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We investigated the ganglionic effects of angiotensin II (Ang II) and the signal transduction involved in the cardiac sympathetic ganglia by the direct administration of agents to the ganglia through the right subclavian artery and monitoring the heart rate as an indicator of the ganglionic function in pithed dogs. Ang II given i.a. caused increases in the heart rate, which was inhibited by the treatment with the AT1-receptor antagonist forasartan, but not by the AT2-receptor antagonist PD-123319. The stimulation by Ang II, but not by acetylcholine, was inhibited after treatment with an inhibitor of phospholipase C, U-73122; a cell-permeant modulator of the Ins(1,4,5)P3 receptors, 2-aminoethoxydiphenyl borate; an intracellular calcium and calcium-associated protein kinase inhibitor, HA-1077; calmodulin (CaM) inhibitor, W-7; Ca2+/CaM-dependent protein kinase II inhibitor, KN-93; a selective protein kinase C inhibitor, calphostin C; and Na+H+ exchange inhibitor, dimethylamiloride. These results suggest that Ang II stimulates the ganglionic transmission at postsynaptic sites via the activation of AT1 receptor coupled to either activation of phospholipase C, phosphoinositide hydrolysis and subsequent increase in intracellular Ca2+ and activation of protein kinase C and Ca2+/CaM kinase II, although this ganglionic stimulation seems to involve, at least in part, the protein kinases-dependent increase of amiloride-sensitive Na+ inflow.

The possible involvement of the local angiotensin system in ganglionic functions was investigated in the canine cardiac sympathetic ganglia. Positive chronotropic responses to preganglionic stellate stimulation at high frequencies, after intravenous administration of pentolinium plus atropine, were inhibited by the nonpeptide angiotensin AT(1) receptor antagonist forasartan or the angiotensin I-converting enzyme inhibitor captopril, whereas the rate increases elicited by the postganglionic stellate stimulation and norepinephrine given intravenously failed to be inhibited by these antagonists. The levels of endogenous immunoreactive angiotensin II, as determined by radioimmunoassay in the incubation medium of the stellate and inferior cervical ganglia, were increased after the high-frequency preganglionic stimulation of the isolated ganglia. The increment of the peptide was also antagonized by the pretreatment with captopril but not by a chymase inhibitor, chymostatin. The expression of angiotensinogen mRNA was observed in the stellate ganglion, adrenal, liver, and lung but not in the ovary and spleen. The expression of the mRNA in the stellate and inferior cervical ganglia increased after high-frequency preganglionic stimulation of the in vivo dogs for a period of 1 hour. These results indicate that an intrinsic angiotensin I-converting enzyme-dependent angiotensin system exists in the cardiac sympathetic ganglia, which is activated by high-frequency preganglionic stimulation.

The effects of angiotensin II and neuro-aminoacids administered through the right subclavian artery (i.a.) to the cardiac sympathetic ganglia were investigated in spinal dogs. Angiotensin II (1--8 micrograms) elicited a dose-dependent positive chronotropic effect which was reduced after i.a. injection of saralasin (100 micrograms). The effect of angiotensin II was not reduced after combined treatment with either hexamethonium (10 mg/kg) plus atropine (0.1 mg/kg) or hemicholinium-3 (5 mg/kg) plus preganglionic stimulation. The dose-dependent response to angiotensin II of heart rate was inhibited by GABA (50, 500 micrograms), GABOB (500 micrograms) and muscimol (50, 100 micrograms). The inhibition of the response to angiotensin II by a small dose of GABA (50 micrograms), but not by a high one (500 micrograms), was antagonized by i.a. injection of picrotoxin (2 mg). The positive chronotropism induced by bethanechol (25, 50 micrograms) and a small dose of acetylcholine (25 micrograms) were significantly inhibited by a high dose (500 micrograms) but not by a low dose (50 micrograms) of GABA. These results confirm that angiotensin II stimulates cardiac chronotropism by acting on the angiotensin II receptor located at the cardiac ganglia and show that this stimulant effect is antagonized by GABA.

Participation of endogenous gamma-aminobutyric acid (GABA) in the functions of dog cardiac ganglia was investigated. The ganglionic stimulants as well as agents affecting GABA system were given directly into the cardiac sympathetic ganglia through the right subclavian artery (i.a.), unless otherwise mentioned. Inhibition of endogenous GABA degradation by the GABA-transaminase inhibitor, aminooxyacetic acid (AOAA) administered 10 mg/kg i.v. 2 hr before completion of surgical procedures did not alter the positive chronotropic responses to bethanecol (25 and 50 micrograms) and acetylcholine (25, 50 and 100 micrograms) but reduced markedly those to angiotensin II (1 and 2 micrograms). This reduction was antagonized by picrotoxin (5 mg). Diazepam given 10 mg/kg i.v. also inhibited the ganglionic responses to angiotensin II in both untreated and AOAA-pretreated dogs, this inhibition by diazepam being more marked in the AOAA-pretreated than in the untreated dogs. The same dose of diazepam did not affect the responses to acetylcholine but reduced to a certain degree of responses to bethanechol in the AOAA-pretreated dogs. These inhibitions by diazepam were also reversed by picrotoxin. After i.v. treatment with the glutamic acid decarboxylase inhibitors, 3-mercaptopropionic acid (50 mg/kg) or isoniazid (200 mg/kg), the ganglionic responses to angiotensin II were not altered, but inhibitory effects of diazepam on the responses to angiotensin II were eliminated after 3-mercaptopropionic acid but not after isoniazid. These results suggest that endogenous GABA may play an inhibitory role in the nonnicotinic ganglionic pathways and that diazepam probably exerts a ganglionic action through endogenous GABAergic mechanisms.