Glioma cells release glutamate causing neuronal excitotoxicity and promoting proliferation and invasion. Over-activation of GluR in the peritumoral brain parenchyma may explain the frequent occurrence of seizures in glioma patients. However, it has not been demonstrated that seizures are caused by tumor released glutamate. Among factors contributing to epileptogenesis, the alteration of GABA reversal potential (EGABA), is considered relevant for neuronal hyperexcitability. We measured EGABA, by perforated patch clamp recordings in cultured hippocampal neurons in control and following coculture with MZC glioma cells. Coculture caused a significant positive shift of EGABA, from -70 to -60 mV, without affecting the neuronal resting potential, suggesting reduced neuronal inhibition and a misregulation of Cl transporters activity. As KCC2 activity is inhibited by intracellular zinc, we reduced [Zn2+]i in neurons using TPEN. In this condition, glioma coculture did not affect EGABA, indicating that the shift might arise from inhibition of KCC2 by [Zn2+]i. APV and NBQX treatment prevented EGABA positive shift induced by coculture, suggesting that [Zn2+]i increase is due to GluR activation. In neurons transfected with a CFP-YFP Cl-Sensor, glioma conditioned medium caused a rise in [Cl]i, strongly reduced by GluR antagonists. Our data suggest that glioma cells might alter neuronal Cl homeostasis by releasing glutamate and inhibiting KCC2 through intracellular Zn2+ rise.