Studies on Regulation of Rat Corpus Luteum Function by Prolactin And Luteinizing Hormone

Abstract

The corpus luteum (CL) is a transient endocrine structure formed from the remnants of an ovulated follicle with the primary purpose of producing progesterone (P4), a hormone vital for the establishment and maintenance of pregnancy. The precise regulation of CL function is essential for normal reproductive cycles and maintenance of early pregnancy. In mammals, the pituitary hormones prolactin (PRL) and luteinizing hormone (LH) function as luteotrophic factors during pregnancy, and these two hormones form a functional luteotrophic complex to control CL function in rodents. The mechanistic underpinnings of the luteotrophic actions of PRL and LH, as well as the interplay between the two hormones are poorly understood, and has been the focus of the current investigation. There are several limitations involved in studying luteal function under cell culture conditions. Hence, in vivo animal models employing dopaminergic receptor agonist, 2-Bromo-α-Ergocryptine Mesylate (CB-154; inhibits pituitary PRL secretion) and GnRH receptor antagonist, cetrorelix (CET; inhibits pituitary LH secretion) have been standardized for purposes of examining the roles of PRL and LH in the regulation of CL structure and function in rats. Administration of CB-154 or CET to pregnant rats caused inhibition of CL function and concomitant loss of conceptuses. The CB-154 treatment induced loss of implants was determined to be the result of inhibition of luteal function, rather than the non-specific effects of CB-154 or requirement of PRL for uterine maintenance of implants. To understand how PRL and LH regulate luteal function, targets of PRL and LH in the rat CL needs to be established; however, this has not been well defined by previous studies. The present study observed that CB-154 induced inhibition of luteal function was gradual in its onset; hence, transcriptional changes of genes involved in steroid genesis were examined. mRNA expression of genes involved in P4 production were found to be down regulated, while 20α-hydroxysteroid dehydrogenate (20α-HSD), a P4 catabolizing enzyme was unregulated by CB-154 treatment. CET treatment also had a similar effect on mRNA expression of steroidogenic genes. Interestingly, mRNA expression of the steroidogenic acute regulatory protein (StAR), a key regulator of steroid genesis was not regulated by CB-154 or CET treatment. The luteolytic factor PGF2α also inhibited CL function in pregnant rats but did not down regulate mRNA expression of StAR. However, examination of phospho-StAR (Ser-195), the activated form of StAR, during CET and PGF2α-induced luteolysis suggested that regulation of StAR in the CL of pregnant rats might primarily be at the level of phosphorylation. PRL has been implicated in maintaining luteal expression of LH/choriogonadotrophin receptor (LH/CGR), the cognate receptor for LH. Hence, the luteotrophic actions of PRL may be indirect, by way of regulating LH signalling. Hence, the importance of the LH/CGR pathway and its regulation were examined. LH/CGR mRNA expression was found to correlate with CL function, with CET and CB-154 treatments resulting in down regulation of LH/CGR mRNA expression. Further, CB-154 treatment down regulated LH/CGR pre-mRNA levels, suggesting a role for PRL in the regulation of LH/CGR transcription. mRNA expression of LRH-1, a constitutively active transcription factor previously reported to be important in CL function was down regulated by both CB-154 and CET treatments and hence correlated with LH/CGR mRNA expression. Further, luciferase assays in HeLA cells transiently expressing LRH-1 suggests its involvement in activating the LH/CGR promoter. Estrogen receptor (ER)-α and ER-β also appear to correlate with LH/CGR expression and may play a role along with LRH-1 in the regulation of LH/CGR mRNA expression in the CL of pregnant rats. To examine mechanisms by which PRL may regulate its downstream targets, pathways employed by PRL in the CL of pregnant rats were analysed. The Akt pathway including downstream targets were down regulated by CB-154 treatment. The pathway was found to be regulated at the level of Akt1 mRNA expression. Hence, actions of PRL may regulate the survival of CL. This study has also made observations of LH playing a similar role in survival of the CL. The results of these studies taken together, shed light on the regulation of CL structure and function by PRL and LH, and provide molecular evidence for the two hormones having similar downstream targets and functioning as a luteotrophic complex in pregnant rats, which could only mean a robust interaction between the signalling pathways employed by the two hormones.