Vol. 277, Issue 4, H1441-H1446, October 1999
NO mediates postjunctional inhibitory effect of neurogenic ACh
in guinea pig small intestinal microcirculation
N.
Kotecha and
F. P.
Coffa
Department of Physiology, Monash University, Clayton, Victoria 3168, Australia
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ABSTRACT |
The present
study was designed to evaluate the role of the endothelium as an
effector organ of neurally mediated inhibition of vascular tone.
Acetylcholine (ACh), either released by stimulation of the submucosal
ganglia or applied exogenously, inhibited phenylephrine (PE)-induced
constrictions in arterioles of the guinea pig intestinal submucosa.
NG-monomethyl-L-arginine
(L-NMMA), an inhibitor of nitric
oxide (NO) synthesis, attenuated the response to superfused ACh by 74% compared with 94% attenuation obtained with
NG-nitro-L-arginine
(L-NNA).
L-NNA attenuated the
response to neurally released ACh by 98% and that to iontophoretically
applied ACh by 92%. L-Arginine reversed the effects of
both L-NMMA and
L-NNA. Functional integrity of
the endothelium was essential for the neurally mediated inhibition of
PE-induced constrictions. However, neurogenic inhibition of neurally
evoked constrictions was preserved despite endothelial disruption. It
was concluded that at the postjunctional level, the mechanism of action
of neurally released ACh was almost exclusively via a NO-dependent
pathway, with the source of NO being the vascular endothelium.
submucosal arterioles; vasodilator nerves; acetylcholine; endothelium
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INTRODUCTION |
SUBMUCOSAL NEURONS of the enteric nervous system of the
gut project to arterioles perfusing the guinea pig small intestine (4,
8, 9). Activation of these neurons by electrical stimulation of
submucosal ganglia produces marked dilation in adjacent preconstricted
submucosal arterioles (1, 14, 23). ACh is the main transmitter
mediating this vasodilation (1, 14, 23), although other putative
neurotransmitters have been implicated, such as dynorphin, galanin, and
vasoactive intestinal polypeptide (14).
Andriantsitohaina and Surprenant (1) showed that neurogenic dilations
of the submucosal arterioles that were sensitive to the muscarinic
antagonist,
4-diphenylacetoxy-N-methylpiperidine methiodide, were blocked by ~70% in the presence of the nitric oxide
(NO) inhibitor
NG-monomethyl-L-arginine
(L-NMMA). This demonstrated that
cholinergic dilations of these vessels were dependent on a NO pathway.
The NO was probably released from the endothelium, because this is the
case in most larger vessels investigated in which dilations were
produced by the exogenous application of ACh (6, 7). Vascular smooth
muscle is unlikely to be the source of NO, because only an inducible NO
synthase (iNOS) has been identified in smooth muscle, which does not
produce NO when exposed to muscarinic agonists (18, 24).
Although the exact location of the cholinergic varicosities innervating
the submucosal arterioles is unknown, morphological studies suggest
that they would be located no deeper than the medioadventitial border
of the arteriole wall (10). The media of arterioles is composed of only
one layer of smooth muscle (26); hence it is more likely that neurally
released ACh can reach the endothelium in this preparation than in
preparations with thicker smooth muscle. Nerve-released ACh reaching
the endothelium to produce dilation provides a rarely reported example
of a neurogenic influence on arteriole tone, mediating its effects at
the level of the intima rather than the adventitia of the arteriole
wall, and warrants further investigation. Cholinergic neurogenic
vasodilation mediated by NO has been demonstrated in the dog hindlimb,
but the source of NO was not determined (17) and the study in guinea pig submucosal arterioles did not definitively demonstrate the endothelium to be the effector organ of NO-mediated neurogenic vasodilation (1). We sought to determine the role of the vascular endothelium as an effector organ of neurally mediated NO-dependent inhibition of vascular tone.
