Extract
Note: Please read the complete
full text with Figures and Tables at
Introduction
Urotensin II (UII) is a vasoactive 'somatostatin-like' cyclic peptide which was originally isolated from fish spinal cords,
and has recently been cloned from
man[1,2]. The orphan receptor GPR14, a G-protein-coupled receptor first cloned from rat,
has been defined as the specific receptor for
UII[3_5]. Positive immunoreactive staining for human UII and GPR14 mRNA was
detected in both atrial and ventricular
tissues[3_6]. These studies indicate that UII might act as one of the endogenous
modulators of cardiac function. The cardiovascular action of UII has been examined in a number of
in vitro and in vivo systems, sometimes with contrasting responses depending on the species and vessel studied. In mammals, UII was first
reported to have a vasoconstrictor action in isolated rat thoracic aorta denuded of
endothelium[7], an action that was later shown
to be more potent than that of
endothelin[3]. UII was also shown to cause vasoconstriction in endothelium-denuded vessels
from various species including cynomolgus monkey thoracic
aorta[3], dog coronary
artery[3], and human radial
artery[8]. In contrast, UII caused vasodilation in human small pulmonary arteries and abdominal adipose tissue
arteries[9]. In addition, the cardiovascular responses to
UII in vivo are equally variable. Systemic administration of human UII to monkeys elicited a
fall in cardiac output, severe myocardial contractility
depression, and fatal circulatory
failure[3]. Both in anesthetized and
conscious rats, UII administered iv decreased arterial
pressure[10,11], and hypotension was associated with mesenteric
and hindquarter vasodilatation[11], while
intracerebroventricular injection UII caused arterial pressure and heart rate increase
in conscious rats[12]. Even less clear is the role of UII in
states of cardiovascular disease, but to date, increased plasma
UII has been described in 3 states of volume overload, such
as kidney failure[6], heart
failure[13], and
diabetes[14]. Thus, it is important to have more detailed understanding of the
actions and the role of UII in cardiovascular control.
It is well known that baroreflex is a major mechanism in
modulating blood pressure (BP). Whether UII affects the
carotid sinus baroreflex (CSB) remains to be clarified. The
aims of our study were to observe the action of UII on
the isolated CSB and to elucidate the mechanism involved.
Materials and methods
Drugs UII, verapamil, BIM-23127, and
L-NAME were purchased from Sigma (St Louis, MO, USA). All drugs were
dissolved in distilled water and prepared freshly.
General surgical procedure Male Sprague-Dawley rats
(320±20 g), obtained from the Experimental Animal Center of
Hebei Province (Hebei, Shijiazhuang, China), were
anesthetized with 25% urethane (1.0 g/kg, ip). The trachea was
cannulated for ventilation. The right femoral artery was
cannulated for recording BP with a transducer (MPU-0.5A, Nihon
Kohden, Tokyo, Japan). Body temperature was maintained
at 37_38 ºC throughout the experiment.
Perfusion of left isolated carotid sinus
The perfusion of the isolated carotid sinus area was carried out with a method
modified by our laboratory[15,16]. Carotid sinus areas were
fully exposed by turning rostrally the trachea and esophagus.
Sternohyoideus muscles and superior laryngeal nerves were
cut, then the bilateral aortic nerves, right carotid sinus nerve,
cervical sympathetic nerves, and recurrent laryngeal nerves
were all sectioned. The common, external and internal
carotid arteries and smaller arteries originating from these
vessels were exposed and ligated, while carefully leaving the
left carotid sinus nerve undisturbed. Ligation of the
occipital artery at its origin from the external carotid artery excluded
chemoreceptors from the isolated carotid sinus, thereby
preventing chemoreceptor activation secondary to the decrease
of carotid sinus pressure. Plastic catheters were inserted
anterograde into the left common carotid artery (inlet tube)
and retrograde into the external carotid artery (outlet tube).
The carotid sinus was then perfused with warm (37 ºC)
oxygenated modified Krebs-Henseleit (K-H) solution (in
mmol/L; NaCl 118.0, KCl 4.7, CaCl2 2.5,
MgSO4 1.6, KH2PO4
1.2, NaHCO3 25, glucose 5.6, pH 7.35_7.45), bubbled with
95% O2 and 5% CO2. The intrasinus pressure (ISP) was
monitored by using a pressure transducer (MPU-0.5A, Nihon
Kohden, Tokyo, Japan) connected to the inlet tube. The ISP
was controlled by using a peristaltic pump.
