Subtypes Of Inhibitory Neurons Research Articles

Single-nucleus gene expression

Neurogenomics Identifying the genes expressed at the level of a single cell nucleus can better help us understand the human brain. Lake et al. developed a single-nuclei sequencing technique, which they applied to cells in classically defined Brodmann areas from a postmortem brain. Clustering of gene expression showed concordance with the area of origin and defining 16 neuronal subtypes. Both excitatory and inhibitory neuronal subtypes show regional variations that define distinct cortical areas and exhibit how gene expression clusters may distinguish between distinct cortical areas. This method opens the door to widespread sampling of the genes expressed in a diseased brain and other tissues of interest. Science , this issue p. [1586][1] [1]: /lookup/doi/10.1126/science.aaf1204

The role of parvalbumin-containing interneurons in the regulation of spontaneous synchronous activity of brain neurons in culture

Subtypes of inhibitory GABAergic neurons containing Ca2+-binding proteins play a pivotal role in the regulation of spontaneous synchronous [Ca2+]i transients in a neuronal network. In this study it is shown that: (1) the interneurons that containing Ca2+-binding proteins at buffer concentration can be identified by the shape of Ca2+-signa1 in response to depolarization or activation of ionotropic glutamate receptors; (2) Ca2+-binding proteins are involved in desynchronization of spontaneous Ca2+ transients. At low frequencies of spontaneous synchronous [Ca2+]i transients (less than 0.2 Hz) neurons show quasi-synchronous pulsations. At higher frequencies, synchronization of spontaneous synchronous [Ca2+]i transients occurs in all neurons; (3) it is established that several synchronous oscillations with different frequencies coexist in the network and the amplitude of their depolarizing pulse also varies. This phenomenon is apparently the mechanism that selectively directs information in separate neurons using the same network; and (4) in one population of interneurons at high frequencies of spontaneous synchronous [Ca2+]i transients the inversion of Cl– concentration gradient is observed. In this case, the inhibition of GABA(A) receptors suppresses the activity of neurons in this population and excites other neurons in the network. Thus, the GABAergic neurons that contain Ca-binding proteins show different mechanisms to regulate the synchronous neuronal activities in cultured rat hippocampal cells.

In Vivo Measurement of Cell-Type-Specific Synaptic Connectivity and Synaptic Transmission in Layer 2/3 Mouse Barrel Cortex

SummaryIntracellular recordings of membrane potential in vitro have defined fundamental properties of synaptic communication. Much less is known about the properties of synaptic connectivity and synaptic transmission in vivo. Here, we combined single-cell optogenetics with whole-cell recordings to investigate glutamatergic synaptic transmission in vivo from single identified excitatory neurons onto two genetically defined subtypes of inhibitory GABAergic neurons in layer 2/3 mouse barrel cortex. We found that parvalbumin-expressing (PV) GABAergic neurons received unitary glutamatergic synaptic input with higher probability than somatostatin-expressing (Sst) GABAergic neurons. Unitary excitatory postsynaptic potentials onto PV neurons were also faster and more reliable than inputs onto Sst neurons. Excitatory synapses targeting Sst neurons displayed strong short-term facilitation, while those targeting PV neurons showed little short-term dynamics. Our results largely agree with in vitro measurements. We therefore demonstrate the technical feasibility of assessing functional cell-type-specific synaptic connectivity in vivo, allowing future investigations into context-dependent modulation of synaptic transmission.

Altered distribution of hippocampal interneurons in the murine Down Syndrome model Ts65Dn.

