H(+)-dependent ATPase and K+ channel activities in the rat thyroid cell strain FRTL-5.
Using the fluorescent indicators 2',7'-bis(2-carboxyethyl)-5'-(6')-carboxyfluorescein and Oxonol V to monitor intracellular pH (pHi) and cell membrane potential respectively, we have investigated the involvement of H(+)-dependent ATPase and H(+)-dependent K+ channels in the recovery of the rat thyroid cell strain FRTL-5 from experimentally induced cytosolic acidification and membrane hyperpolarization events. Following exposure of cells to the weak acid sodium butyrate (24 mmol/l) under bicarbonate-free incubation conditions, cytoplasmic acidification was maximal after 3 min, attaining a pHi of 6.42. The subsequent recovery of pHi was unimpaired by the absence of extracellular K+, but was reduced in the presence of the Na+ antagonist amiloride (1 mmol/l), recovering by 0.11 +/- 0.003 units, compared with 0.27 +/- 0.02 units under amiloride-free conditions. In the presence of the H(+)-dependent ATPase antagonist N,N'-dicyclohexylcarbodiimide (DCC), the pHi recovery observed in amiloride-containing, K(+)-free buffer was abolished. The recovery of pHi in Na(+)- and K(+)-containing buffer was accompanied by hyperpolarization of the cell membrane, the later stage of which was reduced after blockade of K+ channels with BaCl2, implying a major contribution of transmembrane K+ movement to such events. In contrast to its attenuating effect on pHi recovery, DCC was ineffective in reducing butyrate-dependent membrane hyperpolarization, suggesting that H(+)-dependent ATPase may not be a major contributory factor to this event. However, when K+ channels were blocked by addition of BaCl2, addition of DCC abolished the butyrate-induced membrane depolarization.(ABSTRACT TRUNCATED AT 250 WORDS)
Woods
D J
DJ
Department of Medicine, University of Manchester.
Soden
J
J
Bidey
S P
SP
D000818:Animals; D002460:Cell Line; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D008564:Membrane Potentials / Q000187:drug effects; D015221:Potassium Channels / Q000187:drug effects / Q000378:metabolism*; D006180:Proton-Translocating ATPases / Q000009:adverse effects / Q000378:metabolism*; D013961:Thyroid Gland / Q000187:drug effects / Q000378:metabolism*; D014634:Valinomycin / Q000494:pharmacology
D015221:Potassium Channels; D014634:Valinomycin; D004024:Dicyclohexylcarbodiimide; D006180:Proton-Translocating ATPases