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Modification of certain vascular responses to dopamine by morphine
European Journal of Pharmacology 28 (1974) 108-113 © North-Holland Publishing Company
MODIFICATION
OF CERTAIN
VASCULAR
RESPONSES TO DOPAMINE
BY
MORPHINE* William E. DRESSLER, Gilbert D'ALONZO, G. Victor ROSSI and Raymond F. ORZECHOWSKI** Department of Pharmacology, Philadelphia College of Pharmacy and Science, Philadelphia, Pennsylvania, U.S.A.
Received 5 March 1974, accepted 30 May 1974 W.E. DRESSLER, G. D'ALONZO, G.V. ROSSI and R.F. ORZECHOWSKI, Modification of certain vascular responses to dopamine by morphine, European J. Pharmacol. 28 (1974) 108-113. Alteration by morphine of two specific vascular responses to dopamine, systemic hypotension in cats and renal vasodilation in dogs, were characterized in a-adrenergically blocked (phenoxybenzamine) anesthetized animals. Morphine sulfate (15-30 mg/kg, i.v.) converted the vasodepressor response to dopamine (50-100 #g/kg, i.v.) to a pressor response in intact cats, whereas in spinal cats dopamine-induced vasodepression was not significantly altered by morphine (15-60 mg/kg, i.v.). Haloperidol, a putative peripheral dopaminergic receptor antagonist, briefly but significantly attenuated the hypotensive response to dopamine in spinal cats. Canine renal vasodilator responses to intrarenal arterial dopamine (0.09-96 #g) were significantly reduced after morphine (2-6 mg/kg, i.a.). Morphine produced a non-parallel shift in the dopamine dose-response curve, whereas a parallel shift was obtained when haloperidol was used as the antagonist in the canine renal test system. These results do not support the concept that morphine may be a competitive antagonist of dopamine at peripheral dopaminergic receptors in these species. Dopamine receptors
Haloperidol
Morphine
1. Introduction Experimental evidence supports the existence, in certain mammalian vascular beds, of dopamine sensitive receptors which subserve vasodilation (see Goldberg, 1972). Dopamine-induced vasodilation in the renal, mesenteric and coronary circulation of the dog is resistant to pharmacologic antagonism by a- and /3-adrenergic blocking agents, atropine or antihistamines (McNay and Goldberg, 1966; Goldberg, 1966; Schuelke et al., 1971). Van Rossum (1966) reported that the butyrophenone neuroleptic haloperidol blocked systemic hypotensive responses to dopamine in yohimbine-pretreated cats. Subsequently, Yeh et * A preliminary report of this investigation was presented at the Fifteenth National Meeting of the Academy of Pharmaceutical Sciences, San Diego, California, 1973. ** Address reprint requests to: Raymond F. Orzechowski, Ph.D., Phila. College of Pharmacy and Science, 43rd Street, Kingsessing Mall, Philadelphia, Pa. 19104, U.S.A.
Renal blood flow, canine
al. (1969) found that haloperidol produced selective, though transient, attenuation of dopamine-induced renal and mesenteric vasodilation in the dog. Brotzu (1970) and Goldberg and Yeh (1971) reported that chlorpromazine also attenuated canine renal vascular responses to dopamine. In contrast, Dhasmana et al. (1969) were unable to alter dopamine-induced systemic vasodepression in cats with chlorpromazine, but observed that either morphine or codeine converted the dopamine depressor response to a pressor response. They also demonstrated that subsequent administration of the narcotic antagonist nalorphine restored the original depressor response to dopamine. The objectives of this investigation were to determine whether morphine could alter dopamine-induced renal vasodilation in dogs in a manner similar to that previously demonstrated with haloperidol, and to determine whether the reversal by morphine of systemic hypotensive responses to dopamine in cats might involve direct antagonism of peripheral vascular dopaminergic receptors.
