The reticular formation in the brainstem controls motor output via axonal projections to the hindbrain and spinal cord. It remains unclear how individual groups of brainstem neurons contribute to ...specific motor functions. Here, we investigate the behavioral role of the nucleus of the medial longitudinal fasciculus (nMLF), a small group of reticulospinal neurons in the zebrafish midbrain. Calcium imaging revealed that nMLF activity is correlated with bouts of swimming. Optogenetic stimulation of neurons in the left or right nMLF activates the posterior hypaxial muscle and produces a graded ipsilateral tail deflection. Unilateral ablation of a subset of nMLF cells biases the tail position to the intact side during visually evoked swims, while sparing other locomotor maneuvers. We conclude that activity in the nMLF provides postural control of tail orientation and thus steers the direction of swimming. Our studies provide an example of fine-grained modularity of descending motor control in vertebrates.
•A midbrain nucleus with spinal projections is involved in diverse motor behaviors•Optogenetic activation evokes smooth steering of the tail via an identified muscle•Ablation of the nucleus impairs postural control during visually evoked swimming•These data support a fine-grained modularity of motor control in vertebrates
The reticular formation in the brainstem controls motor output via axonal projections to the hindbrain and spinal cord. Using optogenetics, calcium imaging, and behavior, Thiele et al. show that a cluster of midbrain reticulospinal neurons control swim posture in zebrafish.
Non-invasive recording in untethered animals is arguably the ultimate step in the analysis of neuronal function, but such recordings remain elusive. To address this problem, we devised a system that ...tracks neuron-sized fluorescent targets in real time. The system can be used to create virtual environments by optogenetic activation of sensory neurons, or to image activity in identified neurons at high magnification. By recording activity in neurons of freely moving C. elegans, we tested the long-standing hypothesis that forward and reverse locomotion are generated by distinct neuronal circuits. Surprisingly, we found motor neurons that are active during both types of locomotion, suggesting a new model of locomotion control in C. elegans. These results emphasize the importance of recording neuronal activity in freely moving animals and significantly expand the potential of imaging techniques by providing a mean to stabilize fluorescent targets.
Non-cortical visual areas in vertebrate brains extract relevant stimulus features, such as motion, object size, and location, to support diverse behavioral tasks. The optic tectum and pretectum, two ...primary visual areas in zebrafish, are involved in motion processing, and yet their differential neural representation of behaviorally relevant visual features is unclear. Here, we characterize receptive fields (RFs) of motion-sensitive neurons in the diencephalon and midbrain. We show that RFs of many pretectal neurons are large and sample the lower visual field, whereas RFs of tectal neurons are mostly small-size selective and sample the upper nasal visual field more densely. Furthermore, optomotor swimming can reliably be evoked by presenting forward motion in the lower temporal visual field alone, matching the lower visual field bias of the pretectum. Thus, tectum and pretectum extract different visual features from distinct regions of visual space, which is likely a result of their adaptations to hunting and optomotor behavior, respectively.
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•Optic tectum and pretectum sample visual space differently (upper nasal versus lower)•Receptive fields are mainly small-size selective in tectum and large in pretectum•Motion in the lower temporal visual field drives optomotor swimming•The unequal sampling is likely an adaptation to hunting and optomotor behavior
Wang et al. show that the zebrafish optic tectum and pretectum extract visual motion features associated with hunting and optomotor behavior, respectively. The tectum preferably represents small stimuli in the upper nasal visual field, whereas the pretectum is biased toward larger stimuli in the lower visual field.
Zebrafish larvae show characteristic prey capture behavior in response to small moving objects. The neural mechanism used to recognize objects as prey remains largely unknown. We devised a machine ...learning behavior classification system to quantify hunting kinematics in semi-restrained animals exposed to a range of virtual stimuli. Two-photon calcium imaging revealed a small visual area, AF7, that was activated specifically by the optimal prey stimulus. This pretectal region is innervated by two types of retinal ganglion cells, which also send collaterals to the optic tectum. Laser ablation of AF7 markedly reduced prey capture behavior. We identified neurons with arbors in AF7 and found that they projected to multiple sensory and premotor areas: the optic tectum, the nucleus of the medial longitudinal fasciculus (nMLF) and the hindbrain. These findings indicate that computations in the retina give rise to a visual stream which transforms sensory information into a directed prey capture response.
Chemotaxis in Caenorhabditis elegans, like chemotaxis in bacteria, involves a random walk biased by the time derivative of attractant concentration, but how the derivative is computed is unknown. ...Laser ablations have shown that the strongest deficits in chemotaxis to salts are obtained when the ASE chemosensory neurons (ASEL and ASER) are ablated, indicating that this pair has a dominant role. Although these neurons are left-right homologues anatomically, they exhibit marked asymmetries in gene expression and ion preference. Here, using optical recordings of calcium concentration in ASE neurons in intact animals, we demonstrate an additional asymmetry: ASEL is an ON-cell, stimulated by increases in NaCl concentration, whereas ASER is an OFF-cell, stimulated by decreases in NaCl concentration. Both responses are reliable yet transient, indicating that ASE neurons report changes in concentration rather than absolute levels. Recordings from synaptic and sensory transduction mutants show that the ON-OFF asymmetry is the result of intrinsic differences between ASE neurons. Unilateral activation experiments indicate that the asymmetry extends to the level of behavioural output: ASEL lengthens bouts of forward locomotion (runs) whereas ASER promotes direction changes (turns). Notably, the input and output asymmetries of ASE neurons are precisely those of a simple yet novel neuronal motif for computing the time derivative of chemosensory information, which is the fundamental computation of C. elegans chemotaxis. Evidence for ON and OFF cells in other chemosensory networks suggests that this motif may be common in animals that navigate by taste and smell.