Down Syndrome, with an incidence of one in 800 live births, is the most common genetic alteration producing intellectual disability. We have used the Ts65Dn model, that mimics some of the alterations observed in Down Syndrome. This genetic alteration induces an imbalance between excitation and inhibition that has been suggested as responsible for the cognitive impairment present in this syndrome. The hippocampus has a crucial role in memory processing and is an important area to analyze this imbalance. In this report we have analysed, in the hippocampus of Ts65Dn mice, the expression of synaptic markers: synaptophysin, vesicular glutamate transporter-1 and isoform 67 of the glutamic acid decarboxylase; and of different subtypes of inhibitory neurons (Calbindin D-28k, parvalbumin, calretinin, NPY, CCK, VIP and somatostatin). We have observed alterations in the inhibitory neuropil in the hippocampus of Ts65Dn mice. There was an excess of inhibitory puncta and a reduction of the excitatory ones. In agreement with this observation, we have observed an increase in the number of inhibitory neurons in CA1 and CA3, mainly interneurons expressing calbindin, calretinin, NPY and VIP, whereas parvalbumin cell numbers were not affected. These alterations in the number of interneurons, but especially the alterations in the proportion of the different types, may influence the normal function of inhibitory circuits and underlie the cognitive deficits observed in DS.

Transcriptional control of GABAergic neuronal subtype identity in the thalamus.

BackgroundThe thalamus is often defined as the ‘gateway to consciousness’, a feature that is supported by the specific connectivity and electrophysiological properties of its neurons. Inhibitory GABAergic neurons are required for the dynamic gating of information passing through the thalamus. The high degree of heterogeneity among thalamic GABA neurons suggests that, during embryonic development, alternative differentiation programmes exist to guide the acquisition of inhibitory neuron subtype identity.ResultsTaking advantage of the accessibility of the developing chick embryo, we have used in ovo manipulations of gene expression to test the role of candidate transcription factors in controlling GABAergic neuronal subtype identity in the developing thalamus.ConclusionsIn this study, we describe two alternative differentiation programmes for GABAergic neurogenesis in the thalamus and identify Helt and Dlx2 as key transcription factors that are sufficient to direct neuronal progenitors along a specific differentiation pathway at the expense of alternative lineage choices. Furthermore, we identify Calb2, a gene encoding for the GABA subtype marker calretinin as a target of the transcription factor Sox14. This work is a step forward in our understanding of how GABA neuron diversity in the thalamus is achieved during development and will help future investigation of the molecular mechanisms that lead up to the acquisition of different synaptic targets and electrophysiological features of mature thalamic inhibitory neurons.

The influence of extracellular matrix composition on the differentiation of neuronal subtypes in tissue engineered innervated intestinal smooth muscle sheets

Differentiation of enteric neural stem cells into several appropriate neural phenotypes is crucial while considering transplantation as a cellular therapy to treat enteric neuropathies. We describe the formation of tissue engineered innervated sheets, where intestinal smooth muscle and enteric neuronal progenitor cells are brought into close association in extracellular matrix (ECM) based microenvironments. Uniaxial alignment of constituent smooth muscle cells was achieved by substrate microtopography. The smooth muscle component of the tissue engineered sheets maintained a contractile phenotype irrespective of the ECM composition, and generated equivalent contractions in response to potassium chloride stimulation, similar to native intestinal tissue. We provided enteric neuronal progenitor cells with permissive ECM-based compositional and viscoelastic cues to generate excitatory and inhibitory neuronal subtypes. In the presence of the smooth muscle cells, the enteric neuronal progenitor cells differentiated to functionally innervate the smooth muscle. The differentiation of specific neuronal subtypes was influenced by the ECM microenvironment, namely combinations of collagen I, collagen IV, laminin and/or heparan sulfate. The physiology of differentiated neurons within tissue engineered sheets was evaluated. Sheets with composite collagen and laminin had the most similar patterns of Acetylcholine-induced contraction to native intestinal tissue, corresponding to an increased protein expression of choline acetyltransferase. An enriched nitrergic neuronal population, evidenced by an increased expression of neuronal nitric oxide synthase, was obtained in tissue engineered sheets that included collagen IV. These sheets had a significantly increased magnitude of electrical field stimulated relaxation, sensitive maximally to nitric oxide synthase inhibition. Tissue engineered sheets containing laminin and/or heparan sulfate had a balanced expression of contractile and relaxant motor neurons. Our studies demonstrated that neuronal subtype was modulated by varying ECM composition. This observation could be utilized to derive enriched populations of specific enteric neurons in vitro prior to transplantation.

