The Endogenous And Exogenous Regulatory Mechanism Of Seasonal Reproduction Of Male Plateau Pika Ochotona Curzoniae

In mammals,reproductive activities are tightly controlled by the hypothalamic-pituitary-gonad axis.Although pathways regulating gametogenesis in non-seasonal model species have been well described,insights into the molecular network regulating seasonal reproduction are severely limited.Plateau pika is a small rodent species native to the Qinghai-Tibetan plateau and its surrounding areas.April-July is the breeding stage of the adult plateau pika and enters the dormant period in winter.In the present study,the molecular mechanism regulating seasonal breeding of male plateau pika was investigated by using histological examinations,transcriptomic analysis and functional experiments.The results are as follows:(1)Significant differences in the weight of reproductive organs existed between active and dormant adult male plateau pika.Histological staining revealed that a large number of undifferentiated spermatogonia accumulated in the seminiferous tubule of plateau pika in nonbreeding season.(2)Analysis of sex hormone concentrations by ELISA showed that melatonin,GnRH and LH levels were significantly decreased in reproductively dormant animals.As expected,testicular testosterone concentrations was also significantly down-regulated in dormant animals.(3)Functional experiments revealed that testosterone injection was capable of inducing spermatogonial differentiation and reinitiating spermatogenesis in short-day adapted animals.Testosterone secretion was inhibited by melatonin and short daylight exposure.(4)Transcriptome analysis showed that the expression of the genes regulating the biosynthesis and degradation of retinoic acid was differentially regulated.A single injection of all-trans retinoic acid or inhibition the function of RA-degrading enzyme CYP26B1 induced spermatogonial differentiation and resumption of spermatogenesis in 7 days.In conclusion,data from morphological analysis and functional experimentsshowed that spermatogenesis in Plateau pika responsed changes in day length and the level of melatonin.Melatonin controlled the activity of the HPG axis,particularly,the luteinizing hormone and testosterone synthesis to regulate the seasonal spermatogenesis of plateau pika.Retinoic acid signaling served as a master regulator of seasonal spermatogonial differentiation and spermatogenesis,Testosterone works upstream of retinoic acid signaling by providing a permissive environment of retinoic acid biogenesis and actions in testis.Overall,our data revealed that seasonal spermatogenesis of plateau pika is mainly regulated by melatonin-testosterone-retinoic acid signaling.