I¢.E. Dressier et al., Vascular responses to dopamine
2. Materials and methods 2.1. Renal arterial blood flow in dogs
Mongrel dogs ( 7 - 1 3 kg) of either sex were anesthetized with sodium pentobarbital, 35 mg/kg, administered i.v. Arterial blood flow to the left kidney was measured with an electromagnetic flowmeter (Biotronex). The left kidney was exposed by a retroperitoneal incision and a calibrated flow probe of appropriate size was fitted to the renal artery. Intrarenal arterial drug administration was accomplished via an L-shaped 23-gauge needle inserted directly into the vessel proximal to the site of application of the flow probe. Patency of the cannula was maintained by continuous infusion of saline at a rate of approximately 0.2 ml/min. Zero flow was checked periodically throughout the experiment by momentarily occluding blood flow distal to the flow probe. Left renal blood flow and systemic blood pressure (femoral artery) were recorded continuously on a Grass polygraph. Phenoxybenzamine, 5 mg/kg, was infused slowly into the renal artery in order to block the c~-vasoconstrictor effects of dopamine. Dextran, at a dose not exceeding 20 ml/kg, was administered by slow intravenous drip (femoral vein) to maintain systemic blood pressure during the phenoxybenzamine infusion. Dose--response data for dopamine were obtained by introduction of four-fold serial dilutions of dopamine (0.09-96 ~g) into the renal artery in a volume .of 0.2 ml. In one series of experiments, a fixed dose of 1.4 × 10 -7 moles (53/ag) of haloperidol was combined with varying doses of dopamine in the same 0.2 ml solution, as described by Yeh et al. (1969). In another series, either 4.4 or 8.8 × 10-6 moles of morphine (1.25 or 2.5 mg) was administered into the renal artery 30 sec prior to the injection of each dose of dopamine. In a third group of experiments, morphine was administered as 3 intra-arterial doses of 2 mg/kg each, providing cumulative doses of 2, 4 and 6 mg/kg. The series of dopamine doses was repeated after each 2 mg/kg increment of morphine. 2.2. Systemic hypotension in cats
Fasted cats (1.5-4.0 kg) of either sex were anesthetized with sodium pentobarbital, 35 mg/kg, ad-
109
ministered i.v. The trachea was cannulated and those animals which subsequently received morphine were respired mechanically with room air by means of a Harvard respiratory pump. Systemic arterial blood pressure was measured directly from the right common carotid artery by means of a pressure transducer and recorded on a polygraph. The left femoral vein was cannulated for administration of drugs, and a bilateral cervical vagotomy was performed. In one group of cats spinalization was effected by an anterior surgical procedure described by Zarro and DiPalma (1964). Heparin sodium (280 units/kg, i.v.) was injected after completion of all surgical procedures, and the animals were allowed to stabilize for 30 rain prior to drug administration. In intact cats blood pressure responses to dopamine (50-100 #g/kg) were compared before and 30 min after infusion of 10 mg/kg of phenoxybenzamine. Cumulative doses of 15-30 mg/kg of morphine sulfate were then administered in increments of 5 - 1 5 mg/kg. Morphine doses were infused slowly during a 20 min period, and dopamine was administered 15 min after each dose increment when blood pressure stabilized. Subsequently, nalorphine ( 5 - 1 0 mg/kg) was infused and the dopamine challenge was repeated. In any individual experiment the dose of dopamine remained constant. The sequence of drug infusions was the same in spinalized cats except that nalorphine was not administered. Most of these animals received a cumulative dose of 60 mg/kg of morphine; the limit of morphine tolerance was estimated by the vital signs. In a group of spinalized cats, hypotensive responses to dopamine (50-100/ag/kg) were compared prior to and following intravenous infusion of 2 - 4 mg/kg of haloperidol. The drugs and their sources were as follows: dopamine hydrochloride (Calbiochem, Los Angeles, CA), 1-epinephrine bitartrate (K and K Laboratories, Plainview, NY), morphine sulfate (Merck and Co., Rahway, N J), nalorphine hydrochloride (Merck Sharp and Dohme, West Point, PA), phenoxybenzamine hydrochloride (Smith Kline and French Laboratories, Philadelphia, PA), haloperidol (McNeil Laboratories, Fort Washington, PA). Haloperidol was dissolved in 1 ml of propylene glycol with 3 N HC1; the volume of solution was increased with 0.9% NaC1 (saline) and the pH was ad-
110
W.E. Dressier et al., Vascular responses to dopamine MORPHINE 1.25 mg (4.4xlO-6 molesl
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Fig. 1. Dopamine-induced renal vasodilation in phenozybenzamine-pretreated dogs: attenuation by haloperidol. , - • dopamine alone; o o dopamine plus haloperidol. A fixed dose of 53 /ag (1.4 X 10.7 moles) of haloperidol was combined with varying doses of dopamine. Each point represents the mean of 6 values, except at 0.09 ;zg of dopamine where 3 values were obtained. Vertical lines indicate standard errors. Values for p were obtained by the paired t-test. justed to approximately 6.9 with 0.1 N NaOH. Phenoxybenzamine was prepared by dissolving the dose in 5 ml of warm propylene glycol and diluting to 25 ml with saline. All other drugs were dissolved in saline; 0.1% sodium bisulfite was added to solutions of dopamine and epinephrine. Doses of dopamine and epinephrine are expressed as the base; doses of other agents are expressed as the salt.
3. Results 3.1. Renal arterial blood f l o w in dogs Our studies confirmed the findings reported by Yeh et al. (1969) that haloperidol attenuated dopamine-induced renal vasodilation in dogs. Fig. 1 illustrates dose-related increases in left renal blood flow
I 1.5
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DOPAMINE ( p g )
Fig. 2. Effects of morphine on dopamine-induced renal vasodilation in phenoxybenzamine-pretreated dogs. • • dopamine alone; o. . . . . -o dopamine after morphine. Fixed doses of morphine sulfate were administered 30 sec prior to varying doses of dopamine. Each point represents the mean of 6 values. Vertical lines indicate representative standard errors. produced by intrarenal arterial injections of dopamine in phenoxybenzamine-pretreated dogs. When a fixed dose of 53/lg (1.4 X 10-7 moles) of haloperidol was administered simultaneously with dopamine, slight but consistent reductions in the vasodilator responses to dopamine were observed. Dose response curves, obtained by administering alternately dopamine alone followed by the dopamine-haloperidol combination, indicated that halopefidol significantly shifted the dose-response curve of dopamine to the right. Attenuation of dopamine vasodilation by haloperidol was transient ( 2 - 4 min), and repeated administration of haloperidol produced no apparent cumulative effects. In a series of 6 experiments, the effect of morphine on dopamine-induced renal vasodilation was determined by administration of fixed doses of either
W.E. Dressler et al., Vascularresponses to dopamine 60100-
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Fig. 3. Effects of morphine on dopamineqnduced renal vasodilation in phenoxybenzamine-pretreated dogs. • . • before morphine; e . . . . . -o after morphine 2 mg/kg; • . . . . . . .• after morphine 4 mg/kg (cumulative dose); ~___A after morphine 6 mg/kg (cumulative dose). Each point represents the mean of 9 values. Vertical lines indicate representative standard error. *p < 0.05, paired t-test. 1.25 or 2.5 mg of morphine (4.4 or 8.8 × 10 -6 moles) 30 sec before varying doses of dopamine. Fig. 2 shows that, in contrast to haloperidol, prior intrarenal arterial injections of morphine produced no detectable alteration of dopamine renal vasodilation. The doses of morphine employed did not affect baseline renal blood flow. Reversal of systemic vasodepressor responses to dopamine in cats required relatively high i.v. doses of morphine (i.e., 15 to 30 mg/kg); therefore, another group of experiments was performed in which d o s e response curves to dopamine were developed before and after each of 3 incremental doses of 2 mg/kg of morphine had been administered intra-arterially. Fig. 3 shows that renal vasodilator responses to dopamine were reduced significantly following the administration of 2, 4 or 6 mg/kg of morphine. However, in contrast to haloperidol, morphine did not produce a parallel shift in the dopamine d o s e response curve, since only the responses to the 3 highest doses of dopamine (i.e., 6, 24 and 96/~g) were decreased significantly.
