Murine arcuate nucleus kisspeptin neurons communicate with GnRH neurons in utero.

Puberty is a transition period of reproductive development from juvenile stages to adulthood and depends upon the activity of gonadotropin-releasing hormone (GnRH) neurons. GnRH neurons are initially activated in utero but remain quiescent throughout the juvenile period. Premature reactivation of GnRH neurons results in precocious puberty in mice and humans, but the mechanisms underlying developmental control of GnRH neuron activity remain unknown. The neuropeptide kisspeptin, a potent activator of GnRH neurons that is implicated as a critical permissive signal triggering puberty and a major regulator of the adult female hypothalamus-pituitary-gonadal axis, is paradoxically produced by neurons in the developing brain well before puberty onset. Thus, the neural circuits controlling the timing of reproductive maturation remain elusive. Here, we delineate the underlying neural circuitry using conditional genetic transsynaptic tracing in female mouse embryos. We find that kisspeptin-producing neurons in the arcuate nucleus (ARC) already communicate with a specific subset of GnRH neurons in utero. We show that ARC kisspeptin neurons are upstream of GnRH neurons, and that GnRH neuron connectivity to ARC kisspeptin neurons does not depend on their spatial position in the brain. Furthermore, we demonstrate that the neural circuits between ARC kisspeptin and GnRH neurons are fully established and operative before birth. Finally, we find that most GnRH neurons express the kisspeptin receptor GPR54 upon circuit formation, suggesting that the signaling system implicated in gatekeeping puberty becomes operative in the embryo.

In Utero Development of Kisspeptin/GnRH Neural Circuitry in Male Mice.

The neuropeptide kisspeptin is a major regulator of the hypothalamus-pituitary-gonadal axis. Although it has long been known that kisspeptin and its receptor G protein-coupled receptor 54 (GPR54) are expressed in the developing brain well before puberty onset, the potential role of kisspeptin/GPR54 signaling in the embryonic brain has remained mysterious. Recent studies in female mice have shown that kisspeptin neurons in the arcuate nucleus of the hypothalamus (ARC) already communicate with a subset of GnRH neurons in utero. Whether this specific neural circuit is also formed in the developing male brain is not known. Here, we used a combination of different genetic strategies to analyze the ontogeny and development of the kisspeptin/GPR54 system in the male mouse brain. We demonstrate orchestrated onset of kisspeptin and GPR54 expression in the male embryonic mouse brain and find that androgen receptor and estrogen receptor-α immunoreactivity within the male brain delineate the birthplace of kisspeptin neurons in the ARC. Using conditional transsynaptic tracing from kisspeptin neurons, we find that ARC kisspeptin neurons already communicate with a subset of GnRH neurons in utero and that the neural circuits between ARC kisspeptin and GnRH neurons in the male mouse brain are established before birth. Furthermore, we also show that the connectivity between kisspeptin and GnRH neurons does not depend on the spatial position of GnRH neurons. Our data delineate the maturing neural circuits underlying control of the reproductive axis in the male embryonic mouse brain.