After perfusion of the left carotid sinus, the ISP was kept
at 100 mmHg for 20 min and then was lowered to 0 mmHg
rapidly. From this point, the ISP was elevated to 250 mmHg
in the form of a pulsatile ramp by regulating the speed of the
peristaltic pump, which was automatically controlled by a
program designed in our laboratory[15]. It took 0.5 min for the
ISP to be increased from 0 to 250 mmHg. The ISP and BP
were simultaneously recorded on a polygraph (RM-6240,
Chengdu Instrument Factory, Sichuan, China). This
process was repeated at an interval of 5 min to check the
stability of the baroreflex. The reproducibility of the experimental
setup was confirmed by the recurrent drop of BP in response
to the increase in ISP.
Experimental protocols By perfusing the left carotid
sinus with K-H solution and elevating the ISP, a functional
curve for the ISP_BP relation was constructed, and the
functional parameters of baroreflex, such as threshold pressure
(TP), saturation pressure (SP), equilibrium pressure (EP), peak
slope (PS), reflex decrease of BP (RD), and operating range
(OR) were determined. TP was the ISP at which BP began to
decrease in response to the increase of ISP. SP was the ISP
at which BP just showed no further reflex decreases with an
increase in ISP. OR was calculated as the SP minus TP.
Before administration of drugs, the K-H solution was used
as a control. Four experimental treatments were used: (1) to
test the effect of UII on carotid baroreflex
(n=24), the ISP was fixed at 100 mmHg for 20 min with K-H solution as a control,
and the baroreflex parameters were measured. Then K-H
solution containing UII (3, 30, 300, or 3000 nmol/L) was used
to perfuse the isolated carotid sinus for 50 min; afterward the
parameters were measured again. Finally, the carotid sinus
was perfused with K-H solution to wash out the UII; (2) to
test the effect of verapamil (10 µmol/L) on the actions of UII
(n=6), baroreflex parameters were examined following
application of UII (300 nmol/L) before and after pretreatment with
verapamil for 20 min; (3) to test the effect of BIM-23127 (3.0
µmol/L) on the actions of UII (n=6), baroreflex parameters
were examined following the application of UII (300 nmol/L)
before and after pretreatment with BIM-23127 for 20 min;
and (4) to test the effect of
NG-nitro-L-arginine methyl ester
(L-NAME, 100 µmol/L) on the actions of UII
(n=6), L-NAME were added into a K-H solution and used to perfuse the
isolated carotid sinus after the baroreflex parameters of the
control were recorded; 20 min later the baroreflex parameters
were recorded again. Finally, the carotid sinus was perfused
with K-H solution containing L-NAME and UII (300 nmol/L)
to check the carotid baroreflex activity.
Data analysis All data were expressed as mean±SD.
The differences between groups of means were assessed
by one-way ANOVA and further analyzed using the
Student-Newman-Kuels test. P<0.05 was considered statistically
significant.
Results
Effects of UII on carotid sinus baroreflex By perfusing
the left carotid sinus with K-H solution and elevating ISP
from 0 to 250 mmHg, the BP was reflexly decreased. UII
altered the baroreflex parameters, which appeared
approximately 30 min after UII started, and disappeared after
30-60 min perfusion of washout. Compared with the control group,
UII at the concentration of 3 nmol/L had no effect on the
CSB, while at concentrations of 30, 300, and 3000 nmol/L, it
inhibited the CSB, shifting the functional curve of the
baroreflex upward and to the right. There was a marked
decrease in PS and RD in BP. The effects were
concentration dependent (Table 1, Figure 1).
The functional curves were shown in Figure 1. These results indicated that UII
exerted an inhibitory action on the carotid baroreflex
(Figure 2).
Effect of verapamil on the actions of UII
Verapamil (10 µmol/L) did not affect the functional parameters of the
carotid baroreceptor, but it partially blocked the actions of
UII (300 nmol/L, Table 2).
