publications

2025

1. Barcoded Rabies In Situ Connectomics for high-throughput reconstruction of neural circuits

   Alexander Becalick, Antonin Blot, Molly Strom, and Petr Znamenskiy

   bioRxiv, 2025

   Abs DOI

   Sequencing of oligonucleotide barcodes holds promise as a high-throughput approach for reconstructing synaptic connectivity at scale. Rabies viruses can act as a vehicle for barcode transmission, thanks to their ability to spread between synaptically connected cells. However, applying barcoded rabies viruses to map synaptic connections in vivo has proved challenging. Here, we develop Barcoded Rabies In Situ Connectomics (BRISC) for high-throughput connectivity mapping in the mouse brain. To ensure that the majority of post-synaptic "starter" neurons are uniquely labeled with distinct barcode sequences, we first generated libraries of rabies viruses with sufficient diversity to label >1000 neurons uniquely. To minimize the probability of barcode transmission between starter neurons, we developed a strategy to tightly control their density. We then applied BRISC to map inputs of single neurons in the primary visual cortex (V1). Using in situ sequencing, we read out the expression of viral barcodes in rabies-infected neurons, while preserving spatial information. We then matched barcode sequences between starter and presynaptic neurons, mapping the inputs of 385 neurons and identifying 7,814 putative synaptic connections. The resulting connectivity matrix revealed layer- and cell-type-specific local connectivity rules and topographic organization of long-range inputs to V1. These results show that BRISC can simultaneously resolve the synaptic connectivity of hundreds of neurons while preserving spatial information, enabling reconstruction of neural circuits at an unprecedented scale.

2. PhotoNeuro: A compact photodetector for synchronization of visual stimulus presentation during behavioural experiments in neuroscience

   Xavier Cano Ferrer, Marcelo Moglie, George Konstantinou, Antonin Blot, Gaia Bianchini, and 3 more authors

   HardwareX, Jul 2025

   Abs DOI

   Presenting visual stimuli in neuroscience experiments often requires precise temporal alignment between visual events and electrophysiological or behavioural recordings. This is typically achieved by combining analogue signals that convey timing information about the visual cue shown on liquid crystal displays (LCDs), sensed via photodetectors and recorded through analogue-to-digital converter (ADC) acquisition boards. However, most commercial photodetector systems pose limitations such as high voltage requirements, large sensor footprints that interfere with stimulus presentation, and limited compatibility with open-source platforms. Here, we present a compact, low-cost photodetector system designed for compatibility with common 3.3–5 V microcontroller-based development boards (e.g., Arduino) and the open-source visual programming language Bonsai, widely used in neuroscience for experiment control. The circuit consists of a photodiode, an amplification stage, and a low-pass filter, and can optionally incorporate an infrared filter—useful for experiments involving infrared touch displays. To facilitate reproducibility, we provide complete design files, a bill of materials and detailed building and operational instructions. We further introduce a four-channel variant, enabling the detection of four-bit binary signals for more complex synchronization needs. Validation and characterization of the device were performed through grayscale gamma correction analysis of LCD monitors using Bonsai. Additionally, we demonstrate the system’s utility in a head-fixed mouse experiment, synchronizing visual stimulus onset with neuronal recordings acquired via Neuropixels 2.0 probes. Performance comparisons with a commercial photodetector device indicate that our system achieves equivalent signal fidelity at a substantially lower cost, while maintaining a minimal footprint suitable for experimental use.

2024

1. A depth map of visual space in the primary visual cortex

   Yiran He, Antonio Colas Nieto, Antonin Blot, and Petr Znamenskiy

   Sep 2024

   Pages: 2024.09.27.615442 Section: New Results

   Abs DOI

   Depth perception is essential for visually-guided behavior. Computer vision algorithms use depth maps to encode distances in three-dimensional scenes but it is unknown whether such depth maps are generated by animal visual systems. To answer this question, we focused on motion parallax, a depth cue relying on visual motion resulting from movement of the observer. As neurons in the mouse primary visual cortex (V1) are broadly modulated by locomotion, we hypothesized that they may integrate vision- and locomotion-related signals to estimate depth from motion parallax. Using recordings in a three-dimensional virtual reality environment, we found that conjunctive coding of visual and self-motion speeds gave rise to depth-selective neuronal responses. Depth-selective neurons could be characterized by three-dimensional receptive fields, responding to specific virtual depths and retinotopic locations. Neurons tuned to a broad range of virtual depths were found across all sampled retinotopic locations, showing that motion parallax generates a depth map of visual space in V1.

