Astrocytes show dramatic changes at the molecular level in response to brain insult and disease, becoming reactive astrocytes. Among the changes found in reactive astrocytes, we have focused on P2Y1 receptor (P2Y1R), which is activated by extracellular ATP and is upregulated in reactive astrocytes in some neurological disorders including epilepsy. To reveal the pathophysiological significance of P2Y1R upregulation in astrocytes, we have used transgenic mice in which astrocytes specifically overexpress P2Y1R using Tet-Off system (P2Y1OE). P2Y1OE mice were more susceptible to drug-induced seizures and showed more abnormal spikes in EEG recordings, suggesting P2Y1OE triggered neuronal hyperexcitability. We analyzed the cellular mechanism underlying the hyper-excitability by imaging techniques and electrophysiology in the hippocampal slices. We found evidence showing that excitatory synaptic transmission was enhanced in P2Y1OE with the increase in P2Y1R-mediated Ca²⁺ signals in astrocytes. Interestingly, the enhancement of excitatory synaptic transmission was due to a novel excitatory molecule X derived from astrocytes rather than glutamate release from astrocytes. Overall, our data show a novel mechanism of astrocytic regulation of excitatory synapses which could contribute to hyperexcitability in neurological diseases.