ToxSci Advance Access originally published online on May 18, 2005
Toxicological Sciences 2005 86(2):417-426; doi:10.1093/toxsci/kfi208
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Biphasic Lindane-Induced Oxidation of Glutathione and Inhibition of Gap Junctions in Myometrial Cells
* Toxicology Program, Department of Environmental Health, University of Michigan, 1420 Washington Heights, Ann Arbor, Michigan 48109-2029; National Food Safety & Toxicology Center and Department of Pediatrics and Human Development, Michigan State University, 243 Food Safety, East Lansing, Michigan 48824-1302; and National Food Safety & Toxicology Center and Department of Pediatrics and Human Development, Michigan State University, 246 Food Safety, East Lansing, Michigan 48824-1302
Received February 24, 2005; accepted May 13, 2005
The insecticide lindane (
-hexachlorocyclohexane) inhibits gap junction intercellular communication in rat myometrial cells by a mechanism involving oxidative stress. We hypothesized that oxidation of reduced glutathione (GSH) to glutathione disulfide (GSSG) and subsequent S-glutathionylation provide a mechanistic link between lindane-induced oxidative stress and lindane's inhibition of myometrial gap junction communication. Gap junction communication between cultured rat myometrial myocytes was assessed by Lucifer yellow dye transfer after microinjection. A biphasic pattern was confirmed, with dye transfer nearly abolished after 1 h of exposure to 100 µM lindane followed initially by recovery after lindane removal, and then the development 4 h after termination of lindane exposure of a delayed-onset, sustained inhibition that continued for 96 h. As measured by HPLC, cellular GSH varied over a 24-h period in a biphasic fashion that paralleled lindane-induced inhibition of dye transfer, whereas GSSG levels increased in a manner inversely related to GSH. In accordance, GSH/GSSG ratios were depressed at times when GSH and dye transfer were low. Lindane substantially increased S-glutathionylation in a concentration-dependent manner, measured biochemically by GSSG reductase-stimulated release of GSH from precipitated proteins. Furthermore, treatments that promoted accumulation of GSSG (50 µM diamide and 25 µM 1,3-bis(2-chloroethyl)-1-nitrosourea [BCNU]) inhibited Lucifer yellow dye transfer between myometrial cells. Findings that lindane induced GSH oxidation to GSSG with increased S-glutathionylation, together with the diamide and BCNU results, suggest that oxidation of GSH to GSSG is a component of the mechanism by which lindane inhibits myometrial gap junctions.
Key Words: glutathione; lindane; -hexachlorocyclohexane; gap junctions; myometrium; oxidative stress.