Enterotoxigenic Escherichia coli (ETEC) triggers diarrhea in humans and livestock. We have previously showed that ETEC promotes intestinal epithelial cell apoptosis and increases gamma-aminobutyric acid (GABA) concentration in the jejunum, suggesting that GABA might mediate ETEC-induced apoptosis. Here, we found that GABA alleviates ETEC-induced intestinal barrier dysfunctions, including ETEC-induced apoptosis both in vivo and in vitro. Interestingly, the alleviation of GABA on ETEC-induced apoptosis largely depends on autophagy. Mechanistically, GABA attenuates ETEC-induced apoptosis via activating GABA_AR signaling and the AMPK-autophagy pathway. These findings highlight that maintaining intestinal GABA concentration could alleviate intestinal ETEC infection.