Protein phosphorylation in rat hippocampal synaptic plasma membranes in response to neurohypophyseal peptides

The effect of arginine vasopressin (AVP) on protein phosphorylation in rat hippocampal synaptic plasma membranes (SPM) was examined. With a crude SPM preparation, AVP (10(-8)-10(-5) M) stimulated phosphorylation of a number of proteins which included a brain-specific protein of 48 kDa called B50 or protein F1, which is thought to be related to synaptic plasticity. Equimolar levels of oxytocin also stimulated B50/F1 phosphorylation. AVP and oxytocin at higher concentrations (10(-4)-10(-3) M) reduced SPM protein phosphorylation. When SPM was treated with both AVP and oxytocin, the effects were not additive; on the other hand, the effects of the phorbol ester (TPA) and AVP were additive. With SPM, partially purified by sucrose density centrifugation, only the inhibitory effect of AVP on B50/F1 phosphorylation was seen. These results suggest that AVP and oxytocin stimulation of B50/F1 phosphorylation requires cellular factors which are removed from SPM during membrane purification. In contrast, the inhibitory mechanism triggered by AVP and oxytocin appears to be associated with, or an integral part of, the synaptic membrane itself. Because the effects on membrane protein phosphorylation with maximal amounts of AVP and oxytocin were not additive, they must bind to the same sites on the membrane. This conclusion is supported by the additivity of the effects of AVP and phorbol ester, since the phorbol ester can act directly on the kinase and does not require a membrane recognition site.