Effect of clomiphene on Ca 2+ movement in human prostate cancer cells Bang-Ping Jiann a , Yih-Chau Lu b , Hong-Tai Chang a , Jong-Khing Huang a , Chung-Ren Jan c, * a Department of Surgery, Kaohsiung Veterans General Hospital, Kaohsiung 813, Taiwan b Department of Orthopaedic Surgery, Kaohsiung Veterans General Hospital, Kaohsiung 813, Taiwan c Department of Medical Education and Research, Kaohsiung Veterans General Hospital, Kaohsiung 813, Taiwan Received 5 October 2001; accepted 19 December 2001 Abstract The effect of clomiphene, an ovulation-inducing agent, on cytosolic free Ca 2+ levels ([Ca 2+ ] i ) in populations of PC3 human prostate cancer cells was explored by using fura-2 as a Ca 2+ indicator. Clomiphene at concentrations between 10-50 mM increased [Ca 2+ ] i in a concentration- dependent manner. The [Ca 2+ ] i signal was biphasic with an initial rise and a slow decay. Ca 2+ removal inhibited the Ca 2+ signal by 41%. Adding 3 mM Ca 2+ increased [Ca 2+ ] i in cells pretreated with clomiphene in Ca 2+ -free medium, confirming that clomiphene induced Ca 2+ entry. In Ca 2+ -free medium, pretreatment with 50 mM brefeldin A (to permeabilize the Golgi complex), 1 mM thapsigargin (to inhibit the endoplasmic reticulum Ca 2+ pump), and 2 mM carbonylcyanide m-chlorophenylhydrazone (to uncouple mitochondria) inhibited 25% of 50 mM clomiphene- induced store Ca 2+ release. Conversely, pretreatment with 50 mM clomiphene in Ca 2+ -free medium abolished the [Ca 2+ ] i increase induced by brefeldin A, thapsigargin or carbonylcyanide m- chlorophenylhydrazone. The 50 mM clomiphene-induced Ca 2+ release was unaltered by inhibiting phospholipase C with 2 mM 1-(6-((17b-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H- pyrrole-2,5-dione (U73122). Trypan blue exclusion assay suggested that incubation with clo- miphene (50 mM) for 2-15 min induced time-dependent decrease in cell viability by 10-50%. Collectively, the results suggest that clomiphene induced [Ca 2+ ] i increases in PC3 cells by 0024-3205/02/$ – see front matter D 2002 Elsevier Science Inc. All rights reserved. PII:S0024-3205(02)01574-6 * Corresponding author. Tel.: +886-7-3422121-1509; fax: +886-7-3468056. E-mail address: crjan@isca.vghks.gov.tw (C.-R. Jan). Life Sciences 70 (2002) 3167 – 3178