Amiloride-sensitive Na+/H+ exchange in rat NB2 node lymphoma cells. Stimulation by prolactin and other mitogens

Human PRL (hPRL) stimulated an amiloride-sensitive Na+/H+ exchange system, measured as extracellular acidification or rate of H+ efflux, uptake of 22Na+, and intracellular alkalinization (pHi) in rat Nb2 node lymphoma cells. The hPRL-induced increase in H+ efflux rate was dependent on dose, time, and the concentration of extracellular Na+. From 5-100 ng/ml, the hPRL-induced increase in H+ efflux rate was measurable by 3 min and reached a dose-dependent maximum by 15 min. A higher final H+ efflux rate was achieved with a higher concentration of hPRL. Complete replacement of Na+ by choline chloride gave a marginal increase in the rate of H+ efflux upon addition of hPRL. At a maximum noncytotoxic concentration (25 microM), the potent amiloride analog, 5-(N-ethyl-N-isopropyl)amiloride (EP-Am), decreased but did not abolish, the stimulatory effect of hPRL. The hPRL-stimulated increase in H+ efflux rate was accompanied by an EP-Am-sensitive uptake of 22Na+ in acid-loaded (pHi = 6.2) Nb2 cells as well as EP-Am sensitive and extracellular Na+-dependent increase in pHi. One minute after the addition of hPRL (5 ng/ml), there was a significant increase in (P less than 0.01) 22Na+ uptake over the hormonally untreated controls. Addition of hPRL (5 and 10 ng/ml) to Nb2 cells induced a significant (P less than 0.05) increase in pHi within 4 min, while with a higher concentration of 30 ng/ml the increase was significant by 1 min. In all cases, the pHi was raised from a resting value of 7.29 +/- 0.02 (n = 8) to a maximum of 7.45 +/- 0.02 (n = 8). The hPRL-stimulated increase in pHi was abolished in Nb2 cells preincubated for 5-10 min with EP-Am (25 microM) before the addition of the hormone. Activation of Na+/H+ exchange in Nb2 cells was also achieved with human recombinant interleukin 2 and guinea pig antiserum to PRL receptors. In each case, there was a dose- and time-dependent increase in H+ efflux rate which was detectable by 3 min. However, both antiserum to PRL receptor (n = 4) and interleukin 2 (n = 6) raised pHi to a maximum of only 7.35 +/- 0.01. The Na+/H+ exchange system in Nb2 cells was also activated by osmotic shocking, rapidly raising pHi to 7.45 +/- 0.01 (n = 3). Activation was abolished by EP-Am. Contrary to these observations, rat GH which is not mitogenic did not have an effect on pHi in Nb2 cells.(ABSTRACT TRUNCATED AT 400 WORDS)