Gen. Pharmac. Vol. 23, No. 5, pp. 843-845, 1992 0306-3623/92 $5.00+0.00 Printed in Great Britain. All rights reserved Copyright © 1992 Persamon Press Ltd

ALPHAI-ADRENOCEPTOR BLOCKING ACTIVITIES OF BEVANTOLOL HYDROCHLORIDE(NC-1400) AND

LABETALOL IN RAT ISOLATED THORACIC AORTA--DO THEY DISTINGUISH BETWEEN SUBTYPES?

KoJt SHIRAISHI, MAMI MORIYA, NORIHISA MIYAKE* and ISSEI TAKAYANAGI Department of Chemical Pharmacology, Toho University School of Pharmaceutical Sciences,

Miyama 2-2-1, Funabashi, Chiba 274, Japan [Tel. (0474) 72-1141]

(Received 30 January 1992)

Abstract--l. Alpha~-adrenoceptor blocking activity of bevantolol(NC-1400) was tested in the rat thoracic aorta, comparing with that of labetalol.

2. In alphal-adrenoceptor blocking activity, bevantolol(NC-1400) was 1/20th as potent as labetalol. 3. The pA 2 values of bevantolol(NC-1400) and labetaiol estimated in the rat thoracic aorta were

compared with those in the rabbit thoracic aorta, which were reported by Takayauagl et al. [Takayanagi I., Kizawa Y., Iwasaki S. and Nakagnshi A. (1987) Gen. Pharmac. 18, 87-89].

4. The pA2 values of bevantolol(NC-1400) and labetalol were approximately 1 order of magnitude higher in rat aorta than in rabbit aorta, suggesting that both the drugs distinguish between subtypes of alphal-adrenoceptors.

INTRODUCTION

Many beta-adrenoceptor blockers are developed and clinically used as hypertensive drugs. As non-selective beta-adrenoceptor blockers have been reported to elicit some side-effects which are mediated through beta2-adrenoceptors, beta~-adrenoceptor blockers with alpham-adrenoceptor blocking activity, such as labetalol may be promising. Bevantolol hydrochlor- ide(NC-1400) was reported to be a betal-adreno- ceptor selective blocker with alphat-adrenoceptor blocking activity. Its pA2 values (negative logarithm of dissociation constants) for betat- and alphal- adrenoceptor blocking activities were, respectively, 7.75 and 4.77, while both activities of labetalol were 7.48 and 6.22, respectively (Takayanagi et al., 1987).

There are two distinct subtypes of alphat-adreno- ceptors, which are distinguished by their affinities for prazosin and yohimbine in the blood vessels; these are termed alpham and alpha~L by Flavahan and Van- houtte (1986), because of their high and low affinities. These observations were in agreement with the results of Takayanagi et al. (1985) who demonstrated two subtypes of alpha~-adrenoceptors in arteries based on their affinities for prazosin and its derivatives, and also mentioned that rat aorta contains alpham, while rabbit aorta contains alpha~L.

Recently, radioligand binding studies with [3H]prazosin indicated the existence of two separate populations of alphaj-adrenoceptors which were termed alphas^ and alpham, respectively (Morrow and Creese, 1986). A similar subclassification was

*To whom all correspondence should be addressed at: Planning and Investigation Department, Nippon Chemiphar Co. Ltd, 2-3, 2-chome, Iwamoto-cho, Chiy- oda-ku, Tokyo 101, Japan [Fax 03-3865-1378].

proposed by Han et ai. (1987); the alphal^ has a high affinity for 5-methylurapidil and WIMI01, while the alpham subtype has a low affinity for these antagon- ists and is selectively susceptible to chioroethyl- clonidine. It was reported that the rabbit thoracic aorta contains alpha1^ and alpham subtypes (Suzuki et al., 1990; Takayanagi et ai., 1992), while the rat thoracic aorta contains alpha~^, primarily (Han et al., 1990).

In this paper, the pA 2 values of bevantolol(NC- 1400) and labetalol in alphal-adrenoceptor blocking action were estimated in the rat isolated thoracic aorta. And we compared the pA2 values estimated herein with those in the rabbit aorta (Takayanagi et al., 1987) to learn whether or not bevantolol(NC- 1400) and labetalol distinguish subtypes of aipha~- adrenoceptors.

