Cadmium toxicity in rat pheochromocytoma cells: studies on the mechanism of uptake.
The uptake and toxicity of cadmium were compared in two rat pheochromocytoma cell lines: PC12 cells, which express voltage-sensitive calcium channels, and PC18 cells, which do not. PC12 but not PC18 cells responded to depolarization with an increase in 45Ca2+ uptake and an increase in the concentration of cytoplasmic free calcium ion, [Ca2+]i. These responses were blocked by the dihydropyridine calcium channel antagonist nimodipine and amplified by the agonist BAY K8644, drugs selective for L-type channels. Cadmium caused death of PC12 cells with an LC50 of 12 microM. Inclusion of high K+ with the agonist BAY K8644 shifted the kill curve to the left (LC50 = 6 microM). whereas nimodipine protected against cadmium toxicity (LC50 = 30 microM). In contrast, drugs acting on L-type calcium channels did not affect Cd2+ toxicity for PC18 cells (LC50 15 microM). Fura 2 was used to measure intracellular free Cd2+ by fluorescence ratio methods. Addition of 25 microM Cd2+ to both PC12 and PC18 cells caused a rise in the 340/380 fluorescence ratio attributable to the uptake of Cd2+, since it was almost completely reversed by chelating extracellular Cd2+ and adding a membrane-permeant chelator of heavy metals. Cd2+ addition resulted in a gradual increase in Fura 2 fluorescence in both PC12 and PC18 cells, but depolarization with BAY K8644 increased the apparent rate of Cd2+ uptake only for the PC12 cells. Cd2+ fluorescence appeared to be concentrated near the plasma membrane. The results confirm the potential involvement of calcium channels in cadmium transport and extend the use of intracellularly trapped fluorescent dyes to monitor intracellular free cadmium ion concentration.
Hinkle
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PM
Department of Pharmacology, University of Rochester School of Medicine and Dentistry, New York 14642.
Osborne
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ME
D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester / Q000494:pharmacology; D000310:Adrenal Gland Neoplasms / Q000378:metabolism*; D000818:Animals; D002104:Cadmium / Q000493:pharmacokinetics* / Q000633:toxicity*; D015220:Calcium Channels / Q000187:drug effects / Q000502:physiology*; D004594:Electrophysiology; D016257:Fura-2; D017136:Ion Transport; D009553:Nimodipine / Q000494:pharmacology; D010673:Pheochromocytoma / Q000378:metabolism*; D051381:Rats; D014407:Tumor Cells, Cultured / Q000187:drug effects
D015220:Calcium Channels; D002104:Cadmium; D009553:Nimodipine; D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; D016257:Fura-2