This report includes work arising from the following clinical protocol: NCT01434368. We have recruited approximately half of the proposed sample, and several of the children in both the original 12-13-year-old and the 8-year old cohorts have completed the study through to their 18th birthday. Once children are enrolled we have approximately an 87% retention rate, reflected by several of the children who were enrolled at age 8 who have completed over 10 repeated visits and now have turned age 18 years. To date we have studied 62 prepubertal children (37 boys and 25 girls), who are rigorously characterized as pre-gonadarchal by Tanner staging and bone age. These children also have plasma levels of estradiol and testosterone at the lower limits of detectability and have a bimodal distribution of plasma adrenal androgen levels (e.g., DHEA-S, androstenedione) indicating both boys and girls are in either early or later stages of adrenarche. The majority of our current findings relate to the prepubertal (8yo) cohort since we have too few 8 yo children who have completed the full longitudinal study. Longitudinal modeling of cortical surface brain development using structural MRI (sMRI) suggests sex differences in trajectories between the ages of 8-13 years. Longitudinal structural scans from 54 children within the 8-year-old cohort (range: 2-7 visits per child) were analyzed to examine changes in cortical thickness. Developmental trajectories were modeled and estimated within each individual and across the whole cortical surface. Preliminary findings demonstrate sex differences in cortical thickness development in several brain regions. At age 8, prior to the onset of puberty, girls and boys had similar thickness values, but over time, girls showed a greater reduction in thickness in some brain regions compared to boys and a more protracted decline in cortical regions than boys. However, these sex differences were absent by age 13. Future analysis will focus on incorporating the 12-13-year-old longitudinal cohort and addressing developmental trajectories in surface area and volumes (both gray and white matter). Sex differences in resting-state functional connectivity also were identified in a cross-sectional analysis of healthy, typically-developing prepubertal children. Scans from fifty-two 8-year-old children were analyzed using independent component analysis to identify spatially and temporally similar resting-state networks (RSNs) across the whole-brain. A total of 15 RSNs were identified, including the Default Mode (DMN), Salience (SN), and Executive Control (ECN) Networks. Preliminary data showed that girls had greater positive connectivity between the DMN and a region of the dorsolateral prefrontal cortex as well as between the ECN and a region in the lateral prefrontal cortex. Boys showed positive connectivity between the SN and a region in the caudate, while girls showed negative connectivity. These findings suggest the presence of sex differences in cortical connectivity that precede (and, therefore, cannot be attributed to) the re-activation of the hypothalamic-pituitary-gonadal axes (i.e., gonadarche). Future analyses will focus on addressing changes in RSNs across development, including models that address age and individual differences in puberty-related timing and trajectories of development. These preliminary results suggest that there may be neural circuits that exhibit sex-differences prior to gonadarche and HPG axis activation. However, further analysis is needed to determine the factors contributing to sex-differences in this cohort. For example, in addition to possible independent sex-chromosomal effects, sex-differences could reflect greater exposures to adrenal androgens in those children who are in the later stages of adrenarche (which we can examine given the dichotomous distribution of adrenal androgen levels in this sample), or earlier sex steroid exposure secondary to the initial post-natal activation of the HPG axis which lasts 1-2 weeks in boys but up to two years in girls. Initial findings in our combined pre- and post-pubertal cohorts have identified several landmarks in reproductive aging that could have important implications for brain development. We have observed that post-menarcheal girls experience the commencement of fluctuations in estradiol secretion (presumably corresponding to the maturation of the normal architecture of the menstrual cycle). Given our findings of the importance of changes in ovarian steroid levels in RERMDs, these observations could signal an important physiologic event during which functional neurocircuitry undergoes reprogramming. We also have observed in both boys and girls that MRI-measured gonadal volumes increase across the pubertal transition. The individual inflection points in these volume curves identify another important reproductive event (i.e., the expansion of the gonad) that is a sentinel event indicating the initial activation of the GnRH pulse generator, gonadotropin secretion, and the beginning of gonadarche. As enrollment continues and our pre-pubertal children mature, the longitudinal analysis of the development of these sex-differences in brain activity (as well as developmental trajectories in brain that are not sexually dimorphic) could identify specific puberty-related reproductive or metabolic events relevant to normal brain development in both boys and girls. These data also will serve as an important archival data set for studies of children at high risk for the development of behavioral disorders. We also have begun to examine the effects the growth spurt on brain development. Peak height velocity, or the growth spurt, is measured prospectively in each child as change in height over time, with height measured at each visit and divided by the time since the previous visit. Onset of the growth spurt is defined as the visit with the greatest change in height velocity that is followed by a decrease in height velocity in subsequent visits. Age at peak height velocity has been demonstrated at approximately 10-12 for girls and 13-14 in boys, with an earlier peak for black compared to white children. Pubertal related growth is a result of the increased stimulation of the somatotropic axis by both adrenal and gonadal hormones, but also dependent on adequate nutrition, psychosocial environment, and the absence of disease. Adrenal and gonadal steroids act to significantly increase GH secretion from the pituitary and subsequent production of IGF-1 from the liver to generate the growth spurt. Age at peak height velocity is thought to be preceded by a sharp increase in IGF-1 secretion, and both measures are significantly correlated in both boys and girls. Additionally, leptin levels rise in proximity to the growth spurt with girls exhibiting a progressive rise in leptin (which in turn alters adipose tissue distribution) and boys having a peak in leptin proximate to the growth spurt which then declines. Thus, the individuals age at peak height velocity can be employed to assess potential changes in brain neurobiology in association with elevated somatotropic hormones. In our evaluation of stress responsivity in these children (both prepubertal and mid-late pubertal samples) we have observed a sex difference in the morning wakening cortisol response with girls having a greater salivary cortisol response compared to boys that persists into later puberty. Therefore, our findings suggest (albeit preliminarily) that sex differences in HPA axis appear prior to gonadarche. These findings are analogous to previous adult data from our group in which sex differences in HPA axis responsivity were observed after men and women were made hypogonadal by leuprolide treatment, in response to either exercise or corticotropin releasing hormone (CRH) stimulation compared with women.
