The intercellular communication via nitric oxide and its regulation in coupling of cyclic GMP synthesis upon stimulation of muscarinic cholinergic receptors in rat superior cervical sympathetic ganglia

Ando, M.; Tatematsu, T.; Kunii, S.; Nagata, Y.

Brain Research 650(2): 283-288


ISSN/ISBN: 0006-8993
PMID: 7525017
DOI: 10.1016/0006-8993(94)91793-0
Accession: 009593253

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Cyclic GMP (cGMP) production in rat superior cervical sympathetic ganglia (SCG) was markedly increased (ca. 7-9-fold) by the addition of either acetylcholine (ACh; 0.1 mM) or a muscarinic agonist. carbachol (Carb; 0.1 mM), in the presence of an inhibitor (3-isobutyl-1-methylxanthine) for cGMP hydrolytic enzyme during in vitro aerobic incubation at 37 degree C for 5 min. The ACh-induced accumulation of cGMP in SCG was effectively blocked 73%) by the further addition of atropine (10 mu-M), a muscarinic antagonist, whereas a nicotinic blocker, hexamethonium (10 mu-M) partially antagonized (-41%) this ACh stimulation. The inhibitory effect of hexamethonium on ACh-evoked ganglionic cGMP production was effectively augmented (-83%) by addition of N-G-monomethyl-L-arginine (L-NMMA, 50 mu-M), a compound that inhibits nitric oxide (NO) synthesis from L-arginine. Comparable inhibition of cGMP formation was observed following application of L-NMMA to the SCG upon stimulation of Carb. In contrast, L-NMMA had no effect on the decreased level of ACh-evoked cGMP production caused by the muscarinic antagonist. The Carb-induced elevation of ganglionic cGMP synthesis was significantly reduced within 1 min of incubation in the medium containing hemoglobin (Hb; 20 mu-M), an agent that scavenges only the extracellular fraction of NO. Thereafter, the tissue cGMP formation attenuated to the control level by subsequent incubation for several minutes. Addition of protein kinase C (PKC) activator, 12-O-tetradecanoylphorbol 13-acetate (TPA; 1 mu-M) to the medium significantly decreased Carb-evoked cGMP synthesis (-61%) in SCG, whereas superoxide dismutase (SOD: 30 U/ml) only slightly suppressed the Carb stimulation. This finding supports the idea that PKC might play a role in dampening the muscarinic receptor-mediated increase in NO release within ganglionic tissue. In axotomized SCG one week prior to examination, where sympathetic neurons were degenerated and reactive proliferation of glial cells was in progress, no stimulatory effect of Carb-induced cGMP production via NO release was seen. These results provide evidence that a large fraction of NO generated upon stimulation of muscarinic receptors in sympathetic neuronal cells can possibly freely diffuse in extracellular space, and then be taken back into the same group of surrounding cells to stimulate cGMP production.