Abstract
The adipose-derived hormone leptin is important for regulating reproduction. The canonical JAK2/STAT3 pathway is the best-characterised leptin receptor signalling pathway. Neural STAT3 deletion is known to cause obesity; however, its reproductive influence is less understood. This experiment investigated whether STAT3 knockout from brain neurons or AgRP neurons, a neuronal population necessary for normal pubertal timing and reproduction, would unveil the necessity of STAT3 in reproductive function. We also tested the role of ERK2 and mTOR, two alternative leptin signalling molecules.Transgenic mice with a neuronal knockout were created via the Cre-loxP system. CamKinaseIIα-Cre mice were crossed with STAT3, mTOR or ERK2 floxed mice. AgRP-Cre mice were crossed with STAT3 floxed mice for STAT3 knockout in AgRP neurons. All groups were compared to Cre-negative littermate controls (n=6-12/group). Puberty onset was assessed post-weaning by examining genitalia development. Reproductive cyclicity and organ weights were measured in adults. Metabolic effects were evaluated through body weight, abdominal fat and fasting glucose measurements. Brain tissue analysis evaluated cellular responses to leptin for STAT3, ERK2, and mTOR. Data presented as mean ± SEM.Mice with ERK2 KO or mTOR KO showed no reproductive or metabolic deficits compared to controls. In marked contrast, neuronal STAT3 KO mice showed significantly increased body weight by 5 weeks of age and a 6-fold increase in abdominal adiposity as adults (both p<0.001 using two-way ANOVA and Student’s t-test respectively) compared to controls. Males had a significant 5-day delay in age at preputial separation (KO: 32.1±2.09 d; Control: 26.8±0.45 d, p<0.01, Student’s t-test) while females had a significant 7-day delay in vaginal opening (KO: 33.0±1.85 d, Control: 26.6±0.37 d, p<0.05, Student’s t-test), a 9-day delay in first estrus (KO: 37.8±1.89 d; Control: 29.4±0.65 d, p<0.01, Student’s t-test) and pronounced acyclicity. Neuronal STAT3 KO mice also had a >2-fold elevation in fasting glucose levels (p<0.001, Student’s t-test) and regressed reproductive organs. Female mice with STAT3 knockout in AgRP neurons showed significantly increased body weight (by 3 weeks [p<0.001], two-way ANOVA) and a significant 3-day delay in first estrus (KO: 31.1±0.71 d, Control: 28.6±0.45 d, p<0.01, Student’s t-test). These data show that neuronal STAT3 is critical for correctly timed puberty and subsequent fertility in male and female mice, whereas mTOR and ERK2 are not essential. These results have prompted a re-evaluation of previous conclusions about the role of STAT3 from experiments using LepR-Cre mice. Additionally, AgRP neuronal requirements for STAT3 exactly recapitulate AgRP requirements for leptin receptors. A better grasp of leptin signalling pathways will improve our understanding of how infertility arises due to metabolic disease.