Role of β-1 and β-2 adrenoceptors in isoproterenol-induced renin release in conscious dogs

Life Sciences, Vol . 24, pp . 1953-1958 Printed in the U .S .A .

Pergamon Press

ROLE OF ß-1 AND ß-2 ADRENOCEPTORS IN ISOPROTERENOL-INDUCED RENIN RELEASE IN CONSCIOUS DOGS Norio Himori, Suguru Hayakawa* and Tsutomu Ishimori Pharmacological Research Department, and Clinical Laboratories*', Teikoku Hormone Mfg ., Co . Ltd ., Shimosakunobe, gawasaki 213, Japan (Received in final form April 9, 1979)

Summary The present experiments were designed to classify the ß-adrenoceptors pertaining to the renin release induced by isoproterenol in conscious dog. Atenolol (ß-1 adrenoceptor antagonist), in oral dose of 6 mg/kg, produced a significant inhibition of renin release caused by isoproterenol . This dose of atenolol suppressed effectively the tachycardia of isoproterenol . On the other hand, the renin release produced by isoproterenol was not modified significantly by a ß-2 adrenoceptor antagonist, IPS-339, at a oral dose of 3 mg/kg which fully antagonized hypotensive response to isoproterenol . These results strongly suggest that the renin release induced by isoproterenol is largely due to stimulation of ß-1 type adrenoceptors . Lands et al . (1), Furchgott (2) and Arnold and Selberis (3) have propose~c £hat adrenergic receptors mediating cardiac acceleration, lipolyais and inhibition of small intestinal motility (ß-1 adrenoceptors) differ significantly from those in leading to bronchodilation, vasodilation and muscle glycogenolyais (ß-2 adrenoceptors) . There is substantial evidence that circulating catecholamines and sympathetic stimuli increase renin secretion through a ß-adrenergic mechanism within the kidney (4,5,6,7) . However, characteristics of the ß-adrenoceptors leading to renin release have not been fully investigated and there seems to be a discrepancy in regard to a role of ß-adrenoceptors in causing renin release (8,9,10,11,12) . These have stimulated need for classification of the ß-adrenoceptors mediating renin release .

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Thua, the present study with conscious do a was designed to elucidate whether the ß-adrenoceptor agoni isoproterenol, increases plasma renin activity by activation of ß-1 or ß-2 adrenoceptors . Materials and Methods Experiments were carried out pn 8 conscious, mongrel normotenaive dogs of either sex, weighing between 10 - 17 kg . 0024-3205/79/211953-06$02 .00/0 Copyright (c) 1979 Pergamon Press Ltd

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Çonaçious Qreparation An arterial catheter which was previously implanted into the abdominal aorta enabled us to perform conveniently the measurement of arterial blood pressure and heart rate, and blood sampling (13,14) . The catheter(INTRAMEDIC ® , PE-60) was introduced into the abdominal aorta via a branch of the femoral artery under pentobarbital anesthesia (27 mg/kg i .v .) . The other end of the catheter was exteriorized through the skin at the back of the neck . The experiments were started at the 5th day after implantation of the catheter . The arterial blood pressure was recorded directly by a pressure transducer (Nikon Kohden, MPU0 .5) and heart rate was counted from the blood pressure pulse waves . Intravenous infusion of isoproterenol was performed through a catheter acutely inserted into the cephalic vein of either foreleg using an infusion pump (Natsume, KN-202) . Blood for estimation of plasma renin activity was collected in chilled tubes containing EDTA-2Na (1 ml blood/1 mg EDTA-2Na), separated immediately, and the plasma was deep frozen untill all samples During the experiments, the unrestrained animal were analysed . sat quietly on the floor of a sound-proof room near the experimenter . Plasma renin activit Plasma ren n ac ivity was quantified by use of the double antibody method which was first introduced for the radioimmunoFrozen plasma assay of insulin by Morgan and Zazarow (15) . divided into two samples . samples were thawed on ice and then ° One was incubated for 3 hr at 37 C and the other for 3 hr at 4°C . In the presence of enzyme inhibitors (4 .5 mM, 8-hydroxyquinoline and 0 .12 mM, dimercaprol), these two samples were individually incubated for 24 hr at 4°C with I -angiotensin I and antiserum which was produced in rabbits with angiotensin I-BSA, and further, after addition of goat antirabbit IgG serum the samples were Finally the formed angiotensin I incubated for 4 days at 4°C . level in plasma was measured on the basis of the value obtained by counting the radioactivity of precipitates and the standard curve constructed by use of standard angiotensin I (Protein Thus, plasma renin activity Research Foundation, Minoh, Japan) . calculated as the difference of was (ng Angiotensin I/ml/hr) a heated plasma sample (37 ° C - 3 hr) formed angiotensin I between ° and an unheated sample (4 C - 3 hr) . Dru~a

Atenolol (ICI), IPS-339 [(t-butylamino-3-ol-2-propyl) oximino-9 fluorene hydrochloride], dl-isoproterenol hydrochloride (Nikken Kagaku) and propranolol hydrochloride (dextro and racemic Isoproterenol was dissolved types, ICI) were used in this study. and in 0 .9 96 saline solution containing ascorbic acid (0 .2 96) Atenolol, IPS-339 and two isomers of prointravenously infused . pranolol were administered orally in hard gelatin capsules . Doses of all drugs refer to the salts, Unless otherwise noted, propranolol was the racemic type . Statistical evaluation slues are given in terms of mean + s .e, with n being the number of experiments . Statistical significance of the differences between two mean values was evaluated by Student's paired t-test and expressed by the p value .

