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Blocking Effects of OPC-21268 and OPC-31260 (Vasopressin V1- and V2-Receptor Antagonists) on Vasopressin-Induced Constrictions in Isolated, Perfused Dog Femoral Arteries
Blocking Effects of OPC-21268 and OPC-31260 (Vasopressin V1 and V2-Receptor Antagonists) on Vasopressin-Induced Constrictions
in Isolated,
Perfused
Shigetoshi
Chiba
Dog
and Miyoko
Femoral
Arteries
Tsukada
Department of Pharmacology, Shinshu University School of Medicine, Matsumoto 390, Japan Received
December
18, 1991
Accepted
March
9, 1992
ABSTRACT Using a perfusion technique of isolated vessels, constrictor responses to vasopressin (VP) and norepinephrine (NE) were investigated in perfused dog femoral arteries. Both OPC-21268, a selec tive V1-antagonist, and OPC-31260, a selective V2-antagonist, significantly shifted the VP-induced dose response curves to the right without influencing the NE-induced ones. The blocking effects of OPC 31260 were much greater than those of OPC-21268, suggesting that there may probably be functional V1 and V2-receptors in isolated dog femoral arteries that mediate vasoconstriction. Keywords:
Vasopressin
receptor
subtypes,
VI-antagonist,
In vascular smooth muscle, vasopressin (VP) induces a vasoconstriction through V1-receptors by a cAMP-in dependent mechanism that is coupled to phosphoinosi tide turnover (1, 2). As reviewed by Share (3), VP is a potent vasoconstrictor, acting directly on vascular smooth muscle via VI-receptors. He also reported that there may also be vascular V2-receptors, the activation of which causes vasodilation, but the physiological role of these receptors is questionable. Recently, nonpeptide V1 and V2-receptor antago nists were developed, i.e., OPC-21268 and OPC-31260, respectively (4, 5). OPC-21268 selectively antagonized binding to the V1-subtype of the VP-receptor in a com petitive manner (4). In experiments in vivo, it acted as a specific antagonist of the VP-induced pressor re sponse in pithed rats and conscious rats (4). More re cently, it was reported that OPC-31260 selectively antagonized binding to the V2-subtype of the VP recep tor in a competetive manner (5). Thus, in the present study, we attempted to investi gate the effects of two VP antagonists on VP-induced vasoconstrictions in isolated dog femoral arteries, using the cannula inserting method, which was developed and modified by Hongo and Chiba in 1983 (6) and Tsuji and Chiba in 1984 (7). Mongrel dogs, weighing 8-16 kg, were anesthetized with sodium pentobarbital (30 mg/kg, i.v.). After treat ment with sodium heparin (200 units/kg, i.v.), the
V2-antagonist
animals were killed by rapid exsanguination. The femo ral arteries were dissected and carefully removed. Iso lated femoral arteries, which were 1.5 to 2.5 cm in length were selected for study. A stainless steel cannula with small holes 2 mm from the distal sealed end (1.15 to 2.65 mm in outer diameter and 3 cm length) was carefully inserted into each vessel segment to avoid in jury of the intraluminal surface of the isolated vessel. Segments were set up in the bath to maintain the prep aration at 37°C and perfused with Krebs' solution at a constant flow rate (2.0 4.0 ml/min) by means of a pump (the perfusion pressure = 50-100 mmHg). The perfusate contained 114 mM NaCI, 4.7 mM KCI, 2.5 mM CaC12, 1.2 mM KH2PO4, 1.2 mM MgSO4, 25 mM NaHCO3 and 11.1 mM glucose. The perfusion pressure was continuously measured with an electric manometer. The details of the cannula inserting method were de scribed in previous papers (7, 8). The drugs used were arginine vasopressin (VP, Sigma), norepinephrine hy drochloride (NE, Sankyo), OPC-21268 (1-(1-[4-(3-acetyl aminopropoxy)benzoyl]-4-piperidyl)-3,4-dihydro-2(1H) quinolinone, Ohtsuka Co.) and OPC-31260 ([5-dimeth ylamino 1(4-(2-methylbenzoylamino)benzoyl)-2,3,4,5 tetrahydro-1H-benzazepine], Otsuka Co.). OPC-21268 was dissolved in dimethylsulfoxide (DMSO) (Wako Pure Chemicals). The volume of dimethylsulfoxide used (10-30,ul) did not modify VP-induced vasoconstrictions (n = 3-10). The drug solution was administered into
the rubber
tubing
close
to the cannula
in a volume
0.01 to 0.03 ml for a period of 4 sec. The data sented as mean values ± S.E.M.
