Cell Type-specific Connectivity Research Articles

Topographical and cell type-specific connectivity of rostral and caudal forelimb corticospinal neuron populations

Corticospinal neurons (CSNs) synapse directly on spinal neurons, a diverse assortment of cells with unique structural and functional properties necessary for body movements. CSNs modulating forelimb behavior fractionate into caudal forelimb area (CFA) and rostral forelimb area (RFA) motor cortical populations. Despite their prominence, the full diversity of spinal neurons targeted by CFA and RFA CSNs is uncharted. Here, we use anatomical and RNA sequencing methods to show that CSNs synapse onto a remarkably selective group of spinal cell types, favoring inhibitory populations that regulate motoneuron activity and gate sensory feedback. CFA and RFA CSNs target similar spinal neuron types, with notable exceptions that suggest that these populations differ in how they influence behavior. Finally, axon collaterals of CFA and RFA CSNs target similar brain regions yet receive highly divergent inputs. These results detail the rules of CSN connectivity throughout the brain and spinal cord for two regions critical for forelimb behavior.

A reservoir of timescales emerges in recurrent circuits with heterogeneous neural assemblies.

The temporal activity of many physical and biological systems, from complex networks to neural circuits, exhibits fluctuations simultaneously varying over a large range of timescales. Long-tailed distributions of intrinsic timescales have been observed across neurons simultaneously recorded within the same cortical circuit. The mechanisms leading to this striking temporal heterogeneity are yet unknown. Here, we show that neural circuits, endowed with heterogeneous neural assemblies of different sizes, naturally generate multiple timescales of activity spanning several orders of magnitude. We develop an analytical theory using rate networks, supported by simulations of spiking networks with cell-type specific connectivity, to explain how neural timescales depend on assembly size and show that our model can naturally explain the long-tailed timescale distribution observed in the awake primate cortex. When driving recurrent networks of heterogeneous neural assemblies by a time-dependent broadband input, we found that large and small assemblies preferentially entrain slow and fast spectral components of the input, respectively. Our results suggest that heterogeneous assemblies can provide a biologically plausible mechanism for neural circuits to demix complex temporal input signals by transforming temporal into spatial neural codes via frequency-selective neural assemblies.

Retinal input integration in excitatory and inhibitory neurons in the mouse superior colliculus in vivo.

The superior colliculus (SC) is a midbrain structure that receives inputs from retinal ganglion cells (RGCs). The SC contains one of the highest densities of inhibitory neurons in the brain but whether excitatory and inhibitory SC neurons differentially integrate retinal activity in vivo is still largely unknown. We recently established a recording approach to measure the activity of RGCs simultaneously with their postsynaptic SC targets in vivo, to study how SC neurons integrate RGC activity. Here, we employ this method to investigate the functional properties that govern retinocollicular signaling in a cell type-specific manner by identifying GABAergic SC neurons using optotagging in VGAT-ChR2 mice. Our results demonstrate that both excitatory and inhibitory SC neurons receive comparably strong RGC inputs and similar wiring rules apply for RGCs innervation of both SC cell types, unlike the cell type-specific connectivity in the thalamocortical system. Moreover, retinal activity contributed more to the spiking activity of postsynaptic excitatory compared to inhibitory SC neurons. This study deepens our understanding of cell type-specific retinocollicular functional connectivity and emphasizes that the two major brain areas for visual processing, the visual cortex and the SC, differently integrate sensory afferent inputs.

Retinal input integration in excitatory and inhibitory neurons in the mouse superior colliculus in vivo

The superior colliculus (SC) is a midbrain structure that receives inputs from retinal ganglion cells (RGCs). The SC contains one of the highest densities of inhibitory neurons in the brain but whether excitatory and inhibitory SC neurons differentially integrate retinal activity in vivo is still largely unknown. We recently established a recording approach to measure the activity of RGCs simultaneously with their postsynaptic SC targets in vivo, to study how SC neurons integrate RGC activity. Here, we employ this method to investigate the functional properties that govern retinocollicular signaling in a cell type-specific manner by identifying GABAergic SC neurons using optotagging in VGAT-ChR2 mice. Our results demonstrate that both excitatory and inhibitory SC neurons receive comparably strong RGC inputs and similar wiring rules apply for RGCs innervation of both SC cell types, unlike the cell type-specific connectivity in the thalamocortical system. Moreover, retinal activity contributed more to the spiking activity of postsynaptic excitatory compared to inhibitory SC neurons. This study deepens our understanding of cell type-specific retinocollicular functional connectivity and emphasizes that the two major brain areas for visual processing, the visual cortex and the SC, differently integrate sensory afferent inputs.

