AMPHETAMINE INCREASES THE PHOSPHORYLATION OF NEUROMODULIN AND SYNAPSIN-I IN RAT STRIATAL SYNAPTOSOMES

Amphetamine is taken up through the dopamine transporter in nerve term inals and enhances the release of dopamine. We previously found that i ncubation of rat striatal synaptosomes increases phosphorylation of th e presynaptic neural-specific protein, neuromodulin (Gnegy et al., Mel . Brain Res. 20:289-293, 1993). Using a state-specific antibody, we no w demonstrate that incubation of rat striatal synaptosomes with amphet amine increases levels of neuromodulin phosphorylated at ser(41), the protein kinase C substrate site. Phosphorylation was maximal at 5 min at 37 degrees C at concentrations from 100 nM to 10 mu M amphetamine. The effect of amphetamine on the phosphorylation of synapsin I at a si te specifically phosphorylated by Ca2+/calmodulin-dependent protein ki nase II (site 3), was examined using a state-specific antibody for sit e 3-phosphosynapsin I. Incubation with concentrations of amphetamine f rom I to 100 nM increased the level of site 3-phospho-synapsin I at ti mes from 30 sec to 2 min. The effect of amphetamine on synapsin I phos phorylation was blocked by nomifensine. The presence of calcium in the incubating buffer was required for amphetamine to increase the level of site 3-phospho-synapsin I. The amphetamine-mediated increase in the content of phosphoser(41)-neuromodulin was less sensitive to extrasyn aptosomal calcium. The amphetamine-mediated increase in the content of site 3-phospho-synapsin I persisted in the presence of 10 mu M okadai c acid and was not significantly altered by DI or D2 dopamine receptor antagonists. Preincubation of striatal synaptosomes with 10 mu M of t he protein kinase C inhibitor, Ro-31-8220, blocked the amphetamine-med iated increases in the levels of both phosphoser(41)-neuromodulin and site 3-phospho-synapsin I. Our results demonstrate that amphetamine ca n alter phosphorylation-related second messenger activities in the syn aptosome. (C) 1997 Wiley-Liss, Inc.