Effect of phox2a knockout on the molecular profiles of hindbrain vestibular neurons in the larval zebrafish (bulk RNA-Seq)

Sensorimotor reflex circuits engage distinct neuronal subtypes, defined by precise connectivity, to transform sensation into compensatory behavior. Whether and how motor partner populations shape the subtype fate and connectivity of their pre-motor counterparts remains controversial. Here, we discovered that motor partners are dispensable for proper connectivity across an entire vestibular reflex circuit that stabilizes gaze. We first measured activity following vestibular sensation in pre-motor projection neurons after constitutive loss of their extraocular motor neuron partners.We observed normal responses and topography consistent with unchanged functional connectivity between sensory neurons and projection neurons. Next, we show that projection neurons remain anatomically and molecularly poised to connect appropriately with their motor partners. Lastly, we show that the transcriptional signatures of projection neuron subtypes develop independently of motor partners. Our findings comprehensively overturn a long-standing model: that connectivity in the circuit for gaze stabilization is retrogradely determined by motor partner-derived signals. By defining the contribution of motor neurons to canonical sensorimotor circuit assembly, our work speaks to comparable processes in spinal circuits and advances our understanding of general principles of neural development. Overall design: Hindbrain vestibular neurons, labeled by Tg(-6.7Tru.Hcrtr2:GAL4-VP16);Tg(UAS:E1b-Kaede), harvested from zebrafish embryos between 72-74 hpf. Embyros were from two conditions: larvae from a stable line of phox2a-/- mutants, and sibling controls (phox2a+/+ or +/-). Fluorescent neurons were isolated by fluorescence activated cell sorting (FACS) according to Kaede fluorescence. Four experimental repeats were performed, each generating two samples (phox2a-/- and sibling control). Bulk RNA sequencing was performed.