vious response. NE-induced vasoconstrictions were not inhibited by 100 ,ug OPC-21268, but rather slightly, though not significantly, potentiated by it. VP-induced vasoconstrictions were significantly inhibited by rela tively high doses OPC-21268 (100 300 ,ug), a selective V1-receptor blocker. Figure 1 summarizes the data. Even a relatively small dose (10,ug) of OPC-31260, a selective V2-blocker, readily inhibited the vasoconstric tions. These data are summarized in Fig. 2. Since it was proven that OPC-21260 is a selective V1-receptor anta gonist (5), the present results indicate that isolated dog
of
are pre
When NE was administered into the cannulated fem oral artery of the dog, the vasoconstrictor response was readily induced in a dose-related manner as reported previously (8). VP also induced a dose-related vasocon striction. In this arterial preparation, a relatively large amount of 0.01 U VP produced a strong vasoconstric tion but did not induce tachphylaxis when the following dose was applied after a complete cessation of the pre
Fig.
1.
oral
arteries.
Effects
Fig.
2.
oral
arteries.
of OPC-21268 Points
Effects
represent
of OPC-31260 Points
represent
on
vasopressin
the
mean
on the
value
vasopressin mean
value
and and
norepinephrine-induced vertical
and and
bars
represent
norepinephrine-induced vertical
bars
represent
vasoconstrictions
in isolated,
perfused
dog
fem
± S.E.M.
vasoconstrictions ± S.E.M.
in isolated,
perfused
dog
fem
femoral arteries contain vasoconstriction-mediated Vi receptors. However, the selective V2-receptor antago nist OPC-31260 (6) potently inhibited the VP-induced vasoconstrictions in the same preparations. This may in dicate that there are also abundant vasoconstriction mediated V2-receptors in the dog femoral arteries. Liard et al. (9, 10) reported that under certain cir cumstances, activation of what are presumed to be vascular V2-receptors may cause vasodilation. In con scious dogs pretreated with a Vl-antagonist, the in travenous administration of VP caused an increase in cardiac output and a decrease in total peripheral resist ance, effects opposite to those obtained with vasopres sin alone (10). In blood vessels in vivo, an applied vasoactive substance exerts it action on the resistance vessel vasculature, but in blood vessels studied in vitro, the substance affects relative large vessels. Vessels of different sizes may show different responses to VP as demonstrated by the vascular responses to histamine; i.e., the histamine H1-receptor mediates a vasoconstric tion in large vessels but causes vasodilatation in small resistance vessels (11). Although OPC-31260 caused a more potent inhibition than OPC-21268 on VP-induced vasoconstriction in isolated dog femoral arteries, re gional and species differences should be considered when estimating VP receptor subtypes in addition to in terpreting the data from receptor binding studies. Acknowledgment We Co.,
thank Ltd.
for
Dr.
Youichi
his kind
Yabuuchi
gifts
of the
of OPC-21268
Otsuka and
Pharmaceutical
OPC-31260.
