Gen. Pharmac. Vol. 15, No. 2, pp. 149-150. 1984 0306-3623/84 $3.00+0.00 Printed in Great Britain. All rights reserved Copyright ,~? 1984 Pergamon Press Ltd

NICOTINIC STIMULANT, DMPP(1,1-DIMETHYL- 4-PHENYLPIPERAZINIUM IODIDE) ON THE ISOLATED BRONCHIAL SMOOTH MUSCLE

PREPARATION OF GUINEA-PIG

YASUO KIZAWA and ISSEI TAKAYANAGI* Department of Chemical Pharmacology, Toho University School of Pharmaceutical Sciences, Funabashi,

Chiba 274, Japan (Tel: (0474) 72 ll41)

(Received 13 June 1983)

A b s t r a c t - - l . Contractile responses of the isolated bronchial preparation of guinea-pig to DMPP, a nicotinic stimulant, were inhibited by ganglion blockers but not influenced by atropine, suggesting that DMPP did not bring about a contribution by stimulation of cholinergic ganglion cells.

2. The fact that tetrodotoxin did not influence the response to DMPP suggests that a possible site of action of DMPP is not on the nerve cells.

INTRODUCTION

In a prev ious p a p e r (Takayanag i and Kizawa , 1983) we r epo r t e d tha t n icot ine d id no t b r ing a b o u t a c o n t r i b u t i o n by s t imula t ion o f chol inerg ic gang l ion cells in the i sola ted b ronch ia l s tr ips o f guinea-pig . In o rde r to ob ta in add i t iona l evidence, we s tudied m o d e s o f ac t ion o f D M P P ( 1 , 1 - d i m e t h y l - 4 - p h e n y l - p ipe raz in ium iodide), a n icot in ic s t imulan t , in the i so la ted b ronch ia l p r e p a r a t i o n o f guinea-pig .

METHODS AND MATERIALS

Male guinea-pigs (350400 g in body weight) were killed by a blow on the head and main bronchi were isolated and cut helically. The preparations (2 x 30 mm) were suspended in a 20 ml organ bath filled with a physiological solution (NaC1 118, KC1 4.72, CaCI 2 2.56, MgSO4.7H20 0.16, KH2PO 4 1.20, NaHCO 3 25.0, and dextrose 10.0 mM) gassed with carbogen and were recorded isometrically under a resting tension of 0.5 g. In some experiments, two platinum electrodes (2 x 35 ram) were set at intervals of 5 mm and field stimulation of the bronchial preparation was carried out by passing a rectangular pulse of 0.5 msec duration, supramaximal voltage and a frequency of 10 Hz between two electrodes for 10 sec. The experiments were started after the preparations were allowed to develop their spontaneous tone for 60 min. All agonists were applied to the bronchial preparations at intervals of 60 min. To test effects of antag- onists, the preparations were preincubated with one of the antagonists for the amount of time listed in Table 1.

RESULTS

Contractile responses to DMPP were reproducible under the experimental conditions used. DMPP (10 ~ 10 -3 M) induced the contraction of the preparation (Fig. l). No inhibitory response to DMPP (10 s 10-3 M) was observed in all the preparations used, which were considerably re- laxed by papaverine (10 -4 M) (Fig. 1). DMPP, 10 4 M and the equieffective acetylcholine, 10 4 M were used as agonists in the following experiments. The response to DMPP was

*Author to whom all correspondence should be addressed.

greatly reduced by hexamethonium (10-SM) and pen- tolinium (10 6 M), ganglion blockers, the concentrations of which were enough to inhibit the responses of guinea-pig ileum to DMPP, but was not influenced by atropine (10 6M), which inhibited the response to acetylcholine (10-4M), tetrodotoxin (3 × 10 6M) and SX-284 [3 × 10 -7 M; 2-(1,2-benzisoxazol-3-yl)-3-[2-(2-piperidino- ethoxy)phenyl]acrylonitrile], the latter of which was found to inhibit acetylcholine release from the parasympathetic nerve specifically (Takayanagi et al., 1982; Sone and Tak- ayanagi, 1983). DMPP-induced contraction was not influenced by indomethacin (10 -6 M) or diphenhydramine (10 7 M). Field stimulation induced a contraction of the preparation, which was about 45~o of that to DMPP (10-4M). Thirty-minute treatment with physostigmine (10 6M), which potentiated the contractile responses to

I I Pop 10-4M

"

DMPP DMPP DMPP IOOmg

10-5M 10-4M 10-3M

(B)

.-~ 45

g ~ ~° m o 15

/ 1 I J I

10 .5 i 0 - 4 i0 - 3

D M P P (M)

Fig. 1. (A) Responses of the isolated guinea pig bronchial preparation to DMPP and papaverine (Pap). (B) Dose-response curve of DMPP. Ordinate is expressed as % of the contraction of preparation induced by acetylcholine, 10 2 M and abscissa is concentration (M) of DMPP. Each value is presented as a mean with SE (bar) of 4 experiments.

