Radiotracer studies of cystic fibrosis transmembrane conductance regulator expressed in Xenopus oocytes.
We measured fluxes of radiotracers in Xenopus oocytes expressing the cystic fibrosis transmembrane conductance regulator (CFTR). Addition of adenosine 3',5'-cyclic monophosphate (cAMP)-elevating agents [forskolin and 3-isobutyl-1-methylxanthine (I/F)] led to large increases in uptake of 36Cl, 125I, and 82Br into oocytes expressing wild-type CFTR or delta F508 CFTR but not sham-injected oocytes. I/F also stimulated halide efflux from CFTR and delta F508 oocytes in the sequence Cl > Br > I. cAMP-induced increases in 36Cl efflux from delta F508 oocytes were approximately 20% of those in CFTR oocytes. Increases in halide efflux were blocked by diphenylamine-2-carboxylic acid but not by 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid. The phorbol ester, phorbol 12-myristate 13-acetate, also stimulated 36Cl efflux from CFTR oocytes. ATP uptakes into CFTR and sham oocytes were similar, and both were reduced by I/F. However, ATP uptake into I/F-treated CFTR oocytes was slightly greater (approximately 40%) than into I/F-treated sham oocytes. Urea uptake into CFTR and sham oocytes was similar and in both cases was increased by I/F. However, the I/F-induced increase in urea uptake into CFTR oocytes was significantly greater than for sham oocytes. I/F stimulated formate uptake into CFTR oocytes but not into sham oocytes. Fluxes of 22Na, 86Rb, 35SO4, 32PO4, and mannitol were unaltered by expression and activation of CFTR.
Ohrui
T
T
Department of Physiology, University of California, San Francisco 94143.
Skach
W
W
Thompson
M
M
Matsumoto-Pon
J
J
Calayag
C
C
Widdicombe
J H
JH
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000493:pharmacokinetics; D000818:Animals; D000242:Cyclic AMP / Q000502:physiology; D003550:Cystic Fibrosis / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D004573:Electrolytes / Q000378:metabolism; D005561:Formates / Q000493:pharmacokinetics; D008353:Mannitol / Q000493:pharmacokinetics; D008565:Membrane Proteins / Q000378:metabolism*; D009865:Oocytes; D011868:Radioisotopes; D012413:Rubidium / Q000493:pharmacokinetics; D013431:Sulfates / Q000493:pharmacokinetics; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014981:Xenopus
D004573:Electrolytes; D005561:Formates; D008565:Membrane Proteins; D011868:Radioisotopes; D013431:Sulfates; C030544:formic acid; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D008353:Mannitol; D000255:Adenosine Triphosphate; D000242:Cyclic AMP; D012413:Rubidium; D013755:Tetradecanoylphorbol Acetate; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid