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EFFECTS OF ORGANOTINS ON RACTIVE OXYGEN SPECIES AND INTRACELLULAR CALCIUM IN CEREBELLAR GRANULE CELLS IN CULTURE.
Citation:
Mundy, W R. AND T Freudenrich. EFFECTS OF ORGANOTINS ON RACTIVE OXYGEN SPECIES AND INTRACELLULAR CALCIUM IN CEREBELLAR GRANULE CELLS IN CULTURE. Presented at Society of Toxicology, San Francisco, CA, March 25-29, 2001.
Description:
Use of organotins has increased drastically in the past decade, including their use as stabilizers in polyvinylchloride pipes. Monomethyl- (MMT), dimethyl- (DMT), monobutyl- (MBT), and dibutyltin (DBT) have been found in home water samples and in human blood at concentrations up to 1 nM and 600 nM, respectively. While the tri-organotins are clearly neurotoxic, the effects of the mono- and di-organotins have not been well established. The present experiments examined the ability of MMT, DMT, MBT, and DBT to induce reactive oxygen species formation (ROS; measured as oxidation of 2',7'-dichlorodihydrofluorescin), increases in intracellular calcium (measured using fluo-3), and cytotoxicity, in primary neuronal cultures. Cerebellar granule cells were prepared from 7-day-old Long Evans rats and used after 6-8 days in vitro. ROS and intracellular calcium and were determined after 30 or 60 min exposure to organotins (0.001-10 ?M) in Locke's buffer, while cell viability was measured after 24 hr exposure in media. Of the four organotins tested, only DBT produced a significant increase in ROS (1.3-fold at 10 ?M). In comparison, the pro-oxidant Fe(II) increased ROS by 50-fold. Likewise, only DBT produced a significant increase in intracellular calcium (50% increase in basal calcium levels at 10 ?M). Cell viability was determined by live/dead cell counts using the fluorescent probes calcein and propidium iodide. Significant cell death was observed only for DBT at 1 and 10 ?M. Fluorescent staining of nuclei in DBT-treated cells revealed morphologic changes characteristic of apoptosis. Thus, of the organotins tested, only DBT produced significant cellular toxicity after a relatively acute exposure. (This is an abstract of a proposed presentation and does not necessarily reflect EPA policy).