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ß-Adrenoceptors and Renin Release

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Results and Discussion The average arterial renin activity (PRA) of 8 lic)/87 + 3 .7 (diastolic) 0.40 ng Âng I/ml/hr (mean

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blood pressure, heart rate and plasma conscious dogs were 127 + 4.1 (systomm Hg, 93 + 5 .8 beats/min and 1 .74 ± + s .e .), réapectively .

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Influences oî propranolol and d-propranolol on changes in arterial blood pressure, heârt rate and plasma renin activity produced by isoproterenol in conscious dogs . Propranolol (Prop, 3 mg/kg, n=5) and d-propranolol (ciProp, 3 mg/kg, n=4) were administered orally at 120 . min before the start of i .v . infusion of isoproterenol (Iso, 0 .5 }x /kg/min for 20 min) . In the lowest panel the mark ~as) indicates the period of intravenous infusion of isoproterenol (20 min) . Each open bar represents the change in control e$perimenta (isoproterenol alone) and solid bars represent the change .produced by isoproterenol after p. o . administration of propranolol or d-propranolol . Statistical analysis of the change in blood pressure and heart rate was evaluated from the differences between the values just before the start of isoproterenol infusion (0 min) and those after isoproterenol infusion (5,10 and 20 min) and after cessation of infusion (40 min), While the . change in PRA was evaluated statistically from the differences between the two mean values at the corresponding time of .the egperiments of isoproterenol alone and that after p . o . administration of antagonists . * p~0 .05, ~-*~ p<0 .01 .

Vol, 24, No . 21, 1979

a-Adrenoceptors and Renin Release

1956

In the control experiments in 6 animals receiving intravenous infusion of saline (6 .7 ml/20 min) 2 hr after p . o . administration of lactose (200 mg/head), there were no change in arterial blood pressure, heart rate and PRA . On the other hand, intravenous infusion of iaoproterenol (0 .5 yig/kg/min for 20 min, 6 .7 ml in volume) caused a significant decrease in diastolic blood pressure and an increase in heart rate and PRA (Figs . 1 and 2) . Maximum change in blood pressure was observed at 5 min and the decreased diastolic blood pressure gradually returned to the initial value during the infusion period . While the maximum increase in PRA was observed at 20 min after the start of infusion of iaoproterenol (approx, 10 fold elevations) (Figs, 1 and 2), vf . .,w) uo -

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FIG, 2 Influences of atenolol and IPS-339 on changea in arterial blood pressure, heart rate and plasma renin activity produced by iaoproterenol in conscious doge, Atenolol (Ate, 6 mg/kg, n=5) and IPS-339 (3 mg/kg, n=4) were administered orally at 180 and 120 min, respectively, before the start of i,v, infusion of iaoproterenol (Iso, 0 .5 yg /kg/min for 20 min) . Other details as in Fig, 1, These changes in arterial blood pressure, heart rate and PRA were effectively inhibited by the administration of 3 mg/kg p . o . of propranolol . On the contrary, d-propranolol did not modify significantly the response of arterial blood reasure, In anesheart rate and PRA to-iaoproterenol infusion (Fig . 1~ . thetized rats, ß-adrenoceptor blocking activity of d-propranolol

