ACTIVATION OF NO-CGMP PATHWAY BY ACETYLCHOLINE IN BOVINE CHROMAFFIN CELLS - POSSIBLE ROLE OF CA2-REGULATION OF CGMP SIGNALING( IN THE DOWN)

The production of cyclic GMP (cGMP) induced by acetylcholine and other stimuli was studied in bovine chromaffin cells. Acetylcholine increas ed intracellular cGMP in a transitory (peak at 2 min) and concentratio n-dependent manner (estimated half maximal increase, EC(50) = 61 +/- 5 mu M). N-G-nitro-L-arginine methyl ester (NAME) inhibited such a rise in cGMP with a half maximal inhibitory concentration (IC50) of 231 +/ - 55 mu M. The acetylcholine-induced increase in cGMP was also inhibit ed by a calmodulin antagonist (calmidazolium, 30 mu M) and by the abse nce of extracellular calcium. Other agents that strongly increased cyt osolic calcium concentration ([Ca2+](i)) as acetylcholine did, such as the nicotinic-agonist, 1,1-dimethyl-4-phenylpiperazinium (DMPP), high -KCl (50 mM), and ionomycin, also caused a rise in cGMP in cultured bo vine chromaffin cells. Veratridine, an activator of sodium channels, p roduced a slowly developing calcium increase and no significant cGMP p roduction. The muscarinic-agonist, muscarine, failed to increase cytos olic calcium, and was the weakest stimulator of cGMP production. cGMP formation, induced by sodium nitroprusside (SNP, 100 mu M) and by C-ty pe natriuretic peptide (CNP, 100 nM), was inhibited by 30-40% by incre asing [Ca2+](i) with ionomycin. This inhibition was abolished by calmi dazolium (30 mu M) and by the absence of calcium in the extracellular medium. In conclusion, bovine chromaffin cells synthesize nitric oxide (NO) to activate guanylate cyclase in response to several stimuli, wh ich increase [Ca2+](i). Moreover, the increase in [Ca2+](i) also stimu lates a Ca2+/calmodulin phosphodiesterase, which could down-regulate t he levels of cGMP in these cells.