In CA1 pyramidal neurons of the hippocampus protein kinase C regulates calcium-dependent inactivation of NMDA receptors

Abstract

The NMDA subtype of the glutamate-gated channel exhibits a high permeability to Ca2+. The influx of Ca2+ through NMDA channels is limited by a rapid and Ca2+/calmodulin (CaM)-dependent inactivation that results from a competitive displacement of cytoskeleton-binding proteins from the NR1 subunit of the receptor by Ca2+/CaM (Zhang et al., 1998; Krupp et al., 1999). The C terminal of this subunit can be phosphorylated by protein kinase C (PKC) (Tingley et al., 1993). The present study sought to investigate whether PKC regulates Ca2+-dependent inactivation of the NMDA channel in hippocampal neurons. Activation of endogenous PKC by 4β-phorbol 12-myristate 13-acetate enhanced peak (I(p)) and depressed steady-state (I(ss)) NMDA- evoked currents, resulting in a reduction in the ratio of these currents (I(ss)/I(p)). We demonstrated previously that PKC activity enhances I(p) via a sequential activation of the focal adhesion kinase cell adhesion kinase β/proline-rich tyrosine kinase 2 (CAKβ/Pyk2) and the nonreceptor tyrosine kinase Src (Huang et al., 1999; Lu et al., 1999). Here, we report that the PKC-induced depression of I(ss) is unrelated to the PKC/CAKβ/Src-signaling pathway but depends on the concentration of extracellular Ca2+. Intracellular applications of CaM reduced I(ss)/I(p) and occluded the Ca2+- dependent effect of phorbol esters on I(ss). Moreover, increasing the concentration of intracellular Ca2+ buffer or intracellular application of the inhibitory CaM-binding peptide (KY9) greatly reduced the phorbol ester- induced depression of I(ss). Taken together, these results suggest that PKC enhances Ca2+/CaM-dependent inactivation of the NMDA channel, most likely because of a phosphorylation-dependent regulation of interactions between receptor subunits, CaM, and other postsynaptic density proteins.