Fig. 4. Effects of morphine and haioperidol on dopamine vasodepressor responses in a-adrenergically blocked cats. I~ Dopamine (DA) control, 50-100 t~g/kg; [] DA after phenoxybenzamine, 10 mg/kg; ~ DA after morphine, 15-30 mg/kg (intact) of 40-60 mg/kg (spinal); i DA after nalorphine, 5-10 mg/kg; (a) t~ DA 30 sec after haloperidol, 2-4 mg/kg; (b) [] DA 20 min after haloperidol, 2-4 mg/kg. Vertical lines indicate one standard deviation. 3.2. Systemic hypotension in cats
Resting blood pressure values (mean -+ standard deviation) after phenoxybenzamine in 6 intact and 8 spinalized cats that subsequently received morphine, and in 6 spinal cats that subsequently received haloperidol were: 95 -+ 18, 72 + 16 and 91 + 11 m m H g , respectively. These values did not differ significantly from each other ( p > 0 . 0 5 , Wilcoxon Rank Sum Test). Systemic blood pressure responses to dopamine in these 3 groups are summarized in fig. 4. Following c~-adrenergic blockade with phenoxybenzamine, the hypertensive response to dopamine was converted to a vasodepressor response in all animals. In intact cats, the subsequent administration of morphine ( 1 5 - 3 0 mg/kg) resulted in the reappearance of pressor responses to dopamine. However, the magnitude of these pressor responses was reduced in comparison to those obtained prior to phenoxybenzamine. In contrast, morphine failed to attenuate dopamine-induced vasodepression in a-adrenergically blocked, spinalized cats even after cumulative doses (i.e., 4 0 - 6 0 mg/kg) greater than were clearly effective in re-establishing pressor reponses to dopamine in intact cats. Nalorphine antagonized the morphine-induced pressure reversal to dopamine in intact cats, as was noted by the reappearance of hypotensive responses to dopamine. Haloperidol markedly, though very briefly, attenu-
112
I¢.E. Dressler et al., Vascular responses to dopamine
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Fig. 5. Selective reversal by morphine of dopamine vasodepressor responses in a-adrenergicallyblocked cats. DA, dopamine; E, epinephrine. Numbers indicate doses of amines in /~g. ated dopamine-induced vasodepression; in most instances, the duration of inhibition was only a few rain. The selectivity of the morphine-induced reversal of dopamine vasodepressor responses is illustrated in fig. 5. The influence of morphine on depressor responses to epinephrine and dopamine was compared in 3 intact a-adrenergically blocked cats. Morphine failed to attenuate depressor responses to epinephrine at doses which in the same animals reversed dopamine vasodepression.