Vol . 24, No . 21, 1979

a-Adrenoceptors and Renin Release

1957

used in this study was approximately 60 - 70 times leas potent than that of propranolol . These observations strongly suggest that the ß-adrenoceptor blocking action ~er se is a prerequisite for the inhibition of isoproterenol-induce3 renin release . Atenolol is a cardioaelective ß-adrenoceptor blocking drug which is devoid of a significant membrane stabilizing activity. In animal study this compound is approximately equipotent to ro ranolol in inhibiting isoproterenol-induced tachycardia (ß -1) p(16~ . More recently, a new ß-adrenoceptor blocking drug, IPS-339 has become available that, in animal study, has been shown to have a predominant blocking action on the peripheral vaeculature and tracheal smooth muscle (ß -2) (17) . Also in our experiments with conscious dogs, IPS-339 showed a highly selective inhibition for the vasodilator ß-2 adrenoceptors in the peripheral vascular bed ; being approximately equipotent ß-blocking action with propranolol on the peripheral vasculatures, but 8 - 9 times leas potent on the heart (submitted for publication) . Thus in this study, to determine with accuracy the role of ß-1 and ß-2 adrenoceptors underlying the renin releasing action of isoproterenol, approximately equipotent oral doses of atenolol and IPS-339 in comparison with propranolol were appropriately chosen from our previous experimental data concerned with the antagonism of tachycardia (ß -1) and hypotension (ß-2) produced by isoproterenol, respectively. As clearly shown in left panel in Fig . 2, atenolol (6 mg/kg p .o .), a selective ß-1 adrenoceptor antagonist, did not antagonize the hypotensive'Y~esponae to isoproterenol, but did inhibit significantly the isoproterenol-induced tachycardia and renin release . IPS-339 (3 mg/kg p .o.), a selective ß-2 adrenoceptor antagonist, changed the hypotensive response to isoproterenol in the presaor one (right panel in Fig . 2) . This reversal phenomenon may derive from the effective inhibition on peripheral vasodilation (ß-2 effect) caused by isoproterenol and the impotence in inhibiting tachycardia (ß-1 effect) caused by isoproterenol . Under these circumstances, at 10 and 20 min during the infusion period of isoproterenol the renin secretion was not significantly modified by the treatment of IPS-339 . The presaor response to isoproterenol emerged on the ground of effective ß-2 adrenoceptor blockade of IPS-339 might be responsible for statistically significant renin suppression observed at 5 min . Thus we assumed that IPS-339 did not essentially inhibit renin release exerted by isoproterenol . Some authors have proposed that ß-2 adrenoceptors carry out a major role in controlling renin release 8,9) . However, several investigators, e .g . Daviea et al . ~10) who compared the action of salbutamol on plasma renin a~ivity with that of isoproterenol, suggested that ß-1 adrenoceptors may be essential same for renin release . Others also have been conducted to the are Thus, it is obvious tha`~ our rea~lts hypothesis (11,12) at variance .with those reported by Johns and Singer (8) and Weber et al . (9) . At least a part of this inconsistency was_conaidered ~ô ûe due to the following reasons : First, experimental environment, viz ., conscious or anesthetized conditions and in vitro or in v~o experiments, may be subtly influenced the resu-t~ Second, methodological differences for measuring plasma renin activity may exert unexpected influence upon the values . Third,

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a-Adrenoceptors and Renin Release

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species differences may also reside in leading to these conflicting observations, And lastly, a certain action oî anesthetic drugs, less likely, may modify the effect of these ß=adrenoceptor blocking drugs on isoproterenol-induced renin Taken together with our results and other observarelease . tions, the role of ß-2 adrenoceptors in causing renin release Especially in our thus was regarded of little importance . experiments IPS-339 was essentially without effects on renin release, but propranolol and atenolol considerably inhibited . In conclusion, based on these experimental results in conscious dogs we must conclude that ß-adrenoceptors pertaining to renin release produced by isoproterenol were mainly ß-1 type, Acknowledgement We are grateful to Prof . C .G . Wermuth (Laboratoire de Chimie Organique et Therapeutique, Faculte de Pharmacie, France) for his generous supply of IPS=339 . References 1. 2. 3. 4. 5. 6. 7. 8. 9. 10 . 11 . 12 . 13 . 14 . 15 . 16 . 17 .

A,M . LANDS F .P . LUDUENA and H . BUZZO, Life Sci . _6 22412249 (1967j . R .F, FURCHGOTT, Ann. N .Y, Acad . Sci . 139 553-570 (1967) . A . ARNOLD and W .H, SELBERIS, Experience 24 1010-1011 (1967) . J .A, JOHNSON, J .O, RAVIS and R,T . WITTY, Cir . Res . 2~ 646-653 (1971) . R . VANDONGEN, W .S . PEART and G .W . BOYD, Cir . Res . 32 290296 (1973) . W .F, GANONG, Fed . Proc, 32 1782-1784 (1973) . TER, Lancet 2 594-596 (1976) . R . RAVIES and J .D .H . SLAE .J . JOHNS and B . SINGER, Br, J . Pharmac . 52 315-318 (1974) . M,A, WEBER, H,F . OATES and G .S . STOKES, Clin, Sci . Mol. Med . ,~8 89s-91s (1975) . R. RAVIES, D.M . GEDDES, J .D .H . SLAT ER, R .C, WIGGINS and N .N . PAYNE, Clin . Sci . Mol . Med. ~4 14s (1975) . E. DESAULES F . MIESCH and J, SCHWARTZ, Br . J . Pharmac . 63 421-425 1978) . H .F . OATES, L .M . STOKER, J .C . MONAGHAN and G .S . STOKES, Arch, int . Pharmacodyn . 234 205-213 (1978) . N . HIMORI, A. IZUMI and Y,HIRAMATSU, Arch, int . Pharmacodyn. 220 4-18 (1976) . N . HIMORI and . IZUMI, Pharmacometrics 14 593-598 (1978) . C .R . MORGAN and A . LAZAROW, Proc . Soc, exp, Biol . Med. 110 29-32 (1962) . -Ti~ . BARRETT, J . CARTER, J .D . FITZGII2AZD, R . HULL and A Le D . COUNT, Br . J . Pharmac . 48 340p (1973) . J .L . IMBS, F . MIESCH, J . SCHWAI~TZ, J . VELLY, G . LECLERC, A . MANN and C .G, WERMUTH, Br . J . Pharmac . 60 357-362 (1977) .