Downregulation of GABA Receptor Recycling Mediated by HAP1 Contributes to Neuronal Death in In Vitro Brain Ischemia.

Downregulation of GABAergic synaptic transmission contributes to the increase in overall excitatory activity in the ischemic brain. A reduction of GABA receptor (GABAR) surface expression partly accounts for this decrease in inhibitory activity, but the mechanisms involved are not fully elucidated. In this work, we investigated the alterations in GABAR trafficking in cultured rat hippocampal neurons subjected to oxygen/glucose deprivation (OGD), an in vitro model of global brain ischemia, and their impact in neuronal death. The traffic of GABAR was evaluated after transfection of hippocampal neurons with myc-tagged GABAR β3 subunits. OGD decreased the rate of GABAR β3 subunit recycling and reduced the interaction of the receptors with HAP1, a protein involved in the recycling of the receptors. Furthermore, OGD induced a calpain-mediated cleavage of HAP1. Transfection of hippocampal neurons with HAP1A or HAP1B isoforms reduced the OGD-induced decrease in surface expression of GABAR β3 subunits, and HAP1A maintained the rate of receptor recycling. Furthermore, transfection of hippocampal neurons with HAP1 significantly decreased OGD-induced cell death. These results show a key role for HAP1 protein in the downmodulation of GABAergic neurotransmission during cerebral ischemia, which contributes to neuronal demise.