily due to
an increase in muscularis blood flow. Blood flow in the muscularis in-
creased from 0.28 ml/min/gm to 1.24 ml/min/gm. Blood flow was also in-

creased in the submucosa, but mucosal flow did not change. Although there

52

was a significant change in blood flow in the distended segment, the total
wall flow and compartmental blood flow in the remainder of the gastroin-
testinal tract which was left intact within the abdominal cavity was not
altered. This indicates that the effects of distention are local, invol-
ving only the muscle and submucosal layers of distended segment.

These results are consistent with those to other investigators.
Distention of an intestinal segment to an intraluminal pressure of less
than 30 mm Hg has been shown to decrease local blood flow transiently.

The flow then returns to levels at or above control within 1 to 3 minutes
even though the segment is still distended (8, 44, 45). Distention to
higher intraluminal pressures usually produces a decrease in total wall
blood flow (19, 24, 25, 26, 48, 63) although flow has also been shown to
increase with intraluminal pressures as high as 50 mm Hg (44, 45) and

to return to control levels with pressures of 60 mm Hg (32, 33, 34). The
finding is similar to that of others who found that recovery of blood flow
to its control level during distention is rapid, reaching a plateau within
one to three minutes after the beginning of distention (45).

In the present study, the venous outflow of the distended segment
increased after distention and reached a maximum after four minutes. Blood
flow remained above control levels for more than 15 minutes (figure 4),
which is consistent with the finding of other investigators (8, 32, 33, 34,
44, 45).

Since the systemic arterial pressure was not altered, the increase
in submucosal and muscularis flow indicates a decrease in the vascular
resistence of those layers. The mechanism of the decreased resistence was
not investigated in this study.

Wall tension of the intestine is an important factor in determining

vascular resistence (ll, 12, 13). Increased wall tension can increase

53

resistence to blood flow by increasing extra vascular pressure and de-
creasing vascular transmural pressure thereby compressing the vessels.
Conversely, decreased wall tension can decrease the vascular resistence.

It has been postulated by Hanson and Moore (32, 33, 34) that
the recovery of blood flow in the wall during distention may be a result
of stress relaxation of the gut and the subsequent decrease in wall ten-
sion. They observed that with distention the intestine enlarged which
was thought to be evidence of stress relaxation. Lawson and Chumley (44,
45) found that the decreased flow that occured during the first 10 seconds
of distention eventually returned to control levels even though the dis-
tending pressure was maintained in in_§i£u_intestinal segments. However,
when the intestinal segments were encased in plaster to prevent its en-
largement, the recovery of blood flow was not observed (45). These studies
support the theory that wall tension is an important factor in determining
vascular resistance during distention.

Reactive hyperemia was ruled out as a possible mechanism by
Lawson and Chumley (45). It was found that the encased segment was capable
of reactive hyperemia following temporary occlusion of the artery. In
spite of the ability of the intestine to respond to arterial occlusion,
blood flow did not recover during distention in the encased segment,in-
dicating reactive hyperemia was not involved.

Lawson and Chumley (45) also found that denervation did not abol-
ish the recovery of blood flow during distention which indicates that cen-
tral neural reflexes are not involved. However, the administration of a
local anesthetic, 1% cocaine hydrochloride, markedly attenuated the re-
covery of blood flow. The importance of local nerves in regulation of

blood flow was also demonstrated by Yu (66) who found that the hyperemia

54

associated with intraluminal placement of hypertonic glucose could be
blocked bytreating the mucosa with dibucaine before exposure to the glucose.

In addition to decreased wall tension and the action of local
nerves, active hyperemia may be involved in the increased blood flow
during distention. During distention, motor activity was usually observed
to increase (4, 5, 56, 57, 58). Stimulation of intestinal motor activity
may produce an active hyperemia in the muscularis layer (66). The active
hyperemia during exercise in skeletal muscle has long been established.
Such a mechanism would be consistent with the results of the present study
(3). The hyperemia observed after distention was primarily the result of
increased muscularis blood flow as this hypothesis would predict.