Simulating vertical and horizontal inhibition with short-term dynamics in a multi-column multi-layer model of neocortex.

The paper introduces a multi-layer multi-column model of the cortex that uses four different neuron types and short-term plasticity dynamics. It was designed with details of neuronal connectivity available in the literature and meets these conditions: (1) biologically accurate laminar and columnar flows of activity, (2) normal function of low-threshold spiking and fast spiking neurons, and (3) ability to generate different stages of epileptiform activity. With these characteristics the model allows for modeling lesioned or malformed cortex, i.e. examine properties of developmentally malformed cortex in which the balance between inhibitory neuron subtypes is disturbed.

Prdm13 Mediates the Balance of Inhibitory and Excitatory Neurons in Somatosensory Circuits

Generating a balanced network of inhibitory and excitatory neurons during development requires precise transcriptional control. In the dorsal spinal cord, Ptf1a, a basic helix-loop-helix (bHLH) transcription activator, maintains this delicate balance by inducing homeodomain (HD) transcription factors such as Pax2 to specify the inhibitory lineage while suppressing HD factors such as Tlx1/3 that specify the excitatory lineage. We uncover the mechanism by which Ptf1a represses excitatory cell fate in the inhibitory lineage. We identify Prdm13 as a direct target of Ptf1a and reveal that Prdm13 actively represses excitatory cell fate by binding to regulatory sequences near the Tlx1 and Tlx3 genes to silence their expression. Prdm13 acts through multiple mechanisms, including interactions with the bHLH factor Ascl1, to repress Ascl1 activation of Tlx3. Thus, Prdm13 is a key component of a highly coordinated transcriptional network that determines the balance of inhibitory versus excitatory neurons in the dorsal spinal cord.

SINs and SOMs: neural microcircuits for size tuning in the zebrafish and mouse visual pathway

In many animals, a fast and reliable circuit for discriminating between predator-sized objects and edible (prey-sized) objects is necessary for survival. How are receptive fields (RFs) in visual brain areas organized to extract information about size? Recent studies from the zebrafish optic tectum and the mouse visual cortex suggest de novo shaping of RFs by subtypes of inhibitory neurons. Del Bene et al. (2010) describe a population of GABAergic neurons in the zebrafish optic tectum (superficial interneurons, SINs) that are necessary for size filtering during prey capture. Adesnik et al. (2012) describe a somatostatin-expressing interneuron population (SOMs) that confers surround suppression on layer II/III pyramidal cells in mouse V1. Strikingly both the SINs and the SOMs, display size-dependent response properties. Increasing visual stimulus size increases excitatory input to these neurons. Dampening SIN or SOM activity alters tuning of neighboring circuits such that they lose preference for small objects. Both results provide exciting evidence for mechanisms of size filtering in visual circuits. Here we review the roles of the SINs and the SOMs and speculate on the similarity of such spatial filters across species.

Anterior-Posterior Direction Opponency in the Superficial Mouse Lateral Geniculate Nucleus

We show functional-anatomical organization of motion direction in mouse dorsal lateral geniculate nucleus (dLGN) using two-photon calcium imaging of dense populations in thalamus. Surprisingly, thesuperficial 75μm region contains anterior and posterior direction-selective neurons (DSLGNs) intermingled with nondirection-selective neurons, while upward- and downward-selective neurons are nearlyabsent. Unexpectedly, the remaining neurons encode both anterior and posterior directions, forming horizontal motion-axis selectivity. A model of random wiring consistent with these results makes quantitative predictions about the connectivity of direction-selective retinal ganglion cell (DSRGC) inputs to the superficial dLGN. DSLGNs are more sharply tuned than DSRGCs. These results suggest that dLGN maintains and sharpens retinal direction selectivity and integrates opposing DSRGC subtypes in a functional-anatomical region, perhaps forming a feature representation for horizontal-axis motion, contrary to dLGN being a simple relay. Furthermore, they support recent conjecture that cortical direction and orientation selectivity emerge in part from a previously undescribed motion-selective retinogeniculate pathway.

