ATP-sensitive potassium channels in capillaries isolated from guinea-pig heart

1. The full-length cDNAs of two different alpha-subunits (Kir6.1 and Kir6.2 ) and partial cDNAs of three different beta-subunits (SUR1, SUR2A and SUR2B ) of ATP-sensitive potassium (K-ATP) channels of the guinea-pig (gp) were o btained by screening a cDNA library from the ventricle of guinea-pig heart. 2. Cell-specific reverse-transcriptase PCR with gene-specific intron-spanni ng primers showed that gpKir6.1, gpKir6.2 and gpSUR2B were expressed in a p urified fraction of capillary endothelial cells. In cardiomyocytes, gpKir6. 1, gpKir6.2, gpSUR1. and gpSUR2A were detected. 3. Patch-clamp measurements were carried out in isolated capillary fragment s consisting of 3-15 endothelial cells. The membrane capacitance measured i n the whole-cell mode was 19.9 +/- 1.0 pF and was independent of the length of the capillary fragment, which suggests that the endothelial cells were not electrically coupled under our experimental conditions. 4. The perforated-patch technique was used to measure the steady-state curr ent-voltage relation of capillary endothelial cells. Application of K+ chan nel openers (rilmakalim or diazoxide) or metabolic inhibition (250 mu M 2,4 -dinitrophenol plus 10 mM deoxyglucose) induced a current that reversed nea r the calculated K+ equilibrium potential. 5. Rilmakalim (1 mu M), diazoxide (300 mu M) and metabolic inhibition incre ased the slope conductance measured at -55 mV by a factor of 9.0 (+/- 1.8), 2 5 (+/- 0.2) and 3.9 (+/- 1.7), respectively. The effects were reversed b y glibenclamide (1 mu M). 6. Our results suggest that capillary endothelial cells from guinea-pig hea rt express K-ATP channels composed of SUR2B and Kir6.1 and/or Kir6.2 subuni ts. The hyperpolarization elicited by the opening of K-ATP channels may lea d to an increase in free cytosolic Ca2+, and thus modulate the synthesis of NO and the permeability of the capillary wall.