4. Discussion
Morphine was found to inhibit two specific vascular responses to dopamine, i.e., renal vasodilation in dogs and systemic hypotension in cats. However, the characteristics of these morphine-induced alterations were not indicative of competitive antagonism at peripheral dopaminergic receptors. Initial attempts to attenuate dopamine-induced renal vasodilation by administering morphine in a manner similar to that previously shown to be effective for haloperidol (Yeh et al., 1969) were unsuccessful. When morphine was injected 30 sec prior to dopamine, and at molar concentrations approximately 3 0 - 6 0 times higher than was found to be sufficient for haloperidol, no alteration in the dopamine response was observed. Only after relatively high intra-arterial doses of morphine ( 2 - 6 mg/kg) were canine renal vasodilator responses to
dopamine significantly attenuated. Furthermore, the shift in the dopamine dose response curve produced by high doses of morphine was nonparallel and thus not typical of other substances, i.e., haloperidol (Yeh et al., 1969), chlorpromazine (Brotzu, 1970; Goldberg and Yeh, 1971) and bulbocapnine (Goldberg and Musgrave, 1971) which have been considered to act under similar conditions as competitive dopaminergic receptor antagonists. Systemic hypotension following dopamine injection in c~-adrenergically blocked cats has been attributed primarily to splanchnic vasodilation as a consequence of peripheral dopaminergic receptor activation (Hamilton, 1972). Our experiments in intact, anesthetized, a-adrenergically blocked cats confirmed previously reported data (Dhasmana et al., 1969) which indicated that vasodepressor responses to dopamine were converted to pressor responses by the administration of morphine. If the morphine reversal of dopamine-induced hypotension involved a direct effect on peripheral vascular dopaminergic receptors then similar results might have been expected in both intact and spinal cats. However, morphine failed to produce any significant alteration in the vasodepressor response to dopamine in spinalized animals, whereas haloperidol was capable of significantly attenuating the systemic hypotensive response. Reversal of depressor responses to dopamine was selective, inasmuch as epinephrine-induced vasodepression following a-blockade was not affected by morphine. Thus, it appears that the morphinedopamine interaction does not involve a peripheral vascular ~-adrenergic receptor mechanism. In addition to direct vascular receptor activation, dopamine-induced systemic hypotension may be at least partly due to a neurogenic mechanism involving inhibition of ganglionic transmission as described by Bogaert and De Schaepdryver (1967). In this regard, Willems et al. (1972) and Willems and Bogaert (1972) have shown that doses of dopamine which evoked vasodilation in isolated perfused hindlimbs of dogs also inhibited transmission in lumbar paravertebral ganglia. Since attenuation by morphine of the vasodepressor response to dopamine in our experiments was dependent upon an intact nervous system, it is possible that morphine may have prevented that hypotensive component of dopamine which was mediated by ganglionic blockade. Haloperidol has also been shown
W.E. Dressler et al., Vascular responses to dopamine
to antagonize selectively dopaminednduced inhibition of ganglionic transmission (Willems et al., 1972). Therefore, it would be difficult to attribute similar mechanisms to morphine and haloperidol since spinalization differentially antagonized morphine but not haloperidol. Since systemic blood pressure is a reflection of a multitude of simultaneously operant factors, the resuits of these experiments do not permit a conclusion regarding the nature of the apparent central inhibitory mechanism o f morphine. The interpretation of these data is further confounded by apparent species differences in susceptibility o f dopamine vasodilation to antagonism by morphine. Whereas our results confirmed those of Dhasmana et al. (1969) that morphine caused a reversal of dopamine-induced hypotension in intact cats, Furukawa et al. (1970) reported that 1 0 - 1 5 mg/kg of morphine 'hardly blocked' systemic vasodilator responses to dopamine in dogs. Our results with haloperidol are consistent with reports of other investigators that this neuroleptic agent may be a relatively weak and short-acting antagonist at peripheral dopaminergic receptor sites (Yeh et al., 1969; Schuelke et al., 1971; Shanbour and Parker, 1972; Hamilton, 1972).
References Bogaert, M.G. and A.F. De Schaepdryver, 1967, Dopamineinduced neurogenic vasodilation in the hindleg of the dog, Arch. Intern. Pharmacodyn. Therap. 166,203. Brotzu, G., 1970, Inhibition by chlorpromazine of the effects of dopamine on the dog kidney, J. Pharm. Pharmacol. 22, 664. Dhasmana, K.M., K.S. Dixit, K.N. Dhawan and G.P. Gupta, 1969, Blockade of depressor response of dopamine, Jap. J. Pharmacol. 19,168. Furukawa, T., K. Yamada and K. Kushiku, 1970, Reversal
113
of dopamine pressor action after ephedrine tachyphylaxis, Arch. Intern. Pharmacodyn. 186,298. Goldberg, L.I., 1966, Discussion, in: The Second Symposium on Catecholamines, Pharmacol. Rev. 18,539. Goldberg, L.I., 1972, Cardiovascular and renal actions of dopamine-potential clinical applications, Pharmacol. Rev. 24, 1. Goldberg, L.I. and G.E. Musgrave, 1971, Selective attenuation of dopamine-induced renal vasodilation by bulbocapnine and apomorphine, Pharmacologist 13,227. Goldberg, L.I. and B.K. Yeh, 1971, Attenuation of dopamine induced renal vasodilation in the dog by phenothiazines, European J. Pharmacol. 15, 36. Hamilton, T.C., 1972, Effects of dopamine on the conductance of perfused vascular beds of the chloralosed cat, Brit. J. Pharmacol. 44,442. McNay, J.L. and L.I. Goldberg, 1966, Comparison of the effects of dopamine, isoproterenol, norepinephrine and bradykinin on canine renal and femoral blood flow, J. Pharmacol. Exptl. Therap. 151, 23. Schuelke, D.M., A.L. Mark, P.G. Schmid and J.W. Eckstein, 1971, Coronary vasodilation produced by dopamine after adrenergic blockade, J. Pharmacol. Exptl. Therap. 176, 320. Shanbour, L.L. and D. Parker, 1972, Effects of dopamine and other catecholamines on the splanchnic circulation, Can. J. Physiol. Pharmacol. 50,594. Van Rossum, J.M., 1966, The significance of dopaminereceptor blockade for the mechanisms of action of neuroleptic drugs, Arch. Intern. Pharmacodyn. 160,492. Willems, J.L. and M.G. Bogaert, 1972, Neurogenic vasodilation: dopamine and related substances, Arch. Intern. Pharmacodyn. Therap. 197,412. Willems, J.L., A. Hoszowska-Owczarek and M.G. Bogaert, 1972, Dopamine and lumbar ganglionic transmission in the dog, Arch. Intern. Pharmacodyn. Therap. 196,315. Yeh, B.K., J.L. McNay and L.I. Goldberg, 1969, Attenuation of dopamine renal and mesenteric vasodilation by haloperidol: evidence for a specific dopamine receptor, J. Pharmacol. Exptl. Therap. 168,303. Zarro, V.J. and J.R. DiPalma, 1964, A preparation of the spinal cat by an anterior approach, J. Pharm. Pharmacol. 16,427.
