1583
Effects of Purines and Pyrimidines on the Rat
Mesenteric Arterial Bed
Vera Ralevic and Geoffrey Burnstock
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The effects of the purines, adenosine 5'-triphosphate (ATP) and guanosine 5'-triphosphate
(GTP), and the pyrimidines, uridine 5'-triphosphate (UTP), cytidine 5'-triphosphate (CTP),
and thymidine 5'-triphosphate (TTP), on vascular resistance were investigated in the rat
mesenteric arterial bed. In preparations at basal tone, these agents produced dose-related
vasoconstriction with a potency order of ATP>CTP>UTP>>TTP=GTP. When tone was
raised with norepinephrine (30 ,M), these agents caused dose-related vasodilatation with the
potency order of UTP=ATP>ITP=GTP. CTP did not elicit vasodilatation. Removal of the
endothelium with sodium deoxycholate resulted in an increased responsiveness of the mesenteric bed preparation to the vasoconstrictor effects of each of the purines and pyrimidines
tested. The selective P2X-purinoceptor-desensitizing agent a43-methylene ATP inhibited vasoconstrictor responses to ATP and to CTP but had no effect on vasoconstrictor responses elicited
by UTP, TTP, and GTP. In raised-tone preparations, vasodilator responses to ATP, UTP, TTP,
and GTP were abolished after removal of the endothelium with sodium deoxycholate.
Responses to acetylcholine were also abolished; those to sodium nitroprusside were unimpaired. An inhibitor of the formation of nitric oxide from L-arginine, Na-nitro-L-arginine
methyl ester (30 t,M), which antagonizes responses mediated by endothelium-derived relaxing
factor (nitric oxide), attenuated vasodilatation to ATP, UTP, and acetylcholine but not to
sodium nitroprusside. In addition to confirming the presence of P2X- and P2ypurinoceptors in
the rat mesenteric arterial bed, these results demonstrate the presence of discrete "pyrimidinoceptors," which mediate vasoconstriction and vasodilatation by receptors located on the
smooth muscle and on endothelial cells, respectively. Furthermore, it is shown that the vascular
relaxations to ATP and UTP occur largely via production of endothelium-derived relaxing
factor. It is probable that, like P2-purinoceptors, UTP pyrimidinoceptors comprise a heterogeneous population, subtypes of which have been partially characterized according to their
different actions and locations within the vasculature. (Circulation Research 1991;69:1583-1590)
denosine 5'-triphosphate (ATP) produces
powerful systemic effects; it influences many
biological processes, being released from
nerve endings, platelets, and endothelial cells in
physiological and pathophysiological processes.' In
the cardiovascular system its ability to cause vasoconstriction or vasodilatation is mediated through activation of subtypes of the P2-purinoceptor, P2X- and
P2Y-purinoceptors, respectively.2 In many systems,
including the rat mesenteric arterial bed,3 P2X-purinoceptors are present on the vascular smooth muscle;
P2Y-purinoceptors are located on endothelial cells,
A
From the Department of Anatomy and Developmental Biology
and the Centre for Neuroscience, University College London.
Supported by the British Heart Foundation.
Address for correspondence: Prof. G. Burnstock, Department
of Anatomy and Developmental Biology and Centre for Neuroscience, University College London, Gower Street, London WC1E
6BT, UK.
Received November 8, 1990; accepted July 29, 1991.