REFERENCES 1
Tolbert,
M.E.M.,
White,
A.C.,
Aspry,
K.,
Cutts,
J.
and
2
3 4
Fain, J.N.: Stimulation by vasopressin and a-catecholamines of phosphatidylinositol formation in isolated rat liver paren chymal cells. J. Biol. Chem. 225, 1938-1944 (1980) Berridge, M.J. and Irvine, R.F.: Inositol trisphosphate, a novel second messenger in cellular signal transduction. Nature 312, 315 321 (1984) Share, L.: Role of vasopressin in cardovascular regulation. Physiol. Rev. 64, 1248 -1284 (1988) Yamamura, Y., Ogawa, H., Chihara, T., Komdo, K., Ono
gawa, T., Nakamura, S. et al.: OPC-21268, an orally effec tive, nonpeptide vasopressin V1 receptor antagonist. Science 252, 572-574 (1991) 5 Yamamura, Y., Ogawa, H., Yamashita, H., Chihara, T., Miyamoto, H., Nakamura, S. et al.: Characterization of a novel aquaretic agent, OPC-31260, as an orally effective, nonpeptide vasopressin V2 receptor antagonist. Br. J. Phar macol. (in press) 6 Hongo, K. and Chiba, S.: A new method for measuring vascular responsiveness of relatively larger arteries of dogs. J. Pharmacol. Methods 9, 83 91 (1983) 7 Tsuji, T. and Chiba, S.: Potentiating effect of methysergide on norepinephrine-induced constriction of the isolated inter nal carotid artery of the dog. Japan. J. Pharmacol. 34, 95 100 (1984) 8 Kawai, K. and Chiba, S.: Vascular responses of isolated, per fused canine femoral arteries and veins to vasoactive sub stances. Japan. J. Pharmacol. 41, 537-540 (1986) 9 Liard, J.F., Deriaz, 0., Schelling, P. and Thibonnier, M.: Cardiac output distribution during vasopressin infusion or de hydration in conscious dogs. Am. J. Physiol. 243, H663 H669 (1982) 10 Liard, J.F. and Spadone, J.C.: Hemodynamic effects of an tagonists of the vasoconstrictor action of vasopressin in con scious dogs. J. Cardiovasc. Pharmacol. 6, 713 719 (1984) 11 Levi, R., Owen, D.A.A. and Trzeciakowski, J.: Actions of histamine on the heart and vasculature. In Pharmacology of Histamine Receptors, Edited by Ganellin, C.R. and Parsons, M.E., p. 236-297, Wright, PSG, Bristol (1982)
in Isolated,
Perfused
Shigetoshi
Chiba
Dog
and Miyoko
Femoral
Arteries
Tsukada
Department of Pharmacology, Shinshu University School of Medicine, Matsumoto 390, Japan Received
December
18, 1991
Accepted
March
9, 1992
ABSTRACT Using a perfusion technique of isolated vessels, constrictor responses to vasopressin (VP) and norepinephrine (NE) were investigated in perfused dog femoral arteries. Both OPC-21268, a selec tive V1-antagonist, and OPC-31260, a selective V2-antagonist, significantly shifted the VP-induced dose response curves to the right without influencing the NE-induced ones. The blocking effects of OPC 31260 were much greater than those of OPC-21268, suggesting that there may probably be functional V1 and V2-receptors in isolated dog femoral arteries that mediate vasoconstriction. Keywords:
Vasopressin
receptor
subtypes,
VI-antagonist,
In vascular smooth muscle, vasopressin (VP) induces a vasoconstriction through V1-receptors by a cAMP-in dependent mechanism that is coupled to phosphoinosi tide turnover (1, 2). As reviewed by Share (3), VP is a potent vasoconstrictor, acting directly on vascular smooth muscle via VI-receptors. He also reported that there may also be vascular V2-receptors, the activation of which causes vasodilation, but the physiological role of these receptors is questionable. Recently, nonpeptide V1 and V2-receptor antago nists were developed, i.e., OPC-21268 and OPC-31260, respectively (4, 5). OPC-21268 selectively antagonized