Large-conductance Ca(2+)-activated K(+) (BK(Ca)) channels are widely distributed in cellular membranes of various tissues, but have not previously been found in cardiomyocytes. In this study, we cloned a gene encoding the mouse cardiac BK(Ca) channel alpha-subunit (mCardBKa). Sequence analysis of the cDNA revealed an open reading frame encoding 1154 amino acids. Another cDNA variant, identical in amino acid sequence, was also identified by sequence analysis. The nucleotide sequences of the two mCardBKa cDNAs, type 1 (mCardBKa1) and type 2 (mCardBKa2), differed by three nucleotide insertions and one nucleotide substitution in the N-terminal sequence. The amino acid sequence demonstrated that mCardBKa was a unique BK(Ca) channel alpha-subunit in mouse cardiomyocytes, with amino acids 41-1153 being identical to calcium-activated potassium channel SLO1 and amino acids 1-40 corresponding to BK(Ca) channel subfamily M alpha member 1. These findings suggest that a unique BK(Ca) channel alpha-subunit is expressed in mouse cardiomyocytes.