Estradiol-17β inhibits gonadotropin-releasing hormone-induced Ca 2+ in gonadotropes to regulate negative feedback on luteinizing hormone release

Abstract

In pituitary gonadotropes, estrogens have biphasic actions to cause an initial negative feedback followed by a positive feedback on LH secretion, but the mechanisms involved are not clearly understood. To investigate the feedback effects of estrogen, we used mixed ovine pituitary cell cultures (48-72 h), which were treated with 10-9 M estradiol-17β (E2) or vehicle followed by a pulse of 10-9 M GnRH. Medium was collected for LH assay and cells extracted to determine activation of MAPK (phosphorylated ERK-1/2). E2 treatment for 5 min reduced GnRH-induced LH release and caused phosphorylation of ERK-1/2. E2 alone also caused phosphorylation of ERK-1/2, similar to the response evoked by GnRH alone. GnRH increased cytoplasmic intracellular free calcium concentration ([Ca 2+]i) and this was abolished by 2 min pretreatment with E2 or E-bovine serum albumen conjugate. Blockade of Ca2+ channels with nifedipine had no effect on the initial peak of GnRH-induced increase in [Ca2+]i but reduced its duration by 27 ± 6%. Depletion of intracellular Ca2+ stores with thapsigargin prevented GnRH-induced increase in [Ca2+]i. Thapsigargin (10-7 M) or nifedipine (10-5 M) pretreatment (15 min) of cells lowered GnRH-induced LH secretion by 30 ± 6 and 50% ± 4%, respectively. We conclude that inhibition of the GnRH-induced increase in [Ca2+]i in gonadotropes by E2 is a likely mechanism for the negative feedback effect of E2 on LH secretion involving a rapid nongenomic effect of E2. Activation of the MAPK pathway by E2 may be the mechanism for the time-delayed positive feedback effect on LH secretion at the level of the gonadotrope. Copyright © 2009 by The Endocrine Society.