2023

1. Learning shapes cortical dynamics to enhance integration of relevant sensory input

   Angus Chadwick, Adil G. Khan, Jasper Poort, Antonin Blot, Sonja B. Hofer, and 2 more authors

   Neuron, Jan 2023

   Abs DOI

   Adaptive sensory behavior is thought to depend on processing in recurrent cortical circuits, but how dynamics in these circuits shapes the integration and transmission of sensory information is not well understood. Here, we study neural coding in recurrently connected networks of neurons driven by sensory input. We show analytically how information available in the network output varies with the alignment between feedforward input and the integrating modes of the circuit dynamics. In light of this theory, we analyzed neural population activity in the visual cortex of mice that learned to discriminate visual features. We found that over learning, slow patterns of network dynamics realigned to better integrate input relevant to the discrimination task. This realignment of network dynamics could be explained by changes in excitatory-inhibitory connectivity among neurons tuned to relevant features. These results suggest that learning tunes the temporal dynamics of cortical circuits to optimally integrate relevant sensory input.

2022

1. Learning and attention increase visual response selectivity through distinct mechanisms

   Jasper Poort, Katharina A. Wilmes, Antonin Blot, Angus Chadwick, Maneesh Sahani, and 4 more authors

   Neuron, Feb 2022

   Abs DOI

   Selectivity of cortical neurons for sensory stimuli can increase across days as animals learn their behavioral relevance and across seconds when animals switch attention. While both phenomena occur in the same circuit, it is unknown whether they rely on similar mechanisms. We imaged primary visual cortex as mice learned a visual discrimination task and subsequently performed an attention switching task. Selectivity changes due to learning and attention were uncorrelated in individual neurons. Selectivity increases after learning mainly arose from selective suppression of responses to one of the stimuli but from selective enhancement and suppression during attention. Learning and attention differentially affected interactions between excitatory and PV, SOM, and VIP inhibitory cells. Circuit modeling revealed that cell class-specific top-down inputs best explained attentional modulation, while reorganization of local functional connectivity accounted for learning-related changes. Thus, distinct mechanisms underlie increased discriminability of relevant sensory stimuli across longer and shorter timescales.

2021

1. Visual intracortical and transthalamic pathways carry distinct information to cortical areas

   Antonin Blot, Morgane M. Roth, Ioana Gasler, Mitra Javadzadeh, Fabia Imhof, and 1 more author

   Neuron, Jun 2021

   Abs DOI

   Sensory processing involves information flow between neocortical areas, assumed to rely on direct intracortical projections. However, cortical areas may also communicate indirectly via higher-order nuclei in the thalamus, such as the pulvinar or lateral posterior nucleus (LP) in the visual system of rodents. The fine-scale organization and function of these cortico-thalamo-cortical pathways remains unclear. We find that responses of mouse LP neurons projecting to higher visual areas likely derive from feedforward input from primary visual cortex (V1) combined with information from many cortical and subcortical areas, including superior colliculus. Signals from LP projections to different higher visual areas are tuned to specific features of visual stimuli and their locomotor context, distinct from the signals carried by direct intracortical projections from V1. Thus, visual transthalamic pathways are functionally specific to their cortical target, different from feedforward cortical pathways, and combine information from multiple brain regions, linking sensory signals with behavioral context.

