The membrane-bound water channel aquaporin-4 (AQP4) plays a significant role in maintaining brain water homeostasis. In ischemic brain, changes in the expression level of AQP4 have been reported. Previous studies suggest that the internalization of several membrane-bound proteins, including AQP4, may occur with or without lysosomal degradation. In this study, the internalization of AQP4 was detected in the ischemic rat brain via double immunofluorescence labeling. Specifically, AQP4 and early endosome antigen-1 (EEA1) co-localized after 1 h post-ischemic injury. Moreover, the co-expression of AQP4 and lysosomal-associated membrane protein-1 (LAMP1) was observed after 3 h post-ischemia. These findings suggest that AQP4 is internalized and the lysosome is involved in degrading the internalized AQP4 in the ischemic brain. AQP4 is known to be downregulated by the protein kinase C activator phorbol 12-myristate 13-acetate (PMA) in vivo and in vitro. The results in this study displayed that PMA infusion could decrease brain edema accompanied by AQP4 downregulation in ischemic brain. However, compared with vehicle infusion, PKC activator infusion did not increase the ratio of internalized or lysosomal degraded AQP4. That is, we have not found out evidence to prove protein kinase C activator PMA can promote the internalization or lysosomal degradation of AQP4 in the ischemic brain. Crown Copyright (C) 2013 Published by Elsevier B.V. All rights reserved.