although in some vessels P2y-purinoceptors are also
located on the smooth muscle.45
The naturally occurring nucleotides, cytidine 5'triphosphate (CTP), thymidine 5'-triphosphate
(TTP), and uridine 5'-triphosphate (UTP), which are
pyrimidines, and guanosine 5'-triphosphate (GTP),
which is a purine, have also been shown to have
effects on the vasculature. Of these, probably the
most studied has been UTP, which may be released
from blood platelets.6'7 Several differences between
the effects of UTP and ATP have led to a proposal
for the existence of specific "pyrimidinoceptors,"
distinct from purinoceptors.8'9 The basis for this
proposal has come primarily from studies on the
effects of ATP and UTP in the perfused liver, in
HL-60 cells, and in neutrophils and macrophages,
with limited studies on blood vessels. These are
summarized in a review article by Seifert and
Schultz.10 Among other discriminating differences, it
has been shown in the rabbit ear artery that, whereas
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Circulation Research Vol 69, No 6 December 1991
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selective desensitization of the P2X-purinoceptor with
a,13-methylene ATP (a,43-meATP)"1 abolishes contractile responses to ATP, there is potentiation or
only partial desensitization of responses to UTP.9
Differential desensitization has also been observed
after pretreatment with UTP and other agents in the
rabbit basilar artery12 and in the rat tail and femoral
arteries and dog saphenous vein.13
The isolated rat mesenteric arterial bed was used
to study the effects of ATP, UTP, and other purine
and pyrimidine nucleotides on vascular tone. Previous characterization of P2-purinoceptor subtypes has
shown this preparation to be amenable to the study
of smooth muscle-mediated and endothelium-mediated responses.3 In this preparation we aimed to
produce a preliminary characterization of pyrimidinoceptors based on the nature of the responses and
the location in the vasculature of the receptor subtypes. We further aimed to distinguish between purinoceptors and pyrimidinoceptors by looking at the
effects of P2x-purinoceptor desensitization with a,8meATP and at the effects of inhibition of endothelium-derived relaxing factor (EDRF)14 formation using Nw-nitro-L-arginine methyl ester (L-NAME).15
Materials and Methods
Adult male Wistar rats (300-400 g) were used in
the study. Rats were given heparin (1,000 units i.p.)
and killed by stunning and exsanguination. Mesenteric beds were prepared for perfusion essentially as
described by McGregor.16 A midline abdominal incision was made, and the superior mesenteric artery
was exposed and cannulated. The superior mesenteric vein was severed, and perfusion was begun with
Krebs' solution containing (mM) NaCl 133, KCl 4.7,
NaH2PO4 1.35, NaHCO3 16.3, MgSO4 0.61, glucose
7.8, and CaCl2 2.52. Bovine serum albumin was added
(5 g/l) to raise the colloid osmotic pressure and
prevent tissue edema.17 The perfusate was gassed
with 95% 02-5% CO2 and maintained at 37°C. The
vascular bed was isolated by cutting as closely as
possible to the gut, and then perfusion was interrupted briefly to allow the bed to be transferred
onto gauze in a heated chamber. Perfusion was
begun again and adjusted to 4.8 ml/min. The preparation was also superfused at 1 ml/min with Krebs'
solution of the same composition. The preparation
was allowed to equilibrate for 30 minutes before
experimentation. Responses were measured as
changes in perfusion pressure (mm Hg) with a
pressure transducer (model P23, Gould, Cleveland,
Ohio) on a side arm of the perfusion cannula and
were recorded on a polygraph (model 79D, Grass
Instrument Co., Quincy, Mass.).
Removal of Endothelium
The endothelium was removed by perfusing the
preparation with 2 ml of a 2 mg/ml solution of
sodium deoxycholate in saline over a period of 30
seconds. This caused a transient increase in perfusion pressure. The preparation was washed through
for 10-15 minutes after addition of sodium deoxycholate solution. In experiments with raised tone,
norepinephrine (NE) was washed out before removal of the endothelium and reintroduced after
sodium deoxycholate treatment, after washout,
when perfusion pressure had returned to baseline.
The success of this treatment was assessed by using
the endothelium-dependent relaxing agent acetylcholine (ACh). The integrity of the smooth muscle
was checked with the direct smooth muscle relaxant
sodium nitroprusside (SNP).
Drug Administration
Drugs were applied as bolus injections of 50 ,A into
an injection port proximal to the tissue. Vasoconstrictor responses to agents were examined on the preparation at basal tone. Vasodilator responses were
examined after the tone of the preparation had been
raised by the addition of NE to the perfusate to a
final concentration of 30 gM. Pressor responses to
doses of ATP, UTP, CTP, TTP, and GTP were
examined before and after removal of the endothelium and after desensitization of P2X-purinoceptors
with a,4-meATP (1 ,uM, added to the perfusate 15
minutes before drug application). Responses to the
purines and pyrimidines were also investigated in the
raised-tone preparation before and after the endothelium had been removed.
