Determination of the effect of melatonin/kisspeptin interaction on puberty age and hormones responsible for ovulation in female rats in prepubertal period

Determination of the Effect of Melatonin/Kisspeptin Interaction on Puberty Age and Hormones Responsible for Ovulation in Female Rats in Prepubertal Period The aim of this study is to examine the effect of melatonin/kisspeptin interaction on puberty age and hormones responsible for ovulation in pinealectomized female rats in prepubertal period. In the study, 126 female Wistar albino rats at the age of 25 days with 60-80 grams of live weight were used. During the study, animals were kept at 12 hours light and 12 hour dark at room temperature (20-24°C), and fed ad libitum with commercial pelleted rat chow and regular tap water. Pinealectomy operation was performed to 108 of the rats and sham pinealectomy operation was performed to 18 of the rats. The weights of the rats were taken under record when they reached to 29 days of age and divided into 7 groups so that each group consists of 18 rats. Groups were created as; control (Sham), pinealectomy (PE), melatonin (PE+M), kisspeptin (PE+Kiss), kisspeptin antagonist (PE+KissA), melatonin + kisspeptin (PE+M+Kiss) and melatonin + kisspeptin antagonist (PE+M+KissA). 0.1 ml/24 h intraperitoneal serum physiologic to Sham and PE groups, daily 0.5 mg/kg subcutaneous melatonin to PE+M group, daily 1 mg/kg intraperitoneally kisspeptin to PE+Kiss group, daily 1 mg/kg intraperitoneal kisspeptin antagonist to PE+KissA group were administered. In all groups, injections were applied between 21:00 and 23:00 hours. Hormone administrations to 6 rats from each group (except PE and Sham) were continued until prepubertal 33th day to 30 rats, the first follicular period following the start of cyclus to 30 rats, and the first luteal period following the start of cyclus to 30 rats. Rats were checked for vaginal opening every 12 hours, and 6 rats with no vaginal opening at the 33 days of age from each group were taken and euthanized at the 20th minute following the final hormone administration under general anesthesia by taking blood from their hearts. The day of vaginal opening formation was taken under record with vaginal opening control, and oestrus monitoring was carried out every 12 hours by using vaginal douche method. Six rats from every group at follicular and luteal phase euthanized at the 20th minute following the final hormone administration under general anesthesia by taking blood from their hearts. For the control of vaginal douche, progesterone levels in the blood samples taken were determined. In the research, it was found that, on the 33th day, melatonin administration, compared to pinealectomy (PE) group, delayed the vaginal opening and cyclus formation, whilst decreased the serum GnRH and FSH levels. Kisspeptin administration, on the contrary, compared to Sham and PE groups, expedited the vaginal opening and cyclus starting age, while increased the serum GnRH and LH levels. At the same time in prebubertal period (33th day), it was found that kisspeptin shortened the vaginal opening and cyclus starting age delayed by melatonin, and likewise, increased the reduced levels of GnRH and LH. Also in the study, kisspeptin antagonist, compared to PE group, shortened the cyclus starting age, while decreased the prepubertal 33th day GnRH and FSH levels, compared to PE and Sham groups. In the follicular phase, in kisspeptin administered PE+Kiss and PE+M+Kiss groups, serum GnRH and LH levels were found higher than the other groups. As a result, it was found that kisspeptin avoided the negative impact of melatonin on the duration to reach puberty by suppressing GnRH and FSH. Also, it was seen that kisspeptin takes a role in ovulation by stimulating LH secretion in follicular period, and it is contemplated that the results may change depending on both the age the pinealectomy operations were performed and the dose, duration and method of melatonin and kisspeptin administration. Keywords: Kisspeptin, melatonin, puberty, rats, reproduction