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Changes in Granule Cell Representations Correlate with a Behavioral Change in Sensitivity to Sensory Stimuli (A) Upper-left trace: probability of a behavioral response to visual stimuli in the ten trials immediately following injection of 10 mM bicuculline, a GABAA receptor antagonist, into the cerebellum (n = 11). The average behavior is shown with shaded SE. Note that no electrical stimulus was presented in this subset of experiments. Lower-left trace: behavioral responses in these same animals recover as bicuculline washes out. Lower-right trace: sham injections into the cerebellum failed to elicit any increase in luminance-evoked behavior. Upper right: example epifluorescence micrographs showing the extent of the bicuculline (with rhodamine) injections. The scale bar represents 100 mm. The shaded region in behavioral traces indicates the SE. (B) Heatmaps from the same imaging plane through the granule cell layer in a 7-dpf GR152:gal4; UAS:GCaMP6s fish showing the change in pixelwise correlations to luminance (upper) and motor (lower) regressors following the addition of bicuculline. The scale bar represents 50 mm. Colored circles indicate example granule cells analyzed in subsequent panels. (C) Heatmaps of activity from (i) two example granule cells that change their responses following the addition of bicuculline. The upper cell, gc 1, becomes less responsive to luminance and more responsive to motor activity, whereas the bottom cell, gc 2, becomes newly responsive to luminance and motor activity. Location of granule cells is shown in (B). (ii) Heatmap showing the behavioral responses over the experiment. (D) The plotted change in regressor over trials for the two example granule cells, color coded for the two example cells as in (Ci). The gray bar indicates the presence of 30 mm bicuculline.
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A fundamental question in neurobiology is how animals integrate external sensory information from their environment with self-generated motor and sensory signals in order to guide motor behavior and adaptation. The cerebellum is a vertebrate hindbrain region where all of these signals converge and that has been implicated in the acquisition, coordi...
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... Correlate with a Pharmacologically Induced Change in Behavioral Sensitivity In order to investigate the behavioral relevance of the granule cell representations we observed, we developed a pharmacological protocol that elicited swimming responses to previously neutral changes in luminance. Both local unilateral (n = 8) and bilateral (n = 7) injections of 10 mM bicuculline, a GABAA receptor antagonist, led to an immediate increase in luminance-evoked swimming behavior ( Figure 5A). This robust behavioral phenotype emerged rapidly following injection and then behavior returned to baseline following washout of the drug ( Figure 5A). ...
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... local unilateral (n = 8) and bilateral (n = 7) injections of 10 mM bicuculline, a GABAA receptor antagonist, led to an immediate increase in luminance-evoked swimming behavior ( Figure 5A). This robust behavioral phenotype emerged rapidly following injection and then behavior returned to baseline following washout of the drug ( Figure 5A). The same behavioral phenotype was observed and maintained in the continuing presence (20-30 mM bath application) of bicuculline (n = 29; data not shown). ...
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... established a paradigm to acutely change sensorimotor behavior, we next performed two-photon functional imaging of a single cerebellar plane in order to monitor the activity of granule cells over this time course. We observed the widespread activation of granule cells following bicuculline treatment that correlated highly with the increased behavioral responsiveness of fish to visual stimuli ( Figure 5B). Many granule cells acquired responses that correlated with motor activity whereas others showed newly acquired responses to luminance (Figures 5B-5D). ...
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... was true for both luminance-excited ( Figures 6A and 6B) and luminance-inhibited cells (Figures 6C and 6D). Both types of luminance-responsive granule cells also produced bursts of spikes during moving gratings that were fixed to the frequency of luminance transitions in either a forward or backward direction ( Figures S5A-S5C), showing that their stereotyped responses to luminance were not altered by motion. ...
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... cell-attached recordings were obtained from granule cells with other response profiles, including forwardmotion-excited (n = 5) and shock-excited (n = 6) cells ( Figure S5). The spiking of cells to their preferred stimulus within a response type was highly stereotyped with respect to temporal patterning, tuning preference, latency to the first spike, and change in gain of firing during their preferred stimulus ( Figure S5). ...
