Faculty Bibliography

Mature vertebrates maintain posture using vestibulospinal neurons that transform sensed in-stability into reflexive commands to spinal motor circuits. Postural stability improves across development. However, due to the complexity of terrestrial locomotion, vestibulospinal con-tributions to postural refinement in early life remain unexplored. Here we leveraged the relative simplicity of underwater locomotion to quantify the postural consequences of losing vestibulospinal neurons during development in larval zebrafish of undifferentiated sex. By comparing posture at two timepoints, we discovered that later lesions of vestibulospinal neu-rons led to greater instability. Analysis of thousands of individual swim bouts revealed that lesions disrupted movement timing and corrective reflexes without impacting swim kinemat-ics, and that this effect was particularly strong in older larvae. Using a generative model of swimming, we showed how these disruptions could account for the increased postural variability at both timepoints. Finally, late lesions disrupted the fin/trunk coordination observed in older larvae, linking vestibulospinal neurons to postural control schemes used to navigate in depth. Since later lesions were considerably more disruptive to postural sta-bility, we conclude that vestibulospinal contributions to balance increase as larvae mature. Vestibulospinal neurons are highly conserved across vertebrates; we therefore propose that they are a substrate for developmental improvements to postural control.Significance Statement Many animals experience balance improvements during early life. Mature vertebrates use vestibulospinal neurons to transform sensed instability into postural corrections. To under-stand if/how these neurons shape postural development, we ablated them at two develop-mentally important timepoints in larval zebrafish. Loss of vestibulospinal neurons disrupted specific stabilizing behaviors (swim timing, tilt correction, and fin/body coordination) more profoundly in older fish. We conclude that postural development happens in part by changes to vestibulospinal neurons - a significant step towards understanding how developing brains gain the ability to balance.

Astrocytes play crucial roles in regulating neural circuit function by forming a dense network of synapse-associated membrane specializations, but signaling pathways regulating astrocyte morphogenesis remain poorly defined. Here, we show the Drosophila lipid-binding G protein-coupled receptor (GPCR) Tre1 is required for astrocytes to establish their intricate morphology in vivo. The lipid phosphate phosphatases Wunen/Wunen2 also regulate astrocyte morphology and, via Tre1, mediate astrocyte-astrocyte competition for growth-promoting lipids. Loss of s1pr1, the functional analog of Tre1 in zebrafish, disrupts astrocyte process elaboration, and live imaging and pharmacology demonstrate that S1pr1 balances proper astrocyte process extension/retraction dynamics during growth. Loss of Tre1 in flies or S1pr1 in zebrafish results in defects in simple assays of motor behavior. Tre1 and S1pr1 are thus potent evolutionarily conserved regulators of the elaboration of astrocyte morphological complexity and, ultimately, astrocyte control of behavior.

Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons (MNs). MNs are essential to movement and differentially susceptible in disease, but little is known about how MNs acquire functional subtype-specific features during development. Using single-cell RNA profiling in embryonic and larval zebrafish, we identify novel and conserved molecular signatures for MN functional subtypes and identify genes expressed in both early post-mitotic and mature MNs. Assessing MN development in genetic mutants, we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors, Prdm16 and Mecom, are both functional subtype-specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes.

Balance and movement are impaired in many neurological disorders. Recent advances in behavioral monitoring provide unprecedented access to posture and locomotor kinematics but without the throughput and scalability necessary to screen candidate genes/potential therapeutics. Here, we present a scalable apparatus to measure posture and locomotion (SAMPL). SAMPL includes extensible hardware and open-source software with real-time processing and can acquire data from D. melanogaster, C. elegans, and D. rerio as they move vertically. Using SAMPL, we define how zebrafish balance as they navigate vertically and discover small but systematic variations among kinematic parameters between genetic backgrounds. We demonstrate SAMPL's ability to resolve differences in posture and navigation as a function of effect size and data gathered, providing key data for screens. SAMPL is therefore both a tool to model balance and locomotor disorders and an exemplar of how to scale apparatus to support screens.

Vestibulospinal neurons integrate sensed imbalance to regulate postural reflexes. As an evolutionarily conserved neural population, understanding their synaptic and circuit-level properties can offer insight into vertebrate antigravity reflexes. Motivated by recent work, we set out to verify and extend the characterization of vestibulospinal neurons in the larval zebrafish. Using current-clamp recordings together with stimulation, we observed that larval zebrafish vestibulospinal neurons are silent at rest, yet capable of sustained spiking following depolarization. Neurons responded systematically to a vestibular stimulus (translation in the dark); responses were abolished after chronic or acute loss of the utricular otolith. Voltage-clamp recordings at rest revealed strong excitatory inputs with a characteristic multimodal distribution of amplitudes, as well as strong inhibitory inputs. Excitatory inputs within a particular mode (amplitude range) routinely violated refractory period criteria and exhibited complex sensory tuning, suggesting a nonunitary origin. Next, using a unilateral loss-of-function approach, we characterized the source of vestibular inputs to vestibulospinal neurons from each ear. We observed systematic loss of high-amplitude excitatory inputs after utricular lesions ipsilateral, but not contralateral, to the recorded vestibulospinal neuron. In contrast, while some neurons had decreased inhibitory inputs after either ipsilateral or contralateral lesions, there were no systematic changes across the population of recorded neurons. We conclude that imbalance sensed by the utricular otolith shapes the responses of larval zebrafish vestibulospinal neurons through both excitatory and inhibitory inputs. Our findings expand our understanding of how a vertebrate model, the larval zebrafish, might use vestibulospinal input to stabilize posture. More broadly, when compared with recordings in other vertebrates, our data speak to conserved origins of vestibulospinal synaptic input.

