Large-conductance calcium- and voltage- activated potassium (BK) channels play a fundamental role in the signaling pathways regulating mouse anterior pituitary corticotrope function. Here we describe the cloning and functional characterization of the components of mouse corticotrope BK channels. RT-PCR cloning and splice variant analysis of mouse AtT20 D16:16 corticotropes revealed robust expression of mslo transcripts encoding pore-forming alpha-subunits containing the mouse homolog of the 59-amino acid STREX-1 exon at splice site 2. RT-PCR and functional analysis, using the triterpenoid glycoside, DHS-1, revealed that native corticotrope BK channels are not functionally coupled to beta-subunits in vivo. Functional expression of the STREX-1 containing alpha-subunit in HEK 293 cells resulted in BK channels with calcium sensitivity, single-channel conductance, and inhibition by protein kinase A identical to that of native mouse corticotrope BK channels. This report represents the first corticotrope ion channel to be characterized at the molecular level and demonstrates that mouse corticotrope BK channels are composed of alpha-subunits expressing the mouse STREX-1 exon.