Pb2(+)-induced secretion from bovine chromaffin cells: fura-2 as a probe for Pb2+

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Abstract
The effect of Pb2+ on catecholamine release was studied in isolated intact and permeabilized bovine chromaffin cells. Fura-2 was used to monitor intracellular Pb2+. A characterization of Pb2(+)-fura-2 interactions in solutions simulating intracellular ionic composition showed that Pb2+ forms a 1:1 Pb2(+)-fura-2 complex (apparent dissociation constant = 4.2 x 10(-12) M, pH 7.05) whose fluorescence resembles that of the Ca2(+)-fura-2 complex. Spectra recorded from fura-2-loaded cells indicate entry of Pb2+ into the cells. Intracellular Pb2+ concentrations were proportional to extracellular Pb2+ concentrations and were found to be 10(-11)-10(-12) M in cells exposed to micromolar Pb2+ concentrations. Pb2+ elicited the release of tritiated norepinephrine from fura-2-loaded cells, indicating the extraordinary effectiveness of Pb2+ as a releasing agent. Permeabilization of cells with digitonin showed that Pb2+ is able, in the absence of Ca2+, to produce exocytotic release at concentrations of 3.2 x 10(-10) M or higher (3 orders of magnitude lower than Ca2+). These results support the notion that Pb2+ can act as a potent Ca2+ surrogate in triggering secretion.