Na(+)-H+ exchanger kinetics in adrenal glomerulosa cells and its activation by angiotensin II.
We have studied the kinetic properties of basal and angiotensin II (ANG II) stimulated Na(+)-H+ exchange in adrenal glomerulosa cells by measuring changes in cytosolic pH (pHi) and initial rates of 22Na uptake in the presence or absence of dimethylamiloride (DMA). The cells were studied 1) under basal conditions, 2) at constant pHi (6.8) with varied external sodium (Na+o), and 3) at varied pHi with constant Na+o (50 mM). In 2,7-biscarboxyethyl-5(6)-carboxyfluorescein loaded cells under basal conditions, pHi rose from 7.09 +/- 0.02 to 7.19 +/- 0.02 (P less than 0.05) with addition of ANG II (100 nM). Similarly, DMA-sensitive Na influx was enhanced from 9.2 +/- 1.3 to 14.8 +/- 2.1 nmol Na+/mg protein x min (P less than 0.01) by ANG II. In cells acid-loaded by preincubation in Na(+)-free media (pHi 6.8), addition of varying Na+o resulted in a rapid H+ efflux that was markedly inhibited by DMA. DMA-sensitive Na+ influx into these acidified cells with varied Na+o exhibited a Michaelis-Menten constant (Km) of 23 mM and a maximum velocity (Vmax) of 43 nmol Na+/mg protein x min. By varying pHi (from pHi 7.1 to 6.2), DMA-sensitive Na+ influx likewise showed activation with cellular acidification with a pK at pHi 7.09. At pHi 6.8, ANG II decreased the Km for Na+o from 23 to 17 mM and increased the Vmax from 43 to 53 nmol Na+/mg protein x min. The pHi dependence of DMA-sensitive Na+ influx was not affected by ANG II (pK at pHi 7.03). DMA also inhibited AII-stimulated aldosterone secretion and Na+ influx similarly. These results indicate that Na(+)-H+ exchange in adrenal glomerulosa cells is 1) functioning under basal conditions, and 2) is modulated by ANG II with enhanced Na+o affinity and Vmax but without a shift in pHi dependence (similar to ANG II effects on vascular smooth muscle cells). These effects suggest an important role for Na(+)-H+ exchange during ANG II stimulation of aldosterone production by glomerulosa cells.
Conlin
P R
PR
Department of Medicine, Brigham and Women's Hospital, Boston, Massachusetts 02115.
Kim
S Y
SY
Williams
G H
GH
Canessa
M L
ML
D000450:Aldosterone / Q000378:metabolism; D000584:Amiloride / Q000031:analogs & derivatives / Q000494:pharmacology; D000804:Angiotensin II / Q000494:pharmacology*; D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D002417:Cattle; D005452:Fluoresceins; D005456:Fluorescent Dyes; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D012964:Sodium / Q000378:metabolism / Q000494:pharmacology; D012979:Sodium Radioisotopes; D017923:Sodium-Hydrogen Exchangers; D015384:Zona Glomerulosa / Q000187:drug effects / Q000378:metabolism*
D002352:Carrier Proteins; D005452:Fluoresceins; D005456:Fluorescent Dyes; D012979:Sodium Radioisotopes; D017923:Sodium-Hydrogen Exchangers; D000804:Angiotensin II; C039465:5-dimethylamiloride; D000450:Aldosterone; D000584:Amiloride; C043829:2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein; D012964:Sodium