Pannexins are a newly described family of proteins that may form gap junctions. We made antisera against mouse pannexin1 (Panx1). HeLa cells expressing Panx1 have cell surface labeling, but not gap junction plaques, and do not transfer small fluorescent dyes or neurobiotin in a scrape-loading assay. Neuro2a cells expressing Panx1 are not electrophysiologically coupled. Intracellular Panx1-immunoreactivity, but not gap junction plaques, is seen in cultured oligodendrocytes, astrocytes, and hippocampal neurons. Thus, at least in these mammalian cells lines, Panx1 does not form morphological or functional gap junctions, and it remains to be demonstrated that Panx1 forms gap junction-forming protein in the CNS.