The mechanism of the vasodilator response to
ATP and UTP was investigated by using L-NAME.15
After control vasodilator responses had been established in the raised-tone preparation, NE was removed from the perfusate, and the preparation was
perfused with L-NAME (30 ,M) for 40-45 minutes,
after which time the tone was again raised with NE.
Since L-NAME caused a supersensitivity of the preparation to NE, careful titration of NE into the
perfusate was required to avoid excessive and detrimental rises in perfusion pressure but yet to produce
a level of tone equivalent to the control level. The
new lower final concentration required to raise the
tone was found to be 1-8 uM NE.
Drugs
ATP (sodium salt), UTP (sodium salt), GTP (sodium salt), CTP (sodium salt), TTP (sodium salt),
a,4-meATP (lithium salt), SNP, ACh (chloride), NE
(bitartrate), and L-NAME were obtained from
Sigma Chemical Co., Poole, UK. All were made up in
distilled water except for NE, which was made up as
a stock solution of 10 mM in 0.1 mM ascorbic acid.
Statistical Analysis
Results are given as mean+ SEM. Statistical significance was evaluated by Student's t test. A value of
p<0.05 was taken as significant. The rank order of
potency of agents was determined empirically.
Ralevic and Bumstock Vascular Effects of Purines and Pyrimidines
ATP
100-
UTP
CTP
TTP
GTP
68 -63 5
6 -6
6.86.3 5.8
6.8 6.3 5.8
1585
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ATP
c
UTP
CTP
TTP
GTP
.2 1000-
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6.8
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9-3 8-3
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8-3 73 6-3
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8-3 73
63
2 min
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FIGURE 1. Representative tracings showing vasoconstrictor (upper tracings, preparation at basal tone) and vasodilator (lower
tracings, preparation with tone raised with 30 pM norepinephrine) responses ofthe rat mesenteric arterial bed to doses ofATP, CTP,
UTP, TTP, and GTP. Doses (expressed as -log mol) were applied as 50 pl bolus injections. In the upper tracings, each dose is
labeled; in the lower tracings, altemate doses are labeled starting from the lowest dose at the right. Intertnediate doses, which are
not labeled, are, from left to right, as follows (-log mol): 9.3, 8.3, and 7.3 forATP; 7.8 and 6.8 for CTP; 8.8, 7.8, and 6.8 for UTP,
TTP, and GTP. Note that CTP did not produce relaxation.
Results
Isolated rat mesenteric beds were perfused at a
constant flow rate of 4.8 mlmin. Basal perfusion
pressure was 18.2+0.8 mm Hg (n=38).
EE 50-
E
a)
, 40-
c 30._
c0)
The Effect of Purines and Pyrimidines on Rat
Mesenteric Artenial Beds at Basal Tone
In preparations at basal tone, ATP, UTP, and
CTP elicited dose-dependent contractions; TTP
and GTP were without effect except at the highest
doses used (Figures 1 and 2a). The potency order of
these agents, as determined empirically, was
ATP>CTP>UTP> >TTP=GTP. In the absence of
the endothelium, vasoconstrictor responses to each
of these agents were significantly greater than control
responses, as evidenced by a shift to the left of the
dose-response curves (Figure 3). Removal of the
endothelium caused greater potentiation of responses to UTP than to ATP. The tone of the
preparations was subsequently raised to test the
success of endothelium removal; responses to doses
of ACh up to and including 5 x 10- mol were abolished. The preparation produced dose-related relaxations to SNP at doses from 5 x 10`1 to 5 x 10 8 mol.
