Methamphetamine induces GSDME-dependent cell death in hippocampal neuronal cells through the endoplasmic reticulum stress pathway

Methamphetamine (METH) is an illegal amphetamine-typed psychostimulant that is abused worldwide and causes serious public health problems. METH exposure induces apoptosis and autophagy in neuronal cells. However, the role of pyroptosis in METH-induced neurotoxicity is still unclear. Here, we investigate whether pyroptosis is involved in METH-induced hippocampal neurotoxicity and the potential mechanisms of Endoplasmic reticulum (ER) stress in hippocampal neuronal cells. For this purpose, the expression levels of pyroptosis-related proteins, GSDMD and GSDME, were analyzed by immunoblotting and immunohistochemistry in the hippocampal neuron cell line HT-22. Next, we explored METH-induced pyroptosis in HT-22 using immunoblotting, LDH assays and SYTOX green acid staining. Further, the relationship between pyroptosis and ER stress in METH-induced hippocampal neuron damage was studied in HT-22 cells using inhibitors including TUDCA, a specific inhibitor of ER stress, GSK-2656157, a PERK pathway inhibitor and STF-0803010, an inhibitor of IRE1 alpha endoribonuclease activity. This relationship was also studied using siRNAs, including siTRAF2, an siRNA against IRE1 alpha kinase activity and siATF6 against the ATF6 pathway, which were analyzed by immunoblotting, LDH assays and SYTOX green acid staining. GSDME but not GSDMD was found to be expressed in HT-22 cells. METH treatment induced the upregulation of cleaved GSDME-NT and LDH release, as well as the increase of SYTOX green positive cells in HT-22 cells, which was partly reversed by inhibitors and siRNAs, indicating that the ER stress signaling pathway was involved in GSDME-dependent cell death induced by METH. In summary, these results revealed that METH induced ER stress that mediated GSDME-dependent cell death in hippocampal neuronal cells. These findings provide novel insight into the mechanisms of METH-induced neurotoxicity.