Selective Functional Interactions between Excitatory and Inhibitory Cortical Neurons and Differential Contribution to Persistent Activity of the Slow Oscillation

The neocortex depends upon a relative balance of recurrent excitation and inhibition for its operation. During spontaneous Up states, cortical pyramidal cells receive proportional barrages of excitatory and inhibitory synaptic potentials. Many of these synaptic potentials arise from the activity of nearby neurons, although the identity of these cells is relatively unknown, especially for those underlying the generation of inhibitory synaptic events. To address these fundamental questions, we developed an in vitro submerged slice preparation of the mouse entorhinal cortex that generates robust and regular spontaneous recurrent network activity in the form of the slow oscillation. By performing whole-cell recordings from multiple cell types identified with green fluorescent protein expression and electrophysiological and/or morphological properties, we show that distinct functional subpopulations of neurons exist in the entorhinal cortex, with large variations in contribution to the generation of balanced excitation and inhibition during the slow oscillation. The most active neurons during the slow oscillation are excitatory pyramidal and inhibitory fast spiking interneurons, receiving robust barrages of both excitatory and inhibitory synaptic potentials. Weak action potential activity was observed in stellate excitatory neurons and somatostatin-containing interneurons. In contrast, interneurons containing neuropeptide Y, vasoactive intestinal peptide, or the 5-hydroxytryptamine (serotonin) 3a receptor, were silent. Our data demonstrate remarkable functional specificity in the interactions between different excitatory and inhibitory cortical neuronal subtypes, and suggest that it is the large recurrent interaction between pyramidal neurons and fast spiking interneurons that is responsible for the generation of persistent activity that characterizes the depolarized states of the cortex.

Understanding Appropriate Use Criteria in Nuclear Medicine

There has been a significant increase in cardiac radionuclide imaging over the past decade, leading to a corresponding increase in scrutiny from Federal and private health plans questioning the necessity of these tests. In response to efforts by third-party payers to limit all types of cardiovascular imaging studies, the American College of Cardiology Foundation, in conjunction with other professional societies, developed appropriate use criteria. The goal of this article is to explain how the criteria were created and define the 3 categories of indications: appropriate, inappropriate, and uncertain. Tips for using appropriate use criteria and tables, including a definition of several key terms technologists should be familiar with, will be provided. In addition, reimbursement, benchmark data, and practical considerations for implementation will be discussed. Finally, several tools to aid in calculating appropriateness are suggested. With a basic understanding, the appropriate use criteria are relatively easy to apply. It is important for facilities to begin to voluntarily incorporate them into their practice and document levels of appropriateness now as payers are developing 2 trends that are not favorable for nuclear cardiology: preauthorization and denial of payment for inappropriate studies.

Regulation of neuronal input transformations by tunable dendritic inhibition

Transforming synaptic input into action potential output is a fundamental function of neurons. The pattern of action potential output from principal cells of the mammalian hippocampus encodes spatial and nonspatial information, but the cellular and circuit mechanisms by which neurons transform their synaptic input into a given output are unknown. Using a combination of optical activation and cell type-specific pharmacogenetic silencing in vitro, we found that dendritic inhibition is the primary regulator of input-output transformations in mouse hippocampal CA1 pyramidal cells, and acts by gating the dendritic electrogenesis driving burst spiking. Dendrite-targeting interneurons are themselves modulated by interneurons targeting pyramidal cell somata, providing a synaptic substrate for tuning pyramidal cell output through interactions in the local inhibitory network. These results provide evidence for a division of labor in cortical circuits, where distinct computational functions are implemented by subtypes of local inhibitory neurons.