Abstract
Direct tests of gene function have historically been performed in a limited number of model organisms. The CRISPR/Cas system is species-agnostic, offering the ability to manipulate genes in ...a range of models, enabling insights into evolution, development, and physiology.
Astatotilapia burtoni
, a cichlid fish from the rivers and shoreline around Lake Tanganyika, has been extensively studied in the laboratory to understand evolution and the neural control of behavior. Here we develop protocols for the creation of CRISPR-edited cichlids and create a broadly useful mutant line. By manipulating the
Tyrosinase
gene, which is necessary for eumelanin pigment production, we describe a fast and reliable approach to quantify and optimize gene editing efficiency.
Tyrosinase
mutants also remove a major obstruction to imaging, enabling visualization of subdermal structures and fluorophores in situ. These protocols will facilitate broad application of CRISPR/Cas9 to studies of cichlids as well as other non-traditional model aquatic species.
We present BonZeb-a suite of modular Bonsai packages which allow high-resolution zebrafish tracking with dynamic visual feedback. Bonsai is an increasingly popular software platform that is ...accelerating the standardization of experimental protocols within the neurosciences due to its speed, flexibility, and minimal programming overhead. BonZeb can be implemented into novel and existing Bonsai workflows for online behavioral tracking and offline tracking with batch processing. We demonstrate that BonZeb can run a variety of experimental configurations used for gaining insights into the neural mechanisms of zebrafish behavior. BonZeb supports head-fixed closed-loop and free-swimming virtual open-loop assays as well as multi-animal tracking, optogenetic stimulation, and calcium imaging during behavior. The combined performance, ease of use and versatility of BonZeb opens new experimental avenues for researchers seeking high-resolution behavioral tracking of larval zebrafish.
A reliable method for recording evoked synaptic events in identified neurons in Caenorhabditis elegans would greatly accelerate our understanding of its nervous system at the molecular, cellular and ...network levels. Here we describe a method for recording synaptic currents and potentials from identified neurons in nearly intact worms. Dissection and exposure of postsynaptic neurons is facilitated by microfabricated agar substrates, and ChannelRhodopsin-2 is used to stimulate presynaptic neurons. We used the method to analyse functional connectivity between a polymodal nociceptor and a command neuron that initiates a stochastic escape behaviour. We find that escape probability mirrors the time course of synaptic current in the command neuron. Moreover, synaptic input increases smoothly as stimulus strength is increased, suggesting that the overall input-output function of the connection is graded. We propose a model in which the energetic cost of escape behaviours in C. elegans is tuned to the intensity of the threat.
Animal sensory systems are tightly adapted to the demands of their environment. In the visual domain, research has shown that many species have circuits and systems that exploit statistical ...regularities in natural visual signals. The zebrafish is a popular model animal in visual neuroscience, but relatively little quantitative data is available about the visual properties of the aquatic habitats where zebrafish reside, as compared to terrestrial environments. Improving our understanding of the visual demands of the aquatic habitats of zebrafish can enhance the insights about sensory neuroscience yielded by this model system. We analyzed a video dataset of zebrafish habitats captured by a stationary camera and compared this dataset to videos of terrestrial scenes in the same geographic area. Our analysis of the spatiotemporal structure in these videos suggests that zebrafish habitats are characterized by low visual contrast and strong motion when compared to terrestrial environments. Similar to terrestrial environments, zebrafish habitats tended to be dominated by dark contrasts, particularly in the lower visual field. We discuss how these properties of the visual environment can inform the study of zebrafish visual behavior and neural processing and, by extension, can inform our understanding of the vertebrate brain.
Genetic access to small, reproducible sets of neurons is key to an understanding of the functional wiring of the brain. Here we report the generation of a new Gal4- and Cre-driver resource for ...zebrafish neurobiology. Candidate genes, including cell type-specific transcription factors, neurotransmitter-synthesizing enzymes and neuropeptides, were selected according to their expression patterns in small and unique subsets of neurons from diverse brain regions. BAC recombineering, followed by Tol2 transgenesis, was used to generate driver lines that label neuronal populations in patterns that, to a large but variable extent, recapitulate the endogenous gene expression. We used image registration to characterize, compare, and digitally superimpose the labeling patterns from our newly generated transgenic lines. This analysis revealed highly restricted and mutually exclusive tissue distributions, with striking resolution of layered brain regions such as the tectum or the rhombencephalon. We further show that a combination of Gal4 and Cre transgenes allows intersectional expression of a fluorescent reporter in regions where the expression of the two drivers overlaps. Taken together, our study offers new tools for functional studies of specific neural circuits in zebrafish.
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