The kisspeptin receptor GPR54 is required for sexual differentiation of the brain and behavior

Abstract

GPR54 is a G-protein-coupled receptor, which binds kisspeptins and is widely expressed throughout the brain. Kisspeptin-GPR54 signaling has been implicated in the regulation of pubertal and adulthood gonadotropin-releasing hormone (GnRH) secretion, and mutations or deletions of GPR54 cause hypogonadotropic hypogonadism in humans and mice. Other reproductive roles for kisspeptin-GPR54 signaling, including the regulation of developmental GnRH secretion or sexual behavior in adults, have not yet been explored. Using adult wild-type (WT) and GPR54 knock-out (KO) mice, we first tested whether kisspeptin-GPR54 signaling is necessary for male and female sexual behaviors. We found that hormone-replaced gonadectomized GPR54 KO males and females displayed appropriate gender-specific adult sexual behaviors. Next, we examined whether GPR54 signaling is required for proper display of olfactory-mediated partner preference behavior. Testosterone-treated WT males preferred stimulus females rather than males, whereas similarly treated WT females and GPR54 KO males showed no preference for either sex. Because olfactory preference is sexually dimorphic and organized during development by androgens, we assessed whether GPR54 signaling is essential for sexual differentiation of other sexually dimorphic traits. Interestingly, adult testosterone-treated GPR54 KO males displayed "female-like" numbers of tyrosine hydroxylase-immunoreactive and Kiss1 mRNA-containing neurons in the anteroventral periventricular nucleus and likewise possessed fewer motoneurons in the spino-bulbocavernosus nucleus than did WT males. Our findings indicate that kisspeptin-GPR54 signaling is not required for male or female copulatory behavior, provided there is appropriate adulthood hormone replacement. However, GPR54 is necessary for proper male-like development of several sexually dimorphic traits, likely by regulating GnRH-mediated androgen secretion during "critical windows" in perinatal development.

Figure 1.

Representative photomicrographs of GnRH-immunoreactive neurons in the forebrain of gonad-intact WT (A) and GPR54 KO (B) male mice infused intracerebroventricularly with 1 nmol of kisspeptin. Fos immunoreactivity (labeled black) is visible in the nucleus of GnRH cells (labeled brown) of WT but not GPR54 KO mice. Blue arrows denote example GnRH neurons colabeled with Fos; black arrowheads denote GnRH neurons lacking Fos. C, Mean (±SEM) percentage of GnRH neurons colabeled with Fos in WT and GPR54 KO males treated with kisspeptin or vehicle (VEH). *p < 0.05, significantly different from vehicle-treated WT mice. D, Mean (±SEM) concentration of plasma LH in WT and GPR54 KO males 30 min after treatment with kisspeptin or vehicle (VEH). *p < 0.05, significantly different from vehicle-treated WT mice.

Figure 2.

Mean (±SEM) LQs of adult female WT and GPR54 KO mice over four sexual behavior trials. All females were ovariectomized in adulthood and hormone primed with EB and P before testing with a stud male. There were no statistical differences in the degree of female sexual behavior displayed between the two genotypes.

Figure 3.

Mean (±SEM) Olfactory Preference Index of testosterone-treated WT and GPR54 KO males and females. Positive values of the Preference Index reflect an olfactory preference for stimulus females; negative values reflect a preference for stimulus males. No preference for either sex would produce a Preference Index of 0. Testosterone-treated WT males had a higher Preference Index than all of the other testosterone-treated groups; the Preference Indices of testosterone-treated WT females, KO females, and KO males did not differ from each other. *p < 0.05, significantly different from all other groups.

Figure 4.

A, Representative photomicrographs of TH neurons in the AVPV nucleus of testosterone-treated WT and GPR54 KO mice of both sexes. F, Female; M, male; 3V, third ventricle. Scale bar, 50 μm. B, Mean (±SEM) number of TH neurons in the AVPV nucleus of testosterone-treated WT and GPR54 KO male and female mice. *p < 0.05, significantly different from all other groups.

Figure 5.

Photomicrographs of Kiss1 mRNA-containing neurons in the AVPV nucleus of representative testosterone-treated WT and GPR54 KO male mice. GPR54 KO males had significantly more Kiss1 neurons in the AVPV nucleus than WT males, as well as greater numbers of silver grains per Kiss1 cell (see Results for numerical values). White arrows denote sample Kiss1 neurons. 3V, Third ventricle; o.c., optic chiasm. Scale bar, 50 μm.

Figure 6.

A, Photomicrographs of the ventral horn of the spinal cord of representative male (M) and female (F) WT and GPR54 KO mice. Photomicrographs were taken at 20× magnification. White arrows denote example SNB motoneurons, noticeable primarily in the WT males. Scale bar, 50 μm. B, Mean (±SEM) SNB motoneuron counts in adult WT and GPR54 KO mice of both sexes. *p < 0.01, significantly different from all other groups.