MATERIALS AND METHODS

Male Wistar rats (200-250 g) were stunned by a blow on the head and killed by bleeding from the neck. Segments of thoracic aorta were removed and immediately placed in Krebs solution of the following composition (raM): NaCl ll8, KC1 4.7, CaCl 2 2.54, MgSO4 1.17, KI-12PO4 1.19, NaHCO3 25.0 and glucose 11.0. Thoracic aorta were cut into helical strips about l0 mm in length and 2 mm in width.

The strips were suspended in a 20 ml organ bath filled with Krebs solution gassed with a mixture of 95% 02 and 5% CO2 and maintained at 37°C. The response to norepi- nephrine was isometrically recorded under a resting tension of I g for all strips. The strips were allowed to equilibrate for 90 rain, they were then contracted by norepinephrine (10-SM) and allowed to equilibrate for 30min after washout. This was repeated until two successive contrac- tions of approximately equal size had been obtained.

The competitive antagonistic activities were express~ as pA, values (negative logarithms of the dissociation constant). After determination of control concentration-

843

844 KoJ] SHIRAISH1 et al.

o

-1 l] -9 -8 -7 -6 -5 -4

Log[NEI

Fig. 1. Concentration-response curves of norepinephrine in the absence and presence of various concentrations of bevantolol(NC-1400). Ordinate: contraction (%). Abscissa: logarithm of concentration (M) of norepinephrine. Each value is presented as a mean + SE (bar) of 4 strips. (O) Norepinephrine alone, (O) with bevantolol(NC-1400) 3 x 10 -6 M, (A) with bevantolol(NC-1400) 10 -5 M and (A)

with bevantolol(NC-1400) 3 x 10 -5 M.

Table 1. The pA 2 values and slopes of Schild plot of bevantolol(NC- 1400) and labetalol

Antagonist pA 2 Slope

Bevantolol(NC-1400) 5.77 + 0.07 1.29 + 0.20 Labetalol 7.09 _+ 0.11 1.06 + 0.22

Each value is presented as a mean _+ SE of 4 experiments. Slope: slope of Schild plot.

shown in Figs 1 and 2. Schild plots of these results yielded straight lines with a slope of unity (Table 1) suggesting a simple competitive antagonism. The pA2 values for bevantolol(NC-1400) and labetalol were summarized in Table 1, indicating that in alphal-adrenoceptor blocking activity, labetalol is about 20 times as potent as bevantoiol(NC-1400), since a difference between both the pA 2 values is 1.32.

DISCUSSION

response curves, the strips were equilibrated with a competi- tive antagonist for 30 min. Concentration-response curves were then obtained in the presence of the antagonist and the procedure repeated with a high (either 3-fold or 10-fold) concentration of the antagonist in the same preparation. After determination of the control concentration-response curves, two or three successive cumulative concentration- response curves for norepinephrine were determined. Curves were nearly superimposable and changes in sensitivity, sensitization or desensitization were minimal (data not shown). The pA 2 values were calculated according to the method of Arunlakshana and Schild (1959).

Drugs used: bevantolol hydrochloride(NC-1400) was kindly supplied from Nippon Chemiphar Co. Ltd (Tokyo, Japan). Other drugs were labetalol hydrochloride (Sigma Chemical Co., St Louis, Mo.) and norepinephrine hydro- chloride (Wako-Junyaku Co., Tokyo, Japan), all in powder form.

RESULTS

Bevantolol(NC-1400; 3 x 10 -6, 10 -5 and 3 x 10 -5 M) and labetalol (3 x 10 -6, 10 -5 and 3 x 10 -5 M) antagonized competitively a norepinephrine-induced contraction of rat aorta: that is, the concen- trat ion-response curve of norepinephrine showed a parallel shift to the right. The parallel shifts of concentrat ion-response curve of norepinephrine are

6

-10 -9 "-8 -7 -6 -5 -4 -3

Log[N£]

Fig. 2. Concentration-response curves of norepinephrine in the absence and presence of various concentrations of labetalol. Ordinate: contraction (%). Abscissa: logarithm of concentration (M) of norepinephrine. Each value is pre- sented as a mean __+ SE (bar) of 4 strips. (Q) Norepinephrine alone, (O) with labetaloi 3 x 10 -6 M, (&) with labetalol

10 -5 M and (Zk) with labetalol 3 x 10 -5 M.