149

150 YASUO KIZAWA and ISSEI TAKAYANAGI

Table 1. Effects of some drugs on the responses to DMPP, acetylcholine and field stimulation

Incubation time ~o of contraction Treatment (rain) (N)

DMPP, 10 4M 100.0 Hexamethonium, 10 5 M 5 33.5 + 4.1" (4) Pentolinium, 10 6 M 5 26.6 _+ 5.7* (4) Atropine, 10 6 M 5 90.5 _+ 4.9 (4) Tetrodotoxin, 3 x 10 6 M 15 94.0 _+ 9.4 (7) Indomethacin, 10 6 M 30 94.8 _+ 11.3 (7) SX-284, 3 × 10-TM 15 111.3 _+ 13.2(5) Physostigmine, 10 -6 M 30 108.2 _+ 8.1 (7) Diphenhydramine, 10 7 M 5 102.8 +_ 7.7 (8)

Acetylcoline, 10-4M 100.0 Atropine, 10 -6 M 5 12.4 _+ 4.9* (5) Physostigmine, 10-6 M 15 230.0 + 20.1" (4)

Field stimulation 100.0 Tetrodotoxin, 3 x 10-6 M 15 16.4 _+ 4.3* (4) Physostigmine, 10 6 M 30 148.0 -- 6.0* (4)

Each value is presented as a mean _+SEM of the number of experiments in parentheses. *Significant difference from 100~o at P < 0.05.

field stimulation and to acetylcholine (10 4 M), was without any effect on that to DMPP. These results were summarized in Table 1.

DISCUSSION

Ac t ions o f n ico t ine and nicot in ic s t imulan ts on the i sola ted gu inea-p ig t r achea are well k n o w n (Jones et al., 1980; C o b u r n and Tomi t a , 1973; H a w k i n s and Pa ton , 1958). The re are, however , a few repor t s ( H a w k i n s and Pa ton , 1958; T a k a y a n a g i and Kizawa , 1983) a b o u t the ac t ion o f n ico t ine on the isolated b ronch ia l musc le o f guinea-pig . The p resen t results wi th D M P P were s imilar to those with n icot ine r epo r t ed previous ly (Takayanag i and Kizawa, 1983). The cont rac t i l e r esponse to D M P P was inh ib i ted by the gang l ion b lockers , bu t no t inf luenced by a t rop ine , physos t igmine , t e t r o d o t o x i n and SX-284, the specific inh ib i to r o f ace ty lchol ine release f rom pa ra sym- pa the t i c nerve (Takayanag i et al., 1982; Sone and Takayanag i , 1983). These results suggest tha t D M P P did n o t b r ing a b o u t any c o n t r i b u t i o n by s t imula t ion o f chol inergic gang l ion cells in this s tudy. F u r t h e r ev idence tha t t e t r o d o t o x i n did no t inf luence the con- tract i le r e sponse o f b ronch ia l p r e p a r a t i o n to D M P P suggests tha t a poss ible site o f ac t ion o f n icot ine is on the nerve cells. The cont rac t i l e r e sponse o f b ronch ia l p r e p a r a t i o n to D M P P was no t inh ib i ted by in- d o m e t h a c i n and d i p h e n h y d r a m i n e , suggest ing tha t

the response to D M P P was no t due to a release o f p ro s t ag l and i n s or h is tamine . We could not , however , rule ou t a c o n t r i b u t i o n o f chemica l med ia to r s re- leased by D M P P in the cont rac t i le mechan i sms .

Acknowledgement--This research was supported by a grant from the Japan Monopoly Corporation.

REFERENCES

Coburn R. J. and Tomita T. (1973) Evidence for non- adrenergic inhibitory nerves in guinea-pig trachealis mus- cle. Am. J. Physiol. 224, 1072-1077.

Hawkins D. F. and Paton W. D. M. (1958) Responses of isolated bronchial muscle to ganglionically active drugs. J. Physiol. 144, 193-219.

Jones T. R., Lefcoe N. M. and Hamilton J. T. (1980) Studies of the action of nicotine in guinea-pig tracheal smooth muscle: interaction with fl-adrenoceptor antagonists. Eur. J. Pharmac. 67, 53-64.

Sone H. and Takayanagi I. (1983) Inhibitory action of 2- (1,2-benzisoxazol-3-yl)-3-[2-(2-piperidinoethoxy)phenyl] -acrylonitrile (SX-284) on acetylcholine release from va- gus nerve in guinea-pig ileum. J. Pharm. Dyn. 6, 272 274.

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