EXPRESSION AND SEXUAL DIMORPHISM OF GALANIN MESSENGER-RIBONUCLEIC-ACID IN GROWTH HORMONE-RELEASING HORMONE NEURONS OF THE RAT DURING DEVELOPMENT

In the rat, the secretion of GH is episodic and sexually dimorphic. The development and regulation of this patterning of GH secretion are governed by the reciprocal influence of the hypothalamic peptide somatostatin and GH-releasing hormone (GHRH). Galanin is a neuropeptide that is colocalized with GHRH in hypothalamic neurons and is thought to be involved in generating the episodic pattern of GH secretion. We hypothesized that galanin mRNA expression in GHRH neurons increases over development in both sexes, and that in the adults, galanin expression in GHRH neurons is greater in males than in females. To test these hypotheses, we used a double label in situ hybridization procedure to detect and measure galanin mRNA expression in GHRH neurons in the rat brain. GHRH mRNA-positive cells were visualized by an alkaline phosphatase color reaction, and galanin mRNA levels were measured by counting autoradiographic grains over individual GHRH mRNA-positive cells. Galanin mRNA coexpression was found in GHRH mRNA-containing cells of the arcuate nucleus, periarcuate area, and ventromedial hypothalamus. In both males and females there was a significant increase in galanin mRNA in GHRH neurons over development. Galanin mRNA levels in GHRH neurons of 10- and 25-day-old rats were higher in females than in males [10-day-old: females, 12 +/- 2; males, 6 +/- 1 grains/cell (P < 0.05); 25-day-old:females, 28 +/- 4; males, 15 +/- 3 grains/cell (P < 0.02)]. In adults (70 days), galanin mRNA levels in GHRH neurons were significantly higher in males than in females (males, 54 +/- 4; females, 32 +/- 3 grains/ cell; P < 0.005). In the adult rat, galanin mRNA levels in the individual hypothalamic areas exhibited a significant sexual dimorphism in the arcuate nucleus and periarcuate area, with higher levels in the male, whereas no sexual dimorphism was observed in the ventromedial hypothalamus. To determine whether galanin gene expression is influenced by circulating levels of testosterone, we measured galanin mRNA levels in castrated male rats with and without testosterone replacement. Castration reduced galanin message levels in GHRH neurons (intact, 73 +/- 6; castrate, 57 +/- 4 grains/cell), and although this reduction was not statistically significant (P = similar to 0.07), testosterone replacement significantly increased galanin message content (castrate/sham, 58 +/- 4 grains/cell; castrate plus testosterone replacement, 77 +/- 5 grains/ cell; P < 0.02) to intact levels (intact, 73 +/- 6 grains/cell). In summary, galanin message expression in GHRH neurons of both male and female rats increases over development. In adults, male GHRH neurons contain higher levels of galanin mRNA than those of females, and this difference is probably due at least in part to differences in circulating testosterone levels. These observations support the hypothesis that sexual dimorphism in the GH secretory system is related to sex-dependent differences in galanin gene expression of GHRH neurons.