Effect of BIM-23127 on the actions of UII
BIM-23127
(3.0 µmol/L), an antagonist of human and rat UII receptors,
did not affect functional parameters of baroreflex alone, but
it abolished the actions of UII (300 nmol/L) on the CSB.
Effect of L-NAME on the actions of UII
L-NAME (100 µmol/L) did not affect the functional parameters of the
carotid baroreceptor, and did not influence the effects of UII
(300 nmol/L) either (Table 2).
Discussion
The present study showed for the first time that UII could
inhibit the CSB in a dose-dependent manner. By perfusing
the left isolated carotid sinus with UII, the functional curve
of the CSB was shifted upward and to the right, with
decreases in PS and RD and increases in TP, indicating the
inhibitory action of UII on the CSB. It is well established
that the arterial baroreceptors play an important role in the
short-term control of cardiovascular activity. The inhibition
of the CSB might be expected to weaken the ability to
anta-gonize hypertension and destabilize BP.
It was demonstrated that baroreceptors would be excited
when their ending was deformated by tissue
tension[17],
that is, wall tension s, which by the Laplace's relation is:
s=P×r/h (where P is the distending pressure,
r is the radius of the vessel lumen, and
h is the wall thickness). UII might directly induce arterial contraction and thus decrease
r, so s became smaller, and then the sensitivity of the baroreceptor
was decreased. Thus, UII-induced inhibition of the CSB
might have resulted from the contraction of the vessel.
Indeed, it is known that UII is a vasoconstrictor, being even
more potent than endothelin[3]. UII was also shown to
stimulate the constriction of endothelium-denuded rat aorta and
increase cytosolic Ca2+ levels within 10_20 s, which was
partially blocked by the L-type calcium channel antagonist,
verapamil[18]. In order to elucidate the relationship between
the L-type calcium and the action of UII, we used verapamil.
We found in the present study that pretreatment with
verapamil partially blocked the effects of UII on the CSB,
suggesting that the opening of the L-type calcium channel
was involved in the action of UII on the CSB.
BIM-23127 was reported as a potent and competitive
antagonist of both human UII and rat UII receptors, and was
also considered as the most potent agent for reversing
human UII-induced contractile tone in the rat isolated
aorta[19]. In the present study, we found that BIM-23127 abolished
UII action on the CSB. This result indicates that UII inhibits
the CSB through the UII receptor.
UII was also reported as a vasodilator via release of one
or more vasodilator factors such as nitric oxide (NO) in rat
isolated aorta[20]. Since the endothelium is intact in the present
study, UII may induce NO release from the carotid sinus.
Increasing evidence have shown that NO may suppress the
generation of action potential of
baroreceptors[21,22]. NO suppressed
Na+ current in baroreceptor neurons and activated
the calcium-dependent K+ channels localized in vascular
smooth muscles, then hyperpolarized baroreceptor
neurons[21]. In order to test the role of NO plays, we tested the effect of
L-NAME, a nonselective inhibitor of the NO synthase.
Pretreatment with L-NAME did not affect the action of UII, thus
indicating that locally-released NO was not involved in the
effect of UII on the CSB. Thus, we propose that
vasodilatation in response to UII is likely to have resulted from UII
receptors on the endothelium, while constriction responses
are mediated by UII receptors on smooth muscle cells.
In summary, the present study indicates that UII exerts
an inhibitory action on the isolated CSB. Such an action of
UII is predominantly mediated by the UII receptors in
vascular smooth muscles, resulting in the opening of
L-type calcium channels.
References
1 Chartel N, Conlon JM, Collin F, Braun B, Waugh D, Vallarino M,
et al. Urotensin II in the central system of the frog Rana ridibunda:
immunohistochemical localization and biochemical
characteriza-tion. J Comp Neurol 1996; 364: 324_39.
2 Conlon J M, Yano K, Waugh D, Hazon N. Distribution and
molecular forms of urotensin II and its role in cardiovascular
regulation in vertebrates. J Exp Zool 1996; 275: 226_38.
3 Ames RS, Sarau HM, Chambers JK, Willette RN, Aiyar NV,
Romanic AM, et al. Human urotensin-II is a potent
vasoconstrictor and agonist for the orphan receptor GPR14. Nature
1999; 401: 282_6.
