Synergistic control of protein kinase Cγ activity by ionotropic and metabotropic glutamate receptor inputs in hippocampal neurons

Conventional protein kinase C (PKC) isoforms are abundant neuronal signaling proteins with important roles in regulating synaptic plasticity and other neuronal processes. Here, we investigate the role of ionotropic and metabotropic glutamate receptor (iGluR and mGluR, respectively) activation on the generation of Ca2+ and diacylglycerol (DAG) signals and the subsequent activation of the neuron-specific PKC gamma isoform in hippocampal neurons. By combining Ca2+ imaging with total internal reflection microscopy analysis of specific biosensors, we show that elevation of both Ca2+ and DAG is necessary for sustained translocation and activation of EGFP (enhanced green fluorescent protein)-PKC gamma. Both DAG production and PKC gamma translocation were localized processes, typically observed within discrete microdomains along the dendritic branches. Markedly, intermediate-strengthNMDAreceptor (NMDAR) activation or moderate electrical stimulation generated Ca2+ but no DAG signals, whereas mGluR activation generated DAG but no Ca2+ signals. Both receptors were needed for PKC gamma activation. This suggests that a coincidence detection process exists between iGluRs and mGluRs that relies on a molecular coincidence detection process based on the corequirement of Ca2+ and DAG for PKC gamma activation. Nevertheless, the requirement for costimulation with mGluRs could be overcome for maximal NMDAR stimulation through a direct production of DAG via activation of the Ca2+-sensitive PLC delta (phospholipase C delta) isoform. In a second important exception, mGluRs were sufficient for PKC gamma activation in neurons in which Ca2+ stores were loaded by previous electrical activity. Together, the dual activation requirement for PKC gamma provides a plausible molecular interpretation for different synergistic contributions of mGluRs to long-term potentiation and other synaptic plasticity processes.