Semester

Summer

Date of Graduation

2012

Document Type

Dissertation

Degree Type

PhD

College

School of Medicine

Department

Microbiology, Immunology, and Cell Biology

Committee Chair

Robert L Goodman

Abstract

Gonadotropin-releasing hormone (GnRH) is the final common output from the central nervous system for controlling luteinizing hormone (LH) secretion from the anterior pituitary. GnRH, and subsequently LH, secretion for most of the menstrual/estrous cycle occurs in an episodic pattern that is controlled by the inhibitory actions of estradiol and progesterone. However, GnRH neurons are devoid of the appropriate steroid receptors, therefore, estradiol and progesterone must mediate their effects on GnRH/LH secretion through interneurons. Three likely candidates are neurons containing kisspeptin, neurokinin B (NKB) and orphanin FQ (OFQ).;Puberty in the ewe is marked by an increase in GnRH/LH secretion which in part results from a lessening of estradiol negative feedback. Kisspeptin and NKB are coexpressed in the same subsets of neurons in the arcuate nucleus (ARC) of the hypothalamus and are proposed to be critical for normal timing of puberty. Given that greater than 95% of kisspeptin/NKB neurons colocalize with estrogen receptor-alpha (ERalpha) and progesterone receptor in the ewe, we examined changes in kisspeptin- and NKB-positive neurons in the ARC of pre- and postpubertal ewes in the presence and absence of sex steroids. We observed a greater number of kisspeptin, but not NKB, positive neurons in the ARC of postpubertal ewes compared to prepubertal ewes, which corresponded with changes in LH secretion. Also, we showed that an intravenous injection of senktide, an NKB receptor agonist, stimulates LH secretion in prepubertal ewes. These results support a role for kisspeptin and NKB in ovine puberty.;OFQ when given centrally has been shown to inhibit LH secretion in rats and ewes and OFQ localizes to the external zone of the median eminence. Given its effect on LH secretion and its location, we investigated whether OFQ acts at the pituitary to inhibit LH release and/or at the hypothalamus to inhibit GnRH secretion. OFQ had no direct influence on ovine anterior pituitary cells in cell culture. However, we did observe a decrease in GnRH secretion in hypophyseal portal blood samples following infusion of an OFQ agonist into the third cerebroventricle of ovariectomized ewes. From this we conclude that OFQ acts centrally at the hypothalamus to inhibit GnRH secretion.;Given its central inhibitory action on GnRH secretion in the ewe, we examined if OFQ neurons in the preoptic area (POA) and hypothalamus contain steroid receptors. Furthermore, we investigated a role for endogenous OFQ by infusing an OFQ receptor antagonist into steroid-treated ewes. POA OFQ neurons did not contain steroid receptors, while a majority of OFQ neurons in the ARC contained both ERalpha and progesterone receptor. Furthermore, ovariectomized (OVX) ewes with luteal phase concentrations of progesterone and estradiol, but not OVX ewes implanted with only estradiol or ovary-intact anestrous ewes, showed a significant increase in LH pulse frequency during infusion of an OFQ receptor antagonist. Therefore, we conclude that OFQ mediates, at least in part, the negative feedback action of progesterone on GnRH/LH secretion in the ewe.

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