The Effect of Purines and Pyrimidines on
Preconstricted Rat Mesenteric Arterial Beds
Addition of NE (30 ,uM) to the perfusate raised
the tone of the preparations to a final perfusion
pressure of 86.5 +5.9 mm Hg (n=28). In preparations in which tone had been raised with NE,
dose-dependent relaxations were elicited by ATP,
UTP, TTP, and GTP (Figures 1 and 2b). Their rank
order of potency, as determined empirically, was
UTP=ATP>TTP=GTP. CTP did not cause relaxation. ATP and UTP were equally effective in elicit-
- 20v 10
-0
)
C
,a
E
E
ecn
c
0
cl
C:
tom
a)
C)
Q)
C]
-log mol purine or pyrimidine
FIGURE 2. Dose-response curves showing vasoconstrictor
(panel a) and vasodilator (panel b) responses of the rat
mesenteric arterial bed to purines and pyrimidines. Vasoconstrictor responses are shown as increases in perfusion pressure
(mm Hg) and are the means of 14-20 preparations. Vasodilator responses were examined in the raised-tone preparation,
in which tone was raised by the addition of norepinephrine (30
pM) to the perfusate. Shown are responses to ATP (c), UTP
(A), GTP (A), CTP (,), and iTP (o). Vasodilator responses
are shown as decreases in perfusion pressure (mm Hg) and are
the mean of 6-11 preparations. Symbols plus vertical bars
indicate mean +SEM.
1586
Circulation Research Vol 69, No 6 December 1991
preparation to 97.6+8.3 mm Hg (n= 13), which
ot significantly different from the control. Reof the endothelium abolished vasodilator ress to ATP, UTP, TTP, and GTP, and contracwere often produced instead. Relaxations to
at doses up to and including 5 x 10-9 mol were
hed. Dose-dependent relaxations of the prepato SNP (doses of 5 x 10-1to 5 x 10-8 mol) were
tenuated by removal of the endothelium.
10090-
Tl
I 80-
E
E
at
I /
II
70-
,1 Ifi
cn
D(X 60-
1:
a
1,
3-meATP on
to Purines and Pyrimidines
fusion with a,43-meATP (1 gM) abolished rees to lower doses of ATP and greatly reduced
agnitude of the responses to higher doses.
nses to CTP were also abolished by a,c3'P. Responses to UTP, GTP, and TTP were
ffect of Desensitization With a,
o 50c)
r Responses
40-
a)
en
-
20-
cted by
10
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ffect of L-NAME on Vasodilator Responses
085
8
7*5
7
LMesenteric Arterial Beds to ATP, UTP,
and SNP
fusion with L-NAME (30 ,gM) caused an end sensitivity of preparations to NE when this
.applied to raise the tone of the preparations.
ul titration of NE into the perfusate was re1 to allow the tone to be raised to 88.4±11.0
[g (n=8), which was not significantly different
the control preparations. The concentration of
quired to produce this increase was 1-8 ,uM.
ME produced a significant attenuation of rees to ACh at all doses up to and including 5
(Figure 5). Responses to SNP were not attenbut the lower doses were, in fact, enhanced in
resence of L-NAME (results not shown).
ME also attenuated vasodilator responses to
ind UTP (Figures 6 and 7). Probably because of
ifficulty in manipulating the tone to compare
re-L-NAME tone, there was a large variation
en preparations in the degree of effectiveness
qAME as an antagonist. There was a tendency
6
65
-log mol purine or pyrimidine
FIGURE 3. Dose-response curves showing vasoconstrictor
responses of the rat mesenteric arterial bed to purines and
pyrimidines before (solid lines) and after (broken lines) removal of the endothelium with sodium deoxycholate (n=6).
Response curves to ATP (-), UTP (A), CTP (o), GTP (A),
and TTP (o) are shifted to the left in the absence of the
endothelium. Symbols plus vertical bars indicate mean ±SEM.
ing vasodilator responses; pD2 values (the negative
log of the dose per 50 ,ll bolus) of 8.83+0.14 (n= 11)
and 8.39±0.15 (n=8) were obtained for ATP and
UTP, respectively; however, at the highest doses
there was a tendency for responses to ATP to become smaller (Figure 2b). After removal of the
endothelium with sodium deoxycholate (in the preparation at basal tone after NE had been washed out),
reintroduction of NE to the perfusate raised the tone
50
a,4-meATP (Figure 4).
b
a
I
E40-
FIGURE 4. Dose-response curves showing vasoconstrictor responses of the rat mesenteric arterial
bed to purines and pyrimidines before (solid lines)
and after (broken lines) the addition of a,,(3
methyleneATP (1 MM) to the perfusate. Note that
desensitization with a f,3methylene ATP virtually
abolishes responses to ATP (-, n=8) and CTP
(oi, n =6) (panel a) but has no effect on responses
to UTP (A, n=4), GTP (A, n=4), and TTP (o,
n=6) (panel b). Symbols plus vertical bars indi-
30-
20/
10
A
_
cate mean ±SEM.