4 Liu QY, Pong SS, Zeng ZZ. Identification of urotensin II as the
endogenous ligand for the orphan G-protein-coupled receptor
GPR14. Biochem Biophys Res Commun 1999; 266: 174_8.
5 Mori M, Sugo T, Abe M, Shimomura Y, Kurihara M, Kitada C,
et al. Urotensin II is the endogenous ligand of a G-protein-coupled
receptor, SENR (GPR14). Biochem Biophys Res Commun 1999;
265: 123_9.
6 Totsune K, Taskahashi K, Arihara Z, Sone M, Stoh F, Ito S,
et al. Role of urotension II in patients on dialysis. Lancet 2001; 358:
5_9.
7 Douglas SA, Sulpizio AC, Piercy V, Sarau HM, Ames RS, Aiyar
NV, et al. Differential vasoconstrictor activity of human
urotension-II in vascular tissue isolated from the rat, mouse, dog,
pig, marmoset and cynomolgus monkey. Br J Pharmacol 2000;
131: 1262_74.
8 Maguire JJ, Kuc RE, Davenport AP. Orphan-receptor ligand
human urotensin-II: receptor localization in human tissue and
comparison of vasoconstrictor responses with endothelin-1. Br J
Pharmacol 2000; 131: 441_6.
9 Stirrat A, Gallagher M, Douglas SA, Ohlstein EH, Berry C, Kirk
A, et al. Potent vasodilator responses to human urotensin-II in
human pulmonary and abdominal resistance arteries. Am J Physiol
Heart Circ Physiol 2001; 280: 925_8.
10 Gibson A, Wallace P, Bern HA. Cardiovascular effects of urotensin
II in anesthetized and pitched rats. Gen Comp Endocrinol 1986;
64: 435_9.
11 Gardiner SM, March JE, Kemp PA, Davenport AP, Bennet T.
Depressor and regionally selective vasodilator effects of human
and rat urotensin II in conscious rats. Br J Pharmacol 2001; 132:
1625_9.
12 Lin Y, Tsuchihashi T, Matsumura K, Abe I, Iida M. Central
cardiovascular action of urotensin II in conscious rats. J Hypertens
2003; 21: 159_65.
13 Richards AM, Nicholls MG, Lainchbury JG, Fisher S, Yandle TG.
Plasma urotensin II in heart failure. Lancet 2002; 360:
545-6.
14 Totsune K, Takahashi K, Arihara Z, Sone M, Ito S, Muramaki O.
Increased plasma urotensin II levels in patients with diabetes
mellitus. Clin Sci 2003; 104: 1_5.
15 Yi XL, Fan ZZ, He RR. An automatic system controlled by
computer for carotid sinus perfusion. Chin J Appl Physiol 1993;
9: 156_9.
16 Wu YM, He RR. Bradykinin inhibits carotid sinus baroreflex in
anesthetized rats. Acta Physiol Sin 1999; 51: 303_9.
17 Arndt JO. Baroreceptors: morphology and mechanics of
receptor zones and sischarge properties of baroafferents. In: Zucker
IH, Gilmore JP. editors. Reflex control of the circulation. Boston:
CRC Press; 1991. p 103_8.
18 Tasaki K, Hori M, Ozaki H, Karaki H, Wakabayashi I.
Mechanism of human urotension II-induced contraction in rat aorta. J
Pharmacol Sci 2004; 94: 376_83.
19 Herold CL, Behm DJ, Buckley PT, Foley JJ, Wixted MM,
Douglas SA. The neuromedin B receptor antagonist, BIM-23127, is
a potent antagonist at human and rat urotensin-II receptors. Br
J Pharmacol 2003; 139: 203_7.
20 Gibson A. Complex effects of Gillichthys urotensin II on rat
aortic strips. Br J Pharmacol 1987; 91: 205_12.
21 Li Z, Chapleau MW, Bates JN, Bielefeldt K, Lee HC, Abboud FM.
Nitric oxide as an autocrine regulator of sodium currents in
baroreceptor neurons. Neuron 1998; 20: 1039_49.
22 Chapleau MW, Hajduczok G, Sharma RV, Wachtel RE,
Cunningham JT, Sullivan MJ, et al. Mechanisms of baroreceptor
activation. Clin Exp Hypertens 1995; 17: 1_13.
|
