The Kinetics of Norepinephrine-induced Stimulation of Serotonin N-Acetyltransferase in Bovine Pineal Gland

Abstract

Fresh pineal explants were cultured in medium 199 containing 0.1 mg/ml ascorbic acid, 2 mM glutamine, and 0.1% bovine serum albumin in 95% O2 and 5% CO2 from 1-10 h. Norepinephrine (10 .mu.M) increased serotonin N-acetyltransferase (NAT) activity from 2.36 .+-. 0.23 to 6.98 .+-. 0.46 pmol product produced/mg protein per min (unit), without influencing tryptophan hydroxylase, 5-hydroxytryptophan decarboxylase, monoamine oxidase or hydroxyindole-O-methyltransferase. Dibutyryl cAMP (1 mM) also stimulated NAT from 1.39 .+-. 0.12 to 4.33 .+-. 0.65 U. The norepinephrine-induced stimulation of NAT was blocked by GABA (10 .mu.M), by dl-propranolol (20 .mu.M), by cycloheximide (30 .mu.M), by puromycin (20 .mu.M), but not by actinomycin D (10 .mu.M). Comparing the kinetics of native and stimulated NAT, there occurred changes in Vmax with regard to both substrates.sbd.tryptamine (from 0.885 to 3.44 U) and acetyl CoA (from 3.45 to 9.10 U).sbd.without changes in Km with respect to both tryptamine (Km = 5 .mu.M) and acetyl CoA (Km = 50 .mu.M). The norepinephrine-induced stimulation of NAT may be due to enhanced protein synthesis involving translational or post-translational events. The enhanced protein synthesis is coupled through activation of .beta.-receptor and cAMP-mediated actions in pineal gland. The precise interrelationship between GABA and norepinephrine in mediating and perhaps in controlling the biological events in the bovine pineal gland is still unclear.