Across the nervous system, neurons with similar attributes are topographically organized. This topography reflects developmental pressures. Oddly, vestibular (balance) nuclei are thought to be disorganized. By measuring activity in birthdated neurons, we revealed a functional map within the central vestibular projection nucleus that stabilizes gaze in the larval zebrafish. We first discovered that both somatic position and stimulus selectivity follow projection neuron birthdate. Next, with electron microscopy and loss-of-function assays, we found that patterns of peripheral innervation to projection neurons were similarly organized by birthdate. Finally, birthdate revealed spatial patterns of axonal arborization and synapse formation to projection neuron outputs. Collectively, we find that development reveals previously hidden organization to the input, processing, and output layers of a highly conserved vertebrate sensorimotor circuit. The spatial and temporal attributes we uncover constrain the developmental mechanisms that may specify the fate, function, and organization of vestibulo-ocular reflex neurons. More broadly, our data suggest that, like invertebrates, temporal mechanisms may assemble vertebrate sensorimotor architecture.

Animals use information about gravity and other destabilizing forces to balance and navigate through their environment. Measuring how brains respond to these forces requires considerable technical knowledge and/or financial resources. We present a simple alternative: Tilt In Place Microscopy (TIPM). TIPM is a low-cost and non-invasive way to measure neural activity following rapid changes in body orientation. Here we used TIPM to study vestibulospinal neurons in larval zebrafish during and immediately after roll tilts. Vestibulospinal neurons responded with reliable increases in activity that varied as a function of ipsilateral tilt amplitude. TIPM differentiated tonic (i.e. sustained tilt) from phasic responses, revealing coarse topography of stimulus sensitivity in the lateral vestibular nucleus. Neuronal variability across repeated sessions was minor relative to trial-to-trial variability, allowing us to use TIPM for longitudinal studies of the same neurons across two developmental timepoints. There, we observed global increases in response strength, and systematic changes in the neural representation of stimulus direction. Our data extend classical characterization of the body tilt representation by vestibulospinal neurons and establish TIPM's utility to study the neural basis of balance, especially in developing animals.Significance Statement:Vestibular sensation influences everything from navigation to interoception. Here we detail a straightforward, validated and nearly-universal approach to image how the nervous system senses and responds to body tilts. We use our new method to replicate and expand upon past findings of tilt sensing by a conserved population of spinal-projecting vestibular neurons. The simplicity and broad compatibility of our approach will democratize the study of the brain's response to destabilization, particularly across development.

Background: The ability to maintain balance is an evolutionarily-conserved behavior that is frequently disrupted found in patients with neurodegenerative diseases. One of the most prominent balance disorders is found in patients with progressive supranuclear palsy (PSP), a primary tauopathy pathologically characterized by tau over-representation in the brainstem vestibulospinal (VS) nucleus, where they frequently exhibit accidental-backward falls starting from the early stage of the disease. Although pathological features of PSP correlate well with its clinical phenotype, how tau aggregation affects neuronal and circuit functions, which eventually leads to behavioral deficits, remains unclear. Method: To dissect disease mechanisms across molecular, cellular, circuitry, and behavioral levels, we generated tau fish by expressing human 0N/4R-Tau in zebrafish VS nucleus. Tau expression and phosphorylation were validated using immunohistochemistry staining with PHF-1 antibody. To examine the effect of tau on balance behavior, we measured postural control of free-swimming tau fish and compared to that of tau-negative siblings. Moreover, we tested response of VS neurons to nose-down and nose-up tilt stimulus using 2-photon calcium imaging. Result: Ttau-expressing zebrafish exhibit impaired balance control while maintaining normal locomotor ability. Interestingly, we did not observe any neuronal death in the VS nucleus. Functional imaging of the VS nucleus shows impaired directional tuning in tau-expressing neurons in response to tilt stimulus. We also found ectopic accumulation of acidic organelles in the cell bodies of tau-positive neurons, suggesting abnormal lysosomal function. Conclusion: Our results demonstrate how molecular abnormalities disrupt specific behavior in tauopathies before neurodegeneration appeared.

Efference copies of movement-inducing neural signals have been proposed to serve a role in gaze stabilization. Prior work has demonstrated a spino-extraocular motor circuit in the tadpole that relays copies of spinal commands to extraocular motor neurons. A recent study demonstrates the presence of this circuitry in mice, suggesting a unique method of gaze stabilization in the locomoting mouse.

Myelin is classically known for its role in facilitating nerve conduction. However, recent work casts myelin as a key player in both proper neuronal circuit development and function. With this expanding role comes a demand for new approaches to characterize and perturb myelin in the context of tractable neural circuits as they mature. Here we argue that the simplicity, strong conservation, and clinical relevance of the vestibular system offer a way forward. Further, the tractability of the larval zebrafish affords a uniquely powerful means to test open hypotheses of myelin's role in normal development and disordered vestibular circuits. We end by identifying key open questions in myelin neurobiology that the zebrafish vestibular system is particularly well-suited to address.