Functional Specialization of Seven Mouse Visual Cortical Areas

To establish the mouse as a genetically tractable model for high-order visual processing, we characterized fine-scale retinotopic organization of visual cortex and determined functional specialization of layer 2/3 neuronal populations in seven retinotopically identified areas. Each area contains a distinct visuotopic representation and encodes a unique combination of spatiotemporal features. Areas LM, AL, RL, and AM prefer up to three times faster temporal frequencies and significantly lower spatial frequencies than V1, while V1 and PM prefer high spatial and low temporal frequencies. LI prefers both high spatial and temporal frequencies. All extrastriate areas except LI increase orientation selectivity compared to V1, and three areas are significantly more direction selective (AL, RL, and AM). Specific combinations of spatiotemporal representations further distinguish areas. These results reveal that mouse higher visual areas are functionally distinct, and separate groups of areas may be specialized for motion-related versus pattern-related computations, perhaps forming pathways analogous to dorsal and ventral streams in other species.

Broadly Tuned Response Properties of Diverse Inhibitory Neuron Subtypes in Mouse Visual Cortex

Different subtypes of GABAergic neurons in sensory cortex exhibit diverse morphology, histochemical markers, and patterns of connectivity. These subtypes likely play distinct roles in cortical function, but their in vivo response properties remain unclear. We used in vivo calcium imaging, combined with immunohistochemical and genetic labels, to record visual responses in excitatory neurons and up to three distinct subtypes of GABAergic neurons (immunoreactive for parvalbumin, somatostatin, or vasoactive intestinal peptide) in layer 2/3 of mouse visual cortex. Excitatory neurons had sharp response selectivity for stimulus orientation and spatial frequency, while all GABAergic subtypes had broader selectivity. Further, bias in the responses of GABAergic neurons toward particular orientations or spatial frequencies tended to reflect net biases of the surrounding neurons. These results suggest that the sensory responses of layer 2/3 GABAergic neurons reflect the pooled activity of the surrounding population--a principle that may generalize across species and sensory modalities.

Synaptic and Network Mechanisms of Sparse and Reliable Visual Cortical Activity during Nonclassical Receptive Field Stimulation

During natural vision, the entire visual field is stimulated by images rich in spatiotemporal structure. Although many visual system studies restrict stimuli to the classical receptive field (CRF), it is known that costimulation of the CRF and the surrounding nonclassical receptive field (nCRF) increases neuronal response sparseness. The cellular and network mechanisms underlying increased response sparseness remain largely unexplored. Here we show that combined CRF + nCRF stimulation increases the sparseness, reliability, and precision of spiking and membrane potential responses in classical regular spiking (RS(C)) pyramidal neurons of cat primary visual cortex. Conversely, fast-spiking interneurons exhibit increased activity and decreased selectivity during CRF + nCRF stimulation. The increased sparseness and reliability of RS(C) neuron spiking is associated with increased inhibitory barrages and narrower visually evoked synaptic potentials. Our experimental observations were replicated with a simple computational model, suggesting that network interactions among neuronal subtypes ultimately sharpen recurrent excitation, producing specific and reliable visual responses.

LMO4 Controls the Balance between Excitatory and Inhibitory Spinal V2 Interneurons

Multiple excitatory and inhibitory interneurons form the motor circuit with motor neurons in the ventral spinal cord. Notch signaling initiates the diversification of immature V2-interneurons into excitatory V2a-interneurons and inhibitory V2b-interneurons. Here, we provide a transcriptional regulatory mechanism underlying their balanced production. LIM-only protein LMO4 controls this binary cell fate choice by regulating the activity of V2a- and V2b-specific LIM complexes inversely. In the spinal cord, LMO4 induces GABAergic V2b-interneurons in collaboration with SCL and inhibits Lhx3 from generating glutamatergic V2a-interneuons. In LMO4;SCL compound mutant embryos, V2a-interneurons increase markedly at the expense of V2b-interneurons. We further demonstrate that LMO4 nucleates the assembly of a novel LIM-complex containing SCL, Gata2, and NLI. This complex activates specific enhancers in V2b-genes consisting of binding sites for SCL and Gata2, thereby promoting V2b-interneuron fate. Thus, LMO4 plays essential roles in directing a balanced generation of inhibitory and excitatory neurons in the ventral spinal cord.

