INTRODUCTION: Neurons of the arcuate nucleus (ARC) are central integrators of metabolic signals within hypothalamus, with leptin serving as a crucial modulatory hormone. Among these, kisspeptin-expressing neurons play an essential role in regulating the reproductive axis, and their development is particularly sensitive to metabolic status.

AIM(S): To examine how early-life undernutrition impacts this system, we analyzed leptin-induced axonal growth in ARC explants from suckling mice, with a focus on sex-specific responses.

METHOD(S): We crossed two transgenic mouse strains to generate offspring expressing tdTomato fluorescence in kisspeptin neurons using the Cre-loxP system. Newborn pups were assigned to either control group (CON; dams fed ad libitum during lactation) or lactation undernutrition group (LUN; dams received 50% of the control diet). On postnatal day 8, ARC explants were microdissected and cultured separately by sex. After 24 hours in vitro, explants were treated with leptin or vehicle. Following another 24 hours, tissues were fixed, immunostained for neurofilaments and imaged using fluorescence microscopy. Axon length was quantified with Zeiss ZEN Blue Software.

RESULTS: Leptin promoted axonal growth in ARC explants from CON females, an effect not observed in males. Notably, in LUN females, leptin failed to stimulate axon outgrowth. However, when analysis restricted to kisspeptin neurons-specific axons, which were markedly shorter than those of the general ARC population, the leptin response was preserved in LUN females but completely absent in LUN males.

CONCLUSIONS: To our knowledge, this is the first study to directly quantify axon growth in kisspeptin neurons and to reveal sex-dependent differences in this process. Our findings highlight the existence of sex-specific sensitivity in the developing hypothalamus to early-life nutritional stress, with potential long-term consequences for reproductive function and the integration of metabolic and reproductive pathways.

FINANCIAL SUPPORT: Research supported by Polish National Science Centre [2018/31/B/NZ4/03527].