The L-NMMA-resistant component
of the vasodilation found by Andriantsitohaina and Surprenant (1) may
have been caused by 1) ACh acting
directly on the arteriole smooth muscle (3, 13), 2) ACh stimulating the release of
another relaxing factor from the endothelium, such as
endothelium-derived hyperpolarizing factor (EDHF) (21), or
3) the inability of
L-NMMA to completely inhibit NO
synthesis in this preparation.
The NO inhibitor NG-nitro-L-arginine
(L-NNA) has been shown to be
more effective than L-NMMA in a
number of in vitro preparations, including rabbit aorta, rabbit femoral
artery, and rat mesenteric artery (12, 19, 20). Hence, in the present
study, L-NNA was used to provide
insight into the
L-NMMA-resistant vasodilation observed by Andriantsitohaina and Surprenant (1).
The question posed in this study was twofold, i.e., is there another
endothelium-dependent or -independent mechanism mediating cholinergic
neurogenic inhibition and is the vascular endothelium the effector
organ of neurally mediated postjunctional inhibition of vascular tone?
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METHODS |
Tissue preparation.
Guinea pigs (Monash bred, either sex, 200-300 g) were
killed by stunning with a blow to the back of the head followed by
exsanguination. Use of these guinea pigs and all procedures undertaken
were approved by the Monash standing committee on ethics in animal
experimentation. A small piece of ileum was removed from the animal,
cut open, and pinned out tightly, mucosal side up. The mucosa was
carefully peeled away, revealing several arteriole trees and submucosal plexus embedded within translucent connective tissue. A small piece of
connective tissue containing one or two arteriole trees was cut and
peeled away from the underlying circular muscle layer. This piece of
tissue was then pinned onto a small organ bath with a transparent base,
with the mucosal side facing down. The preparation was continuously
superfused with warmed physiological saline composed of (mM) 146 Na+, 5 K+, 2.5 Ca2+, 2 Mg2+, 134 Cl
, 25 HCO3, 1 HPO24, and 11 D-glucose and equilibrated with
carbogen gas (95% O2-5% CO2).
The organ bath containing the submucosal arteriole preparation was
mounted onto an inverted microscope equipped with a television camera.
A magnified image (×100) of the preparation was projected onto a
television screen, and arteriole diameter was monitored by image
analysis using the Diamtrak software program (22).
The submucosal arterioles were transiently constricted by iontophoretic
application of phenylephrine (PE) from a micropipette. A bevelled glass
pipette with a tip diameter of 80-100 µm, filled with
physiological saline, was used to stimulate the ganglion or the
perivascular nerves. It was connected to a standard pulse stimulator
with the earth electrode in the recording chamber. Arterioles were
exposed to ACh by superfusion, iontophoresis, or ganglion stimulation.
Vascular endothelium was disrupted by cannulating the parent mesenteric
artery, which gives rise to an arteriole network, at its point of entry
to the intestine. Disruption of the endothelium was obtained by
perfusing arterioles with physiological saline containing 0.1%
3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) (13).
Protocol.
Submucosal arterioles were constricted every 2 min by
iontophoretic application of 10 mM PE from a glass micropipette (10 Hz,
1-2 s). The arterioles constricted transiently within 1 s of PE
application and remained constricted for 3-5 s. By altering stimulus parameters for the iontophoretic release of PE, we adjusted the amplitude of constrictions so that they were 30-50% of the resting diameter of the arteriole; this ensured that the constrictions were equivalent to the maximally obtainable constrictions as determined by exposing the arterioles to 100 mM isotonic KCl. Once adjusted, the
amplitude of constrictions remained constant over at least 0.5 h. We
investigated changes in the amplitude of these constrictions to ACh
applied by superfusion, ganglion stimulation, or iontophoresis (see
Fig. 1A). When the amplitude of
constrictions had returned to control levels, the tissue was rested for
~20 min before further testing.
Analysis and statistics.
