Bumetanide-sensitive ion fluxes in vascular smooth muscle cells: lack of functional Na+, K+, 2 Cl- cotransport.
To examine the involvement of Na+,K+,2Cl- cotransport in monovalent ion fluxes in vascular smooth muscle cells (VSMC), we compared the effect of bumetanide on 86Rb, 36Cl and 22Na uptake by quiescent cultures of VSMC from rat aorta. Under basal conditions, the values of bumetanide-sensitive (BS) inward and outward 86Rb fluxes were not different. Bumetanide decreased basal 86Rb uptake by 70-75% with a Ki of approximately 0.2-0.3 microM. At concentrations ranging up to 1 microM, bumetanide did not affect 36Cl influx and reduced it by 20-30% in the range from 3 to 100 microM. In contrast to 86Rb and 36Cl influx, bumetanide did not inhibit 22Na uptake by VSMC. BS 86Rb uptake was completely abolished in Na(+)- or Cl(-)-free media. In contrast to 86Rb, basal BS 36Cl influx was not affected by Nao+ and Ko+. Hyperosmotic and isosmotic shrinkage of VSMC increased 86Rb and 36Cl influx to the same extent. Shrinkage-induced increments of 86Rb and 36Cl uptake were completely abolished by bumetanide with a Ki or approximately 0.3 microM. Shrinkage did not induce BS 86Rb and 36Cl influx in (Na+ or Cl-)- and (Na+ or K+)-depleted media, respectively. In the presence of an inhibitor of Na+/H+ exchange (EIPA), neither hyperosmotic nor isosmotic shrinkage activated 22Na influx. Bumetanide (1 microM) did not modify basal VSMC volume and intracellular content of sodium, potassium and chloride but abolished the regulatory volume increase in isosmotically-shrunken VSMC. These data demonstrate the absence of the functional Na+,K+,2Cl- cotransporter in VSMC and suggest that in these cells basal and shrinkage-induced BS K+ influx is mediated by (Nao+ + Clo-)-dependent K+/K+ exchange and Nao(+)-dependent K+,Cl- cotransport, respectively.
Orlov
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SN
Céntre de Recherche Hôtel-Dieu de Montréal, Université de Montréal, Québec, Canada.
Tremblay
J
J
Hamet
P
P
D000818:Animals; D002034:Bumetanide / Q000494:pharmacology*; D002712:Chlorides / Q000493:pharmacokinetics; D004305:Dose-Response Relationship, Drug; D017136:Ion Transport; D007477:Ions; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000187:drug effects* / Q000378:metabolism; D009995:Osmosis; D051381:Rats; D011914:Rats, Inbred BN; D012414:Rubidium Radioisotopes / Q000493:pharmacokinetics; D012979:Sodium Radioisotopes / Q000493:pharmacokinetics; D014867:Water
D002712:Chlorides; D007477:Ions; D012414:Rubidium Radioisotopes; D012979:Sodium Radioisotopes; D014867:Water; D002034:Bumetanide