_
0
A
7
6:5 6 5.5
-log mol purine or pyrimidine
8
75
8
7.5
65
6
-log mol purine or pyrimidine
55
Ralevic and Bumstock Vascular Effects of Purines and Pyrimidines
-log mol ACh
-log mol purine or pyrimidine
10
E
1587
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9
8
7
I
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FIGURE 5. Dose-response curves showing vasodilator responses of the rat mesenteric arterial bed to acetylcholine
(ACh) in the absence (o) and presence (o) of Nw-nitro-Larginine methyl ester (30 gAl) (n= 7). Symbols plus vertical
bars indicate mean ±SEM. *p<0.05, **p<0.01, and
***p<0.001 vs. the corresponding value in the presence of
N'O-nitro-L-arginine methyl ester.
for it to be more effective against ATP than UTP,
although this was not significant. There was a more
pronounced tendency for ATP to produce vasocon,1 t50
ATP
a
0
FIGURE 7. Dose-response curves showing attenuation of
vasodilator responses of the rat mesenteric arterial bed to A TP
(circles, n=6) and UTP (triangles, n= 7) by 30 ,M N0-nitroL-arginine methyl ester (L-NAME). Closed symbols indicate
control responses; open symbols indicate responses in the
presence of L-NAME. Symbols plus vertical bars indicate
mean+±SEM. *p<0.05, **p<0.0], and ***p<0.001 vs. control responses.
striction of the raised-tone preparation in the presence of L-NAME (Figure 6).
:
E
E
100-
E
0
0
c
0
50-
.0
0
..
S
*
S
0
98 88 78
6-8
a0a
0
60
8.8 78
68
L-NAME
UTP
b 150E
E
0
1 00 -
0
c
5
,to
:3
i
*.
S
98 88 78
*
0
S
68
06a50
60
9-8 88 78
L-NAME
0
0
68
2 min
FIGURE 6. Representative tracings showing responses of the rat
mesenteric bed to doses (denoted by solid circles and expressed as
-log mol) ofATP and UTP before and after addition of 30 pM
NW_nitro-L-aiginine methyl ester (L-NAME) (denoted by horizontal bar). Alternate doses are labeled startingffrom the top dose
at the left. Intermediate doses, which are not labeled, are, from
left to right, as follows (-log mol): 9.3, 8.3, and 7.3. Note the
prominent biphasic response produced byATP, but not by UTP,
in the presence of L-NAME.
Discussion
The results of this study demonstrate the presence
of the recently described pyrimidinoceptors,8,9 distinct from purinoceptors, in the rat mesenteric arterial bed. In this preparation, the low perfusion pressures obtained at "basal tone" are not representative
of normal conditions in vivo but allow assessment of
vasoconstrictor mechanisms with minimum contribution of opposing dilator effects. NE added to the
perfusate to raise the tone allowed us to look selectively at vasodilator mechanisms. In vivo, the net
response is the resultant of vasodilator and vasoconstrictor effects, the relative contribution of which will
vary according to vascular tone. As with ATP, the
pyrimidines UTP and TTP and the purine GTP can
elicit both vasoconstriction and vasodilatation in the
mesenteric arterial bed, after action at receptors
located on the vascular smooth muscle and on the
endothelium, respectively. CTP produces only contraction. Desensitization to a,/3-meATP was used to
distinguish between the P2x-purinoceptors and pyrimidinoceptors mediating vasoconstriction, whereas
EDRF was shown to be common to the mechanism of
action of the vasodilator activity of UTP and ATP.
The many potent and diverse effects of ATP have
long been recognized (see Reference 1 for review).