Inhibition of a transient arteriole constriction was expressed as a
proportion of the control constriction and was calculated as follows
where
a is control PE-induced arteriole
constriction, b is constriction to PE
after application of ACh, and c is
inhibition of constriction produced by ACh (see Fig.
1A).
Values are expressed as means ± SE and were compared using a paired
or unpaired Student's t-test or
ANOVA. A probability value of <0.05 was considered significant. Drugs
used were ACh chloride, CHAPS,
L-arginine, PE hydrochloride,
and pirenzepine dihydrochloride (Sigma),
L-NMMA (Calbiochem),
L-NNA (Novabiochem), and
tetrodotoxin (Research Biochemicals International).
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RESULTS |
Arteriole response to superfused ACh.
ACh superfused into the organ bath attenuated the amplitude of
arteriole constrictions induced by PE. Increase in concentration of
superfused ACh produced an increase in inhibition of arteriole constrictions to PE (Fig.
1B). An
EC50 of 17.7 ± 8.0 nM
(n = 4) was obtained from individual
values of EC50 calculated from
sigmoid curves fitted to each set of data.
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Fig. 1.
A: protocol used to assess inhibitory
effect of ACh on phenylephrine (PE)-induced constrictions. Transient
diameter changes of guinea pig submucosal arteriole to iontophoretic
application of PE (10 Hz, 1 s) were obtained every 2 min. Effects of
ACh (applied exogenously, either by superfusion or iontophoresis, or
released from nerves) on PE-induced constrictions were measured. In
example shown, ACh was applied iontophoretically (at 10 Hz for 1 s) 1 s
before application of PE. Values obtained for analysis were control
PE-induced constriction of submucosal arteriole
(a), constriction of submucosal
arteriole to PE after application of ACh
(b), and inhibition of constriction
produced by ACh (c).
B: summary of inhibition produced by
superfused ACh on constrictions to iontophoretically applied PE in
guinea pig submucosal arteriole. Relationship between change in size of
PE-induced constrictions (plotted as fractional inhibition) and
concentration of superfused ACh
(EC50 17.7 ± 8.0 nM) is shown;
each point is mean ± SE of 4 observations. An apparent
EC50 taken from this curve with
pooled data will not match value above because true value quoted was
obtained from sigmoid curves fitted to each set of data.
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Effects of L-NNA and
L-NMMA on arteriole response to
superfused ACh.
These experiments were performed to enable us to compare the results of
our study with those obtained by Andriantsitohaina and Surprenant (1),
because there are methodological differences between the two. A known
concentration of ACh (5-20 nM) was superfused into the organ bath
such that the amplitude of constrictions induced by PE was attenuated
by 0.60-0.80 (fractional inhibition) to match the maximum
inhibitory effect produced by ganglion stimulation (see
Effects of
L-NNA and
L-arginine on
pirenzepine-sensitive arteriole response to ganglion
stimulation). The
experiment was then repeated in the presence of either
L-NMMA (300 µM) or
L-NNA (100 µM). Both
L-NMMA and
L-NNA significantly attenuated
the ACh-induced inhibition of PE constrictions
[L-NMMA: 0.69 ± 0.06 to 0.19 ± 0.07, n = 4 (Fig.
2A);
L-NNA: 0.79 ± 0.04 to 0.05 ± 0.02, n = 4 (Fig.
2B)]. The reversal of ACh
response in L-NMMA (74 ± 7%) was in accordance with previous findings (1) but lower than that
obtained in L-NNA (94 ± 2%), indicating that L-NNA was
a more effective NO synthesis inhibitor in this preparation.
L-Arginine (10 mM) significantly
reversed the effects of both
L-NMMA and L-NNA (Fig. 2).
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Fig. 2.
Effects of NG-monomethyl-L-arginine
(L-NMMA),
NG-nitro-L-arginine
(L-NNA), and
L-arginine on ACh-induced
inhibition of PE constrictions in guinea pig submucosal arterioles.
A: effects of
L-NMMA and
L-arginine on superfused ACh.
