Rapid effects of pesticides on human granulosa-lutein cells

Abstract

Following our previous demonstration thatp,p′-DDE (dichlorodiphenylchloroethylene), at environmentally relevant concentrations, can rapidly increase intracellular calcium [Ca2+]iconcentrations 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+]iconcentrations 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+]iconcentrations 2–6 times higher than baseline values within minutes of exposure. Methoxychlor at concentrations of 2.8–280 nmol/ml failed to alter [Ca2+]ilevels consistently in cells from 10 patients. However, at 0.28 and 1.4 nmol/ml, increases in [Ca2+]iconcentrations could be elicited by methoxychlor. The isomero,p-DDE at 3 nmol/ml increased [Ca2+]iin granulosa cells of 11/20 patients. Pertussis toxin treatment inhibited the [Ca2+]iincreases 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+]iincreases induced by Kepone also occurred in Ca2+-free medium, suggesting that [Ca2+]imobilization occurred from the smooth endoplasmic reticulum. Thapsigargin and cyclopiazonic acid, two inhibitors of the endoplasmic reticulum Ca2+pump, also stimulated [Ca2+]iincreases 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