binding to the V1-subtype of the VP-receptor in a com petitive manner (4). In experiments in vivo, it acted as a specific antagonist of the VP-induced pressor re sponse in pithed rats and conscious rats (4). More re cently, it was reported that OPC-31260 selectively antagonized binding to the V2-subtype of the VP recep tor in a competetive manner (5). Thus, in the present study, we attempted to investi gate the effects of two VP antagonists on VP-induced vasoconstrictions in isolated dog femoral arteries, using the cannula inserting method, which was developed and modified by Hongo and Chiba in 1983 (6) and Tsuji and Chiba in 1984 (7). Mongrel dogs, weighing 8-16 kg, were anesthetized with sodium pentobarbital (30 mg/kg, i.v.). After treat ment with sodium heparin (200 units/kg, i.v.), the
V2-antagonist
animals were killed by rapid exsanguination. The femo ral arteries were dissected and carefully removed. Iso lated femoral arteries, which were 1.5 to 2.5 cm in length were selected for study. A stainless steel cannula with small holes 2 mm from the distal sealed end (1.15 to 2.65 mm in outer diameter and 3 cm length) was carefully inserted into each vessel segment to avoid in jury of the intraluminal surface of the isolated vessel. Segments were set up in the bath to maintain the prep aration at 37°C and perfused with Krebs' solution at a constant flow rate (2.0 4.0 ml/min) by means of a pump (the perfusion pressure = 50-100 mmHg). The perfusate contained 114 mM NaCI, 4.7 mM KCI, 2.5 mM CaC12, 1.2 mM KH2PO4, 1.2 mM MgSO4, 25 mM NaHCO3 and 11.1 mM glucose. The perfusion pressure was continuously measured with an electric manometer. The details of the cannula inserting method were de scribed in previous papers (7, 8). The drugs used were arginine vasopressin (VP, Sigma), norepinephrine hy drochloride (NE, Sankyo), OPC-21268 (1-(1-[4-(3-acetyl aminopropoxy)benzoyl]-4-piperidyl)-3,4-dihydro-2(1H) quinolinone, Ohtsuka Co.) and OPC-31260 ([5-dimeth ylamino 1(4-(2-methylbenzoylamino)benzoyl)-2,3,4,5 tetrahydro-1H-benzazepine], Otsuka Co.). OPC-21268 was dissolved in dimethylsulfoxide (DMSO) (Wako Pure Chemicals). The volume of dimethylsulfoxide used (10-30,ul) did not modify VP-induced vasoconstrictions (n = 3-10). The drug solution was administered into
the rubber
tubing
close
to the cannula
in a volume
0.01 to 0.03 ml for a period of 4 sec. The data sented as mean values ± S.E.M.
vious response. NE-induced vasoconstrictions were not inhibited by 100 ,ug OPC-21268, but rather slightly, though not significantly, potentiated by it. VP-induced vasoconstrictions were significantly inhibited by rela tively high doses OPC-21268 (100 300 ,ug), a selective V1-receptor blocker. Figure 1 summarizes the data. Even a relatively small dose (10,ug) of OPC-31260, a selective V2-blocker, readily inhibited the vasoconstric tions. These data are summarized in Fig. 2. Since it was proven that OPC-21260 is a selective V1-receptor anta gonist (5), the present results indicate that isolated dog
of
are pre
When NE was administered into the cannulated fem oral artery of the dog, the vasoconstrictor response was readily induced in a dose-related manner as reported previously (8). VP also induced a dose-related vasocon striction. In this arterial preparation, a relatively large amount of 0.01 U VP produced a strong vasoconstric tion but did not induce tachphylaxis when the following dose was applied after a complete cessation of the pre
Fig.
1.
oral
arteries.
Effects
Fig.