Nitric oxide synthase inhibitor
L-NMMA (300 µM) significantly
attenuated ACh-induced inhibition (plotted as fractional inhibition) of
PE-induced constrictions of submucosal arterioles (see text for
details). L-Arginine (10 mM) antagonized effect of L-NMMA. B: effects
of L-NNA and L-arginine on superfused ACh.
Nitric oxide synthase inhibitor
L-NNA (100 µM) significantly
attenuated ACh-induced inhibition of PE-induced constrictions of
submucosal arterioles.
L-Arginine (10 mM) antagonized
effect of L-NNA.
C: effects of
L-NNA and
L-arginine on iontophoretic ACh.
L-NNA (100 µM) significantly
attenuated iontophoretic ACh-induced inhibition of PE constrictions.
L-Arginine (10 mM) antagonized
effects of L-NNA.
* Significantly different from control.
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Effects of L-NNA and
L-arginine on arteriole response to
iontophoretically applied ACh.
ACh (0.1 µM) applied to the surface of an arteriole by iontophoresis
(10 Hz for 1-2 s), 1 s before the application of PE, inhibited the
subsequent arteriole constriction to PE. Stimulus parameters for the
iontophoretic release of the ACh were adjusted to obtain an inhibition
of constriction to PE in the range of 0.60 to 0.80 (Fig.
2C), to match the maximum inhibition
produced by ganglion stimulation (see Effects of
L-NNA and
L-arginine on pirenzepine-sensitive arteriole response to ganglion
stimulation). These experiments were performed to
mimic the effects of neurally released ACh, both in time and amplitude
of response obtained.
Incubation for 10 min in 100 µM
L-NNA resulted in a significant
attenuation of the inhibition produced by ACh from 0.75 ± 0.07 to
0.05 ± 0.03 (n = 6, Fig.
2C). In the presence of 10 mM L-arginine, the inhibition to
ACh increased from 0.05 ± 0.03 (in L-NNA alone) to 0.63 ± 0.01 (n = 5, Fig.
2C).
Effects of L-NNA and
L-arginine on pirenzepine-sensitive
arteriole response to ganglion stimulation.
The submucosal ganglia selected for stimulation were within a 500-µm
radius of the arteriole being investigated. Ganglion stimulation (10 Hz
for 10 s), 1 s before application of PE, inhibited the subsequent
arteriole constriction to PE. These stimulus parameters were chosen to
obtain maximal neural response (23). The inhibition produced by
ganglion stimulation was in the range of 0.60 to 0.80 (Fig.
3) of control PE-induced constrictions.
Neurally mediated inhibitory responses were completely abolished in the
presence of 1 µM tetrodotoxin, thus confirming the sole involvement
of nerve conduction.
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Fig. 3.
Effects of pirenzepine, L-NNA,
and L-arginine on ACh (neurally
released)-induced inhibition of PE constrictions in guinea pig
submucosal arteriole. Pirenzepine (1 µM, a muscarinic antagonist)
significantly and reversibly reduced inhibition produced by stimulation
of submucosal ganglia, thus confirming that response was due to release
of ACh from intrinsic submucosal nerves. L-NNA (100 µM)
significantly attenuated pirenzepine-sensitive (and therefore ACh
mediated) component of neurally inhibited PE constrictions.
L-Arginine (10 mM) antagonized effects of
L-NNA (see text for details). * Significantly
different from control.
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To ensure that the response to ganglion stimulation was mediated by
ACh, its sensitivity to pirenzepine (a muscarinic antagonist) was
tested. In the presence of 1 µM pirenzepine (a concentration that
blocks all muscarinic receptors), the inhibition of arteriole constriction produced by ganglion stimulation was significantly reduced
from 0.73 ± 0.07 to 0.07 ± 0.03 (n = 5, Fig. 3), indicating that the
response being investigated was mediated by ACh. After pirenzepine was
washed out, the inhibition of arteriole constriction to ganglion
stimulation increased from 0.07 ± 0.03 (in pirenzepine) to 0.62 ± 0.07 (n = 5), which was not
significantly different from control inhibition (0.73 ± 0.07;
n = 5, Fig. 3).
