A Role for Estrogen in Lower Urinary Tract Function via Brainstem Neurons

Objectives: Barrington’s nucleus (Bar), a brainstem nucleus with estrogen receptor-expressing neurons, drives micturition by innervating motor neurons that promote synergistic contraction of the bladder’s detrusor muscle and relaxation of the external urethral sphincter (EUS). Bar is integral to the neurologic circuit that allows for switches between urine storage and voiding phases. Loss of ovarian function and lower estradiol levels after menopause are associated with increased prevalence of urinary incontinence, yet the mechanism of action of estrogen as a modulator of lower urinary tract (LUT) function remains underexplored. This research characterizes proenkephalin (Penk)-expressing neurons, a unique estrogen receptor expressing neuronal subpopulation in Bar, and compares quantities of estrogen receptor-alpha (Esr1) positive and negative Penk neurons between male and female mice at developmentally critical timepoints. The objective of this research is to better understand how estrogen signaling in the brainstem affects lower urinary tract function and to uncover the mechanisms by which changes in hormone signaling influence female urogynecologic health.

Methods: We collected brain tissue at the level of Bar from male and female Penk-IRES2-Cre transgenic mice crossed to a reporter mouse line (H2B-mCherry) at 8, 14, and 17.5 weeks of age. We performed immunohistochemistry using an antibody directed against the Esr1 receptor. With fluorescence imaging, we quantified the red Penk-mCherry labeled neurons contained within Bar and the number of overlapping blue Esr1-expressing neurons.

Results: The total number of Penk+ neurons in Bar increases with age in both male and female mice. Furthermore, the average absolute number of Penk+ neurons expressing Esr1 in Bar increases with age in male and female mice while the absolute number of Penk+ neurons negative for Esr1 expression remains constant. Lastly, the average absolute number of Penk+ neurons expressing Esr1 in Bar is significantly greater in male mice compared to female mice.

Conclusions: Penk+ neurons with the capacity to respond to estrogen signaling via Esr1 increase in population size in early development while the number of Penk+ neurons without the capacity to respond to estrogen signaling via Esr1 remains constant, suggesting a crucial role for estrogen modulation in Bar neurogenesis in both male and female mice and an association between maintenance and proliferation of Penk+ neurons of Bar and Esr1 signaling capacity. Furthermore, the pattern of increase from ages 8 to 17.5 weeks could signify a role for estrogen in both developmental learning of neurological control of the LUT and in altered micturition behavior with aging and menopause. This study uncovers sex-dependent differences in neuron subpopulations of Bar and postulates a dynamic role for estrogen modulation of the neurologic circuits controlling LUT function. Future studies will aim to quantify the total number of Esr1-expressing neurons in Bar at different ages, including menopausal and post-menopausal stages.