Molecular and neuronal basis for sexual dimorphism in a vocal-motor control circuitry

Oregon Health & Science University, Portland OR

Investigators

Abstract

Project Summary Defining how hormonal and genetic factors modulate neurodevelopment is key to understanding the origins of sex differences in brain function and behavior, addressing sex biases in neurological and psychiatric disorders, and developing effective treatment for disorders of sex development, and gender-affirming hormone replacement therapies. The zebra finch robust arcopallial (RA) nucleus exhibits extreme sexual dimorphism and offers unique opportunities for defining how hormonal and sex-linked genetic factors mediate sex differences in cortical circuits that subserve complex behaviors of relevance to human cognition. Only males sing, and RA neuronal size and complexity undergo dramatic increases during vocal development in males, while regressing in females. Analogous to the human laryngeal cortex, RA integrates inputs from cortical and basal ganglia vocal-motor areas, provides the sole descending output of the cortical vocal circuit, and plays key roles in the motor encoding and production of vocalizations. Notably, early exposure to estradiol (E2) masculinizes the song circuitry and leads to song emergence in females, indicating high sensitivity to sex steroids, whereas other evidence points to the role of sex-linked genes with sex-biased expression. Our recent cellular, physiological and molecular studies have led us to hypothesize that RA's sexual dimorphism relates to sex differences in cell types, intrinsic excitable and synaptic properties, and sex-linked genes with sex- biased expression during vocal learning, and that the masculinizing action of E2 in females shifts RA's cell composition, transcriptome, and neuronal excitability towards males. To test these hypotheses, our Specific Aims will: SA1- Determine how sex, age and hormonal factors affect cell type composition in RA. We will use single nucleus (sn)RNA-seq to define RA cell types at key stages of vocal development. We predict that RA's unique cell types emerge in males but not females, and that E2 shifts cell types in females towards males. SA2- Determine how masculinization affects neuronal excitable and synaptic properties in female RA. We will use whole-cell patch clamp recordings of single neurons in slices to test if female masculinization by E2 involves modulation of RA excitability and synaptic function. We predict that E2 shifts RA excitability towards males, enabling the high-frequency firing of ultranarrow spikes required for the emergence of song. SA3- Determine the role of sex-biased Z-linked genes in modulation of RA excitability We will use viral-based gene expression to test the prediction that sex chromosome (Z-linked) transcription factors (TCF4, KLF4) and downstream targets (SCN4B/3B, KCNC1) determine sex differences in RA excitability. This overall effort will lead to in-depth understanding of how hormonal vs. genetic factors act to regulate the dimorphism of a cortical circuitry that captures key aspects of human brain function and communication behavior. This mechanistic knowledge will in turn provide key insights into the potential and limitations of hormone-based therapies, and a solid basis for designing therapies for sex development disorders and gender-affirming hormone therapies.