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... cell-attached recordings were obtained from granule cells with other response profiles, including forwardmotion-excited (n = 5) and shock-excited (n = 6) cells ( Figure S5). The spiking of cells to their preferred stimulus within a response type was highly stereotyped with respect to temporal patterning, tuning preference, latency to the first spike, and change in gain of firing during their preferred stimulus ( Figure S5). The temporal profile of these responses were furthermore consistent across repeated trials for a given granule cell ( Figure S6). ...
Citations
... In contrast with predictions of classical theories, modern calcium imaging approaches have shown that granule cell responses can be dense and redundant in some conditions [15][16][17][18] . These studies have indicated that complex behaviors requiring task engagement, learning ...
... In contrast with predictions of classical theories, modern calcium imaging approaches have shown that granule cell responses can be dense and redundant in some conditions [15][16][17][18] . These studies have indicated that complex behaviors requiring task engagement, learning video was used to detect whisker and facial movements between trials and isolate responses related to these movements 32 ( Supplementary Fig. 4). ...
... Finally, in agreement with our imaging data suggesting that inhibition serves a central role in establishing discrete sensory representations, we find that granule cell inhibition is required for accurate sensorimotor behavior in a cerebellum-dependent task. Recent work has shown that, during complex behaviors, granule cell activity can be denser than was predicted by classical Marr-Albus models [15][16][17][18] . In contrast, our goal was to isolate discrete sensory responses, independent of motor-related signals and contextual modulation 26 . ...
The cerebellar cortex has a key role in generating predictive sensorimotor associations. To do so, the granule cell layer is thought to establish unique sensorimotor representations for learning. However, how this is achieved and how granule cell population responses contribute to behavior have remained unclear. To address these questions, we have used in vivo calcium imaging and granule cell-specific pharmacological manipulation of synaptic inhibition in awake, behaving mice. These experiments indicate that inhibition sparsens and thresholds sensory responses, limiting overlap between sensory ensembles and preventing spiking in many granule cells that receive excitatory input. Moreover, inhibition can be recruited in a stimulus-specific manner to powerfully decorrelate multisensory ensembles. Consistent with these results, granule cell inhibition is required for accurate cerebellum-dependent sensorimotor behavior. These data thus reveal key mechanisms for granule cell layer pattern separation beyond those envisioned by classical models.
... In our experiments, we sampled ~27% of GCs in one hemisphere per fish. Our observation of dense activation of the GC population in response to optic flow stimuli (Fig. 6, C to G) is consistent with the recent findings from larval zebrafish (39,40) and other species (17,41). We also found response profiles in the GC population exhibiting stimulus-tuned but persistent elevation in calcium activity (Fig. 6, D and G). ...
The ability to predict the future based on past experience lies at the core of the brain’s ability to adapt behavior. However, the neural mechanisms that participate in generating and updating predictions are not clearly understood. Further, the evolutionary antecedents and the prevalence of predictive processing among vertebrates are even less explored. Here, we show evidence of predictive processing via the involvement of cerebellar circuits in larval zebrafish. We presented stereotyped optic flow stimuli to larval zebrafish to evoke swims and discovered that lesioning the cerebellum abolished prediction-dependent modulation of swim latency. When expectations of optic flow direction did not match with reality, error signals arrive at Purkinje cells via the olivary climbing fibers, whereas granule cells and Purkinje cells encode signals of expectation. Strong neural representations of expectation correlate with faster swim responses and vice versa. In sum, our results show evidence for predictive processing in nonmammalian vertebrates with the involvement of cerebellum, an evolutionarily conserved brain structure.
... In light of these basic properties of the cerebellar circuit and its principal plasticity mechanisms, recent granule cell and climbing fiber findings pose several mysteries. Empirical observation of the activation of many granule cells at once (Giovannucci et al., 2017;Knogler et al., 2017;Wagner et al., 2017) is at least superficially incompatible with using spike coincidence-based synaptic modification via climbing fiber-directed LTD as a means to modify small and specific sets of granule cell synapses. Similarly, granule cell signals that are sustained for extended periods (Wagner et al., 2019;Lin et al., 2020) appear to challenge the temporal specificity of climbing fiber-directed LTD based on spiking coincidences. ...
