Rapid effects of pesticides on human granulosa-lutein cells

Abstract

Following our previous demonstration that p,p'-DDE (dichlorodiphenylchloroethylene), at environmentally relevant concentrations, can rapidly increase intracellular calcium [Ca2+]i concentrations in human granulosa-lutein cells, we examined whether other pesticides, such as Kepone, o,p-DDE and methoxychlor, have similar effects. Cultured human granulosa-lutein cells were loaded with Fura-2 AM, and changes in [Ca2+]i concentrations within small areas of single cells were studied with a dynamic digital Ca2+ imaging system. Kepone, at concentrations of 0.2-2 nmol/ml, consistently increased [Ca2+]i concentrations 2-6 times higher than baseline values within minutes of exposure. Methoxychlor at concentrations of 2.8-280 nmol/ml failed to alter [Ca2+]i levels consistently in cells from 10 patients. However, at 0.28 and 1.4 nmol/ml, increases in [Ca2+]i concentrations could be elicited by methoxychlor. The isomer o,p-DDE at 3 nmol/ml increased [Ca2+]i in granulosa cells of 11/20 patients. Pertussis toxin treatment inhibited the [Ca2+]i increases induced by estradiol, p,p'-DDE, o,p-DDE and methoxychlor, but not by Kepone or progesterone, indicating that Kepone and progesterone may act through an insensitive G protein-coupled receptor. The [Ca2+]i increases induced by Kepone also occurred in Ca2+-free medium, suggesting that [Ca2+]i mobilization occurred from the smooth endoplasmic reticulum. Thapsigargin and cyclopiazonic acid, two inhibitors of the endoplasmic reticulum Ca2+ pump, also stimulated [Ca2+]i increases but did not inhibit the Ca2+ response to all the pesticides. These results demonstrate that pesticides can have a rapid effect on human granulosa-lutein cells, and a nongenomic mechanism of action is suggested.