In the rat thoracic aorta, bevantolol(NC-1400) and labetalol caused a parallel shift of the concentra t ion- response curve of norepinephrine, as reported pre- viously in the rabbit thoracic aorta (Figs 1 and 2). The pA 2 values of bevantolol(NC-1400) and labetalol estimated in the rat aorta (Table 1) indicates that labetalol is about 20 times as potent as bevan- tolol(NC-1400) in alphal-adrenoceptor blocking ac- tivity. This potency ratio of bevantolol(NC-1400) relative to labetalol is similar to the ratio estimated in the rabbit aorta by Takayanagi et al. (1987), who reported that bevantolol(NC-1400) is about 1/30th as potent as labetalol.

The pA 2 values of bevantoloi(NC-1400) and labetalol (Table 1) were approximately 1 order of magnitude higher in the rat thoracic aorta than in the rabbit aorta. These results are similar to the findings of Takayanagi et al. (1985) reported that the pA2 values of prazosin and its derivatives were all approximately 1 logarithm unit higher in the rat aorta than in the rabbit aorta, and also to the observations of Digges and Summers (1983), who mentioned that the pA 2 values of prazosin, phentolamine and rauwol- scine were about 1 logarithm unit less in rat portal veins than in rat aortic strips. Further, it was reported that the rabbit aorta contains primarily alpham (Suzuki et al., 1990; Takayanagi et al., 1992), while the rat aorta contains primarily alphalA (Hart et al., 1990). Therefore, a difference between the pA 2 values of bevantolol(NC-1400) and labetalol in the rat and rabbit thoracic aortic strips suggests that both the drugs might distinguish between alphalA and alphaiB or between alphalL and alpham.

REFERENCES

Arunlakshana O. and Schild H. O. (1959) Some quantitative uses of drug antagonists. Br. J. Pharmac. 14, 48-58.

Digges K. G. and Summers R. J. (1983) Characterization of postsynaptic ~t-adrenoceptors in rat aortic strips and portal veins. Br. J. Pharmac. 79, 655-665.

Flavahan N. A. and Vanhoutte P. M. (1986) ~tt-Adrenoce p- tor subclassification in vascular smooth muscle. Trends Pharmac. Sci. 7, 347-349.

Han C., Adel P. W. and Minneman K. P. (1987) Heterogen- eity of cq-adrenergic receptors revealed by chloroethyl- clonidine. Molec. Pharmac. 32, 505-510.

Alpham-adrenoceptor blocker activities of bevantolol and labetalol 845

Han C., Li J. and Minneman K. P. (1990) Subtypes of ~,-adrencccptors in rat blood vessels. Eur. J. Pharmac. 190, 97-104.

Morrow A. L. and Creese I. (1986) Characterization of ~l-adrenergic receptor subtypes in rat brain: a reevalu- ation of [3H]WB4101 and [3H]prazosin binding. Molec. Pharmac. 29, 321-330.

Suzuki E., Tsujimoto G., Tamura K. and Hashimoto K. (1990) Two pharmacologically distinct ~-adrenoceptor subtypes in the contraction of rabbit aorta: each subtype couples with a different Ca 2+ signalling mechanism and plays a different physiological role. Molec. Pharmac. 38, 725-736.

Takayanagi I., Harada M., Koike K. and Satoh M. (1992) Differences in aradrenoceptor mechanisms for

phenylephrine and tizanidine in rabbit thoracic aorta and common iliac artery. Can. J. Physiol. Pharmac. 67. In press.

Takayanagi I., Kizawa Y., Iwasaki S. and Nakagoshi A. (1987) ( + )- l-[[2-(3,4-Dimethoxyphenyl)ethyl]amino]-3- (3-methylphenoxy)-2-propanol hydrochloride (bevanto- 1ol, NC-1400) as a//l-selective adrenoceptor blocker with al-adrenoceptor blocking activity. Gen. Pharmac. 18, 87-89.

Takayanagi I., Konno F., Arai H., Chimoto K. and Kitada T. (1985) Newly synthesized ~radrenoceptor blocker (SM911 and SM2470) and characterization of a-adrenoceptors in rat aortic strips, rat vas deferens preparations and rabbit aortic strips. Gen. Pharmac. 16, 613-615.