Incubation in 100 µM L-NNA for
10 min significantly attenuated the pirenzepine-sensitive inhibition of
arteriole constriction from 0.62 ± 0.07 to 0.02 ± 0.02 (n = 5, Fig. 3). This attenuation was
reversed in the presence of 10 mM
L-arginine, with an increase in
inhibitory response to ganglion stimulation from 0.02 ± 0.02 (in
L-NNA alone) to 0.47 ± 0.05 (n = 5, Fig. 3).
In the presence of L-NNA,
attenuation of responses to neurally released ACh (98 ± 2%;
n = 5) and iontophoretically applied ACh (92 ± 5%; n = 6) were not
significantly different. Hence, a NO-dependent pathway was almost
exclusively involved in the mechanisms of neurally released and matched
exogenous ACh action.
Effects of endothelial disruption on arteriole response to ganglion
stimulation.
In four experiments, the inhibitory effect of ganglion stimulation on
PE-induced arteriole constrictions was evaluated before and after the
endothelium was disrupted using CHAPS (13). The protocol used was the
same as that in the experiments described in Effects
of L-NNA and
L-arginine on
pirenzepine-sensitive arteriole response to ganglion
stimulation. Disruption of the endothelium resulted in
a significant and complete abolition of neurally mediated inhibition (from 0.70 ± 0.14 to 
0.16 ± 0.09, n = 4; Fig.
4).
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Fig. 4.
Effects of endothelial disruption on neurally mediated inhibition of
arteriole constrictions in guinea pig small intestine. In
endothelium-intact arterioles, PE-induced constrictions were
significantly attenuated by ganglion stimulation. Disruption of
endothelium abolished inhibitory effect of ganglion stimulation of
PE-induced constrictions. However, constrictions evoked by perivascular
nerve stimulation (PVNS) were attenuated by ganglion stimulation
despite endothelial damage.
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Effect of ganglion stimulation on arteriole constrictions evoked by
perivascular nerve stimulation.
Interaction between ganglion stimulation and perivascular nerves was
tested in three arterioles, after endothelial disruption, from the
experiments described in Effects of endothelial
disruption on arteriole response to ganglion
stimulation. Arteriole constrictions were evoked by
applying a train of stimuli (10 Hz, 1 s), once every 2 min, to the
perivascular nerves. Evoked constrictions were abolished in the
presence of 1 µM tetrodotoxin. Ganglion stimulation (10 Hz, 10 s, 1 s
before an evoked constriction) significantly inhibited the subsequent
constriction (0.69 ± 0.16, n = 3;
Fig. 4).
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DISCUSSION |
In agreement with previous investigations, the results obtained in the
present study suggest that nerve-released ACh mediates its response by
stimulating the release of NO in guinea pig submucosal arterioles (1).
A discrepancy between the effects of
L-NNA and
L-NMMA has been observed in a
number of vessels similar to that observed in this preparation (12, 19,
20). For instance, in the rabbit aorta
L-NMMA produced a 65%
inhibition of ACh-induced dilations, whereas
L-NNA produced a 90% inhibition (12, 25).
Although both L-NNA and
L-NMMA are competitive
inhibitors of NO synthase (the enzyme that catalyzes the synthesis of
NO from L-arginine), they differ
in that L-NNA is effectively
irreversible and may bind covalently to the enzyme, whereas
L-NMMA, but not
L-NNA, is metabolized within
endothelial cells, thus lowering the effective intraendothelial
concentration of L-NMMA (5).
Hence, using the more effective NO synthesis inhibitor,
L-NNA, as opposed to L-NMMA (see
RESULTS), we found a near-maximal
block of the response to neurally released ACh (98%) and
iontophoretically applied ACh (92%), which were not significantly
different. Inhibition of superfused ACh-induced response by
L-NMMA obtained in this study
(~74%) correlates closely with that obtained by Andriantsitohaina
and Surprenant (1) (~70%), indicating that the difference in
methodology between the two studies cannot account for a more effective
block obtained using L-NNA.
