Chronic blockade of glutamate receptors enhances presynaptic release and downregulates the interaction between synaptophysin-synaptobrevin-vesicle-associated membrane protein 2

During development of neuronal circuits, presynaptic and postsynaptic funct ions are adjusted in concert, to optimize interneuronal signaling. We have investigated whether activation of glutamate receptors affects presynaptic function during synapse formation, when constitutive synaptic vesicle recyc ling is downregulated. Using primary cultures of hippocampal neurons as a m odel system, we have found that chronic exposure to both NMDA and non-NMDA glutamate receptor blockers during synaptogenesis produces an increase in m iniature EPSC (mEPSC) frequency, with no significant changes in mEPSC ampli tude or in the number of synapses. Enhanced synaptic vesicle recycling, sel ectively in glutamatergic nerve terminals, was confirmed by the increased u ptake of antibodies directed against the lumenal domain of synaptotagmin. N o increased uptake was detected in neuronal cultures grown in the chronic p resence of TTX, speaking against an indirect effect caused by decreased ele ctrical activity. Enhanced mEPSC frequency correlated with a reduction of s ynaptophysin-synaptobrevin-vesicle-associated membrane protein 2 (VAMP2) co mplexes detectable by immunoprecipitation. Intracellular perfusion with a p eptide that inhibits the binding of synaptophysin to synaptobrevin-VAMP2 in duced a remarkable increase of mEPSC frequency in control but not in glutam ate receptor blocker-treated neurons. These findings suggest that activatio n of glutamate receptors plays a role in the downregulation of the basal ra te of synaptic vesicle recycling that accompanies synapse formation. They a lso suggest that one of the mechanisms through which this downregulation is achieved is an increased interaction of synaptophysin with synaptobrevin-V AMP2.