ATP-sensitive potassium channels in capillaries isolated from guinea-pig heart.
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 (KATP) channels of the guinea-pig (gp) were obtained by screening a cDNA library from the ventricle of guinea-pig heart. Cell-specific reverse-transcriptase PCR with gene-specific intron-spanning primers showed that gpKir6.1, gpKir6.2 and gpSUR2B were expressed in a purified fraction of capillary endothelial cells. In cardiomyocytes, gpKir6.1, gpKir6.2, gpSUR1 and gpSUR2A were detected. Patch-clamp measurements were carried out in isolated capillary fragments consisting of 3-15 endothelial cells. The membrane capacitance measured in 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. The perforated-patch technique was used to measure the steady-state current-voltage relation of capillary endothelial cells. Application of K+ channel openers (rilmakalim or diazoxide) or metabolic inhibition (250 microM 2,4-dinitrophenol plus 10 mM deoxyglucose) induced a current that reversed near the calculated K+ equilibrium potential. Rilmakalim (1 microM), diazoxide (300 microM) and metabolic inhibition increased 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 by glibenclamide (1 microM). Our results suggest that capillary endothelial cells from guinea-pig heart express KATP channels composed of SUR2B and Kir6.1 and/or Kir6.2 subunits. The hyperpolarization elicited by the opening of KATP channels may lead to an increase in free cytosolic Ca2+, and thus modulate the synthesis of NO and the permeability of the capillary wall.
Mederos y Schnitzler
M
M
Institut fur Normale und Pathologische Physiologie, Universitat Marburg, Deutschhausstrasse 2, D-35037 Marburg, Germany.
Derst
C
C
Daut
J
J
Preisig-Müller
R
R
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D000963:Antimetabolites / Q000494:pharmacology; D001483:Base Sequence; D002196:Capillaries / Q000378:metabolism; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D004730:Endothelium, Vascular / Q000378:metabolism; D006168:Guinea Pigs; D066298:In Vitro Techniques; D008564:Membrane Potentials / Q000187:drug effects; D009206:Myocardium / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D014018:Tissue Distribution
D000963:Antimetabolites; D017931:DNA Primers; D018076:DNA, Complementary; D015221:Potassium Channels; D012333:RNA, Messenger; D000255:Adenosine Triphosphate