MODIFICATION
OF CERTAIN
VASCULAR
RESPONSES TO DOPAMINE
BY
MORPHINE* William E. DRESSLER, Gilbert D'ALONZO, G. Victor ROSSI and Raymond F. ORZECHOWSKI** Department of Pharmacology, Philadelphia College of Pharmacy and Science, Philadelphia, Pennsylvania, U.S.A.
Received 5 March 1974, accepted 30 May 1974 W.E. DRESSLER, G. D'ALONZO, G.V. ROSSI and R.F. ORZECHOWSKI, Modification of certain vascular responses to dopamine by morphine, European J. Pharmacol. 28 (1974) 108-113. Alteration by morphine of two specific vascular responses to dopamine, systemic hypotension in cats and renal vasodilation in dogs, were characterized in a-adrenergically blocked (phenoxybenzamine) anesthetized animals. Morphine sulfate (15-30 mg/kg, i.v.) converted the vasodepressor response to dopamine (50-100 #g/kg, i.v.) to a pressor response in intact cats, whereas in spinal cats dopamine-induced vasodepression was not significantly altered by morphine (15-60 mg/kg, i.v.). Haloperidol, a putative peripheral dopaminergic receptor antagonist, briefly but significantly attenuated the hypotensive response to dopamine in spinal cats. Canine renal vasodilator responses to intrarenal arterial dopamine (0.09-96 #g) were significantly reduced after morphine (2-6 mg/kg, i.a.). Morphine produced a non-parallel shift in the dopamine dose-response curve, whereas a parallel shift was obtained when haloperidol was used as the antagonist in the canine renal test system. These results do not support the concept that morphine may be a competitive antagonist of dopamine at peripheral dopaminergic receptors in these species. Dopamine receptors
Haloperidol
Morphine
1. Introduction Experimental evidence supports the existence, in certain mammalian vascular beds, of dopamine sensitive receptors which subserve vasodilation (see Goldberg, 1972). Dopamine-induced vasodilation in the renal, mesenteric and coronary circulation of the dog is resistant to pharmacologic antagonism by a- and /3-adrenergic blocking agents, atropine or antihistamines (McNay and Goldberg, 1966; Goldberg, 1966; Schuelke et al., 1971). Van Rossum (1966) reported that the butyrophenone neuroleptic haloperidol blocked systemic hypotensive responses to dopamine in yohimbine-pretreated cats. Subsequently, Yeh et * A preliminary report of this investigation was presented at the Fifteenth National Meeting of the Academy of Pharmaceutical Sciences, San Diego, California, 1973. ** Address reprint requests to: Raymond F. Orzechowski, Ph.D., Phila. College of Pharmacy and Science, 43rd Street, Kingsessing Mall, Philadelphia, Pa. 19104, U.S.A.
Renal blood flow, canine
al. (1969) found that haloperidol produced selective, though transient, attenuation of dopamine-induced renal and mesenteric vasodilation in the dog. Brotzu (1970) and Goldberg and Yeh (1971) reported that chlorpromazine also attenuated canine renal vascular responses to dopamine. In contrast, Dhasmana et al. (1969) were unable to alter dopamine-induced systemic vasodepression in cats with chlorpromazine, but observed that either morphine or codeine converted the dopamine depressor response to a pressor response. They also demonstrated that subsequent administration of the narcotic antagonist nalorphine restored the original depressor response to dopamine. The objectives of this investigation were to determine whether morphine could alter dopamine-induced renal vasodilation in dogs in a manner similar to that previously demonstrated with haloperidol, and to determine whether the reversal by morphine of systemic hypotensive responses to dopamine in cats might involve direct antagonism of peripheral vascular dopaminergic receptors.
