Differences In Expression Of RFRP-3 / GPR147 In Black Line Hamsters

Seasonal reproduction is the phenomenon that animal breeding only occurs in a certain season.Seasonal reproduction animals integrate optical signal(photo-period) and other environmental factors(food abundance, temperature, etc.) to regulate their breeding activities. Physiological state of the animal changes with the external environment signal changed. In order to acquire maximum reproductive success, seasonal reproduction animals carry out reproductive activities only in the most appropriate period of a year to ensure the maximum survival rate of maternal and offspring. This also results in fluctuations in abundance of the animal species among seasons.Cricetulus barabensis is typical animal with seasonal reproduction. It it widely distributed in North China plain. Its reproduction activities are regulated by photo-period, temperature, food and other factors. Hamster has the reproduction-peak in spring and autumn. Cricetulus barabensis is one of agricultural rodents, eating seeds. In the periods of crop germinates and grows, Cricetulus barabensis eat crop stalks and leaves leading to reduce the yields of crops.Wheat and corn as main food crops were mainly planted in North China Plain. Hamster, except for causing the loss of the crop production, also carry transmissible viruses causing threats to people’s health. So the study of the reproductive mechanisms and controlling reproduction activity is very important.In this study, the Cricetulus barabensis captured from Linyi City, Shandong Province were used as experimental materials.The methods of different photo-periods, food restriction and estrogen injection were used to deal with the sampled hamsters, and the Real-time PCR technology was used to analyze the expression level of genes related to reproduction. First, we cloned a partial cDNA sequence of GPR147 in hypothalamus, and the sequence is 355 bp and belongs to the coding region. The expression levels of GPR147 in hypothalamus, testis and ovary of the hamster from various seasons were examined. The results show that the expression level of GPR147 in the hypothalamus, testis are significantly higher in summer and winter than that in spring and autumn(P <0.05), which is consistent with the intensity of the reproductive activity of the Cricetulus barabensis. These results indicate that RFRP-3 may inhibit the reproductive activity by the role of GPR147 in the non-breeding season. However, the expression pattern of GPR147 in ovarian is opposite to that of GPR147 in testis. RFRP-3 could be more effective on reproduction in ovarian than in testis.Secondly, for the different photo-periods(16L: 8D, 12L: 12 D, 8L: 16D) treated hamsters, the relative expression levels of RFRP-3 / GPR147 and GnRH were examined by real-time PCRtechnology. RFRP-3 expressed in hypothalamus at significant higher level(P <0.05) in hamsters treated by long light than by short light. The expression level of GnRH in hypothalamus increases with day time and the expression levels is under significantly higher in long photo-period than in short photo-period(P <0.05). In hypothalamus, the expression level of estrogen receptor(ER) increased with the order of middle,long, and short photo-periods, and the expression level in short photo-period was significantly higher than that in long photo-period(P<0.05). The expression levels of GPR147 in hypothalamus is significantly higher in short photo-period than in long photo-period(P <0.05), which is opposite to the expression mode of RFRP-3. These results show that the expression level of RFRP-3 was up-regulated under long photo-period, which may be related to the secretion of melatonin.Thirdly, the weight and the expression mode of RFRP-3 / GPR147 in hypothalamus and the gonads were examined in this study. The results show that the weight of group with limited food supply was decreased(P <0.05) and the expression level of RFRP-3 / GPR147 were increased(P<0.05). The results show that RFRP-3 / GPR147 system may be involved in the energy balance that is related to reproduction activity.Finally, the expression mode of RFRP-3 / GPR147 in the hypothalamus and gonads of the hamsters with injection of estrogen were examined by real-time PCR technology. The results show that the expression level of the RFRP-3 in hypothalamus was significantly increased(P<0.05) in the group injected with estrogen than that in the control group, which suggests that estrogen can act its negative feedback role by up-regulating the expression level of RFRP-3. The expression level of ER in the hypothalamus was also significantly increased, indicating that estrogen could significantly increased the expression of estrogen receptor(P <0.05). The estrogen could act on RFRP-3 neurons to elevate RFRP-3 expression level by binding to its receptor, which leads to the activity of HPG is suppressed. In ovarian, exogenous estrogen could increase the expression level of ER and RFRP-3 / GPR147. Hypothalamic GnRH at low concentrations were significantly higher(P <0.05), at high concentrations of estrogen its expression was lower, but in ovarian GnRH expression was not significant. The results show that under the conditions of different concentrations of estrogen, different feedback RFRP-3 /GPR147 or Kiss-1 / GPR54 system,estrogen plays different functions. At low concentration of estrogen may be through a separate regulatory pathways, causing GnRH expression level increased. As the concentration increased, the inhibition was enhanced.