Effect of Cocaine on Intracellular Calcium Regulation in Myometrium From Pregnant Women V. P. Fomin, PhD, D. M. Singh, BSc, H. L. Brown, MD, V. Natarajan, PhD, and W. W. Hurd, MD OBJECTIVE: To evaluate the effect of cocaine on intracellularfree calcium (fCa2+1) regulation in human myometrial cells by determining the sources of Ca2+ it might mobilize, as well as assess the role cocaine might play in the catecholamine's effect on the cell's fCa2+1i. METHODS: Primary culture of myometrial cells from pregnant women was used as an experimental model. [Ca2+±i relative changes in response to cocaine and norepinephrine were measured with fura-2 fluorometry and analyzed by means of one-way analysis of variance. RESULTS: Cocaine alone (10-8 to 10~ 3mol/L) increased [Ca2+]i by up to 43 ± 18% over basal level in a dose-dependent manner. Norepinephrine also elevated [Ca2+±i in a concentration-dependent manner (202 ± 24% over basal level at 10-4 mol/L). The norepinephrine-evoked increase was inhibited in Ca2+-free media by 48%, whereas the cocaine response was not affected. The Ca2+-channel antagonist nfedipine caused decrease in the [Ca2+±i response to 10-5 mol/L of norepinephrine by 84%, whereas the [Ca22+ji rise to 10 5 mol/L cocaine was not significantly changed. Inhibitor of the sarcoplasmic reticulum Ca2+ pump, thapsigargin, completely blocked cocaine-evoked increases in [Ca2+±i, whereas norepinephrine responses were greatly reduced. At the same time, cocaine (10-8 to 10-3 mol/L) did not potentiate norepinephrine-evoked Ca2+±i increases in the cells. CONCLUSION: These results indicate that cocaine increases [Ca 2±i in pregnant human myometrial cells, primarily by stimulating release of Ca2+ from intracellular stores rather than by direct stimulation of Ca2+ influx. (J Soc Gynecol Invest 1999;6:147-52) Copyright © 1999 by the Society for Gynecologic Investigation. coca KEY WORDS: Cocaine, myometrium, intracellular calcium, human, cell culture. ocaine use in human pregnancy is associated with trium has been shown to be devoid of functional nerve endings poor reproductive outcome including premature la- in pregnancy.9 Unfortunately, the cellular mechanisms of co- bor, spontaneous abortions, and fetal hypoxia.1 It has caine on the human myometrium are not well understood. been shown that in vitro, cocaine increases spontaneous myo- In 1966, it was suggested that cocaine might also affect the metrial contractions and also augments contractile responses of system of Ca2+ binding and transport in myometrium by myometrium to catecholamines.24 The mechanis for this non-receptor-mediated effects.'0 It was demonstrated that co- has been theorized to be inhibition of catecholamine uptake caine increases myometrial contractility in rabbits independent and metabolism by nerve endings, resulting in increased alpha- of adrenergic or other receptor activation.1 It was hypothe- 1-adrenergic receptor activation.5 However, cocaine retains its sized that cocaine allows Ca2+ that is superficially bound to ability to potentiate responses to norepinephrine in both de- plasma membrane to enter the cell and promote contracture, nervated tissues6 and tissues devoid of sympathetic innerva- while not affecting intracellular Ca2+ stores. tion. 8 This suggests that inhibition of neuronal uptake may More recently it has been reported that cocaine stimulates only partially explain cocaine's effect in some tissues. This inositol 1,4,5-triphosphate production with subsequent release ability may be especially important for understanding the ef- of Ca2+ from sarcoplasmic reticulum in cultured human nmyo- fects of cocaine on pregnant myometrium because myome- metrial cells.12'13 In these studies, prolonged preincubation with cocaine led to a decrease in cyclic adenosine monophos- Fro t Ii prtmut oTh~eticsan Gnecloy ad ~ici~,Indan ~phate generation by the cells. These findings together with :ity Medical Center, Indiarnapoli:, Indiana those on the potentiating effects of cocaine on spontaneous Suppc rted by NIL A Grant nlo. DA064930. 3 4 tha The authors thank D~r. Rolert Bigbb and Andre Caper il-Clrant ftor their hep11 a otatoso ua ndrtmoeru ugs inm-munocstainling of human m~yoretria cells and H[EY cell line with a-s-moo~th-muscle cocaine interferes with the system of signal transduction and actinl antibodies. 2+ Address con :j on~dence and reprint re~quests to): Victo~r P. FominL, Phi), MF11O, 1001 Ca signlaling. Based ont the ability of cocainle to increase W. Walnut ftreet, Indianlapolis, IN 46202. E-mrail: vf~orrninicsiupui.edu ml~yomgetrial contractility, and considering the essential role of Copyright (C) 1999 by the Society fo~r Gynecologic Inlvestigation. 107 -5576/99/$20.00 Published by Elsevier Science Inc. P11 S1071-5576(99)00008-8