Sustained depolarizing shift of the GABA reversal potential by glutamate receptor activation in hippocampal neurons

The inhibitory action of GABA is a consequence of a relatively hyperpolarized Cl − reversal potential ( E Cl), which results from the activity of K +–Cl − cotransporter (KCC2). In this study we investigated the effects of glutamate and glutamatergic synaptic activity on E Cl. In dissociated culture of mature hippocampal neurons, the application of glutamate caused positive E Cl shifts with two distinct temporal components. Following a large transient depolarizing state, the sustained depolarizing state ( E Cl- sustained) lasted more than 30 min. The E Cl- sustained disappeared in the absence of external Ca 2+ during glutamate application and was blocked by both AP5 and MK801, but not by nifedipine. The E Cl- sustained was also induced by NMDA. The E Cl- sustained was blocked by furosemide, a blocker of both KCC2 and NKCC1, but not bumetanide, a blocker of NKCC1. On the other hand, in immature neurons having less expression of KCC2, NMDA failed to induce the sustained depolarizing E Cl shift. In organotypic slice cultured neurons, repetitive activation of glutamatergic afferents also generated a sustained depolarizing E Cl shift. These results suggest that Ca 2+ influx through NMDA receptors causes the down-regulation of KCC2 and gives rise to long lasting positive E Cl shifts, which might contribute to hyperexcitability, LTP, and epileptiform discharges.