Upregulation of gap junction connexin 32 with epileptiform activity in theisolated mouse hippocampus

Gap junctions which serve as intercellular channels providing direct cytopl asmic continuity and ionic current flow between adjacent cells, are constit uted by connexin proteins. Using an in vitro model of bicuculline-induced e pileptiform activity, we asked whether increased connexin levels occur duri ng epileptiform activity in the intact whole hippocampus, freshly isolated from young (15-day-old) mouse brain. Exposure to bicuculline (10 muM), for 2-10 h, induced persistent changes in electrical activities that included e nhanced spontaneous field activity (4 h), an epileptiform response to singl e electrical stimulation (6 h), and spontaneous epileptiform activity (6 h) . These electrophysiological changes were not reversed by up to 60 nun perf usion with normal artificial cerebrospinal fluid, but were greatly depresse d by the gap junction uncoupler, carbenoxolone (120 muM, 10 min). Data from RNase protection assay and immunoblotting showed that among several detect ed gap junctions, only connexin 32 was affected. After 2-6 h exposure to bi cuculline, the connexin 32 mRNA expression was upregulated to 2-3-fold cont rol (P < 0.01), and its protein level was significantly elevated the follow ing 6 h (P < 0.01), at which time electrophysiologically measured evidence of clearly epileptiform activity was apparent. In addition, the transcripti on factor, c-fos protein, but not the cAMP response element-binding protein , was also found to be increased at the early stage of bicuculline exposure (2 h) compared to control (P < 0.05). Thus we have found that exposing the acutely isolated hippocampus to bicucu lline, induced increased c-fos protein, followed by increased connexin 32 t ranscript and protein, and concurrently, persistent epileptiform activity t hat was depressed by carbenoxolone. (C) 2001 IBRO. Published by Elsevier Sc ience Ltd. All rights reserved.