The increase in submucosal flow may be explained on the basis of
Greenway and Murthy's model of intestinal blood flow (29). According to
Greenway and Murthy the mucosal and submucosal vascular beds are arranged
in series. If mucosal resistance were slightly increased by intraluminal
pressure there would be a greater number of microspheres trapped in the
submucosal layer because of the series arrangement of the vasculatures.
Since the mucosa forms a larger part of the wall than does the submucosa,
small increases in mucosal resistance could produce apparently large in-
creases in submucosal blood flow. In the present study a decrease,'al-
though not significant, in mucosal blood flow was found and there was a
large increase in submucosal flow. These findings are consistent with
Greenway and Murthy's model of intestinal blood flow.

Regardless of the model of intestinal blood flow used to inter-
pret the data, it is clear that distention of an intestinal segment to an
intraluminal pressure of 20-30 mm Hg causes an increase in steady state
total blood flow to the wall. The increase is primarily the result of

increased muscularis blood flow. Mucosal blood flow was not significantly

55

altered. The mechanism of the increased flow in the muscularis was not
determined in this study, but may involve a decrease in wall tension from
stress relaxation, the actions of local nerves or an active hyperemia of
the visceral smooth muscle.

The effects of manipulation on the blood flow of the intestinal
wall have not been widely studied. Yu (66) postulated that manipulation
of the intestinal wall may produce a transient hyperemia via an increase
in intestinal motility.

In the present study it was observed that manipulation of the
exteriorized segments was followed by an increase in the motor activity
of the segment. Total blood flow to the wall of the segment was signifi-
cantly increased (Figure l) as the result of increased flow in the muscu-
laris and submucosal layers. Mucosal blood flow was not altered, however.
Because manipulation and distention produced similar changes in compart-
mental blood flow and motor activity, it is possible that the same mech-
anisms may by involved in the response to each.

Since the major factor in increasing total wall blood flow was
the increase in muscularis flow, a probable mechanism may involve the
release of vasoactive metabolites from the contracting muscle. Although
the submucosa is not extensively muscular, its proximity to the muscularis
could allow the diffusion of the metabolites to it and produce the hyper-
emia observed in this layer.

Another possible mechanism which may explain the hyperemia was
studied by Lawson and Chumley (44, 45). They found that stretching of an
intestinal segment tranversely produced an increase in blood flow. It is
possible that manipulation of the wall in the present study caused stretch-
ing of the visceral muscle which would account for the increased muscularis

blood flow. Since Lawson and Chumley (44, 45) found that local application

56

of atropine to the segment blocked the response to stretch, it is likely
that local nerves may be involved in this response.

Response to Physostigmine

 

Acetylcholine has often been used to stimulate intestinal motor
activity and investigators have reported a dual effect upon intestinal
blood flow depending upon the dose. When acetylcholine was infused at
low doses (4 pg/min) the vascular resistance of the intestine decreased
but at higher doses the effect of acetylcholine was to increase intestinal
motor activity and decrease blood flow (11, 12, 13, 54). Other researchers
however, have found that stimulation of intestinal motor activity by acy-
tylcholine increases intestinal blood flow (7, 51, 67) while simultane-
ously increasing intestinal motor activity. Although the effects of ace-
tylcholine on intestinal blood flow and motility have been studied by
many investigators, its effects on blood flow distribution within the gut
wall has not been studied.

In the present study infusion of physostigmine, an acetylcholine
esterase inhibitor, produced a significant increase in intestinal motor
activity as indicated by an increase in intraluminal pressure (+32 mm Hg).
Blood flow to the gut wall was significantly decreased in the stomach,
duodenum, jejunum and colon. The decrease was primarily the result of
decreased mucosal flow in these segments. Although not statistically sig-
nificant, both submucosa and muscularis flow tended to increase in the
duodenum, jejunum and ileum.

The decrease in mucosal blood flow may be related to the increase
in intestinal intraluminal pressure, stimulation of intestinal nerves or
compression of the vasculature within the gut wall. Similar intraluminal

pressures were produced by distention.inthis study with little effect on

57

the mucosal blood flow, thus it is doubtful that increased intraluminal
pressure played a significant role in decreasing mucosal flow.