Action-specific P2-purinoceptor subtypes have been
described that, in the cardiovascular system, can
mediate contraction (the P2X-purinoceptor) and re-
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Circulation Research Vol 69, No 6 December 1991
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laxation (the P2Y-purinoceptor). In contrast there are
comparatively few reports of the vascular effects of
the other naturally occurring nucleotides. As with
ATP, both vasoconstrictor and vasodilator effects
have been described for UTP; UTP causes vasoconstriction in several preparations including the rabbit
ear artery,9 isolated rat mesenteric vessels,18 and the
isolated perfused rat liver,19 whereas endotheliumdependent vasodilatations have been described in the
pig aorta20 and human pial vessels.21 Both vasoconstriction and vasodilatation due to UTP have been
shown in the perfused rat hind limb.22,23 Since UTP is
a constituent of platelets and brain tissue, it has been
suggested that this agent may have a role as a
mediator of cerebral vasospasm.724 The effects of
CTP, GTP, and TTP on the vasculature tend to be
less pronounced. A lack of effect of TTP and CTP on
human pial vessels has been described,21 although
CTP and GTP have been shown to cause vasoconstriction of the rat hind limb22 and endotheliumdependent relaxation of the pig aorta.20 GTP and
CTP potentiate vasoconstriction to exogenous NE in
the rat mesenteric arterial bed,25 a process that has
also been described for ATP in this preparation.26
One of the first studies to investigate specifically
the possibility of discrete vascular purinoceptors and
pyrimidinoceptors was that of von Kugelgen et a19 in
the rabbit ear artery. These workers used the selective P2X-desensitizing agent a,f3-meATP11 to demonstrate that, although vasoconstrictor responses to
ATP were reduced by 88%, those to UTP were
enhanced or only partially reduced. They concluded
that, in their preparation, UTP elicited vasoconstriction by receptors distinct from the P2-purinoceptor.
In the rat mesenteric arterial bed, we also found that,
although vasoconstrictor responses to ATP were virtually abolished by prior desensitization with a,3meATP, those to UTP, GTP, and TTP were unaffected, which implies a vasoconstrictor action at
receptors other than at the P2x-purinoceptor. Vasoconstrictor responses to CTP, however, were abolished by a,4-meATP. It is possible that the stereochemical similarity of ATP and CTP at the amino
group (N6 for ATP, N4 for CTP) is a crucial link
between the antagonistic effect of a,f3-meATP and
the pressor responses elicited by these agents. CTP
differed from the other purines and pyrimidines in
that it elicited powerful contractions, but only at
relatively high doses, as reflected by its steep doseresponse curve. It is possible that its effects were
mediated indirectly after release of ATP by high
concentrations of this agent, which would account for
the fact that its responses were abolished by a,/3meATP. If this was so, the major source of the
released ATP is likely to be the smooth muscle or
nerves rather than endothelial cells, since removal of
the endothelium did not impair the ability of CTP to
cause vasoconstriction. However, this remains to be
investigated. On the other hand, prolonged exposure
to CTP markedly reduced ATP-induced contractions
of the guinea pig bladder,27 suggesting that in this
tissue CTP and ATP act at the same site (the
P2x-purinoceptor), a possibility that cannot be excluded in our studies.
In a previous study,3 we have shown that, as in
many other isolated vessels and vascular beds, the
receptors for ATP-mediated vasoconstriction of the
rat mesenteric arterial bed are located on the vascular smooth muscle. The present study confirmed
these findings and showed that this was also the case
for receptors mediating vasoconstriction to UTP,
GTP, TTP, and CTP, because responses were not
abolished after removal of the endothelium. In the
absence of the endothelium, pressor responses to all
of these agents were significantly greater than in the
controls, as reflected in a shift to the left of the
dose-response curves. An increased responsiveness
to various agents after removal of the endothelium
has previously been observed in this and in other
preparations.3,28,29 The enhanced responsiveness may
be due to either 1) an increased local concentration
of these agents at their smooth muscle receptors by
virtue of the removal of a diffusion barrier (the
endothelium)30 or 2) the removal of modulatory
effects of the endothelium.28 Since responses to CTP,
which does not elicit relaxation in this preparation,
were also enhanced, it is less likely that this effect was
due to the removal of an opposing relaxant effect
exerted at endothelial receptors.
