Ca(2+) -independent vesicular catecholamine release in PC12 cells by nanomolar concentrations of Pb(2+).
Effects of Pb(2+) on vesicular catecholamine release in intact and ionomycin-permeabilized PC12 cells were investigated using carbon fibre microelectrode amperometry. Changes in intracellular Pb(2+) and Ca(2+) were measured from indo-1 fluorescence by confocal laser scanning microscopy. Depolarization of intact cells and superfusion of permeabilized cells with saline containing > or = 100 microm Ca(2+) rapidly evokes quantal catecholamine release. Superfusion with up to 10 microm Pb(2+) -containing saline evokes release of similar catecholamine quanta after a concentration-dependent delay. Thresholds to induce exocytosis within 30 min of exposure are between 1 and 10 microm Pb(2+) in intact cells and between 10 and 30 nm Pb(2+) in permeabilized cells. Additional inhibition of exocytosis occurs in permeabilized cells exposed to 10 microm Pb(2+). Using membrane-impermeable and -permeable chelators it is demonstrated that intracellular Ca(2+) is not required for Pb(2+) -induced exocytosis. In indo- 1-loaded cells Pb(2+) reduces the fluorescence intensity after a concentration-dependent delay, whereas the fluorescence ratio, indicating intracellular Ca(2+) concentration, remains unchanged. The delay to detect an increase in free intracellular Pb(2+) (> or = 30 nm) is much longer than the delay to Pb(2+) -induced exocytosis, indicating that cytoplasmic components buffer Pb(2+) with high affinity. It is concluded that Pb(2+) acts as a high-affinity substitute for Ca(2+) to trigger essential steps leading to vesicular catecholamine release, which occurs when only approximately 20% of the intracellular high-affinity binding capacity ( approximately 2 attomol/cell) is saturated with Pb(2+).
Westerink
Remco H S
RH
Institute for Risk Assessment Sciences, Utrecht University, Utrecht, The Netherlands. R.Westerink@iras.uu.nl
Vijverberg
Henk P M
HP
D000818:Animals; D002118:Calcium / Q000032:analysis / Q000378:metabolism*; D002395:Catecholamines / Q000378:metabolism*; D002454:Cell Differentiation / Q000187:drug effects; D002463:Cell Membrane Permeability / Q000187:drug effects; D019439:Chromaffin Cells / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism*; D003907:Dexamethasone / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology; D007424:Intracellular Fluid / Q000737:chemistry / Q000378:metabolism; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D007854:Lead / Q000032:analysis / Q000494:pharmacology*; D016716:PC12 Cells; D051381:Rats; D011930:Reaction Time / Q000187:drug effects; D022142:Secretory Vesicles / Q000378:metabolism
D002395:Catecholamines; D007476:Ionophores; D007854:Lead; D015759:Ionomycin; D003907:Dexamethasone; D002118:Calcium