The cerebellum, traditionally associated with motor coordination and balance, also plays a crucial role in various aspects of higher-order function and dysfunction. Emerging research has shed light on the cerebellum's broader contributions to cognitive, emotional, and reward processes. The cerebellum's influence on autonomic function further highlights its significance in regulating motivational and emotional states. Perturbations in cerebellar development and function have been implicated in various neurodevelopmental disorders, including autism spectrum disorder and attention deficit hyperactivity disorder. An increasing appreciation for neuropsychiatric symptoms that arise from cerebellar dysfunction underscores the importance of elucidating the circuit mechanisms that underlie complex interactions between the cerebellum and other brain regions for a comprehensive understanding of complex behavior. By briefly discussing new advances in mapping cerebellar function in affective, cognitive, autonomic, and social processing and reviewing the role of the cerebellum in neuropathology beyond the motor domain, this Mini-Symposium review aims to provide a broad perspective of cerebellar intersections with the limbic brain in health and disease.
... Zebrafish display a mediolateral compartmentalization of the cerebellum characterized by distinct response properties and output targets [22][23][24][25][26] . Multimodal representations were found in both cerebellar granule cells 27,28 and Purkinje cells, the output neurons of the cerebellar cortex 25,29 . Functional assays established a role for the larval zebrafish cerebellum in motor control and sensorimotor integration [30][31][32][33][34][35][36] . ...
Cerebellar dysfunction leads to postural instability. Recent work in freely moving rodents has transformed investigations of cerebellar contributions to posture. However, the combined complexity of terrestrial locomotion and the rodent cerebellum motivate development of new approaches to perturb cerebellar function in simpler vertebrates. Here, we used a powerful chemogenetic tool (TRPV1/capsaicin) to define the role of Purkinje cells — the output neurons of the cerebellar cortex — as larval zebrafish swam freely in depth. We achieved both bidirectional control (activation and ablation) of Purkinje cells while performing quantitative high-throughput assessment of posture and locomotion. Activation disrupted postural control in the pitch (nose-up/nose-down) axis. Similarly, ablations disrupted pitch-axis posture and fin-body coordination responsible for climbs. Postural disruption was more pronounced in older larvae, offering a window into emergent roles for the developing cerebellum in the control of posture. Finally, we found that activity in Purkinje cells could individually and collectively encode tilt direction, a key feature of postural control neurons. Our findings delineate an expected role for the cerebellum in postural control and vestibular sensation in larval zebrafish, establishing the validity of TRPV1/capsaicin-mediated perturbations in a simple, genetically-tractable vertebrate. Moreover, by comparing the contributions of Purkinje cell ablations to posture in time, we uncover signatures of emerging cerebellar control of posture across early development. This work takes a major step towards understanding an ancestral role of the cerebellum in regulating postural maturation.
... Several recent studies have reported dense activity in cerebellar granule cells in response to sensory stimulation or during motor control tasks (Jörntell and Ekerot, 2006;Knogler et al., 2017;Wagner et al., 2017;Giovannucci et al., 2017;Badura and De Zeeuw, 2017;Wagner et al., 2019), at odds with classical theories (Marr, 1969;Albus, 1971). Moreover, there is evidence that granule cell firing rates differ across cerebellar regions (Heath et al., 2014;Witter and De Zeeuw, 2015). ...
... While sparse representations are suitable for learning to categorize inputs into random categories, as predicted by classic theories, tasks involving structured input-output mappings benefit from denser representations ( Figure 2). This reconciles such theories with the observation of dense granule cell activation during movement (Knogler et al., 2017;Wagner et al., 2017;Giovannucci et al., 2017;Badura and De Zeeuw, 2017;Wagner et al., 2019). We also show that, in contrast to the task-dependence of optimal coding level, optimal anatomical values of granule cell and Purkinje cell connectivity are largely task-independent ( Figure 6). ...
... While dense activity has been taken as evidence against theories of combinatorial coding in cerebellar granule cells (Knogler et al., 2017;Wagner et al., 2019), our theory suggests that the two are not incompatible. Instead, the coding level of cerebellum-like regions may be determined by behavioral demands and the nature of the input to granule-like layers (Muscinelli et al., 2022). ...
