Identification of exchange mechanisms for the regulation of intracellular pH in rat thyroid FRTL-5 cells.
pH is maintained in cells by plasma membrane exchange mechanisms. In the absence of HCO3- ions, FRTL-5 cells regulate intracellular pH (pHi) by an Na+/H+ antiport but HCO3(-)-dependent exchangers cannot operate. We have investigated pHi regulation (by microfluorimetry and the pH sensitive dye 2',7'-bis(2-carboxyethyl)-5(6')-carboxyfluorescein) in small groups (five to six cells) of FRTL-5 thyroid cell monolayers held in Krebs-Ringer buffer (pH 7.4) with or without HCO3- ions. The exchangers were investigated with inhibitors (amiloride or its derivative dimethylamiloride for the Na+/H+ antiporter and the stilbene derivative disodium 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS) for HCO3(-)-dependent mechanisms), ionic substitution and by NH4+/NH3 (10 mM) acid loading. Basal pHi was lower in the presence (7.3 +/- 0.058, mean +/- S.D., n = 14) than in the absence (7.59 +/- 0.078, n = 10) of HCO3- ions. In HCO3(-)-free media, cells recovered from acid load by 0.34 +/- 0.04 pH units in the first 2 min and finally reached a pHi of 7.35 +/- 0.06. This recovery was Na(+)-dependent and blocked by dimethylamiloride during the 15 min following intracellular acidification. In HCO3(-)-containing media, cells recovered from an acid load at a similar rate, but reached 99 +/- 10% (n = 9) of the baseline pH; this recovery was also dependent on Na+ ions. Moreover, although dimethylamiloride and DIDS reduced the rate of recovery to 0.06 +/- 0.02 and 0.18 +/- 0.04 pH units respectively during the 2-min period, the cells returned to the basal pHi within 15 min.(ABSTRACT TRUNCATED AT 250 WORDS)
Wood
A M
AM
Department of Medicine (Clinical Biochemistry), Hope Hospital, Salford, U.K.
Warhurst
G
G
Bidey
S P
SP
Soden
J
J
Taylor
R
R
Robertson
W R
WR
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D001639:Bicarbonates / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007424:Intracellular Fluid / Q000378:metabolism; D007474:Ion Exchange; D051381:Rats; D017923:Sodium-Hydrogen Exchangers; D013961:Thyroid Gland / Q000187:drug effects / Q000378:metabolism*
D001639:Bicarbonates; D002352:Carrier Proteins; D017923:Sodium-Hydrogen Exchangers; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D000584:Amiloride; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid