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Purpose  Hypothalamic kisspeptin neurons are considered to play a critical role in regulating mammalian reproduction and integrating humoral and neuronal inputs that control gonadotropin‐releasing hormone (Gn RH )/gonadotropin release. The present study aimed to investigate the upstream regulator candidates for kisspeptin neurons.    Methods  Visualized kisspeptin neurons that were taken from the arcuate nucleus ( ARC ) of  Kiss1 ‐tdTomato rats were subjected to next‐generation sequencing ( NGS ) analysis. In situ hybridization ( ISH ) for the calcitonin receptor gene ( Calcr ) was performed throughout the whole forebrain of ovariectomized wild‐type female rats that had been implanted with a negative feedback level of estrogen, because the  Calcr  expression was evident in the  ARC  kisspeptin neurons from the  NGS  analysis. Then, a double  ISH  was performed for the  Calcr  and kisspeptin gene ( Kiss1 ) in the brain regions, containing either the anteroventral periventricular nucleus ( AVPV ) or  ARC  of the female rats.    Results  The  NGS  analysis revealed that the  Calcr  was highly expressed in the  ARC  kisspeptin neurons. It was found that the  Calcr  was co‐expressed in 12% and 22% of the  Kiss1 ‐expressing cells in the  ARC  and  AVPV , respectively.    Conclusion  The present study suggests that calcitonin receptor signaling could be involved in the regulation of reproductive function through the direct control of the  ARC  and/or  AVPV  kisspeptin neurons, and then Gn RH /gonadotropin release.

Co‐expression of the calcitonin receptor gene in the hypothalamic kisspeptin neurons in female rats