Aim: This study was designed to investigate whether the activation of the phosphatidylinositol 3-kinase (PI3-K)/Akt pathway is required for thermal pre-conditioning to protect rat cerebellar granule neurons (CGN) against apoptosis induced by low potassium, and to explore the possibility of a link between the upregulated heat shock protein (HSP)70 expression and Akt activation in the acquisition of neuroprotection induced by thermal preconditioning. Methods: CGN cultured for 8 d in vitro were switched to 5K medium for 24 h after thermal preconditioning (TP; 43.5 ℃ for 90 rain, then 37 ℃ for 1 h). To study the role of thePI3-K/Akt pathway, a PI3-K inhibitor, LY294002 (20 μmol/L) was added into the cultures 1 h before TP. 3-(4,5-dimethylthiazol-2-yl)2,5-diphenyltetrazolium bro-mide (MTT) assay and fluorescein diacetate staining were used to determine cell viability. Hoechst 33258 staining and agar gel electrophoresis were used to test the morphological and biological characters of CGN. Weste blot analysis was employed to detect the levels of phospho-Akt, phospho-glycogen synthase ki-nase 3β (GSK3β) Akt, GSK3β, and HSP70. Results: TP protected CGN against apoptosis induced by low potassium. LY294002 inhibited the neuroprotective effect on CGN induced by TP. TP induced a robust activation of Akt and the inactivation of GSK3β via PI3-K. Furthermore, the activation of the PI3-K/Aktpathway by TP persisted for 24 h in the 5K cultures. LY294002 (20 μmol/L) failed to inhibit the upregulated HSP70 expression induced by TP. Conclusion: The activation of the PI3-K/Akt pathway is required for TP to protect CGN against apoptosis induced by low potassium, but the neuroprotective effect by Akt activation is not mediated through the downstream induction of HSP70 expression.