Voltage-gated sodium channels are members of a multigene family of transmembrane proteins that are important determinants of electrical excitability in cell membranes. These proteins are typically composed of a large alpha-subunit and one or two beta-subunits. The primary structure of alpha-subunits is highly conserved among different subtypes and different species. Based on the conserved sequences and application of the rapid amplification of cDNA ends (RACE) reaction, we have isolated three overlapping clones from human brain. These sequences share highest homology (89%) to the rat brain subtype III gene and cover a 4.2-kb expanse of the transcript. The 5'-most clone has a translation start site located in the same region as other mammalian brain sodium channel genes. A 92-nucleotide insert was found in domain I at a location previously demarcated by published splice sites in rat brain sodium channels IIN/IIA and IIIN/IIIA. It is most likely that this transcript represents the two isoforms (neonatal and adult) of the human brain sodium channel gene, SCN3A (GenBank accession numbers AF035685 and AF035686). As is the case for rat brain sodium channels IIN/IIA and IIIN/IIIA, these isoforms are generated through an alternative splicing mechanism. The conservation of the exon structure suggests that alternative RNA splicing is a common feature for sodium channel mRNA processing and may play an important role in modulating the channel function.