Effect of cytoplasmic acidification on the membrane potential of T-lymphocytes: role of trace metals.

The effect of lowering intracellular pH on the membrane potential (Em) of rat thymic lymphocytes was studied using the potential-sensitive dye bis-oxonol. Cells were acid loaded by addition of the electroneutral K+/H+ exchanging ionophore nigericin. Acidification to pH 6.3 in Na(+)-free solution resulted in a biphasic change in Em: an early transient hyperpolarization followed by a sustained depolarization. These changes were associated with a rise in cytosolic free Ca2+ ([Ca2+]i). The hyperpolarization was eliminated when the change in [Ca2+]i was prevented using BAPTA, an intracellular Ca2+ chelator. Moreover, a similar hyperpolarization was elicited by elevation of [Ca2+]i at physiological pHi using ionomycin, suggesting involvement of Ca2(+)-activated K+ channels. In contrast, the depolarization phase could not be mimicked by raising [Ca2+]i with ionomycin. However, intracellular BAPTA effectively inhibited the acidification-induced depolarization. Inhibition was also obtained by extracellular addition of EGTA or dithiothreitol, even when the external free Ca2+ concentration remained unaltered. These observations suggested a possible role of contaminating trace metals. Cytosolic acidification is envisaged to induce intracellular accumulation of one or more trace metals, which induces the observed changes in Em. Accordingly, similar changes in Em can be induced without acidification by the addition of small amounts of Cu2+ to the medium. The ionic basis of the Em changes induced by acidification and the significance of these observations are discussed.