Short promoters in viral vectors drive selective expression in mammalian inhibitory neurons, but do not restrict activity to specific inhibitory cell-types

Short cell-type specific promoter sequences are important for targeted gene therapy and studies of brain circuitry. We report on the ability of short promoter sequences to drive fluorescent protein expression in specific types of mammalian cortical inhibitory neurons using adeno-associated virus (AAV) and lentivirus (LV) vectors. We tested many gene regulatory sequences derived from fugu (Takifugu rubripes), mouse, human, and synthetic composite regulatory elements. All fugu compact promoters expressed in mouse cortex, with only the somatostatin (SST) and the neuropeptide Y (NPY) promoters largely restricting expression to GABAergic neurons. However these promoters did not control expression in inhibitory cells in a subtype specific manner. We also tested mammalian promoter sequences derived from genes putatively coexpressed or coregulated within three major inhibitory interneuron classes (PV, SST, VIP). In contrast to the fugu promoters, many of the mammalian sequences failed to express, and only the promoter from gene A930038C07Rik conferred restricted expression, although as in the case of the fugu sequences, this too was not inhibitory neuron subtype specific. Lastly and more promisingly, a synthetic sequence consisting of a composite regulatory element assembled with PAX6 E1.1 binding sites, NRSE and a minimal CMV promoter showed markedly restricted expression to a small subset of mostly inhibitory neurons, but whose commonalities are unknown.

Similarity Effect and Optimal Control of Multiple-Choice Decision Making

Decision making with several choice options is central to cognition. To elucidate the neural mechanisms of such decisions, we investigated a recurrent cortical circuit model in which fluctuating spiking neural dynamics underlie trial-by-trial stochastic decisions. The model encodes a continuous analog stimulus feature and is thus applicable to multiple-choice decisions. Importantly, the continuous network captures similarity between alternatives and possible overlaps in their neural representation. Model simulations accounted for behavioral as well as single-unit neurophysiological data from a recent monkey experiment and revealed testable predictions about the patterns of error rate as a function of the similarity between the correct and actual choices. We also found that the similarity and number of options affect speed and accuracy of responses. A mechanism is proposed for flexible control of speed-accuracy tradeoff, based on a simple top-down signal to the decision circuit that may vary nonmonotonically with the number of choice alternatives.

Selective, State-Dependent Activation of Somatostatin-Expressing Inhibitory Interneurons in Mouse Neocortex

The specific functions of subtypes of cortical inhibitory neurons are not well understood. This is due in part to a dearth of information about the behaviors of interneurons under conditions when the surrounding circuit is in an active state. We investigated the firing behavior of a subset of inhibitory interneurons, identified using mice that express green fluorescent protein (GFP) in a subset of somatostatin-expressing inhibitory cells ("GFP-expressing inhibitory neuron" [GIN] cells). The somata of the GIN cells were in layer 2/3 of somatosensory cortex and had dense, layer 1-projecting axons that are characteristic of Martinotti neurons. Interestingly, GIN cells fired similarly during a variety of diverse activating conditions: when bathed in fluids with low-divalent cation concentrations, when stimulated with brief trains of local synaptic inputs, when exposed to group I metabotropic glutamate receptor agonists, or when exposed to muscarinic cholinergic receptor agonists. During these manipulations, GIN cells fired rhythmically and persistently in the theta-frequency range (3-10 Hz). Synchronous firing was often observed and its strength was directly proportional to the magnitude of electrical coupling between GIN cells. These effects were cell type specific: the four manipulations that persistently activated GIN cells rarely caused spiking of regular-spiking (RS) pyramidal cells or fast-spiking (FS) inhibitory interneurons. Our results suggest that supragranular GIN interneurons form an electrically coupled network that exerts a coherent 3- to 10-Hz inhibitory influence on its targets. Because GIN cells are more readily activated than RS and FS cells, it is possible that they act as "first responders" when cortical excitatory activity increases.