Freshly isolated bovine coronary endothelial cells do not express the BKCa channel gene

Recent reports have suggested that different types of Ca2+-activated K+ channels may be selectively expressed either in the vascular endothelial cells (ECs) or smooth muscle cells (SMCs) of a single artery. In this study, we directly compared mRNA, protein and functional expression of the high-conductance Ca2+-activated K+ (BKCa) channel between freshly isolated ECs and SMCs from bovine coronary arteries. Fresh ECs and SMCs were enzymatically isolated, and their separation verified by immunofluorescent detection of alpha-actin and platelet/endothelium cell adhesion molecule (PECAM) proteins, respectively. Subsequently, studies using a sequence-specific antibody directed against the pore-forming a-subunit of the BKCa channel only detected its expression in the SMCs, whereas PECAM-positive ECs were devoid of the alpha-subunit protein. Additionally, multicell RTPCR performed using cDNA derived from either SMCs or ECs only detected mRNA encoding the BKCa alpha-subunit in the SMCs. Finally, whole-cell recordings of outward K+ current detected a prominent iberiotoxin-sensitive BKCa current in SMCs that was absent in ECs, and the BKCa channel opener NS 1619 only enhanced K+ current in the SMCs. Thus, bovine coronary SMCs densely express BKCa channels whereas adjacent ECs in the same artery appear to lack the expression of the BKCa channel gene. These findings indicate a cell-specific distribution of Ca2+-activated K+ channels in SMCs and ECs from a single arterial site.