The amiloride-sensitive epithelial sodium channel (ENaC) is composed of three subunits: α, β, and γ. The human α-ENaC subunit is expressed as at least two transcripts (N. Voilley, E. Lingueglia, G. Champigny, M. G. Mattei, R. Waldmann, M. Lazdunski, and P. Barbry. Proc. Natl. Acad. Sci. USA91: 247–251, 1994). To determine the origin of these transcripts, we characterized the 5′ end of the α-ENaC gene. Four transcripts that differ at their first exon were identified. Exon 1A splices to exon 2 to form the 5′ end of α-ENaC1, whereas exon 1B arises separately and continues into exon 2 to form α-ENaC2. Other variant mRNAs, α-ENaC3 and α-ENaC4, are formed by activating 5′ splice sites within exon 1B. Although α-ENaC3 and -4 did not change the open reading frame for α-ENaC, α-ENaC2 contains upstream ATGs that add 59 amino acids to the previous (α-ENaC1) protein. To address the significance of these isoforms, both proteins were expressed in Xenopus oocytes. The cRNA for each α-ENaC transcript when combined with β- and γ-ENaC cRNA reconstituted a low-conductance ion channel with amiloride-sensitive currents of similar characteristics. We have thus identified variant α-ENaC mRNAs that lead to functional ENaC peptides.