A substantial body of evidence now exists which implicates a role for the hypothalamic neuropeptide galanin in the regulation of pituitary reproductive hormone secretion. Galanin neurons regulate pituitary lutenizing hormone (LH) secretion at two distinct levels: 1). at the level of the hypothalamus, where galanin indirectly regulates LH by stimulating the production of the hypothalamic releasing factor gonadotropin-releasing hormone (GnRH), and 2). at the level of the pituitary gland, where galanin directly stimulates the secretion of LH from gonadotropes and enhances GnRH-stimulated LH secretion. While the hypothalamic effects of galanin on GnRH secretion are relatively well-characterized, little is known about galanin regulation of LH secretion directly at the level of the pituitary gland. Experiments outlined in this proposal were designed to characterize the pituitathe hypothalamic releasing factor gonadotropin-releasing hormone (GnRH), and 2). at the level of the pituitary gland, where galanin directly stimulates the secretion of LH from gonadotropes and enhances GnRH-stimulated LH secretion. While the hypothalamic effects of galanin on GnRH secretion are relatively well-characterized, little is known about galanin regulation of LH secretion directly at the level of the pituitary gland. Experiments outlined in this proposal were designed to characterize the pituitary effects of galanin on reproductive hormone secretion in both males and females, using the rat as an animal model. Specific aim number 1 of this proposal in to determine whether gonadal steroids alter gonadotrope sensitivity to galanin stimulation. This hypothesis will be tested in a series of in vivo infusion experiments examining LH secretory responses to galanin in 1) intact, castrated, and castrated + testosterone-replaced male rats; and 2) female rats at different timepoints during the four day estrous cycle. Studies in females will specifically address whether gonadotrope sensitivity to galanin stimulation is upregulated within the context of the proestrous (i.e. preovulatory) endocrine milieu. Experiments of Specific aim number 2 are designed to characterize the galanin receptor subtype that is expressed in gonadotropes in the anterior pituitary gland. Double-label in situ hybridization experiments will be performed to determine if galanin type 2 receptor (GalR2) messenger RNA (mRNA) expression is present in anterior pituitary cells that respond to GnRH, and if so, to determine the relative levels of GalR2 mRNA expression in gonadotropes vs. other anterior pituitary cell types that respond to galanin. Specific aim number 3 of this proposal is to investigate the effects of galanin on reproductive hormone gene expression in the anterior pituitary gland. An in vitro perifusion system will be used to determine if galanin, in addition to stimulating the secretion of LH, also has an effect on the mRNA expression of the Lhbeta subunit. Furthermore, the effects of galanin of GnRH receptor mRNA expression will be assessed. Results from these studies will provide important information regarding the role of galanin in regulating LH synthesis and secretion from pituitary gonadotropes, events that are critically important for maintaining reproductive competence in both males and females. Additionally, participation in the execution of experiments outlined in this proposal will provide a unique opportunity for undergraduate students to learn fundamental techniques in reproductive physiology and molecular biology, experiences that will serve them well as they pursue their graduate studies.