In Vitro Stimulation of the Prepubertal Rat Gonadotropin-Releasing Hormone Pulse Generator by Leptin and Neuropeptide Y through Distinct Mechanisms

Leptin may act as a negative feedback signal to the brain in the control of appetite through suppression of neuropeptide Y (NPY) secretion and stimulation of cocaine- and amphetamine-regulated transcript (CART), a new anorectic peptide. We aimed at studying whether leptin, NPY, and CART have related effects on the hypo- thalamic control of the pituitary-gonadal system and the develop- mental changes in NPY and CART effects. Using retrochiasmatic hypothalamic explants from prepubertal 15-day-old male rats, the GnRH interpulse interval (mean 6 SD :6 26 5 min) was significantly reduced by 1027 M of leptin (46 6 3.3 min) as well as 1027 M of NPY (47 6 4.4 min) and 1026 M of CART (46 6 2.7 min), whereas the GnRH pulse amplitude was not affected. The stimulatory effects of different NPY receptor agonists (human PYY 3-36, porcine NPY 13-36, human (D-Trp 32) NPY, porcine (Leu 31 Pro 34) NPY, human pancreatic polypeptide (PP)), as well as the absent effects of rat PP were con- sistent with the involvement of the Y5-receptor subtype in mediation of NPY effects. Incubation with 1027 M of a Y5-receptor selective antagonist prevented the effect of NPY (61 6 4 vs. 46 6 2 min), whereas leptin and CART effects were not (47 6 3 vs. 46 6 3 min and 46 6 3 vs. 46 6 2 min, respectively), suggesting that NPY was not involved in leptin and CART effects. Using an anti-CART antiserum (1:1000), the reduction of GnRH interpulse interval caused by leptin was partially prevented (56.2 6 4 vs. 47.9 6 3.8 min), whereas the reduction of GnRH interval caused by NPY was not affected (45.9 6 2.5 vs. 47.8 6 3.7). The GnRH interpulse interval was decreased by 1027 M of NPY at 5 days (72 6 3.8 vs. 91.9 6 3.5) as well as at 15 days, whereas such an effect was not observed anymore at 25 and 50 days. Similar effects were observed using 1026 M of CART-peptide. Using 1026 M of the Y5-receptor antagonist, the GnRH interpulse interval was significantly increased at 15 days (66.6 6 2.7 min), 25 days (56.5 6 39.9 min), and 50 days (52.5 vs. 38.2 min), whereas no change was observed at 5 days. Using the anti-CART antiserum, a significant increase of GnRH interpulse interval was observed at 25 days only. In conclusion, the stimulatory effects of leptin and NPY on the fre- quency of pulsatile GnRH secretion before puberty involve two dis- tinct mechanisms. NPY causes acceleration of GnRH pulsatility via the Y5-receptor subtype, which is not involved in leptin effects while the CART is involved in leptin effects on GnRH secretion but not in NPY effects. The reduction of pulsatility by the Y5 antagonist pro- vides evidence of endogenous NPY involvement in the control of GnRH secretion from the time of onset of puberty. (Endocrinology 141: 1464 -1469, 2000)