I¢.E. Dressier et al., Vascular responses to dopamine
2. Materials and methods 2.1. Renal arterial blood flow in dogs
Mongrel dogs ( 7 - 1 3 kg) of either sex were anesthetized with sodium pentobarbital, 35 mg/kg, administered i.v. Arterial blood flow to the left kidney was measured with an electromagnetic flowmeter (Biotronex). The left kidney was exposed by a retroperitoneal incision and a calibrated flow probe of appropriate size was fitted to the renal artery. Intrarenal arterial drug administration was accomplished via an L-shaped 23-gauge needle inserted directly into the vessel proximal to the site of application of the flow probe. Patency of the cannula was maintained by continuous infusion of saline at a rate of approximately 0.2 ml/min. Zero flow was checked periodically throughout the experiment by momentarily occluding blood flow distal to the flow probe. Left renal blood flow and systemic blood pressure (femoral artery) were recorded continuously on a Grass polygraph. Phenoxybenzamine, 5 mg/kg, was infused slowly into the renal artery in order to block the c~-vasoconstrictor effects of dopamine. Dextran, at a dose not exceeding 20 ml/kg, was administered by slow intravenous drip (femoral vein) to maintain systemic blood pressure during the phenoxybenzamine infusion. Dose--response data for dopamine were obtained by introduction of four-fold serial dilutions of dopamine (0.09-96 ~g) into the renal artery in a volume .of 0.2 ml. In one series of experiments, a fixed dose of 1.4 × 10 -7 moles (53/ag) of haloperidol was combined with varying doses of dopamine in the same 0.2 ml solution, as described by Yeh et al. (1969). In another series, either 4.4 or 8.8 × 10-6 moles of morphine (1.25 or 2.5 mg) was administered into the renal artery 30 sec prior to the injection of each dose of dopamine. In a third group of experiments, morphine was administered as 3 intra-arterial doses of 2 mg/kg each, providing cumulative doses of 2, 4 and 6 mg/kg. The series of dopamine doses was repeated after each 2 mg/kg increment of morphine. 2.2. Systemic hypotension in cats
Fasted cats (1.5-4.0 kg) of either sex were anesthetized with sodium pentobarbital, 35 mg/kg, ad-
109
ministered i.v. The trachea was cannulated and those animals which subsequently received morphine were respired mechanically with room air by means of a Harvard respiratory pump. Systemic arterial blood pressure was measured directly from the right common carotid artery by means of a pressure transducer and recorded on a polygraph. The left femoral vein was cannulated for administration of drugs, and a bilateral cervical vagotomy was performed. In one group of cats spinalization was effected by an anterior surgical procedure described by Zarro and DiPalma (1964). Heparin sodium (280 units/kg, i.v.) was injected after completion of all surgical procedures, and the animals were allowed to stabilize for 30 rain prior to drug administration. In intact cats blood pressure responses to dopamine (50-100 #g/kg) were compared before and 30 min after infusion of 10 mg/kg of phenoxybenzamine. Cumulative doses of 15-30 mg/kg of morphine sulfate were then administered in increments of 5 - 1 5 mg/kg. Morphine doses were infused slowly during a 20 min period, and dopamine was administered 15 min after each dose increment when blood pressure stabilized. Subsequently, nalorphine ( 5 - 1 0 mg/kg) was infused and the dopamine challenge was repeated. In any individual experiment the dose of dopamine remained constant. The sequence of drug infusions was the same in spinalized cats except that nalorphine was not administered. Most of these animals received a cumulative dose of 60 mg/kg of morphine; the limit of morphine tolerance was estimated by the vital signs. In a group of spinalized cats, hypotensive responses to dopamine (50-100/ag/kg) were compared prior to and following intravenous infusion of 2 - 4 mg/kg of haloperidol. The drugs and their sources were as follows: dopamine hydrochloride (Calbiochem, Los Angeles, CA), 1-epinephrine bitartrate (K and K Laboratories, Plainview, NY), morphine sulfate (Merck and Co., Rahway, N J), nalorphine hydrochloride (Merck Sharp and Dohme, West Point, PA), phenoxybenzamine hydrochloride (Smith Kline and French Laboratories, Philadelphia, PA), haloperidol (McNeil Laboratories, Fort Washington, PA). Haloperidol was dissolved in 1 ml of propylene glycol with 3 N HC1; the volume of solution was increased with 0.9% NaC1 (saline) and the pH was ad-
110
W.E. Dressier et al., Vascular responses to dopamine MORPHINE 1.25 mg (4.4xlO-6 molesl
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Fig. 1. Dopamine-induced renal vasodilation in phenozybenzamine-pretreated dogs: attenuation by haloperidol. , - • dopamine alone; o o dopamine plus haloperidol. A fixed dose of 53 /ag (1.4 X 10.7 moles) of haloperidol was combined with varying doses of dopamine. Each point represents the mean of 6 values, except at 0.09 ;zg of dopamine where 3 values were obtained. Vertical lines indicate standard errors. Values for p were obtained by the paired t-test. justed to approximately 6.9 with 0.1 N NaOH. Phenoxybenzamine was prepared by dissolving the dose in 5 ml of warm propylene glycol and diluting to 25 ml with saline. All other drugs were dissolved in saline; 0.1% sodium bisulfite was added to solutions of dopamine and epinephrine. Doses of dopamine and epinephrine are expressed as the base; doses of other agents are expressed as the salt.
3. Results 3.1. Renal arterial blood f l o w in dogs Our studies confirmed the findings reported by Yeh et al. (1969) that haloperidol attenuated dopamine-induced renal vasodilation in dogs. Fig. 1 illustrates dose-related increases in left renal blood flow
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Fig. 2. Effects of morphine on dopamine-induced renal vasodilation in phenoxybenzamine-pretreated dogs. • • dopamine alone; o. . . . . -o dopamine after morphine. Fixed doses of morphine sulfate were administered 30 sec prior to varying doses of dopamine. Each point represents the mean of 6 values. Vertical lines indicate representative standard errors. produced by intrarenal arterial injections of dopamine in phenoxybenzamine-pretreated dogs. When a fixed dose of 53/lg (1.4 X 10-7 moles) of haloperidol was administered simultaneously with dopamine, slight but consistent reductions in the vasodilator responses to dopamine were observed. Dose response curves, obtained by administering alternately dopamine alone followed by the dopamine-haloperidol combination, indicated that halopefidol significantly shifted the dose-response curve of dopamine to the right. Attenuation of dopamine vasodilation by haloperidol was transient ( 2 - 4 min), and repeated administration of haloperidol produced no apparent cumulative effects. In a series of 6 experiments, the effect of morphine on dopamine-induced renal vasodilation was determined by administration of fixed doses of either
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Fig. 3. Effects of morphine on dopamineqnduced renal vasodilation in phenoxybenzamine-pretreated dogs. • . • before morphine; e . . . . . -o after morphine 2 mg/kg; • . . . . . . .• after morphine 4 mg/kg (cumulative dose); ~___A after morphine 6 mg/kg (cumulative dose). Each point represents the mean of 9 values. Vertical lines indicate representative standard error. *p < 0.05, paired t-test. 1.25 or 2.5 mg of morphine (4.4 or 8.8 × 10 -6 moles) 30 sec before varying doses of dopamine. Fig. 2 shows that, in contrast to haloperidol, prior intrarenal arterial injections of morphine produced no detectable alteration of dopamine renal vasodilation. The doses of morphine employed did not affect baseline renal blood flow. Reversal of systemic vasodepressor responses to dopamine in cats required relatively high i.v. doses of morphine (i.e., 15 to 30 mg/kg); therefore, another group of experiments was performed in which d o s e response curves to dopamine were developed before and after each of 3 incremental doses of 2 mg/kg of morphine had been administered intra-arterially. Fig. 3 shows that renal vasodilator responses to dopamine were reduced significantly following the administration of 2, 4 or 6 mg/kg of morphine. However, in contrast to haloperidol, morphine did not produce a parallel shift in the dopamine d o s e response curve, since only the responses to the 3 highest doses of dopamine (i.e., 6, 24 and 96/~g) were decreased significantly.
