The connexins are the protein subunits of the gap junction intercellular channels. In the present study a new rat connexin was cloned by degenerate reverse transcription-polymerase chain reaction and its gene isolated from a mouse genomic library. The nucleotide sequence encodes a protein of 321 amino acids (called Cx36) with highly significant homology to the members of the connexin family. In situ hybridization analysis of rat brain and retina showed the strongest expression in neurons of the inferior olive, the olfactory bulb, the CA3/CA4 hippocampal subfields and several brain-stem nuclei. An intense expression was also found in the pineal gland and in the retinal ganglion cell and inner nuclear layers. Experiments with neurotoxins, locally injected in the hippocampus or specifically acting on inferior olivary neurons, confirmed the neuronal localization of Cx36. It is the first connexin to be expressed predominantly in mammalian neurons and its identification paves the way for a molecular approach in the study of the role played by gap junctions in the physiology and the pathology of the mammalian brain.