Determination of transmitter function by neuronal activity

Abstract

The role of neuronal activity in the determination of transmitter function was studied in cultures of dissociated sympathetic neurons from newborn rat superior cervical ganglia. Cholinergic and adrenergic differentiation were assayed by incubating the cultures with radioactive choline and tyrosine and determining the rate of synthesis and accumulation of labeled acetylcholine and catecholamines. Pure neuronal cultures grown in control medium displayed much lower ratios of acetylcholine synthesis to catecholamine synthesis than sister cultures grown in medium previously conditioned by incubation on appropriate nonneuronal cells (conditioned medium). Neurons treated with the depolarizing agents elevated K+ or veratridine, or stimulated directly with electrical current, either before or during application of conditioned medium, displayed up to 300-fold lower acetylcholine/catecholamine ratios than they would have without depolarization, and thus remained primarily adrenergic. Elevated K+ and veratridine produced this effect on cholinergic differentiation without significantly altering neuronal survival. Because depolarization causes Ca2+ entry in a number of cell types, the effects of several Ca2+ agonists and antagonists were investigated. In the presence of the Ca2+ antagonists D600 [.alpha.-isopropyl-.alpha.-[N-methyl-N-homoveratryl-.gamma.-aminopropyl]-3,4,5-trimethoxyphenylacetonitrile] or Mg2+, K+ did not prevent the induction of cholinergic properties by conditioned medium. Depolarization, either steady or accompanying activity, is one of the factors determining whether cultured sympathetic neurons become adrenergic or cholinergic, and this effect may be mediated by Ca2+.