A single transmitter regulates gene expression through two separate mechanisms: cholinergic regulation of phenylethanolamine N-methyltransferase mRNA via nicotinic and muscarinic pathways.

ACh regulates the gene encoding phenylethanolamine N-methyltransferase (PNMT) in bovine adrenal chromaffin cells. In addition to stimulating catecholamine release from these cells, cholinergic agents elevate transcription of the PNMT gene. Carbachol, which activates both nicotinic and muscarinic receptors, produces 12-19-fold increases in PNMT mRNA and a 22-fold increase in epinephrine release. Selective nicotinic and muscarinic antagonists (hexamethonium and atropine) each partially reduce carbachol-stimulated increases in PNMT mRNA while a combination of both eliminates > 90% of the carbachol response, thus indicating that separable nicotinic and muscarinic components contribute to the cholinergic increase in PNMT mRNA. Muscarine alone produces a dose-dependent increase (mean sixfold) in steady state PNMT mRNA levels and stimulates the rate of transcription fivefold. Only atropine and the m3-m4-selective muscarinic antagonist 4-diphenylacetoxy-4-methyl-piperidine (4-DAMP) reduce the response to muscarine, strongly suggesting that the m4 receptor is crucial for PNMT mRNA activation. In these chromaffin cells, muscarine inhibits adenylate cyclase, antagonist bind with affinities characteristic of m4 receptors, and cDNA hybridization detects only m4 mRNAs (Fernando et al., 1991). Nicotine also induces a dose-dependent increase (mean of 8.5-fold) in PNMT mRNA levels. The importance of voltage-gated Ca2+ channels in the nicotine effect is demonstrated by the stimulatory effects of calcium ionophores on PNMT mRNA levels (two-to fivefold increase) and the ability of the L- and N-type channel blockers nifedipine and omega-conotoxin to decrease the nicotine response (by 60% and 40%, respectively). Nuclear "run-on" assays further reveal that nicotine enhances transcription of the PNMT gene (approximately fourfold). Thus, this study provides the first demonstration that both nicotinic and muscarinic stimulation modify genomic responses of bovine adrenergic chromaffin cells and identifies possible mechanisms.