Pubertal changes in gonadotropin-releasing hormone and proopiomelanocortin gene expression in the brain of the male rat

Pubertal development in mammals is in part attributable to a brain-dependent process, whereby increased pulsatile GnRH secretion leads to the awakening of the entire reproductive system. However, the brain mechanisms controlling this event are unknown. The apparent increase in GnRH secretion at puberty could reflect an autonomous change in the activity of GnRH neurons themselves or in the afferent networks leading to GnRH neurons. If there were a significant increase in the secretion of GnRH with puberty onset, we hypothesized that there would be a commensurate increase in the biosynthetic capacity of GnRH neurons to meet the increasing demand. We tested this hypothesis by comparing the level of cellular prepro-GnRH mRNA (GnRH mRNA) observed between prepubertal (25-day-old; n = 5) and adult (75-day-old; n = 4) male rats by in situ hybridization. We detected no significant change with puberty in GnRH mRNA signal levels in any of the anatomical areas examined, which included the vertical limb of the diagonal band of Broca, medial septum, lateral preoptic area, and medial preoptic area. Given the variance of our analytical technique, we determined that there was a greater than 90% probability that we would have detected a 20% increase in GnRH mRNA had there been one. Endogenous opioid peptides have been implicated in timing the onset of puberty in the rat, with the argument being that a loss in opioid tone could effect a disinhibition of GnRH secretion. One opioid peptide, beta-endorphin, is among several peptides cleaved from the precursor POMC. We hypothesized that with puberty, POMC neurons in the arcuate nucleus would have an attenuated capacity to produce beta-endorphin. We tested this hypothesis by comparing cellular pre-POMC mRNA (POMC mRNA) levels in the arcuate nuclei of prepubertal (n = 6) and adult (n = 7) male rats with in situ hybridization. We observed an increase in POMC mRNA levels with puberty; prepubertal rats had relative POMC mRNA signal levels of 119 +/- 10 grains/cell, while adult rats contained 167 +/- 12 grains/cell (P less than 0.02). This increase in cellular POMC mRNA was confined to the rostral portion of the arcuate nucleus. We conclude that the GnRH gene is fully expressed well before the time of normal puberty onset and that the increase in POMC mRNA that occurs with the onset of puberty may be important for the development of pulsatile GnRH secretion

Gonadotropin-releasing hormone messenger ribonucleic acid levels are unaltered with changes in the gonadal hormone milieu of the adult male rat

Testicular function is regulated by the negative feedback effect of sex hormones acting at the brain and pituitary to inhibit the secretion of LH and FSH. An important component of this feedback axis is presumed to involve regulation of secretion and possibly synthesis of GnRH by the brain. We tested the hypothesis that the castration-induced increase in gonadotropin secretion is subserved, at least in part, by increased synthesis of GnRH. Using in situ hybridization and an oligonucleotide probe to pro-GnRH messenger RNA (GnRH mRNA), we compared the level of cellular GnRH mRNA and the relative number of GnRH mRNA-containing neurons between intact and 21-day castrate adult male rats. To derive estimates of the number of GnRH cells and the cellular GnRH mRNA content, coronal sections from each animal were anatomically matched between intact and castrate groups. All identifiable cells within these sections were counted and analyzed with the aid of a computerized image analysis system, by an observer unaware of the animal's experimental group and were assigned an anatomical location for reference. In an initial experiment, we observed no difference in cellular GnRH mRNA signal level between intact (n = 4) and castrate (n = 5) animals (129 +/- 8 vs. 139 +/- 5 grains per cell); however, we did find a statistical difference between the intact and castrated groups in the relative number of GnRH mRNA-containing cells (intact: 212 +/- 15 vs. castrate: 320 +/- 18). To confirm this observation, we repeated the experiment by again comparing the number of GnRH mRNA-positive cells between intact (n = 4) and castrate (n = 4) rats. In this second experiment, we found no difference in the number of identifiable GnRH mRNA-containing cells between intact and castrate animals (272 +/- 14 vs. 274 +/- 36, respectively); this was the case for the total cell count as well as when the data were analyzed by anatomical region. To clarify the conflicting results on cell counts of Exps 1 and 2, we repeated the experiment a third time, again comparing both the number of GnRH mRNA-containing cells and the cellular content of GnRH mRNA. In this experiment, we observed that neither cell number nor content of GnRH mRNA differed between the intact and castrate groups. Again, this was the case for total cell count, as well as when the data were analyzed by anatomical region.(ABSTRACT TRUNCATED AT 400 WORDS

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