Effects of alpha-adrenergic agonists on intracellular and intramitochondrial pH in rat proximal nephrons.
Rat proximal tubular segments were used to examine alpha-adrenoceptor alterations in Na+-H+ exchange by monitoring intracellular pH (pHi) and mitochondrial matrix pH (pHm). To obtain pHi, tubules were incubated with the cell-permeant fluorescent probe, 2',7'-bis(2-carboxyethyl)-5(6) carboxyfluorescein acetoxymethyl ester in a HCO3--free Na+ buffer. The intracellular distribution of the weak acid [2-14C] 5,5-dimethyloxazolidine-2,4-dione was used to calculate pHm, using values of medium pH, pHi, cell volume, and matrix content. Several selective alpha 1- and alpha 2-adrenoceptor agonists and the endogenous mixed agonist, norepinephrine, all produced dose-related increases in pHi. With each of the agonists tested, a maximum increase in pHi was observed at 1 microM final concentrations, with peak effects occurring in less than 1 min. Pretreatment with ethylisopropyl amiloride (EIPA, 10 microM), a specific inhibitor of proximal Na+-H+ exchange, blocked receptor-stimulated increases in pHi, as well as stimulation of Na+-H+ exchange by phorbol ester (PMA, 0.1 microM). Similarly, selective alpha 1- (prazosin, 0.1 microM) and alpha 2-(idazoxan, 0.1 microM) adrenoceptor antagonists inhibited alterations in agonist-induced pHi changes, whereas PMA-stimulated increases in pHi remained unaffected. Neither alpha 1- nor alpha 2-adrenoceptor agonists produced differences in pHm. Adrenoceptor agonist-induced pHi changes were also assessed at various concentrations of external Na+ (0-135 mM). It was observed that 0 and 10 mM external Na+ concentrations significantly reduced alpha 1- and alpha 2-adrenoceptor-stimulated pHi changes; Km values for the alpha 1-agonist phenylephrine and the alpha 2-agonist B-HT 933 were 18.0 +/- 2.1 and 22.7 +/- 2.6, respectively. In summary, stimulation by alpha-adrenergic agonists may be blocked at the receptor level with specific alpha-antagonists or at the exchanger with EIPA. The increase in cellular pH induced by these agonists is sensitive to external Na+ and reflects alpha-adrenoceptor activation of the Na+-H+ exchanger.
Gesek
F A
FA
Department of Internal Medicine, Medical College of Virginia, Virginia Commonwealth University, Richmond 23298-0160.
Schoolwerth
A C
AC
D000316:Adrenergic alpha-Agonists / Q000494:pharmacology*; D000584:Amiloride / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007687:Kidney Tubules, Proximal / Q000187:drug effects / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D009399:Nephrons / Q000187:drug effects / Q000378:metabolism*; D011224:Prazosin / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D017923:Sodium-Hydrogen Exchangers; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D000316:Adrenergic alpha-Agonists; D002352:Carrier Proteins; D017923:Sodium-Hydrogen Exchangers; D000584:Amiloride; D013755:Tetradecanoylphorbol Acetate; C039614:ethylisopropylamiloride; D011224:Prazosin