Highlights • Modern microscopy and genetic techniques enable large-scale imaging of small brains. • Imaging has generated insights into processes such as visual function, motor control and network ...dynamics. • Mapping out the behavioral repertoire of small model organisms will be crucial.
To execute accurate movements, animals must continuously adapt their behavior to changes in their bodies and environments. Animals can learn changes in the relationship between their locomotor ...commands and the resulting distance moved, then adjust command strength to achieve a desired travel distance. It is largely unknown which circuits implement this form of motor learning, or how. Using whole-brain neuronal imaging and circuit manipulations in larval zebrafish, we discovered that the serotonergic dorsal raphe nucleus (DRN) mediates short-term locomotor learning. Serotonergic DRN neurons respond phasically to swim-induced visual motion, but little to motion that is not self-generated. During prolonged exposure to a given motosensory gain, persistent DRN activity emerges that stores the learned efficacy of motor commands and adapts future locomotor drive for tens of seconds. The DRN’s ability to track the effectiveness of motor intent may constitute a computational building block for the broader functions of the serotonergic system.
Display omitted
Display omitted
•The dorsal raphe nucleus (DRN) mediates short-term locomotor learning•DRN responses convey the visual outcomes of swim motor commands•Learning induces a motor memory signal in the DRN that modulates future swimming•DRN ablation/activation abolishes/extends the effect of learning
Animals use environmental feedback to learn the effectiveness of their movements: in larval zebrafish, this motor learning process is mediated by sensorimotor responses and persistent activity in serotonergic neurons in the dorsal raphe nucleus.
Genetically encoded calcium indicators (GECIs) allow measurement of activity in large populations of neurons and in small neuronal compartments, over times of milliseconds to months. Although ...GFP-based GECIs are widely used for in vivo neurophysiology, GECIs with red-shifted excitation and emission spectra have advantages for in vivo imaging because of reduced scattering and absorption in tissue, and a consequent reduction in phototoxicity. However, current red GECIs are inferior to the state-of-the-art GFP-based GCaMP6 indicators for detecting and quantifying neural activity. Here we present improved red GECIs based on mRuby (jRCaMP1a, b) and mApple (jRGECO1a), with sensitivity comparable to GCaMP6. We characterized the performance of the new red GECIs in cultured neurons and in mouse, Drosophila, zebrafish and C. elegans in vivo. Red GECIs facilitate deep-tissue imaging, dual-color imaging together with GFP-based reporters, and the use of optogenetics in combination with calcium imaging.
Increasing the volumetric imaging speed of light-sheet microscopy will improve its ability to detect fast changes in neural activity. Here, a system is introduced for brain-wide imaging of neural ...activity in the larval zebrafish by coupling structured illumination with cubic phase extended depth-of-field (EDoF) pupil encoding. This microscope enables faster light-sheet imaging and facilitates arbitrary plane scanning-removing constraints on acquisition speed, alignment tolerances, and physical motion near the sample. The usefulness of this method is demonstrated by performing multi-plane calcium imaging in the fish brain with a 416×832×160 μm field of view at 33 Hz. The optomotor response behavior of the zebrafish is monitored at high speeds, and time-locked correlations of neuronal activity are resolved across its brain.
•Neural activity can now be recorded in all cells of larval zebrafish brains.•Neurons involved in sensation, motor control, and learning have been identified.•Recently, this capability has been ...extended to free-swimming larvae.•Generally, these neurons’ structures and circuit contributions remain uncharacterized.•Combining activity with anatomical and histochemical data will increase future studies’ impact.
Due to their small size and transparency, zebrafish larvae are amenable to a range of fluorescence microscopy techniques. With the development of sensitive genetically encoded calcium indicators, this has extended to the whole-brain imaging of neural activity with cellular resolution. This technique has been used to study brain-wide population dynamics accompanying sensory processing and sensorimotor transformations, and has spurred the development of innovative closed-loop behavioral paradigms in which stimulus–response relationships can be studied. More recently, microscopes have been developed that allow whole-brain calcium imaging in freely swimming and behaving larvae. In this review, we highlight the technologies underlying whole-brain functional imaging in zebrafish, provide examples of the sensory and motor processes that have been studied with this technique, and discuss the need to merge data from whole-brain functional imaging studies with neurochemical and anatomical information to develop holistic models of functional neural circuits.
Current techniques for monitoring GABA (γ-aminobutyric acid), the primary inhibitory neurotransmitter in vertebrates, cannot follow transients in intact neural circuits. To develop a GABA sensor, we ...applied the design principles used to create the fluorescent glutamate receptor iGluSnFR. We used a protein derived from a previously unsequenced Pseudomonas fluorescens strain and performed structure-guided mutagenesis and library screening to obtain intensity-based GABA sensing fluorescence reporter (iGABASnFR) variants. iGABASnFR is genetically encoded, detects GABA release evoked by electric stimulation of afferent fibers in acute brain slices and produces readily detectable fluorescence increases in vivo in mice and zebrafish. We applied iGABASnFR to track mitochondrial GABA content and its modulation by an anticonvulsant, swimming-evoked, GABA-mediated transmission in zebrafish cerebellum, GABA release events during interictal spikes and seizures in awake mice, and found that GABA-mediated tone decreases during isoflurane anesthesia.
A fundamental question in neuroscience is how entire neural circuits generate behaviour and adapt it to changes in sensory feedback. Here we use two-photon calcium imaging to record the activity of ...large populations of neurons at the cellular level, throughout the brain of larval zebrafish expressing a genetically encoded calcium sensor, while the paralysed animals interact fictively with a virtual environment and rapidly adapt their motor output to changes in visual feedback. We decompose the network dynamics involved in adaptive locomotion into four types of neuronal response properties, and provide anatomical maps of the corresponding sites. A subset of these signals occurred during behavioural adjustments and are candidates for the functional elements that drive motor learning. Lesions to the inferior olive indicate a specific functional role for olivocerebellar circuitry in adaptive locomotion. This study enables the analysis of brain-wide dynamics at single-cell resolution during behaviour.
In the absence of salient sensory cues to guide behavior, animals must still execute sequences of motor actions in order to forage and explore. How such successive motor actions are coordinated to ...form global locomotion trajectories is unknown. We mapped the structure of larval zebrafish swim trajectories in homogeneous environments and found that trajectories were characterized by alternating sequences of repeated turns to the left and to the right. Using whole-brain light-sheet imaging, we identified activity relating to the behavior in specific neural populations that we termed the anterior rhombencephalic turning region (ARTR). ARTR perturbations biased swim direction and reduced the dependence of turn direction on turn history, indicating that the ARTR is part of a network generating the temporal correlations in turn direction. We also find suggestive evidence for ARTR mutual inhibition and ARTR projections to premotor neurons. Finally, simulations suggest the observed turn sequences may underlie efficient exploration of local environments.
Highlights ► The transparency of the larval zebrafish allows for non-invasive optogenetic detection and modulation of neural activity throughout the brain. ► Monitoring of neural activity identifies ...neuronal populations that might be involved in diverse behavior. ► A fictive virtual reality system for larval zebrafish allows for neuronal recording and optical perturbation during behavioral assays. ► Perturbation of the activity of selected groups of neurons in freely swimming animals remains a challenge.
PDF
