Directionally biased neuronal responses to pitch-axis vestibular stimulation in larval zebrafish compared to roll-axis responses.

The vestibular apparatus plays a pivotal role in maintaining postural equilibrium and processing movement signals, underscoring its importance in studying certain neurological disorders. Here, we present a rotating light-sheet microscope, enabling brain-wide functional recordings during dynamic vestibular stimulation along the pitch axes in addition to the roll axis in head-restrained zebrafish larvae. The system incorporates a double galvanometer mirror configuration, is amenable to 3D printing, and enhances scanning efficiency. Employing this apparatus, we have successfully conducted the first comprehensive mapping of zebrafish brain responses to dynamic pitch-tilt vestibular stimulation. Through Fourier and regression analyses, we report an asymmetry in neuronal recruitment during nose-up versus nose-down pitch tilts within critical regions, including the cerebellum, oculomotor nucleus, caudal hindbrain, and vestibular nucleus, highlighting physiological adaptations to downward motion. We identified specific brain regions, notably the cerebellum and medial-rostral rhombencephalon, that respond to roll- but not pitch-tilt vestibular stimuli. Furthermore, we have identified a transgenic line that closely correlates with our functional mappings and demonstrates a significant response to vestibular stimulation. The elucidation of brain-wide neuronal circuits involved in vestibular processing establishes a foundational framework for subsequent detailed investigations into the molecular and genetic mechanisms underlying postural control and motion perception.