Chloride channels in apical and basolateral membranes of CCD cells (RCCT-28A) in culture.
Previously, we found that isoproterenol activates whole cell Cl- conductance by a pathway involving adenosine 3',5'-cyclic monophosphate and protein kinase A (PKA) in a renal cell line (RCCT-28A) derived from the cortical collecting duct. The goal of the present study was to determine whether PKA activates Cl- channels in the apical and/or basolateral membrane. Using the patch clamp technique we found a 305-pS Cl- channel, described previously (22), located exclusively in the apical membrane and an outwardly rectifying Cl- channel (13/96 pS) located exclusively in the basolateral membrane. The outward rectifier was highly selective to Cl- versus cations, was inhibited by 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid and 5-nitro-2-(3-phenylpropylamino)-benzoic acid, but was not regulated by cytoplasmic pH or Ca2+. Neither isoproterenol nor PKA activated the 305-pS Cl- channel. In contrast, PKA activated a subset of outwardly rectifying channels in inside-out patches. In another subset of outwardly rectifying channels, formation of the inside-out configuration increased channel activity. These channels, however, were not sensitive to PKA. In conclusion, these experiments show that isoproterenol increases the Cl- conductance of RCCT-28A cells by activating a subset of outwardly rectifying Cl- channels located in the basolateral membrane.
Dietl
P
P
Department of Physiology, Dartmouth Medical School, Hanover, New Hampshire 03756.
Stanton
B A
BA
D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D018118:Chloride Channels; D002712:Chlorides / Q000378:metabolism; D004553:Electric Conductivity; D007685:Kidney Tubules, Collecting / Q000378:metabolism* / Q000648:ultrastructure; D008565:Membrane Proteins / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D011494:Protein Kinases / Q000494:pharmacology; D014018:Tissue Distribution
D018118:Chloride Channels; D002712:Chlorides; D008565:Membrane Proteins; D011494:Protein Kinases