Ventral Subiculum Inputs to Nucleus Accumbens Medial Shell Preferentially Innervate D2R Medium Spiny Neurons and Contain Calcium Permeable AMPARs.

Ventral subiculum (vSUB) is the major output region of ventral hippocampus (vHIPP) and sends major projections to nucleus accumbens medial shell (NAcMS). Hyperactivity of the vSUB-NAcMS circuit is associated with substance use disorders and the modulation of vSUB activity alters drug seeking and drug reinstatement behavior in rodents. However, to the best of our knowledge, the cell type-specific connectivity and synaptic transmission properties of the vSUB-NAcMS circuit have never been directly examined. Instead, previous functional studies have focused on total ventral hippocampal (vHIPP) output to NAcMS without distinguishing vSUB from other subregions of vHIPP, including ventral CA1 (vCA1). Using ex vivo electrophysiology, we systematically characterized the vSUB-NAcMS circuit with cell type- and synapse-specific resolution in male and female mice and found that vSUB output to dopamine receptor type-1 (D1R) and type-2 (D2R) expressing medium spiny neurons (MSNs) displays a functional connectivity bias for D2R MSNs. Furthermore, we found that vSUB-D1R and vSUB-D2R MSN synapses contain calcium-permeable AMPA receptors in drug-naive mice. Finally, we find that, distinct from other glutamatergic inputs, cocaine exposure selectively induces plasticity at vSUB-D2R synapses. Importantly, we directly compared vSUB and vCA1 output to NAcMS and found that vSUB synapses are functionally distinct and that vCA1 output recapitulated the synaptic properties previously ascribed to vHIPP. Our work highlights the need to consider the contributions of individual subregions of vHIPP to substance use disorders and represents an important first step toward understanding how the vSUB-NAcMS circuit contributes to the etiologies that underlie substance use disorders.SIGNIFICANCE STATEMENT Inputs to nucleus accumbens (NAc) dopamine receptor type 1 (D1R) and D2R medium spiny neurons (MSNs) are critically involved in reward seeking behavior. Ventral subiculum (vSUB) provides robust synaptic input to nucleus accumbens medial shell (NAcMS) and activity of this circuit is linked to substance use disorders. Despite the importance of the vSUB to nucleus accumbens circuit, the functional connectivity and synaptic transmission properties have not been tested. Here, we systematically interrogated these properties and found that basal connectivity and drug-induced plasticity are biased for D2R medium spiny neurons. Overall, we demonstrate that this circuit is distinct from synaptic inputs from other brain regions, which helps to explain how vSUB dysfunction contributes to the etiologies that underlie substance use disorders.

The impact of neuron morphology on cortical network architecture.

SummaryThe neurons in the cerebral cortex are not randomly interconnected. This specificity in wiring can result from synapse formation mechanisms that connect neurons, depending on their electrical activity and genetically defined identity. Here, we report that the morphological properties of the neurons provide an additional prominent source by which wiring specificity emerges in cortical networks. This morphologically determined wiring specificity reflects similarities between the neurons’ axo-dendritic projections patterns, the packing density, and the cellular diversity of the neuropil. The higher these three factors are, the more recurrent is the topology of the network. Conversely, the lower these factors are, the more feedforward is the network’s topology. These principles predict the empirically observed occurrences of clusters of synapses, cell type-specific connectivity patterns, and nonrandom network motifs. Thus, we demonstrate that wiring specificity emerges in the cerebral cortex at subcellular, cellular, and network scales from the specific morphological properties of its neuronal constituents.