2. Learning and attention increase visual response selectivity through distinct mechanisms

   Jasper Poort, Katharina A. Wilmes, Antonin Blot, Angus Chadwick, Maneesh Sahani, and 4 more authors

   bioRxiv, Feb 2021

   Publisher: Cold Spring Harbor Laboratory Section: New Results

   Abs DOI

   \textlessp\textgreaterSelectivity of cortical neurons for sensory stimuli can increase across days as animals learn their behavioral relevance, and across seconds when animals switch attention. While both phenomena are expressed in the same cortical circuit, it is unknown whether they rely on similar mechanisms. We imaged activity of the same neuronal populations in primary visual cortex as mice learned a visual discrimination task and subsequently performed an attention switching task. Selectivity changes due to learning and attention were uncorrelated in individual neurons. Selectivity increases after learning mainly arose from selective suppression of responses to one of the task relevant stimuli but from selective enhancement and suppression during attention. Learning and attention differentially affected interactions between excitatory and PV, SOM and VIP inhibitory cell classes. Circuit modelling revealed that cell class-specific top-down inputs best explained attentional modulation, while the reorganization of local functional connectivity accounted for learning related changes. Thus, distinct mechanisms underlie increased discriminability of relevant sensory stimuli across longer and shorter time scales.\textless/p\textgreater

3. A database and deep learning toolbox for noise-optimized, generalized spike inference from calcium imaging

   Peter Rupprecht, Stefano Carta, Adrian Hoffmann, Mayumi Echizen, Antonin Blot, and 6 more authors

   Nature Neuroscience, Sep 2021

   Number: 9 Publisher: Nature Publishing Group

   Abs DOI

   Inference of action potentials (’spikes’) from neuronal calcium signals is complicated by the scarcity of simultaneous measurements of action potentials and calcium signals (‘ground truth’). In this study, we compiled a large, diverse ground truth database from publicly available and newly performed recordings in zebrafish and mice covering a broad range of calcium indicators, cell types and signal-to-noise ratios, comprising a total of more than 35 recording hours from 298 neurons. We developed an algorithm for spike inference (termed CASCADE) that is based on supervised deep networks, takes advantage of the ground truth database, infers absolute spike rates and outperforms existing model-based algorithms. To optimize performance for unseen imaging data, CASCADE retrains itself by resampling ground truth data to match the respective sampling rate and noise level; therefore, no parameters need to be adjusted by the user. In addition, we developed systematic performance assessments for unseen data, openly released a resource toolbox and provide a user-friendly cloud-based implementation.

2019

1. Cerebellar Contribution to Preparatory Activity in Motor Neocortex

   Francois P. Chabrol, Antonin Blot, and Thomas D. Mrsic-Flogel

   Neuron, Jun 2019

   Abs DOI

   Summary In motor neocortex, preparatory activity predictive of specific movements is maintained by a positive feedback loop with the thalamus. Motor thalamus receives excitatory input from the cerebellum, which learns to generate predictive signals for motor control. The contribution of this pathway to neocortical preparatory signals remains poorly understood. Here, we show that, in a virtual reality conditioning task, cerebellar output neurons in the dentate nucleus exhibit preparatory activity similar to that in anterolateral motor cortex prior to reward acquisition. Silencing activity in dentate nucleus by photoactivating inhibitory Purkinje cells in the cerebellar cortex caused robust, short-latency suppression of preparatory activity in anterolateral motor cortex. Our results suggest that preparatory activity is controlled by a learned decrease of Purkinje cell firing in advance of reward under supervision of climbing fiber inputs signaling reward delivery. Thus, cerebellar computations exert a powerful influence on preparatory activity in motor neocortex.

2018

1. Cerebellar learning using perturbations

   Guy Bouvier, Johnatan Aljadeff, Claudia Clopath, Célian Bimbard, Jonas Ranft, and 5 more authors

   eLife, Nov 2018

   Abs DOI

   The cerebellum aids the learning of fast, coordinated movements. According to current consensus, erroneously active parallel fibre synapses are depressed by complex spikes signalling movement errors. However, this theory cannot solve the credit assignment problem of processing a global movement evaluation into multiple cell-specific error signals. We identify a possible implementation of an algorithm solving this problem, whereby spontaneous complex spikes perturb ongoing movements, create eligibility traces and signal error changes guiding plasticity. Error changes are extracted by adaptively cancelling the average error. This framework, stochastic gradient descent with estimated global errors (SGDEGE), predicts synaptic plasticity rules that apparently contradict the current consensus but were supported by plasticity experiments in slices from mice under conditions designed to be physiological, highlighting the sensitivity of plasticity studies to experimental conditions. We analyse the algorithm’s convergence and capacity. Finally, we suggest SGDEGE may also operate in the basal ganglia.