Compression of the intramural vasculature by the muscle contrac-
tion may be involved in the decreased mucosal flow. Sidky and Bean (57,
58) have reported attenuation of intestinal blood flow when tonic contrac-
tions of the gut were elicited. It is possible that the active contrac-
tions of the muscularis passively compressed the arteries perfusing and
veins draining the mucosal layer thereby limiting blood flow to the mucosa.
Compression of the veins alone by muscularis activity may also be respon-
sible for'the decrease in mucosal blood flow. Increased venous pressure
may elicit arteriolar constriction via axmmo-arteriolar response and thus
decrease mucosal blood flow.

Stimulation of intestinal nerves may also account for the changes
in blood flow observed. Davenport (15) reports that acetylcholine pref-
erentially constricts the arterioles of the muscularis-mucosa layer of
the wall. It is likely then, physostigmine may actively constrict the
arterioles of the mucosa to decrease its flow. The slight increase in
muscularis and submucosal blood flow may be the result of an active hyper-
emia in those layers as a result of the stimulation of motor activity by
physostigmine and the release of vasoactive metabolites. The increased
flow may also be the result of the vasodilatory effects of physostigmine
alone, however.

The slight increase in submucosal blood flow, if not due to the
vasoactive agents, may be due to vaso—constriction in the mucosa. Accor-
ding to Greenway and Murthy (29), the constriction of the arterioles of
the mucosa would cause a greater proportion of the microspheres to become
lodged in the submucosa. The increased radioactivity would then be inter-

preted as an increase in submucosal flow. The results of the present

58

study are then consistent with the hypothesis of Greenway and Murthy.

The results of this series of experiments indicate that infusion
of physostigmine produces an increase in intraluminal pressure and a de-
crease in total blood flow to the wall of the intestine. The decrease
in total wall flow is primarily the result of decreased mucosal blood
flow. It is postulated that the decreased flow in this layer is either
the result of active arteriolar constriction or the result of compression
of mucosal vasculature by muscularis motor activity. Muscularis and sub-
mucosal flows were increased slightly, although not significantly, pos-
sibly as the result of an active hyperemia in those layers or from the

vasodilatory effects of physostigmine.

CHAPTER VI

summary

The present study has investigated the effects of manipulation,
distention and infusion of physostigmine on blood flow and its distribu-
tion in the wall of the canine gastrointestinal tract. The distribution
of radioactive microspheres within the intestinal wall was used to deter-
mine compartmental blood flow. Two types of spheres, both 15 T 5 u in
diameter, labeled with either 8SSr or ll”Ce were used.

To provide an independent verification of the microsphere method,
the experimental procedures were repeated and blood flow was determined
by timed collection of the venous outflow from an intestinal segment. The
results are summarized as follows:

1. All of the experimental procedures, i.e., manipulation, dis-
tention and the administration of physostigmine produced an
increase in intestinal motor activity as judged by visual ob-
servation and recordings of intestinal lumen pressure.

2. Distention of a jejunal segment to an intraluminal pressure
of 20-30 mm Hg by luminal placement of saline significantly
increased blood flow to that segment. The increase in blood
flow was not shared equally by all three layers within the
wall, but was primarily the result of increased muscularis
and submucosal blood flow. Mucosal blood flow was not signifi-

cantly altered.

59

3.

60

Manipulation of the wall of a jejunal segment significantly
increased total blood flow to that segment. The increase

is primarily the result of an increase in muscularis and sub-
mucosal blood flow. Again, mucosal blood fIOW'WaS not signifi-
cantly altered.

Neither manipulation nor distention significantly altered
systemic blood pressure. Neither blood flow nor flow distri-
bution were altered in the remainder of the gastrointestinal
tract which was left within the abdominal cavity.

The administration of physostigmine significantly increased
intraluminal pressure and decreased total blood flow to the
wall of the stomach, duodenum, jejunum and colon. The decrease
was primarily the result of decreased mucosal blood flow.
Neither submucosal nor muscularis blood flow were signifi-
cantly altered. Nor was systemic pressure significantly
altered by the infusion of physostigmine.

The measurements of total blood flow to the wall as determined
by microsphere method were not significantly different than
the measurements made by the collection of venous outflow.

The results of the present study indicate that increased in-
testinal motor activity redistributes the total flow to the
wall in favor of the muscularis layer whether total flow to

the wall was increased or decreased.

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