Journal of Neuroscience, Vol 14, 2106-2116, Copyright © 1994 by Society for Neuroscience
A single transmitter regulates gene expression through two separate mechanisms: cholinergic regulation of phenylethanolamine N- methyltransferase mRNA via nicotinic and muscarinic pathways
MJ Evinger, P Ernsberger, S Regunathan, TH Joh and DJ Reis
Department of Pediatrics, SUNY at Stony Brook 11794-8111.
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.
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