2. Distinct learning-induced changes in stimulus selectivity and interactions of GABAergic interneuron classes in visual cortex

   Adil G. Khan, Jasper Poort, Angus Chadwick, Antonin Blot, Maneesh Sahani, and 2 more authors

   Nature Neuroscience, Jun 2018

   Abs DOI

   Khan et al. simultaneously measured activity from excitatory cells and three classes of inhibitory interneurons in visual cortex and show that learning differentially shapes the stimulus selectivity and interactions of multiple cell classes.

2016

1. Time-invariant feed-forward inhibition of Purkinje cells in the cerebellar cortex in vivo

   Antonin Blot, Camille de Solages, Srdjan Ostojic, German Szapiro, Vincent Hakim, and 1 more author

   The Journal of Physiology, Jun 2016

   Abs DOI

   Key points We performed extracellular recording of pairs of interneuron–Purkinje cells in vivo. A single interneuron produces a substantial, short-lasting, inhibition of Purkinje cells. Feed-forward inhibition is associated with characteristic asymmetric cross-correlograms. In vivo, Purkinje cell spikes only depend on the most recent synaptic activity. Abstract Cerebellar molecular layer interneurons are considered to control the firing rate and spike timing of Purkinje cells. However, interactions between these cell types are largely unexplored in vivo. Using tetrodes, we performed simultaneous extracellular recordings of neighbouring Purkinje cells and molecular layer interneurons, presumably basket cells, in adult rats in vivo. The high levels of afferent synaptic activity encountered in vivo yield irregular spiking and reveal discharge patterns characteristic of feed-forward inhibition, thus suggesting an overlap of the afferent excitatory inputs between Purkinje cells and basket cells. Under conditions of intense background synaptic inputs, interneuron spikes exert a short-lasting inhibitory effect, delaying the following Purkinje cell spike by an amount remarkably independent of the Purkinje cell firing cycle. This effect can be explained by the short memory time of the Purkinje cell potential as a result of the intense incoming synaptic activity. Finally, we found little evidence for any involvement of the interneurons that we recorded with the cerebellar high-frequency oscillations promoting Purkinje cell synchrony. The rapid interactions between interneurons and Purkinje cells might be of particular importance in fine motor control because the inhibitory action of interneurons on Purkinje cells leads to deep cerebellar nuclear disinhibition and hence increased cerebellar output.

2014

1. Ultra-rapid axon-axon ephaptic inhibition of cerebellar Purkinje cells by the pinceau

   Antonin Blot and Boris Barbour

   Nature Neuroscience, Feb 2014

   Abs DOI

   Excitatory synaptic activity in the brain is shaped and balanced by inhibition. Because inhibition cannot propagate, it is often recruited with a synaptic delay by incoming excitation. Cerebellar Purkinje cells are driven by long-range excitatory parallel fiber inputs, which also recruit local inhibitory basket cells. The axon initial segment of each Purkinje cell is ensheathed by basket cell axons in a structure called the pinceau, which is largely devoid of chemical synapses. In mice, we found at the single-cell level that the pinceau mediates ephaptic inhibition of Purkinje cell firing at the site of spike initiation. The reduction of firing rate was synchronous with the presynaptic action potential, eliminating a synaptic delay and allowing granule cells to inhibit Purkinje cells without a preceding phase of excitation. Axon-axon ephaptic intercellular signaling can therefore mediate near-instantaneous feedforward and lateral inhibition.