Additionally, the EC50 value for
the effect of ACh on PE-induced constriction (~18 nM) in this study
is almost identical to that obtained for ACh-induced dilation of
arterioles preconstricted with U-46619 (done as part of a study on
diabetic guinea pigs; K. Eede and N. Kotecha, unpublished observation);
the latter being the method used by Andriantsitohaina and Surprenant
(1). Overall, it would appear that there are no
fundamental problems associated with comparing the effects of ACh on
constrictions obtained using the two different methods. Hence, we
conclude that the inhibitory response of neurally released ACh is
mediated almost exclusively by a NO-dependent pathway; this is in
accord with recent work that demonstrates that NO accounts for all
endothelium-dependent inhibitory effects of superfused ACh (13).
The movement of neurally released ACh through the media is feasible
because, unlike the endothelium, vascular smooth muscle cells are not
connected by tight junctions, which may hinder the movement of
lipophobic molecules (16), but are electrically coupled by gap
junctions (11) allowing ready access of adventitially applied ACh to
the abluminal side of the endothelial cells (16). The only diffusion
barrier is physical, posed by the interleaving layers of smooth muscle
cells. However, in the terminal submucosal arterioles, there is only a
single layer of smooth muscle cells, which is 4 µm at its widest
point (11). As such, the media of the arteriole wall does not pose a
barrier to the movement of ACh from the adventitia to the endothelium.
Experiments using detergent to remove the endothelium confirm that the
functional integrity of the endothelium is essential for the inhibitory
effects of intrinsic vasodilator nerves, implying that the source of NO in the present study must be the endothelium. We have shown in a
previous study (15) that the cholinergic vasodilator nerves interact
with the perivascular constrictor nerves to modulate the release of
sympathetic transmitter by a prejunctional mechanism. The inhibitory
effect of ganglion stimulation on arteriole constrictions obtained by
stimulation of perivascular nerve was preserved after the endothelium
was damaged; this confirmed that the lack of response of neurally
released ACh on PE constrictions was not a result of neural dysfunction
related to the protocol used to disrupt the endothelium.
The inhibitory effects of exogenous ACh are known to be mediated by
other NO-independent, endothelium-dependent and -independent pathways.
A factor released from the endothelium has been identified in many
vascular beds that hyperpolarizes the smooth muscle of blood vessels
(21). This EDHF has also been shown to contribute to ACh-induced
vasodilations along with NO and prostacyclin (21). Additionally, ACh
can act directly on the smooth muscle to produce vasodilation, as is
the case in many vessels of the cephalic circulation, such as the
lingual artery of the rabbit, the posterior auricular artery of the
cat, and the submucosal arterioles of the guinea pig ileum (2, 3, 13).
Although these various mechanisms can mediate the inhibitory effects of
ACh, the focus of the current study was to elucidate the mechanisms
mediating the effects of neurally released ACh from the intrinsic
vasodilator nerves within the submucosa of the guinea pig. The
near-maximal block (98%) produced by
L-NNA in our study would suggest
that NO is the main and perhaps the sole mediator of the
endothelium-dependent inhibitory effects of neurally released ACh and
is in agreement with recent findings in that cholinergic
endothelium-dependent inhibition is fully accounted for by NO in the
guinea pig submucosal arterioles (13). Kotecha (13) also demonstrated
that ACh can cause endothelium-independent direct inhibition of the
vascular smooth muscle. However, the concentrations required for this
direct action are relatively higher
(EC50 > 100 nM) compared with
the endothelium-dependent inhibitory effects
(EC50 ~20 nM). Because the
maximum inhibitory effect produced by neurally released ACh corresponds
to a concentration of 5-20 nM (see
RESULTS), only the
endothelium-dependent inhibitory effects are evident and any direct
effect on the vascular smooth muscle does not make a significant contribution.
