PROTEIN-KINASE-C ACTIVATION ATTENUATES N-METHYL-D-ASPARTATE-INDUCED INCREASES IN INTRACELLULAR CALCIUM IN CEREBELLAR GRANULE CELLS

Activation of the N-methyl-D-aspartate (NMDA) subtype of glutamate rec eptor increases levels of intracellular calcium and can lead to stimul ation of protein kinase C activity. Several reports have demonstrated that stimulation of protein kinase C can, in turn, increase electrophy siological responses to NMDA in certain cells or in oocytes expressing certain NMDA receptor subunits. In the present study, the effects of protein kinase C activation on NMDA receptor-mediated increases in int racellular Ca2+ levels were investigated in primary cultures of rat ce rebellar granule cells using fura-2 fluorescence spectroscopy. Pretrea tment of the cells with the protein kinase C activator phorbol 12-myri state 13-acetate (PMA), but not the inactive analogue 4 alpha-phorbol 12-myristate 13-acetate, inhibited NMDA-induced increases in intracell ular Ca2+ levels. Coincubation of cells with PMA and the kinase inhibi tor staurosporine or calphostin C blocked the PMA effect. The potency of NMDA was reduced twofold, and the potency of the NMDA receptor coag onist, glycine, to enhance the response to NMDA was decreased fourfold by pretreatment of cells with PMA. The effect on glycine was mimicked by pretreatment with okadaic acid, a protein phosphatase inhibitor. P MA treatment did not significantly alter Mg2+ inhibition of the NMDA r esponse but decreased the potency of the competitive antagonist CGS-19 755. These data suggest that, in cerebellar granule cells, the functio n of the NMDA receptor may be subject to feedback inhibition by protei n kinase C stimulation. Under physiological conditions, this inhibitio n may result from a decreased effectiveness of the endogenous coagonis ts, glutamate and glycine.