NOSA, an Analytical Toolbox for Multicellular Optical Electrophysiology.

Understanding how neural networks generate activity patterns and communicate with each other requires monitoring the electrical activity from many neurons simultaneously. Perfectly suited tools for addressing this challenge are genetically encoded voltage indicators (GEVIs) because they can be targeted to specific cell types and optically report the electrical activity of individual, or populations of neurons. However, analyzing and interpreting the data from voltage imaging experiments is challenging because high recording speeds and properties of current GEVIs yield only low signal-to-noise ratios, making it necessary to apply specific analytical tools. Here, we present NOSA (Neuro-Optical Signal Analysis), a novel open source software designed for analyzing voltage imaging data and identifying temporal interactions between electrical activity patterns of different origin. In this work, we explain the challenges that arise during voltage imaging experiments and provide hands-on analytical solutions. We demonstrate how NOSA’s baseline fitting, filtering algorithms and movement correction can compensate for shifts in baseline fluorescence and extract electrical patterns from low signal-to-noise recordings. NOSA allows to efficiently identify oscillatory frequencies in electrical patterns, quantify neuronal response parameters and moreover provides an option for analyzing simultaneously recorded optical and electrical data derived from patch-clamp or other electrode-based recordings. To identify temporal relations between electrical activity patterns we implemented different options to perform cross correlation analysis, demonstrating their utility during voltage imaging in Drosophila and mice. All features combined, NOSA will facilitate the first steps into using GEVIs and help to realize their full potential for revealing cell-type specific connectivity and functional interactions.

Fluorescence-Based Quantitative Synapse Analysis for Cell Type-Specific Connectomics

Anatomical methods for determining cell type-specific connectivity are essential to inspire and constrain our understanding of neural circuit function. We developed genetically-encoded reagents for fluorescence-synapse labeling and connectivity analysis in brain tissue, using a fluorogen-activating protein (FAP)-coupled or YFP-coupled, postsynaptically-localized neuroligin-1 (NL-1) targeting sequence (FAP/YFPpost). FAPpost expression did not alter mEPSC or mIPSC properties. Sparse AAV-mediated expression of FAP/YFPpost with the cell-filling, red fluorophore dTomato (dTom) enabled high-throughput, compartment-specific detection of putative synapses across diverse neuron types in mouse somatosensory cortex. We took advantage of the bright, far-red emission of FAPpost puncta for multichannel fluorescence alignment of dendrites, FAPpost puncta, and presynaptic neurites in transgenic mice with saturated labeling of parvalbumin (PV), somatostatin (SST), or vasoactive intestinal peptide (VIP)-expressing neurons using Cre-reporter driven expression of YFP. Subtype-specific inhibitory connectivity onto layer 2/3 (L2/3) neocortical pyramidal (Pyr) neurons was assessed using automated puncta detection and neurite apposition. Quantitative and compartment-specific comparisons show that PV inputs are the predominant source of inhibition at both the soma and the dendrites and were particularly concentrated at the primary apical dendrite. SST inputs were interleaved with PV inputs at all secondary-order and higher-order dendritic branches. These fluorescence-based synapse labeling reagents can facilitate large-scale and cell-type specific quantitation of changes in synaptic connectivity across development, learning, and disease states.

Maintenance of cell type-specific connectivity and circuit function requires Tao kinase

Sensory circuits are typically established during early development, yet how circuit specificity and function are maintained during organismal growth has not been elucidated. To gain insight we quantitatively investigated synaptic growth and connectivity in the Drosophila nociceptive network during larval development. We show that connectivity between primary nociceptors and their downstream neurons scales with animal size. We further identified the conserved Ste20-like kinase Tao as a negative regulator of synaptic growth required for maintenance of circuit specificity and connectivity. Loss of Tao kinase resulted in exuberant postsynaptic specializations and aberrant connectivity during larval growth. Using functional imaging and behavioral analysis we show that loss of Tao-induced ectopic synapses with inappropriate partner neurons are functional and alter behavioral responses in a connection-specific manner. Our data show that fine-tuning of synaptic growth by Tao kinase is required for maintaining specificity and behavioral output of the neuronal network during animal growth.