In conclusion, the cholinergic vasodilator nerves innervating the
guinea pig submucosal arterioles mediate inhibition at both the pre-
and postjunctional levels. The postjunctional effects are almost
exclusively a result of stimulating the release of NO. Although
previous studies have implicated the vascular endothelium as the source
of NO, this is the first study to directly demonstrate this in the
microvasculature of the guinea pig small intestine. As far as we know,
this is also the only study in which the role of the vascular
endothelium as an effector organ mediating neurogenic inhibition has
been definitively determined.
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ACKNOWLEDGEMENTS |
This work was supported by the National Health and Medical Research
Council of Australia.
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FOOTNOTES |
The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement"
in accordance with 18 U.S.C. §1734 solely to indicate this fact.
Address for reprint requests and other correspondence: N. Kotecha,
Dept. of Physiology, Monash Univ., Clayton, Victoria 3168, Australia
(E-mail: Neela.Kotecha{at}med.monash.edu.au).
Received 24 March 1999; accepted in final form 26 May 1999.
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REFERENCES |
1.
Andriantsitohaina, R.,
and
A. Surprenant.
Acetylcholine released from guinea-pig submucous neurones dilates arterioles by releasing nitric oxide from endothelium.
J. Physiol. (Lond.)
453:
493-502,
1992[Abstract/Free Full Text].
2.
Brayden, J. E.,
and
J. A. Bevan.
Neurogenic muscarinic vasodilation in the cat. An example of endothelial cell-independent cholinergic relaxation.
Circ. Res.
56:
205-211,
1985[Abstract/Free Full Text].
3.
Brayden, J. E.,
and
W. A. Large.
Electrophysiological analysis of neurogenic vasodilation in the isolated lingual artery of the rabbit.
Br. J. Pharmacol.
89:
163-171,
1986[Medline].
4.
Brookes, S. J. H.,
P. A. Steele,
and
M. Costa.
Calretinin immunoreactivity in cholinergic motoneurones, interneurones and vasomotor neurones of the guinea-pig small intestine.
Cell Tissue Res.
263:
471-481,
1991[Medline].
5.
Cocks, T. M.
Endothelium-dependent vasodilator mechanisms.
In: Pharmacology of Vascular Smooth Muscle, edited by C. J. Garland,
and J. A. Angus. Oxford, UK: Oxford Univ. Press, 1996, p. 233-251.
6.
Furchgott, R. F.
The role of the endothelium in the responses of vascular smooth muscle to drugs.
Annu. Rev. Pharmacol. Toxicol.
24:
175-197,
1984[Medline].
7.
Furchgott, R. F.,
and
P. M. Vanhoutte.
Endothelium-derived relaxing and contracting factors.
FASEB J.
3:
2007-2018,
1989[Abstract].
8.
Furness, J. B.,
and
M. Costa.
The Enteric Nervous System. New York: Churchill Livingston, 1987.
9.
Galligan, J. J.,
M. Costa,
and
J. B. Furness.
Changes in surviving nerve fibres associated with submucosal arteries following extrinsic denervation of the small intestine.
Cell Tissue Res.
253:
647-656,
1988[Medline].
10.
Hirst, G. D. S.
Neuromuscular transmission in intramural blood vessels.
In: Handbook of Physiology. The Gastrointestinal System. Motility and Circulation. Bethesda, MD: Am. Physiol. Soc., 1989, sect. 6, vol. I, pt. 2, ch. 45, p. 1635-1666.
11.
Hirst, G. D. S.,
and
F. R. Edwards.
Sympathetic neuroeffector transmission in arteries and arterioles.
Physiol. Rev.
69:
546-604,
1989[Free Full Text].
12.
Kobayashi, Y.,
and
K. Hattori.
Nitroarginine inhibits endothelium-derived relaxation.
Jpn. J. Pharmacol.
52:
167-169,
1990[Medline].
