SAT-036 Gonadal Hormones Direct the Emergence of Sex-Variable Neuronal Excitability Through Gene Regulation During Adolescence

Abstract Disclosure: S.D. Sun: None. K. Yamada: None. B. Gegenhuber: None. M. Wu: None. V. Kulik: None. R. Anderson: None. M. Johnson: None. J. Tollkuhn: None. Adolescence is a neurodevelopmental critical period marked by the pubertal increase in gonadal steroid hormones (GSHs) resulting in the emergence of sex-variable behaviors. While the role of GSHs in regulating these behaviors is well described, the neurological mechanisms through which they act to direct sex-variability during adolescence remain poorly understood. We discovered that during adolescence, the GSH surge reestablishes sex-variable chromatin accessibility in brain areas that mediate several sex-variable behaviors. To assay the developmental dynamics of chromatin accessibility, we employed ATAC-seq on nuclei purified from neurons in the posterior Bed Nucleus of the Stria Terminalis (BNSTp) expressing Estrogen Receptor (ER) alpha (ERα) from mice, before (postnatal day 28), during (P35), and after (P50) the pubertal hormone surge. 3 biological replicates of 50,000 nuclei pooled from 3-5 animals were taken for each timepoint, and from multiple gonadal states (intact ovaries or testes, and prepubertal-gonadectomy controls). We found the most significantly sex-variable chromatin regions (ANOVA-like generalized linear model, FDR<0.001) exhibit several adolescently dynamic trajectories by k-means clustering. Male-biased chromatin is driven primarily by Androgen Receptor (AR) signaling and is augmented by ERα signaling. Female-biased chromatin accessibility is predominantly associated with activity-dependent signaling, not ovarian GSHs. By comparing with similar ATAC-seq experiments from gonadally-intact neonatal and juvenile mice, we identified key differences in sex-variable chromatin accessibility between the neonatal testosterone surge and adolescent activation of the gonads. Adolescently-emerging sex-variable sites are enriched for genes associated with ion channels, some of which are known to be sex-variable in expression levels during adulthood. To determine if adolescent GSH-mediated gene regulation underlies sex-variability in neurocircuit function, we employed whole-cell patch clamp ex vivo slice electrophysiology. We recorded voltage responses to current injection from a specific ERα-expressing BNSTp transcriptomic cell-type that exhibits significant male-bias in abundance and gene expression. We found that these neurons can fire action potentials at substantially higher rates in adult males, compared to adult females and prepubertal juveniles (n>4, per condition). Preliminary experiments in prepubertally gonadectomized mice suggest the adolescent emergence of high-frequency firing capacity of BNSTp neurons in male animals requires the pubertal GSH surge. Our studies describe a potential mechanism by which GSHs, through their regulation of genes that tune neuronal excitability and responsiveness, can “activate” circuits mediating sex-variable behaviors that emerge during adolescence. Presentation: Saturday, July 12, 2025