The α-subunit of the amiloride-sensitive epithelial Na+ channel (αENaC) is critical in forming an ion conductive pore in the membrane. We have identified the wild-type and three splice variants of the human αENaC (hαENaC) from the human lung cell line H441, using RT-PCR. These splice variants contain various structures in the extracellular domain, resulting in premature truncation (hαENaCx), 19-amino acid deletion (hαENaC-19), and 22- amino acid insertion (hαENaC+22). Wild-type hαENaC and splice variants were functionally characterized in Xenopus oocytes by coexpression with hENaC β- and γ-subunits. Unlike wild-type haENaC, undetectable or substantially reduced amiloride-sensitive currents were observed in oocytes expressing these splice variants. Wild-type hαENaC was the most abundantly expressed hαENaC mRNA species in all tissues in which its expression was detected. These findings indicate that the extracellular domain is important to generate structural and functional diversity of hαENaC and that alternative splicing may play a role in regulating hENaC activity.