Influence of pentobarbital-Na on stimulation-evoked catecholamine secretion in the perfused rat adrenal gland

Abstract

Objectives: The present study was attempted to investigate the effects of pentobarbital-Na, one of the barbiturates which are known to depress excitatory synaptic transmission in the central nervous system at concentrations similar to those required for the induction and maintenance of anesthesia, on catecholamines (CA) secretion evoked by cholinergic stimulation and membrane-depolarization from the isolated perfused rat adrenal gland, and to clarify the mechanism of its action.

Methods: Mature male Sprague-Dawley rats were anesthetized with thiopenal-Na (40 mg/kg, s.c.). The adrenal gland was isolated by the methods of Wakade. A cannula used for perfusion of the adrenal gland was inserted into the distal end of the renal vein. The adrenal gland was carefully removed from the animal and placed on a platform of a leucite chamber.

Results: The perfusion of pentobarbital-Na(30-300 uM) into an adrenal vein for 20 min produced relatively dose-dependent inhibition in CA secretion evoked by ACh(5.32 mM), DMPP(100 uM for 1 min), McN-A-343(200 uM for 2 min), Bay-K-8644(10 uM) and high potassium(56 mM), while it did not affect the CA secretion of cyclopiazonic acid(10 uM). Also, in the presence of thiopental-Na (100 uM), CA secretory responses evoked by ACh, DMPP, McN-A-343 and high K+ were markedly depressed. Moreover, in adrenal glands preloaded with ketamine(100 uM for 20 min), which is known to be a dissociative anesthetic, CA secretion evoked by ACh, DMPP, McN-A-343 and high K+ were significantly attenuated.

Conclusion: Taken together, these experimental results suggest that pentobarbital-Na depresses CA release evoked by both cholinergic stimulation and membrane-depolarization from the isolated rat adrenal medulla and that this inhibitory activity may be due to the result of the direct inhibition of Ca++ influx into the chromaffin cells without any effect on the calcium mobilization from the intracellular store.

Fig. 1.

Schematic drawing of the preparation used to study secretion of catecholamines in the isolated perfused rat adrenal gland.

Fig. 2.

Influence of 30uM pentobarbital-Na on nicotinic, muscarinic stimulation- and membrane depolarization-induced catecholamine (CA) secretory responses from the isolated perfused rat adrenal glands. CA secretion was induced by a single injection of ACh (5.32mM) and excess K⁺ (56mM), and by the perfusion of McN-A-343 (200uM) and DMPP (100uM) for 1 min, respectively, after perfusion with normal Krebs solution for one hour prior to initiation of the experimental protocol. “BEFORE” and “AFTER” denote CA secretion evoked by ACh, excess K, McN-A-343 and DMPP before and after preloading with 30 uM pentobarbital-Na for 20 min, respectively. Numbers in the parenthesis indicate number of experimental rat adrenal glands. Vertical bars represent the standard error of the mean (S.E.M.). Ordinate the amounts of CA secreted from the adrenal gland in ng. Abscissa: secretogogues. Statistical difference was obtained by comparing the control with the pretreated group. Each perfusate was collected for 4 minutes, but DMPP-induced perfusates was collected twice successively for each 4 minutes. ACh: acetylcholine. McN-A: McN-A-343. *:p<0.05, ***:p<0.01.

Fig. 3.

Influence of 100uM pentobarbital-Na on nicotinic, muscarinic stimulation- and membrane depolarization-induced catecholamine (CA) secretory responses from the inolated perfused rat adrenal glands. Other legends are the same as in Fig. 2. ***:p<0.01.

Fig. 4.

Influence of 300 uM pentobarbital-Na on nicotinic, muscarinic stimulation- and membrane depolarization-induced catecholamine (CA) secretory responses from the inolated perfused rat adrenal glands. Other legends are the same as in Fig. 2. ***: p<0.01

Fig. 5.

Influence of 100uM pentobarbital-Na on CA secretion evoked by Bay-K-8644 and cyclopiazonic acid. Bay-K-8644 (Bay-K, 10 uM) and cyclopiazonic acid (CPA, 10uM) were perfused into an adrenal vein for 4 min before and after the preloading with 100uM pentobarbital-Na for 4 min before and after the preloading with 100uM pentobarbital-Na for 20 min. Its pefusate was collected for 4 min. Other legends are the same as in Fig. 2. ***: p<0.01.

Fig. 6.

Influence of 100uM thiopental-Na on nicotinic, muscarinic stimulation- and membrane depolarization-induced catecholamine (CA) secretory responses from the isolated perfused rat adrenal glands. ACh (5.32mM), excess K (56mM), McN-A-343 (200uM) and DMPP (100uM) were introduced before and after preloading with 100uM thiopental-Na for 20 min, respectively. Other legends are the same as in Fig. 2. ***: p<0.01.

Fig. 7.

Influence of 100uM ketamine on nicotinic, muscarinic stimulation- and membrane depolarization-induced catecholamine (CA) secretory responses from the isolated perfused rat adrenal glands. ACh (5.32mM), excess K (56 mM), McN-A-343 (200uM) and DMPP (100 uM) were introduced before and after preloading with 100uM ketamine for 20 min, respectively. Other legends are the same as in Fig. 2. ***: p<0.01