The cerebellar granule cell layer has inspired numerous theoretical models of neural representations that support learned behaviors, beginning with the work of Marr and Albus. In these models, granule cells form a sparse, combinatorial encoding of diverse sensorimotor inputs. Such sparse representations are optimal for learning to discriminate random stimuli. However, recent observations of dense, low-dimensional activity across granule cells have called into question the role of sparse coding in these neurons. Here, we generalize theories of cerebellar learning to determine the optimal granule cell representation for tasks beyond random stimulus discrimination, including continuous input-output transformations as required for smooth motor control. We show that for such tasks, the optimal granule cell representation is substantially denser than predicted by classical theories. Our results provide a general theory of learning in cerebellum-like systems and suggest that optimal cerebellar representations are task-dependent.
... Whether a topographic map projection between cell subpopulations of inferior olive and cerebellum occurs in zebrafish like in mammals is not known yet. Nevertheless, physiological studies in the zebrafish larvae revealed regionally distinguishable sub-populations of PCs [63,97,98], as well as granule cell clusters in the different parts of the cerebellum [99]. This could suggest distinctive efferent projections, as well as afferent connections (via climbing and/or mossy fibers) ending on specific areas of the corpus cerebelli in the zebrafish. ...
... The main responsible neurons, which are required for the correct processing of neuronal activity within the cerebellum, are the highly conserved Purkinje cells [33,105]. These cells are also by far the most studied and best understood neuronal cell type in the zebrafish cerebellum regarding their electrophysiological properties [97,99,101,104,106,107]. As for other zebrafish cerebellar neurons, almost no electrophysiological data are available, this review will focus on the physiological activity of PCs. ...
... These findings all correlate perfectly with the time when zebrafish larvae are freely swimming and have to begin hunting their prey and avoid predators to survive [125] and demonstrate that the zebrafish cerebellum reaches functional maturity relatively early during development. This rapid time course makes zebrafish advantageous for electrophysiological in vivo studies of cerebellar development and functions in awake animals, which is relatively easy in this model organism and requires only a minimal invasive surgery [97,99,101,106,126]. In contrast, in rats, cerebellar cortical layers are not evident and Purkinje cells are not functionally mature until ~ 2-3 postnatal weeks, which corresponds to ~ 5-6 weeks post-fertilization [127]. ...
The cerebellum represents a brain compartment that first appeared in gnathostomes (jawed vertebrates). Besides the addition of cell numbers, its development, cytoarchitecture, circuitry, physiology, and function have been highly conserved throughout avian and mammalian species. While cerebellar research in avian and mammals is extensive, systematic investigations on this brain compartment in zebrafish as a teleostian model organism started only about two decades ago, but has provided considerable insight into cerebellar development, physiology, and function since then. Zebrafish are genetically tractable with nearly transparent small-sized embryos, in which cerebellar development occurs within a few days. Therefore, genetic investigations accompanied with non-invasive high-resolution in vivo time-lapse imaging represents a powerful combination for interrogating the behavior and function of cerebellar cells in their complex native environment.
... This is expected, given that multiple forms of plasticity have been shown within the synapses in this layer. For example, the mossy fibre-granule cell synapse undergoes both LTP and LTD [96], and evidence has shown that granule cell activity adapts over time during eyeblink conditioning [97] and other types of learning [98,99]. In addition, the induction of LTP by theta-burst stimulation in acute cerebellar slices activates a cAMP-responsive element-binding protein cascade which, in turn, activates c-Fos expression [100]. ...
Delay eyeblink conditioning has been extensively used to study associative learning and the cerebellar circuits underlying this task have been largely identified. However, there is a little knowledge on how factors such as strain, sex and innate behaviour influence performance during this type of learning. In this study, we used male and female mice of C57BL/6J (B6) and B6CBAF1 strains to investigate the effect of sex, strain and locomotion in delay eyeblink conditioning. We performed a short and a long delay eyeblink conditioning paradigm and used a c-Fos immunostaining approach to explore the involvement of different brain areas in this task. We found that both B6 and B6CBAF1 females reach higher learning scores compared to males in the initial stages of learning. This sex-dependent difference was no longer present as the learning progressed. Moreover, we found a strong positive correlation between learning scores and voluntary locomotion irrespective of the training duration. c-Fos immunostainings after the short paradigm showed positive correlations between c-Fos expression and learning scores in the cerebellar cortex and brainstem, as well as previously unreported areas. By contrast, after the long paradigm, c-Fos expression was only significantly elevated in the brainstem. Taken together, we show that differences in voluntary locomotion and activity across brain areas correlate with performance in delay eyeblink conditioning across strains and sexes.
