Tiltable objective microscope visualizes selectivity for head motion direction and dynamics in zebrafish vestibular system

Abstract

Spatio-temporal information about head orientation and movement is fundamental to the sense of balance and motion. Hair cells (HCs) in otolith organs of the vestibular system transduce linear acceleration, including head tilt and vibration. Here, we build a tiltable objective microscope in which an objective lens and specimen tilt together. With in vivo Ca2+ imaging of all utricular HCs and ganglion neurons during 360° static tilt and vibration in pitch and roll axes, we reveal the direction- and static/dynamic stimulus-selective topographic responses in larval zebrafish. We find that head vibration is preferentially received by striolar HCs, whereas static tilt is preferentially transduced by extrastriolar HCs. Spatially ordered direction preference in HCs is consistent with hair-bundle polarity and is preserved in ganglion neurons through topographic innervation. Together, these results demonstrate topographically organized selectivity for direction and dynamics of head orientation/movement in the vestibular periphery.

Single-cell transcriptomic profiling of the zebrafish inner ear reveals molecularly distinct hair cell and supporting cell subtypes

“…Our study supports zebrafish possessing distinct types of striolar and extrastriolar hair cells in the maculae and cristae, with molecular differences between these subtypes implying different physiological properties. In the zebrafish utricle, vibration is preferentially transduced by striolar cells while static tilt is received by extrastriolar cells ( Tanimoto et al, 2022 ). Consistent with use of a s100s-hs:tdTomato transgene to mark striolar cells in this previous study, s100s is a highly specific marker for our striolar hair cell cluster (Figure 3E).…”

Biomechanics and neural circuits for vestibular-induced fine postural control in larval zebrafish

“…We aimed to perform Ca 2+ imaging in TAN neurons during roll tilts, using the imaging system in which an objective lens and a fish were tilted together using a motorized rotation stage 20 (Fig. 4b; Supplementary Fig.…”

“…In this imaging system, two-color ratiometric imaging is used to reduce the motion-related artificial fluorescence intensity change during the tilt. Because this study is the first to use a wide-field version of the tiltable objective microscope (note that a confocal microscopy is used in the original version 20 ), we evaluated the level of artifacts upon roll tilts using green/red-Dendra2 (the green Dendra2 was partially photoconverted to red-Dendra2 by ultraviolet light such that TAN neurons contained both green and red-Dendra2; Supplementary Fig. 2e).…”

“…Collectively, the ablation and optogenetic activation experiments indicate that TAN neurons play a critical role in producing the VBR in the contralateral direction. neurons at single-cell resolution using a tiltable objective microscope with a spinning-disk confocal unit 20 . For this purpose, nMLF neurons were retrogradely labeled with a green chemical fluorescent Ca 2+ indicator, Cal-520 dextran, and a red chemical fluorescent dye, rhodamine dextran.…”

“…By 5 days postfertilization, larvae start maintaining a dorsal-up posture 17 , indicating that neuronal circuits for postural control are functional by this stage. The transparent brain enables functional imaging in the animals that are subjected to vestibular stimuli [18][19][20] .…”