Fig. 4. Effects of morphine and haioperidol on dopamine vasodepressor responses in a-adrenergically blocked cats. I~ Dopamine (DA) control, 50-100 t~g/kg; [] DA after phenoxybenzamine, 10 mg/kg; ~ DA after morphine, 15-30 mg/kg (intact) of 40-60 mg/kg (spinal); i DA after nalorphine, 5-10 mg/kg; (a) t~ DA 30 sec after haloperidol, 2-4 mg/kg; (b) [] DA 20 min after haloperidol, 2-4 mg/kg. Vertical lines indicate one standard deviation. 3.2. Systemic hypotension in cats
Resting blood pressure values (mean -+ standard deviation) after phenoxybenzamine in 6 intact and 8 spinalized cats that subsequently received morphine, and in 6 spinal cats that subsequently received haloperidol were: 95 -+ 18, 72 + 16 and 91 + 11 m m H g , respectively. These values did not differ significantly from each other ( p > 0 . 0 5 , Wilcoxon Rank Sum Test). Systemic blood pressure responses to dopamine in these 3 groups are summarized in fig. 4. Following c~-adrenergic blockade with phenoxybenzamine, the hypertensive response to dopamine was converted to a vasodepressor response in all animals. In intact cats, the subsequent administration of morphine ( 1 5 - 3 0 mg/kg) resulted in the reappearance of pressor responses to dopamine. However, the magnitude of these pressor responses was reduced in comparison to those obtained prior to phenoxybenzamine. In contrast, morphine failed to attenuate dopamine-induced vasodepression in a-adrenergically blocked, spinalized cats even after cumulative doses (i.e., 4 0 - 6 0 mg/kg) greater than were clearly effective in re-establishing pressor reponses to dopamine in intact cats. Nalorphine antagonized the morphine-induced pressure reversal to dopamine in intact cats, as was noted by the reappearance of hypotensive responses to dopamine. Haloperidol markedly, though very briefly, attenu-
112
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Fig. 5. Selective reversal by morphine of dopamine vasodepressor responses in a-adrenergicallyblocked cats. DA, dopamine; E, epinephrine. Numbers indicate doses of amines in /~g. ated dopamine-induced vasodepression; in most instances, the duration of inhibition was only a few rain. The selectivity of the morphine-induced reversal of dopamine vasodepressor responses is illustrated in fig. 5. The influence of morphine on depressor responses to epinephrine and dopamine was compared in 3 intact a-adrenergically blocked cats. Morphine failed to attenuate depressor responses to epinephrine at doses which in the same animals reversed dopamine vasodepression.
4. Discussion
Morphine was found to inhibit two specific vascular responses to dopamine, i.e., renal vasodilation in dogs and systemic hypotension in cats. However, the characteristics of these morphine-induced alterations were not indicative of competitive antagonism at peripheral dopaminergic receptors. Initial attempts to attenuate dopamine-induced renal vasodilation by administering morphine in a manner similar to that previously shown to be effective for haloperidol (Yeh et al., 1969) were unsuccessful. When morphine was injected 30 sec prior to dopamine, and at molar concentrations approximately 3 0 - 6 0 times higher than was found to be sufficient for haloperidol, no alteration in the dopamine response was observed. Only after relatively high intra-arterial doses of morphine ( 2 - 6 mg/kg) were canine renal vasodilator responses to
dopamine significantly attenuated. Furthermore, the shift in the dopamine dose response curve produced by high doses of morphine was nonparallel and thus not typical of other substances, i.e., haloperidol (Yeh et al., 1969), chlorpromazine (Brotzu, 1970; Goldberg and Yeh, 1971) and bulbocapnine (Goldberg and Musgrave, 1971) which have been considered to act under similar conditions as competitive dopaminergic receptor antagonists. Systemic hypotension following dopamine injection in c~-adrenergically blocked cats has been attributed primarily to splanchnic vasodilation as a consequence of peripheral dopaminergic receptor activation (Hamilton, 1972). Our experiments in intact, anesthetized, a-adrenergically blocked cats confirmed previously reported data (Dhasmana et al., 1969) which indicated that vasodepressor responses to dopamine were converted to pressor responses by the administration of morphine. If the morphine reversal of dopamine-induced hypotension involved a direct effect on peripheral vascular dopaminergic receptors then similar results might have been expected in both intact and spinal cats. However, morphine failed to produce any significant alteration in the vasodepressor response to dopamine in spinalized animals, whereas haloperidol was capable of significantly attenuating the systemic hypotensive response. Reversal of depressor responses to dopamine was selective, inasmuch as epinephrine-induced