The rank order of vasodilator potency of the
purines and pyrimidines in the rat mesenteric bed
was found to be UTP=ATP>TTP=GTP. CTP had
no vasodilator effects in this preparation or in the rat
hind limb preparation,22 in contrast to the pig aorta,
in which it elicits endothelium-dependent relaxation.20 ATP and UTP were equipotent in eliciting
vasodilatation; however, at the highest doses, responses to ATP showed a tendency to become
smaller. It is likely that at these doses the balance of
the response shifts in the favor of the P2x-purinoceptor-mediated contraction, opposing the P2Y-purinoceptor-mediated vasodilatation. Since UTP is less
potent as a vasoconstrictor than ATP, this opposing
effect is less pronounced, resulting in more potent
relaxations compared with ATP at high doses. In the
present study we found that, as in the pig aorta20 and
human pial vessels,21 relaxation of the rat mesenteric
arterial bed by UTP was endothelium dependent.
Relaxations to ATP, TTP, and GTP were also dependent on an intact endothelium.
ATP can stimulate prostaglandin and prostacyclin
production from perfused vascular beds and from
endothelial cells in culture,31 as well as cause the
production of EDRF.32 UTP has also been shown to
stimulate prostacyclin production in endothelial
cells33 and in the rat liver.'9 The vascular relaxations
produced by ATP and UTP could, therefore, be
mediated by production of either of these endothelium-derived substances, although there is evidence
that in the case of ATP relaxation is more likely to
proceed via EDRF.34 We used this information to
investigate, and possibly distinguish between, the
Ralevic and Bumstock Vascular Effects of Purines and Pyrimidines
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vasodilator mechanisms of ATP and UTP by using
L-NAME, an inhibitor of the pathway of conversion
of L-arginine to nitric oxide15 and, hence, an inhibitor
of relaxations mediated by EDRF (identified as nitric
oxide35). A more direct approach to distinguish between P2Y-purinoceptor-mediated and pyrimidinoceptor-mediated relaxations could not be adopted
because of the absence of specific antagonists to
pyrimidinoceptors and because the P2Y-purinoceptor
antagonist, reactive blue 2,36 caused a drop in tone of
the raised-tone preparation.
L-NAME has been successfully used in the rat
mesenteric bed to antagonize relaxant responses to
ACh (Reference 15 and the present study). In the
current study, it significantly attenuated relaxations to
both ATP and UTP, suggesting that, for both of these,
relaxation takes place largely through the formation of
EDRF and not through prostacyclin. The tendency for
L-NAME to produce a greater antagonism of vasodilatation to ATP than UTP may be due to an unmasking of the more potent effects of ATP as a contractile
agent at P2X-purinoceptors, although it may also be
because a more significant proportion of the relaxation to UTP is mediated through a non-EDRF
pathway. The potentiation of responses to NE in the
presence of L-NAME may be due to inhibition of a
continuous basal release of endogenous EDRF, allowing constriction to occur with greater potency. Responses to the endothelium-independent vasodilator
SNP were enhanced by L-NAME. The reason for the
enhancement of responses to SNP is not known but
has also been observed by us in the isolated perfused
rabbit liver preparation and is the subject of a separate
communication.37
In conclusion, we have demonstrated the presence
of pyrimidinoceptors, distinct from P2-purinoceptors,
in the rat mesenteric arterial bed. As with P2-purinoceptors, activation of action-specific pyrimidinoceptors located on the smooth muscle and on
endothelial cells elicits vasoconstriction and vasodilatation, respectively. Further characterization of
these receptors awaits the development of pyrimidine
agonists and antagonists in the same way as has been
done for ATP and other purines. However, there is
strong evidence that pyrimidinoceptors are a heterogeneous population comprising at least two subtypes.
Although the physiological relevance of pyrimidines
in the regulation of rat mesenteric resistance vessels
is not clear from the present study, it is feasible that
they may be released under certain physiological or
pathophysiological conditions to contribute to the
regulation of vascular tone. A preliminary characterization has been made of the receptors involved, and
a model system has been described for testing agonists and antagonists of these agents.
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KEY WORDS
purines * pyrimidines * mesenteric bed
endothelium * smooth muscle
Effects of purines and pyrimidines on the rat mesenteric arterial bed.
V Ralevic and G Burnstock
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Circ Res. 1991;69:1583-1590
doi: 10.1161/01.RES.69.6.1583
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