2.
oral
arteries.
of OPC-21268 Points
Effects
represent
of OPC-31260 Points
represent
on
vasopressin
the
mean
on the
value
vasopressin mean
value
and and
norepinephrine-induced vertical
and and
bars
represent
norepinephrine-induced vertical
bars
represent
vasoconstrictions
in isolated,
perfused
dog
fem
± S.E.M.
vasoconstrictions ± S.E.M.
in isolated,
perfused
dog
fem
femoral arteries contain vasoconstriction-mediated Vi receptors. However, the selective V2-receptor antago nist OPC-31260 (6) potently inhibited the VP-induced vasoconstrictions in the same preparations. This may in dicate that there are also abundant vasoconstriction mediated V2-receptors in the dog femoral arteries. Liard et al. (9, 10) reported that under certain cir cumstances, activation of what are presumed to be vascular V2-receptors may cause vasodilation. In con scious dogs pretreated with a Vl-antagonist, the in travenous administration of VP caused an increase in cardiac output and a decrease in total peripheral resist ance, effects opposite to those obtained with vasopres sin alone (10). In blood vessels in vivo, an applied vasoactive substance exerts it action on the resistance vessel vasculature, but in blood vessels studied in vitro, the substance affects relative large vessels. Vessels of different sizes may show different responses to VP as demonstrated by the vascular responses to histamine; i.e., the histamine H1-receptor mediates a vasoconstric tion in large vessels but causes vasodilatation in small resistance vessels (11). Although OPC-31260 caused a more potent inhibition than OPC-21268 on VP-induced vasoconstriction in isolated dog femoral arteries, re gional and species differences should be considered when estimating VP receptor subtypes in addition to in terpreting the data from receptor binding studies. Acknowledgment We Co.,
thank Ltd.
for
Dr.
Youichi
his kind
Yabuuchi
gifts
of the
of OPC-21268
Otsuka and
Pharmaceutical
OPC-31260.
REFERENCES 1
Tolbert,
M.E.M.,
White,
A.C.,
Aspry,
K.,
Cutts,
J.
and
2
3 4
Fain, J.N.: Stimulation by vasopressin and a-catecholamines of phosphatidylinositol formation in isolated rat liver paren chymal cells. J. Biol. Chem. 225, 1938-1944 (1980) Berridge, M.J. and Irvine, R.F.: Inositol trisphosphate, a novel second messenger in cellular signal transduction. Nature 312, 315 321 (1984) Share, L.: Role of vasopressin in cardovascular regulation. Physiol. Rev. 64, 1248 -1284 (1988) Yamamura, Y., Ogawa, H., Chihara, T., Komdo, K., Ono
gawa, T., Nakamura, S. et al.: OPC-21268, an orally effec tive, nonpeptide vasopressin V1 receptor antagonist. Science 252, 572-574 (1991) 5 Yamamura, Y., Ogawa, H., Yamashita, H., Chihara, T., Miyamoto, H., Nakamura, S. et al.: Characterization of a novel aquaretic agent, OPC-31260, as an orally effective, nonpeptide vasopressin V2 receptor antagonist. Br. J. Phar macol. (in press) 6 Hongo, K. and Chiba, S.: A new method for measuring vascular responsiveness of relatively larger arteries of dogs. J. Pharmacol. Methods 9, 83 91 (1983) 7 Tsuji, T. and Chiba, S.: Potentiating effect of methysergide on norepinephrine-induced constriction of the isolated inter nal carotid artery of the dog. Japan. J. Pharmacol. 34, 95 100 (1984) 8 Kawai, K. and Chiba, S.: Vascular responses of isolated, per fused canine femoral arteries and veins to vasoactive sub stances. Japan. J. Pharmacol. 41, 537-540 (1986) 9 Liard, J.F., Deriaz, 0., Schelling, P. and Thibonnier, M.: Cardiac output distribution during vasopressin infusion or de hydration in conscious dogs. Am. J. Physiol. 243, H663 H669 (1982) 10 Liard, J.F. and Spadone, J.C.: Hemodynamic effects of an tagonists of the vasoconstrictor action of vasopressin in con scious dogs. J. Cardiovasc. Pharmacol. 6, 713 719 (1984) 11 Levi, R., Owen, D.A.A. and Trzeciakowski, J.: Actions of histamine on the heart and vasculature. In Pharmacology of Histamine Receptors, Edited by Ganellin, C.R. and Parsons, M.E., p. 236-297, Wright, PSG, Bristol (1982)