Viral labeling of neurons synaptically connected to nucleus accumbens somatostatin interneurons.

The nucleus accumbens, a key brain reward region, receives synaptic inputs from a range of forebrain and brainstem regions. Many of these projections have been established using electrophysiology or fluorescent tract tracing. However, more recently developed viral tracing techniques have allowed for fluorescent labeling of synaptic afferents in a cell type-specific manner. Since the NAc is comprised of multiple cell types, these methods have enabled the delineation of the cell type-specific connectivity of principal medium spiny neurons in the region. The synaptic connectivity of somatostatin interneurons, which account for <5% of the neurons in the region, has been inferred from electrophysiological and immunohistochemical data, but has not yet been visualized using modern viral tracing techniques. Here, we use the pseudorabies virus (PRV)-Introvert-GFP virus, an alphaherpes virus previously shown to label synaptic afferents in a cell type-specific manner, to label first order afferents to NAc somatostatin interneurons. While we find GFP(+) labeling in several well established projections to the NAc, we also observe that several known projections to NAc did not contain GFP(+) cells, suggesting they do not innervate somatostatin interneurons in the region. A subset of the GFP(+) afferents are c-FOS(+) following acute administration of cocaine, showing that NAc somatostatin interneurons are innervated by some cells that respond to rewarding stimuli. These results provide a foundation for future studies aimed toward elucidating the cell type-specific connectivity of the NAc, and identify specific circuits that warrant future functional characterization.

Conserved neural circuit structure across Drosophila larval development revealed by comparative connectomics.

During postembryonic development, the nervous system must adapt to a growing body. How changes in neuronal structure and connectivity contribute to the maintenance of appropriate circuit function remains unclear. Previously , we measured the cellular neuroanatomy underlying synaptic connectivity in Drosophila (Schneider-Mizell et al., 2016). Here, we examined how neuronal morphology and connectivity change between first instar and third instar larval stages using serial section electron microscopy. We reconstructed nociceptive circuits in a larva of each stage and found consistent topographically arranged connectivity between identified neurons. Five-fold increases in each size, number of terminal dendritic branches, and total number of synaptic inputs were accompanied by cell type-specific connectivity changes that preserved the fraction of total synaptic input associated with each pre-synaptic partner. We propose that precise patterns of structural growth act to conserve the computational function of a circuit, for example determining the location of a dangerous stimulus.

Determination of the connectivity of newborn neurons in mammalian olfactory circuits.

The mammalian olfactory bulb is a forebrain structure just one synapse downstream from the olfactory sensory neurons and performs the complex computations of sensory inputs. The formation of this sensory circuit is shaped through activity-dependent and cell-intrinsic mechanisms. Recent studies have revealed that cell-type specific connectivity and the organization of synapses in dendritic compartments are determined through cell-intrinsic programs already preset in progenitor cells. These progenitor programs give rise to subpopulations within a neuron type that have distinct synaptic organizations. The intrinsically determined formation of distinct synaptic organizations requires factors from contacting cells that match the cell-intrinsic programs. While certain genes control wiring within the newly generated neurons, other regulatory genes provide intercellular signals and are only expressed in neurons that will form contacts with the newly generated cells. Here, the olfactory system has provided a useful model circuit to reveal the factors regulating assembly of the highly structured connectivity in mammals.

PathCellNet: Cell-type specific pathogen-response network explorer

Pathogen specific immune response is a complex interplay between several innate and adaptive immune cell-types. Innate immune cells play a critical role in pathogen recognition and initiating the antigen specific adaptive immune response. Despite specific functional roles of the innate immune cells, they share several anti-viral pathways. The question then becomes, what is the overlap in the transcriptional changes induced upon viral infections across different cell-types? Here we investigate the extent to which gene signatures are conserved across innate immune cell-types by performing a comparative analysis of transcriptomic data. Particularly, we integrate transcriptomic datasets measuring response of two innate immune cells (epithelial and dendritic cells) to influenza virus. The study reveals cell-type specific regulatory genes and a conserved network between the two cell-types. Additionally, novel functionally associated gene clusters are identified which are robustly defined across multiple independent studies. These gene clusters can be used in future investigation, and to facilitate their use we release PathCellNet (version 0), a cloud based tool to explore cell-type specific connectivity of user-defined genes. In the future, expansion of PathCellNet will allow exploration of cell-type specific responses across a variety of pathogens and cell-types.