vasodepression following a-blockade was not affected by morphine. Thus, it appears that the morphinedopamine interaction does not involve a peripheral vascular ~-adrenergic receptor mechanism. In addition to direct vascular receptor activation, dopamine-induced systemic hypotension may be at least partly due to a neurogenic mechanism involving inhibition of ganglionic transmission as described by Bogaert and De Schaepdryver (1967). In this regard, Willems et al. (1972) and Willems and Bogaert (1972) have shown that doses of dopamine which evoked vasodilation in isolated perfused hindlimbs of dogs also inhibited transmission in lumbar paravertebral ganglia. Since attenuation by morphine of the vasodepressor response to dopamine in our experiments was dependent upon an intact nervous system, it is possible that morphine may have prevented that hypotensive component of dopamine which was mediated by ganglionic blockade. Haloperidol has also been shown
W.E. Dressler et al., Vascular responses to dopamine
to antagonize selectively dopaminednduced inhibition of ganglionic transmission (Willems et al., 1972). Therefore, it would be difficult to attribute similar mechanisms to morphine and haloperidol since spinalization differentially antagonized morphine but not haloperidol. Since systemic blood pressure is a reflection of a multitude of simultaneously operant factors, the resuits of these experiments do not permit a conclusion regarding the nature of the apparent central inhibitory mechanism o f morphine. The interpretation of these data is further confounded by apparent species differences in susceptibility o f dopamine vasodilation to antagonism by morphine. Whereas our results confirmed those of Dhasmana et al. (1969) that morphine caused a reversal of dopamine-induced hypotension in intact cats, Furukawa et al. (1970) reported that 1 0 - 1 5 mg/kg of morphine 'hardly blocked' systemic vasodilator responses to dopamine in dogs. Our results with haloperidol are consistent with reports of other investigators that this neuroleptic agent may be a relatively weak and short-acting antagonist at peripheral dopaminergic receptor sites (Yeh et al., 1969; Schuelke et al., 1971; Shanbour and Parker, 1972; Hamilton, 1972).
References Bogaert, M.G. and A.F. De Schaepdryver, 1967, Dopamineinduced neurogenic vasodilation in the hindleg of the dog, Arch. Intern. Pharmacodyn. Therap. 166,203. Brotzu, G., 1970, Inhibition by chlorpromazine of the effects of dopamine on the dog kidney, J. Pharm. Pharmacol. 22, 664. Dhasmana, K.M., K.S. Dixit, K.N. Dhawan and G.P. Gupta, 1969, Blockade of depressor response of dopamine, Jap. J. Pharmacol. 19,168. Furukawa, T., K. Yamada and K. Kushiku, 1970, Reversal
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of dopamine pressor action after ephedrine tachyphylaxis, Arch. Intern. Pharmacodyn. 186,298. Goldberg, L.I., 1966, Discussion, in: The Second Symposium on Catecholamines, Pharmacol. Rev. 18,539. Goldberg, L.I., 1972, Cardiovascular and renal actions of dopamine-potential clinical applications, Pharmacol. Rev. 24, 1. Goldberg, L.I. and G.E. Musgrave, 1971, Selective attenuation of dopamine-induced renal vasodilation by bulbocapnine and apomorphine, Pharmacologist 13,227. Goldberg, L.I. and B.K. Yeh, 1971, Attenuation of dopamine induced renal vasodilation in the dog by phenothiazines, European J. Pharmacol. 15, 36. Hamilton, T.C., 1972, Effects of dopamine on the conductance of perfused vascular beds of the chloralosed cat, Brit. J. Pharmacol. 44,442. McNay, J.L. and L.I. Goldberg, 1966, Comparison of the effects of dopamine, isoproterenol, norepinephrine and bradykinin on canine renal and femoral blood flow, J. Pharmacol. Exptl. Therap. 151, 23. Schuelke, D.M., A.L. Mark, P.G. Schmid and J.W. Eckstein, 1971, Coronary vasodilation produced by dopamine after adrenergic blockade, J. Pharmacol. Exptl. Therap. 176, 320. Shanbour, L.L. and D. Parker, 1972, Effects of dopamine and other catecholamines on the splanchnic circulation, Can. J. Physiol. Pharmacol. 50,594. Van Rossum, J.M., 1966, The significance of dopaminereceptor blockade for the mechanisms of action of neuroleptic drugs, Arch. Intern. Pharmacodyn. 160,492. Willems, J.L. and M.G. Bogaert, 1972, Neurogenic vasodilation: dopamine and related substances, Arch. Intern. Pharmacodyn. Therap. 197,412. Willems, J.L., A. Hoszowska-Owczarek and M.G. Bogaert, 1972, Dopamine and lumbar ganglionic transmission in the dog, Arch. Intern. Pharmacodyn. Therap. 196,315. Yeh, B.K., J.L. McNay and L.I. Goldberg, 1969, Attenuation of dopamine renal and mesenteric vasodilation by haloperidol: evidence for a specific dopamine receptor, J. Pharmacol. Exptl. Therap. 168,303. Zarro, V.J. and J.R. DiPalma, 1964, A preparation of the spinal cat by an anterior approach, J. Pharm. Pharmacol. 16,427.