Depolarizing stimuli increase tyrosine hydroxylase in the mouse locus coeruleus in culture

The influence of membrane depolarization on the development and regulation of brain noradrenergic neurons was studied in explant cultures of the mouse locus coeruleus (l.c.). Exposure to the depolarizing agents veratridine or elevated K+ significantly increased the catalytic activity of tyrosine hydroxylase (TH), the rate-limiting enzyme in catecholamine biosynthesis. The effects of veratridine were prevented by tetrodotoxin, suggesting that transmembrane Na+ influx was necessary for the rise in TH. Morphometric analysis indicated that the rise in TH activity was not accompanied by altered TH-positive cell number or cell diameter. Rather, TH fluorescence intensity increased in each neuron, suggesting that depolarization increased TH per neuron. Immunoblot and densitometric analysis indicated that depolarization did, indeed, increase TH immunoreactive protein. Moreover, depolarization elevated enzyme activity in cultured neurons expressing the normal developmental increase in TH, as well as those in which plateau levels had already been attained. We conclude that depolarization and/or Na+ influx regulates a critical transmitter macromolecule in brain neurons, as in the periphery, by altering enzyme molecule number.