2013

1. Analysis of the study of the cerebellar pinceau by Korn and Axelrad

   Antonin Blot and Boris Barbour

   bioRxiv, Jan 2013

   The axon initial segment of each cerebellar Purkinje cell is ensheathed by basket cell axons in a structure called the pinceau, which is largely devoid of chemical synapses and gap junctions. These facts and ultrastructural similarities with the axon cap of the teleost Mauthner cell led to the conjecture that the pinceau mediates ephaptic (via the extracellular field) inhibition. Korn and Axelrad published a study in 1980 in which they reported confirmation of this conjecture. We have analysed their results and show that most are likely to be explained by an artefactual signal arising from the massive stimulation of parallel fibres they employed. We reproduce their experiments and confirm that all of their results are consistent with this artefact. Their data therefore provide no evidence regarding the operation of the pinceau.

   Abs DOI

   The axon initial segment of each cerebellar Purkinje cell is ensheathed by basket cell axons in a structure called the pinceau, which is largely devoid of chemical synapses and gap junctions. These facts and ultrastructural similarities with the axon cap of the teleost Mauthner cell led to the conjecture that the pinceau mediates ephaptic (via the extracellular field) inhibition. Korn and Axelrad published a study in 1980 in which they reported confirmation of this conjecture. We have analysed their results and show that most are likely to be explained by an artefactual signal arising from the massive stimulation of parallel fibres they employed. We reproduce their experiments and confirm that all of their results are consistent with this artefact. Their data therefore provide no evidence regarding the operation of the pinceau.

2. Functions of cerebellar molecular layer interneurones

   Antonin Blot

   Paris 6, Jan 2013

   Abs

   In the central nervous system, numerous functions have been proposed for inhibition. However, due to the large heterogeneity of inhibitory interneurones, it remains difficult to attribute one defined cellular substrate to these functions. The relative simplicity of the cerebellar cortex, and notably of its molecular layer, which contains only two types of interneurones, offers the possibility to tackle this question. During my thesis, I demonstrated that the pinceau, whichd basket cells, one type of molecular layer interneurone, form around the Purkinje cell axon initial segment, allows quasi-instantaneous ephaptic inhibition of Purkinje cells. Basket cell action potentials propagate passively to the pinceau and induce a capacitive and a potassium current, which produce a positivity in the extracellular field. This ∼200 μV positivity is synchronous with the basket cell spike. It delays Purkinje cell action potentials by one millisecond on average, suppressing any synaptic delay, and thus creates a functional equivalent of inhibitory synapses between granule cells and Purkinje cells. This thesis also presents an analysis of molecular layer interneurone activity in vivo and the first results of an electrophysiological caracterisation in adult animals of these interneurones and of the connections they establish on Purkinje cells.

2009

1. Functional expression of two system A glutamine transporter isoforms in rat auditory brainstem neurons

   A. Blot, D. Billups, M. Bjørkmo, A.Z. Quazi, N.M. Uwechue, and 2 more authors

   Neuroscience, Dec 2009

   Abs DOI

   Glutamine plays multiple roles in the CNS, including metabolic functions and production of the neurotransmitters glutamate and GABA. It has been proposed to be taken up into neurons via a variety of membrane transport systems, including system A, which is a sodium-dependent electrogenic amino acid transporter system. In this study, we investigate glutamine transport by application of amino acids to individual principal neurons of the medial nucleus of the trapezoid body (MNTB) in acutely isolated rat brain slices. A glutamine transport current was studied in patch-clamped neurons, which had the electrical and pharmacological properties of system A: it was sodium-dependent, had a non-reversing current-voltage relationship, was activated by proline, occluded by N-(methylamino)isobutyric acid (MeAIB), and was unaffected by 2-aminobicyclo-[2.2.1]-heptane-2-carboxylic acid (BCH). Additionally, we examined the expression of different system A transporter isoforms using immunocytochemical staining with antibodies raised against system A transporter 1 and 2 (SAT1 and SAT2). Our results indicate that both isoforms are expressed in MNTB principal neurons, and demonstrate that functional system A transporters are present in the plasma membrane of neurons. Since system A transport is highly regulated by a number of cellular signaling mechanisms and glutamine then goes on to activate other pathways, the study of these transporters in situ gives an indication of the mechanisms of neuronal glutamine supply as well as points of regulation of neurotransmitter production, cellular signaling and metabolism in the native neuronal environment.