Background Astrocyte swelling is an important consequence of hepatic encephalopathy, and aquaporin-4 has been reported to play a vital role in this swelling. Ammonia causes astrocyte swelling and is also known to modulate aquaporin-4 expression in the astrocyte foot processes. The purpose of this study was to explore the mechanism of ammonia-induced aquaporin-4 expression, which has been suggested to involve the p38 mitogen-activated protein kinase pathway.Methods We exposed cultured astrocytes to ammonium chloride, an in vitro model of hepatic encephalopathy. The purity of cultured astrocytes was evaluated by fluorescent glial fibrillary acidic protein labeling; cell morphology was assessed by light microscopy; the expression of aquaporin-4, phospho-p38, and p38 were detected by Western blotting analysis. Statistical analysis was performed by one-way factorial analysis of variance, and the relationship between variables was calculated by linear regression using SPSS version 13.0 program for Windows (SPSS, Chicago, IL, USA).Results The purity of cultured astrocytes was (96.6 ±1.4)%. Astrocytes swelled significantly when exposed to 5 mmol/L ammonium chloride for 24 hours as compared to non-exposed astrocytes. Co-treatment with 10 μmol/L SB203580 (an inhibitor of p38) attenuated the degree of ammonium chloride induced astrocyte swelling. Western blotting analysis revealed that the expression levels of phospho-p38 and aquaporin-4 in ammonium chloride treated cells were significantly increased relative to the control group (P