The role of the melanocortin system in the control of reproduction in the ewe

Altered metabolic status and steroid milieu are relayed to the brain to regulate the secretion of gonadotropin releasing hormone (GnRH), the gatekeeper of the neuroendocrine reproductive axis, but the means by which signals from the periphery are relayed to GnRH cells is unclear. The melanocortins, encoded by the proopiomelanocortin (POMC) gene, are poised to act as a conduit for metabolic and sex steroid feedback to GnRH cells. Although data from the rodent species indicate a role for the melanocortins as a positive regulator of the reproductive axis, little work has been carried out in the ovine species. Furthermore, the neuronal circuitry by which the melanocortins exert their effect on the reproductive axis is yet to be delineated. The first study (Chapter 2) aimed to test the hypothesis that adipose hormone, leptin, is capable of restoring gonadotropic function by up regulating central acetylated melanocortin production in the ewe. The results showed that intracerebroventricular (icv) infusion of leptin was able to correct the hypogonadotropic state in the lean condition with a concomitant up regulation of POMC. Furthermore, administration of a melanocortin agonist, MTII, into the brain by icv infusion activated pulsatile LH secretion (reflecting GnRH secretion) in our lean ewe model. A particularly salient finding of Chapter 2 was the decrease in acetylated α-MSH in the terminal beds of lean hypogonadotropic ewes, and it is proposed that leptin treatment increases the transport and acetylation of melanocortins to the synaptic boutons of POMC neurons in the brain. In Chapter 3, the objective was to further substantiate the stimulatory role of the melanocortins on the reproductive axis in the ewe, and the associated aim was to delineate how ‘dialogue’ is achieved between melanocortins and GnRH cells. It was revealed that central treatment with a melanocortin agonist was able to overcome the negative feedback effects of sex steroids in the luteal phase of the estrous cycle, to stimulate pulsatile LH secretion. The same treatment was unable to induce an LH surge or ovulation in seasonally anestrous ewes, but a transient rise in basal LH secretion was evident. Few melanocortin producing cells of the arcuate nucleus (ARC) were found to project to the preoptic area (POA) where GnRH neurons are located, and an interneuronal pathway is proposed, consistent with previous work from this laboratory. Orexin (ORX) mRNA in the dorsomedial hypothalamus (DMH) and kisspeptin (Kiss1) mRNA in the POA were up regulated following melanocortin treatment in the luteal phase, illustrating potential novel conduits for melanocortin regulation of GnRH cells. The studies conducted in Chapter 4 were undertaken to decipher the neuronal circuitry through which the melanocortins might act to stimulate the reproductive system. The results confirmed and extended data obtained in rodent species that illustrate leptin regulation of the kisspeptin system. Using real-time polymerase chain reaction (RT-PCR) of single cells, the presence of the long form leptin receptor (Ob-Rb) was demonstrated in kisspeptin neurons of the ARC and POA. This allows for facilitation of the actions of melanocortins to regulate metabolic and reproductive homeostasis, via kisspeptin cells. Novel reciprocal pathways between the kisspeptin and the POMC/NPY systems were shown using Zeiss Apotome technology. The mutual communication between these systems may aid in the transmission of sex steroids and leptin to regulate reproductive and metabolic function within the brain, and a model is proposed for this. Collectively, the body of work presented in this thesis presents evidence that the melanocortins act within the brain to positively regulate the neuroendocrine reproductive axis in the ewe. Thus, melanocortin cells relay information regarding leptin status and sex steroid feedback to the brain via an interneuronal pathway to regulate both reproductive and metabolic homeostasis. Understanding the involvement of the melanocortin system in the stimulation of the reproductive axis during times of reproductive suppression, will aid in the therapeutic intervention of hypothalamic amenorrhea