Increased hypothalamic GPR54 signaling: a potential mechanism for initiation of puberty in primates

Abstract

To further study the role of GPR54 signaling in the onset of primate puberty, we used the monkey to examine the ability of kisspeptin-10 to elicit the release of gonadotropin-releasing hormone (GnRH) precociously, and we describe the expression of GPR54 and KiSS-1 in the hypothalamus during the peripubertal period. Agonadal juvenile male monkeys were implanted with a lateral cerebroventricular cannula and a jugular vein catheter. The responsiveness of the juvenile pituitary to endogenous GnRH release was heightened with a chronic pulsatile i.v. infusion of synthetic GnRH before kisspeptin-10 (112-121) injection. Intracerebroventricular (30 microg or 100 microg) or i.v. (100 microg) bolus injections of kisspeptin-10 elicited a robust GnRH discharge, as reflected by luteinizing hormone secretion, which was abolished by pretreatment with a GnRH-receptor antagonist. RNA was isolated from the hypothalamus of agonadal males before (juvenile) and after (pubertal) the pubertal resurgence of pulsatile GnRH release and from juvenile, early pubertal, and midpubertal ovary-intact females. KiSS-1 mRNA levels detected by real-time PCR increased with puberty in both male and female monkeys. In intact females, but not in agonadal males, GPR54 mRNA levels in the hypothalamus increased approximately 3-fold from the juvenile to midpubertal stage. Hybridization histochemistry indicated robust KiSS-1 and GPR54 mRNA expression in the region of the arcuate nucleus. These findings are consistent with the hypothesis that GPR54 signaling by its cognate ligand in the primate hypothalamus may be activated at the end of the juvenile phase of development and may contribute to the pubertal resurgence of pulsatile GnRH release, the central drive for puberty.

Fig. 1.

Effect of central (A; n = 3-4) or peripheral (B; n = 2) administration of 100 μg of kisspeptin-10 (•), vehicle (▴), or 100 μg of kisspeptin-10 with acyline pretreatment (○) on GnRH release, as reflected by plasma LH levels (mean ± SEM), in agonadal juvenile male monkeys. Kisspeptin was injected at time 0 (arrow). The LH-releasing action of i.c.v. or i.v. kisspeptin was abolished by acyline pretreatment. Statistical analysis revealed significant effects of treatment, time, and treatment × time interaction.

Fig. 2.

Changes in KiSS-1 and GPR54 mRNA content (mean ± SEM) determined by real-time PCR in MBH of agonadal male monkeys before (Juv) or after (Pub) the pubertal resurgence of GnRH release. KiSS-1 mRNA content in the MBH was significantly greater (*, P < 0.005) after the pubertal resurgence of GnRH, whereas changes in GPR54 mRNA content during these developmental stages were unremarkable. Three animals were examined per group.

Fig. 3.

Changes in KiSS-1 and GPR54 mRNA content (mean ± SEM) determined by real-time PCR in MBH of intact female monkeys during pubertal development. KiSS-1 and GPR54 mRNA content increased in MBH with pubertal development. Numbers in parentheses indicate the number of animals per group. *, P < 0.005 (ANOVA), ♦, P < 0.05 (χ² with threshold for bins set at 6,000 fg of KiSS-1 mRNA per μg of total RNA). Juv, juvenile; eP, early pubertal; mP, midpubertal.

Fig. 4.

Selective expression of KiSS-1 mRNA in neurons of the medial ARC of the female monkey (ovariectomized and estrogen-treated) hypothalamus. (A) Low-magnification view. (Scale bar: 400 μm.) (B) Merged bright- and darkfield higher-magnification image showing the presence of KiSS-1 mRNA transcripts in neurons of the ARC (arrows). (Scale bar: 20 μm.)

Fig. 5.

GPR54 mRNA is expressed in cells of the ARC. (A-C) Medial ARC (A; MARC), lateral ARC (B; LARC), and section hybridized with a sense RNA probe (C). (Scale bar: 200 μm.) V, third ventricle; ME, median eminence. (D) Higher-magnification image showing expression of GPR54 mRNA in neurons of the ARC (arrows). (Scale bar: 20 μm.)