The presence of astrocyte-to-neuron signal transmission has been suggested, but direct evidence has been lacking. Our previous research showed that astrocytic stimulation by photoactivation of channelrhodopsin-2 (ChR2) induces glutamate release and this astrocytic glutamate can modulate neuronal activity. However, the mechanism of its release remained unclear. In neuron, glutamate release is known to be released by calcium-dependent vesicular release. In contrast, ChR2-photoactivated astrocytic glutamate release was not affected by suppression of astrocytic intracellular calcium-increase. In astrocytes, factors other than calcium could be the actual trigger for glutamate release. It is known that ChR2 has much more permeability to H+ than Na+ or Ca2+. We found that lowering intracellular buffering capacity of pH produced stronger intracellular acidification in astrocytes in response to ChR2-photoactivation and this stronger acidification caused larger amount of glutamate release from astrocytes. This result suggests that acidification is a trigger for astrocytic glutamate release. We next studied the astrocytic contribution to excitotoxicity because the form of astrocyte-to-neuron signaling that we found would likely manifest most under pathological conditions. Brain ischemia is a situation that occurs upon cessation of blood supply to the brain. Two major events occur; acidosis and excess glutamate release. However, the cellular source of excess glutamate and the mechanism of its release have not been well determined. We simply hypothesized that the extreme acidification in astrocytes causes excess glutamate release. Optogenetic countering of astrocytic acidification with archaerhodopsin (ArchT) dramatically inhibited glutamate release from astrocytes upon brain ischemia. This result suggests that pH-dependent astrocytic glutamate release is one of the major cause of excitotoxicity. Previous studies have considered fluctuation of intracellular calcium as the only key signal that controls astrocytic activity. Here, we showed the importance of previously underrated signal, the intracellular pH.