13.
Kotecha, N.
Mechanisms underlying ACh induced modulation of neurogenic and exogenous ATP constrictions in the submucosal arterioles of the guinea-pig small intestine.
Br. J. Pharmacol.
126:
1625-1633,
1999[Medline].
14.
Kotecha, N.,
and
T. O. Neild.
Vasodilatation and smooth muscle membrane potential changes in arterioles from the guinea-pig small intestine.
J. Physiol. (Lond.)
482:
661-667,
1995[Medline].
15.
Kotecha, N.,
and
T. O. Neild.
Actions of vasodilator nerves on arteriolar smooth muscle and sympathetic nerves in the guinea-pig small intestine.
J. Physiol. (Lond.)
489:
849-855,
1995[Medline].
16.
Lew, M. J.,
R. J. Rivers,
and
B. R. Duling.
Arteriolar smooth muscle responses are modulated by an intramural diffusion barrier.
Am. J. Physiol.
257 (Heart Circ. Physiol. 26):
H10-H16,
1989[Abstract/Free Full Text].
17.
Loke, K. E.,
C. G. Sobey,
G. J. Dusting,
and
O. L. Woodman.
Cholinergic neurogenic vasodilatation is mediated by nitric oxide in the dog hind limb.
Cardiovasc. Res.
28:
542-547,
1994[Abstract/Free Full Text].
18.
Moncada, S.,
R. M. Palmer,
and
E. A. Higgs.
Nitric oxide: physiology, pathophysiology and pharmacology.
Pharmacol. Rev.
43:
109-142,
1991[Medline].
19.
Moore, P. K.,
O. A. al-Swayeh,
N. W. Chong,
R. A. Evans,
and
A. Gibson.
L-NG-nitro arginine (L-NOARG), a novel, L-arginine-reversible inhibitor of endothelium-dependent vasodilatation in vitro.
Br. J. Pharmacol.
99:
408-412,
1990[Medline].
20.
Mülsch, A.,
and
R. Busse.
NG-nitro-L-arginine (N5-[imino(nitroamino)methyl]-L-ornithine) impairs endothelium-dependent dilations by inhibiting cytosolic nitric oxide synthesis from L-arginine.
Naunyn Schmiedebergs Arch. Pharmacol.
341:
143-147,
1990[Medline].
21.
Nagao, T.,
and
P. M. Vanhoutte.
Endothelium-derived hyperpolarizing factor and endothelium-dependent relaxations.
Am. J. Respir. Cell Mol. Biol.
8:
1-6,
1993.
22.
Neild, T. O.
Measurement of arteriole diameter changes by analysis of television images.
Blood Vessels
26:
48-52,
1989[Medline].
23.
Neild, T. O.,
K.-Z. Shen,
and
A. Surprenant.
Vasodilatation of arterioles by acetylcholine released from single neurones in the guinea-pig submucous plexus.
J. Physiol. (Lond.)
420:
247-265,
1990[Abstract/Free Full Text].
24.
Rees, D. D.,
S. Cellek,
R. M. Palmer,
and
S. Moncada.
Dexamethasone prevents the induction by endotoxin of a nitric oxide synthase and the associated effects on vascular tone: an insight into endotoxin shock.
Biochem. Biophys. Res. Commun.
173:
541-547,
1990[Medline].
25.
Rees, D. D.,
R. M. Palmer,
H. F. Hodson,
and
S. Moncada.
Specific inhibitor of nitric oxide formation from L-arginine attenuates endothelium dependent relaxation.
Br. J. Pharmacol.
96:
418-424,
1989[Medline].
26.
Rhodin, J. A. G.
Architecture of the vessel wall.
In: Handbook of Physiology. The Cardiovascular System, Vascular Smooth Muscle. Bethesda, MD: Am. Physiol. Soc., 1980, sect. 2, vol. II, chapt. 1, pt. 1-32.
Am J Physiol Heart Circ Physiol 277(4):H1441-H1446
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