Cell-type specific connectivity accounts for diverse in vivo functional roles of inhibitory neurons in V1

Inhibitory neurons have a large diversity [1], however functional roles of diverse subtypes have not been elucidated. We aim to help systematically addressing this question using large scale modeling and coarse graining between scales. Mesoscopic models of in vivo activity starting from structure have only recently become feasible making use of development of cortical column models [2], knowledge of connectivity [3,4], scalable simulation tools [NEST] and robust population statistic techniques [5]. Vasoactive intestinal peptide-positive (VIP+) interneurons are one of the major inhibitory cell types in the cortex [1], and their excitability has been found to be associated with various endogenous factors such as brain states and top-down signals. Zhang et al. [6] recently found that top-down signals originated in the cingulate (Cg) strongly innervate VIP + interneurons of V1, indicating the critical role of VIP+ cells in attentional gain modulation. We studied their effect on top-down signals using both analytical and computational models. Using a coarse-grained firing rate model for the cell types in the superficial layers we found that the reported cell-type specific connectivity allows top-down inputs to VIP+ interneurons to effectively disinhibit pyramidal cell activity. Due to the limitations of the firing rate model, we utilized large scale computational models to investigate mechanisms underlying attentional gain modulation in an entire network. Specifically, we adopted the multiple column model proposed by Wagatsuma et al. [4] and modified them by incorporating three inhibitory cell types into the superficial layers of the model. In our model we considered parvalbumin-positive (PV+), SST+ and VIP+ inhibitory cells. We also constructed the data-driven cell-type specific intercolumnar connections between columns. Our model was capable of reproducing multiplicative gain modulation (see Figure ​Figure1),1), and our simulation results suggest that non-specific activation of VIP cells is sufficient for generating multiplicative gain modulation. Figure 1 Simulated tuning curve for pyramidal neurons as a function of top-down inputs to VIP cells. These inputs allow a multiplicative gain modulation of the bottom-up input. We also note that PV+ cell-mediating inhibition across columns is critical for regulating the responses over small spatial scales: e.g. induced by non-preferred stimulus (orientation). SST+ cell mediate inhibition across columns and allow contextual visual processing in V1.

Transition to chaos in random networks with cell-type-specific connectivity.

In neural circuits, statistical connectivity rules strongly depend on cell-type identity. We study dynamics of neural networks with cell-type-specific connectivity by extending the dynamic mean-field method and find that these networks exhibit a phase transition between silent and chaotic activity. By analyzing the locus of this transition, we derive a new result in random matrix theory: the spectral radius of a random connectivity matrix with block-structured variances. We apply our results to show how a small group of hyperexcitable neurons within the network can significantly increase the network's computational capacity by bringing it into the chaotic regime.

The role of cell adhesion molecules in visual circuit formation: from neurite outgrowth to maps and synaptic specificity.

ABSTRACTThe formation of visual circuitry is a multistep process that involves cell–cell interactions based on a range of molecular mechanisms. The correct implementation of individual events, including axon outgrowth and guidance, the formation of the topographic map, or the synaptic targeting of specific cellular subtypes, are prerequisites for a fully functional visual system that is able to appropriately process the information captured by the eyes. Cell adhesion molecules (CAMs) with their adhesive properties and their high functional diversity have been identified as key actors in several of these fundamental processes. Because of their growth‐promoting properties, CAMs play an important role in neuritogenesis. Furthermore, they are necessary to control additional neurite development, regulating dendritic spacing and axon pathfinding. Finally, trans‐synaptic interactions of CAMs ensure cell type‐specific connectivity as a basis for the establishment of circuits processing distinct visual features. Recent discoveries implicating CAMs in novel mechanisms have led to a better general understanding of neural circuit formation, but also revealed an increasing complexity of their function. This review aims at describing the different levels of action for CAMs to shape neural connectivity, with a special focus on the visual system. © 2015 Wiley Periodicals, Inc. Develop Neurobiol 75: 569–583, 2015

