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Progesterone production by the corpus luteum is a process vital for reproduction. In humans its secretion is stimulated by the placental hormone human chorionic gonadotropin (hCG), and this stimulatory action can also be observed in cultured human luteinized granulosa cells (GCs). We now provide evidence that opening of a Ca(2+)-activated K(+) channel, the BK(Ca), is crucially involved in this process. Immunohistochemistry and RT-PCR revealed the presence of the pore-forming alpha-subunit in human luteinized GCs and in luteal cells of human, macaque, and rat, implying that BK(Ca) channels are important throughout species. Blocking of BK(Ca) channels by iberiotoxin attenuated hCG-induced progesterone secretion. The inhibitory action of iberiotoxin suggests that BK(Ca) channels are activated in the course of hCG-induced steroidogenesis. In search of physiological activators we used an electrophysiological approach and could preclude a direct regulation of channel activity by hCG or GC-derived steroids (progesterone and 17beta-estradiol). Instead, the peptide hormone oxytocin and an acetylcholine (ACh) agonist, carbachol, evoked transient BK(Ca) currents and membrane hyperpolarization. These two molecules are both secreted by GCs and act via raised intracellular Ca(2+) levels. The release of oxytocin is stimulated by hCG, and a similar mechanism is likely in the case of ACh. We conclude that BK(Ca) channel activity in GCs is mediated by components of the intraovarian signaling system, thereby interlinking a systemic hormonal and a local neuroendocrine system in control of steroidogenesis.

An endocrine type of voltage-activated sodium channel (eNaCh) was identified in the human ovary and human luteinized granulosa cells (GC). Whole-cell patch-clamp studies showed that the eNaCh in GC is functional and tetrodotoxin (TTX) sensitive. The luteotrophic hormone human CG (hCG) was found to decrease the peak amplitude of the sodium current within seconds. Treatment with hCG for 24-48 h suppressed not only eNaCh mRNA levels, but also mean Na+ peak currents and resting membrane potentials. An unexpected role for eNaChs in regulating cell morphology and function was indicated after pharmacological modulation of presumed eNaCh steady-state activity in GC cultures for 24-48 h using TTX (NaCh blocker) and veratridine (NaCh activator). TTX preserved a highly differentiated cellular phenotype. Veratridine not only increased the number of secondary lysosomes but also led to a significantly reduced progesterone production. Importantly, endocrine cells of the nonhuman primate corpus luteum (CL), which represent in vivo counterparts of luteinized GC, also contain eNaCh mRNA. Although the mechanism of channel activity under physiological conditions is not clear, it may include persistent Na+ currents. As observed in GC in culture, abundant secondary lysosomes were particularly evident in the regressing CL, suggesting a functional link between eNaCh activity and this form of cellular regression in vivo. Our results identify eNaCh in ovarian endocrine cells and demonstrate that their expression is under the inhibitory control of hCG. Activation of eNaChs in luteal cells, due to loss of gonadotropin support, may initiate a cascade of events leading to decreased CL function, a process that involves lysosomal activation and autophagy. These results imply that ovarian eNaChs are involved in the physiological demise of the temporary endocrine organ CL in the primate ovary during the menstrual cycle. Because commonly used drugs, including phenytoin, target NaChs, these results may be of clinical relevance.

Intra-ovarian regulation of follicular maturation is modulated by various factors. Among these, growth factors are important local actors. We examined the immunohistochemical localization of transforming growth factor alpha (TGF-alpha), epidermal growth factor (EGF) and epidermal growth factor receptor (EGF-R) in 18 human ovaries. The concentration of TGF-alpha and EGF in follicular fluid was measured by ELISA. TGF-alpha was detected in oocytes of primordial and early preantral follicles. Furthermore, staining was observed in granulosa cells of preantral follicles and in theca cells of preantral, antral and preovulatory follicles. EGF showed a similar distribution as TGF-alpha. Atretic follicles were strongly positive for TGF-alpha and EGF. In the corpus luteum, theca lutein cells were strongly positive for TGF-alpha and EGF. Immunoreactivity for EGF-R was observed only in the granulosa cells of antral follicles. Follicular fluids from patients undergoing in-vitro fertilization (IVF) were examined for their content of TGF-alpha and EGF. TGF-alpha was detected in 37% of the samples. The concentration ranged from 43 pg/ml to 602 pg/ml. EGF was not detected in any of the follicular fluids tested. These observations support the participation of EGF/TGF-alpha in follicular maturation. Furthermore, the presence of TGF-alpha in follicular fluid and the simultaneous absence of EGF suggests that TGF-alpha plays a more pronounced role than EGF in oocyte maturation during late follicular phase.