... This result suggests that a disentangled movement encoding at the PC level emerges through substantial amplification of those seemingly insignificant variabilities in MF responses by the cerebellar network. Highly correlated activity, resulting in an apparently small dimensionality, has been widely observed in work on the cerebellar input layer [46][47][48] (but see also ref. 44). We found the same in our MF data, but our analysis together with the PC data suggests that enhancing small input variabilities is a fundamental information processing principle of the cerebellar network. ...
Both the environment and our body keep changing dynamically. Hence, ensuring movement precision requires adaptation to multiple demands occurring simultaneously. Here we show that the cerebellum performs the necessary multi-dimensional computations for the flexible control of different movement parameters depending on the prevailing context. This conclusion is based on the identification of a manifold-like activity in both mossy fibers (MFs, network input) and Purkinje cells (PCs, output), recorded from monkeys performing a saccade task. Unlike MFs, the PC manifolds developed selective representations of individual movement parameters. Error feedback-driven climbing fiber input modulated the PC manifolds to predict specific, error type-dependent changes in subsequent actions. Furthermore, a feed-forward network model that simulated MF-to-PC transformations revealed that amplification and restructuring of the lesser variability in the MF activity is a pivotal circuit mechanism. Therefore, the flexible control of movements by the cerebellum crucially depends on its capacity for multi-dimensional computations.
... Specifically, a very large number of GCs and sparse synaptic connectivity from MFs to GCs have been theoretically shown to be beneficial to demix input signals entering through MFs. However, recent studies using in vivo calcium imaging have challenged this idea by showing higher densities of GC activations than predicted [45][46][47] . Our finding of preferential connections, in which MFs originating from a certain precerebellar nucleus preferentially innervate GCs having neighboring PFs, implies higher chances for the simultaneous activation of two or more dendrites of individual GCs when a specific type of input signal enters. ...
The long-standing hypothesis that synapses between mossy fibers (MFs) and cerebellar granule cells (GCs) are organized according to the origins of MFs and locations of GC axons, parallel fibers (PFs), is supported by recent findings. However, the mechanisms of such organized synaptic connections remain unknown. Here, using our technique that enabled PF location-dependent labeling of GCs in mice, we confirmed that synaptic connections of GCs with specific MFs originating from the pontine nucleus (PN-MFs) and dorsal column nuclei (DCoN-MFs) were gently but differentially organized according to their PF locations. We then found that overall MF-GC synaptic connectivity was biased in a way that dendrites of GCs having nearby PFs tended to connect with the same MF terminals, implying that the MF origin- and PF location-dependent organization is associated with the overall biased MF-GC synaptic connectivity. Furthermore, the development of PN-MFs preceded that of DCoN-MFs, which matches the developmental sequence of GCs that preferentially connect with each type of these MFs. Thus, our results revealed that overall MF-GC synaptic connectivity is biased in terms of PF locations, and suggested that such connectivity is likely the result of synaptic formation between developmental timing-matched partners.
... In addition, there were experimental results implying GrC's Gaussian tuning for specific input patterns. Zebrafish GrCs demonstrated a receptive field spanning 5-25% of the visual field (Knogler et al., 2017), and mouse GrCs exhibited narrow tuning to whisker set point at the level of individual cell while populations seemed to linearly encode a broad range of movement (Chen et al., 2017). ...
Sensorimotor information provided by mossy fibers (MF) is mapped to high-dimensional space by a huge number of granule cells (GrC) in the cerebellar cortex’s input layer. Significant studies have demonstrated the computational advantages and primary contributor of this expansion recoding. Here, we propose a novel perspective on the expansion recoding where each GrC serve as a kernel basis function, thereby the cerebellum can operate like a kernel machine that implicitly use high dimensional (even infinite) feature spaces. We highlight that the generation of kernel basis function is indeed biologically plausible scenario, considering that the key idea of kernel machine is to memorize important input patterns. We present potential regimes for developing kernels under constrained resources and discuss the advantages and disadvantages of each regime using various simulation settings.