The Cell-Type Specific Cortical Microcircuit: Relating Structure and Activity in a Full-Scale Spiking Network Model

In the past decade, the cell-type specific connectivity and activity of local cortical networks have been characterized experimentally to some detail. In parallel, modeling has been established as a tool to relate network structure to activity dynamics. While available comprehensive connectivity maps ( Thomson, West, et al. 2002; Binzegger et al. 2004) have been used in various computational studies, prominent features of the simulated activity such as the spontaneous firing rates do not match the experimental findings. Here, we analyze the properties of these maps to compile an integrated connectivity map, which additionally incorporates insights on the specific selection of target types. Based on this integrated map, we build a full-scale spiking network model of the local cortical microcircuit. The simulated spontaneous activity is asynchronous irregular and cell-type specific firing rates are in agreement with in vivo recordings in awake animals, including the low rate of layer 2/3 excitatory cells. The interplay of excitation and inhibition captures the flow of activity through cortical layers after transient thalamic stimulation. In conclusion, the integration of a large body of the available connectivity data enables us to expose the dynamical consequences of the cortical microcircuitry.

Activity-Induced Long-Term Potentiation of Excitatory Synapses in Developing Zebrafish Retina In Vivo

Neural activity-induced long-term potentiation (LTP) of synaptic transmission is believed to be one of the cellular mechanisms underlying experience-dependent developmental refinement of neural circuits. Although it is well established that visual experience and neural activity are critical for the refinement of retinal circuits, whether and how LTP occurs in the retina remain unknown. Using in vivo perforated whole-cell recording and two-photon calcium imaging, we find that both repeated electrical and visual stimulations can induce LTP at excitatory synapses formed by bipolar cells on retinal ganglion cells in larval but not juvenile zebrafish. LTP induction requires the activation of postsynaptic N-methyl-D-aspartate receptors, and its expression involves arachidonic acid-dependent presynaptic changes in calcium dynamics and neurotransmitter release. Physiologically, both electrical and visual stimulation-induced LTP can enhance visual responses of retinal ganglion cells. Thus, LTP exists in developing retinae with a presynaptic locus and may serve for visual experience-dependent refinement of retinal circuits.

Development of Cell Type-Specific Connectivity Patterns of Converging Excitatory Axons in the Retina

To integrate information from different presynaptic cell types, dendrites receive distinct patterns of synapses from converging axons. How different afferents invivo establish specific connectivity patterns with the same dendrite is poorly understood. Here, we examine the synaptic development of three glutamatergic bipolar cell types converging ontoa common postsynaptic retinal ganglion cell. We find that after axons and dendrites target appropriate synaptic layers, patterns of connections among these neuronsdiverge through selective changes in the conversion of axo-dendritic appositions to synapses. This process is differentially regulated by neurotransmission, which is required for the shift from single to multisynaptic appositions of one bipolar cell type but not for maintenance and elimination, respectively, of connections from the other two types. Thus, synaptic specificity among converging excitatory inputs in theretina emerges via differential synaptic maturation of axo-dendritic appositions and is shaped by neurotransmission in a cell type-dependent manner.

Parallel processing strategies of the primate visual system

Incoming sensory information is sent to the brain along modality-specific channels corresponding to the five senses. Each of these channels further parses the incoming signals into parallel streams to provide a compact, efficient input to the brain. Ultimately, these parallel input signals must be elaborated on and integrated in the cortex to provide a unified and coherent percept. Recent studies in the primate visual cortex have greatly contributed to our understanding of how this goal is accomplished. Multiple strategies including retinal tiling, hierarchical and parallel processing and modularity, defined spatially and by cell type-specific connectivity, are used by the visual system to recover the intricate detail of our visual surroundings.