ROLE OF INTRA-NEURONAL 5-HT AND OF TRYPTOPHAN-HYDROXYLASE ACTIVATION IN THE CONTROL OF 5-HT SYNTHESIS IN RAT-BRAIN SLICES INCUBATED IN K+-ENRICHED MEDIUM

Abstract— The incubation of brain stem slices from adult rats in a K+‐enriched medium containing a 5‐HT uptake inhibitor (fluoxetine) significantly increased their capacity to synthesize 5‐HT from tryptophan. The K+‐induced stimulation of 5‐HT synthesis was at least partly dependent on the depletion of the indoleamine in tissues since: (1) a good correlation was found between the respective changes in 5‐HT release and synthesis evoked by high K+ concentrations in the presence of various 5‐HT uptake inhibitors; (2) the modifications in endogenous 5‐HT levels produced by in vim treatments with drugs (reserpine, pargyline) or by incubating slices with 5‐HT altered the stimulating effect of high K+ concentrations and fluoxetine on 5‐HT synthesis; (3) the replacement of Ca2+ by Co2+ (4 mM) or EGTA (0.1 mM) in the incubating medium completely prevented the increased 5‐HT release and synthesis evoked by high K+ concentrations and fluoxetine. The extraction of tryptophan hydroxylase from incubated tissues revealed that the increased 5‐HT synthesis occurring in K+‐enriched medium was associated with an activation of this enzyme. Kinetic analyses indicated that this activation resulted from an increase in the Vmax of tryptophan hydroxylase, its apparent affinities for both tryptophan and 6‐MPH4 being not significantly affected. In contrast to the tryptophan hydroxylase from tissues incubated in normal physiological medium, the activated enzyme from tissues depolarized by K+ was hardly stimulated by Ca2+‐mediated phosphorylating conditions. This led to the proposition of a hypothetical model by which the Ca2+ influx produced by the neuronal depolarization would trigger the activity of a Ca2+‐dependent protein kinase capable of activating tryptophan hydroxylase. Although this sequence is still largely speculative it must be emphasized that, as expected from such a model, the regional differences in the K+‐evoked activation of tryptophan hydroxylase in slices (cerebral cortex > brain stem > spinal cord) were parallel to those of the Ca2+‐dependent protein phosphorylation (r= 0.92) and those of the activating effect of phosphorylating conditions on soluble tryptophan hydroxylase (r= 0.96). Copyright © 1979, Wiley Blackwell. All rights reserved