Cholinergic input to mouse visual cortex signals a movement state and acutely enhances layer 5 responsiveness.

Acetylcholine is released in visual cortex by axonal projections from the basal forebrain. The signals conveyed by these projections and their computational significance are still unclear. Using two-photon calcium imaging in behaving mice, we show that basal forebrain cholinergic axons in the mouse visual cortex provide a binary locomotion state signal. In these axons, we found no evidence of responses to visual stimuli or visuomotor prediction errors. While optogenetic activation of cholinergic axons in visual cortex in isolation did not drive local neuronal activity, when paired with visuomotor stimuli, it resulted in layer-specific increases of neuronal activity. Responses in layer 5 neurons to both top-down and bottom-up inputs were increased in amplitude and decreased in latency, whereas those in layer 2/3 neurons remained unchanged. Using opto- and chemogenetic manipulations of cholinergic activity, we found acetylcholine to underlie the locomotion-associated decorrelation of activity between neurons in both layer 2/3 and layer 5. Our results suggest that acetylcholine augments the responsiveness of layer 5 neurons to inputs from outside of the local network, possibly enabling faster switching between internal representations during locomotion.

Functional Dynamics and Selectivity of Two Parallel Corticocortical Pathways from Motor Cortex to Layer 5 Circuits in Somatosensory Cortex.

In the rodent whisker system, active sensing and sensorimotor integration are mediated in part by the dynamic interactions between the motor cortex (M1) and somatosensory cortex (S1). However, understanding these dynamic interactions requires knowledge about the synapses and how specific neurons respond to their input. Here, we combined optogenetics, retrograde labeling, and electrophysiology to characterize the synaptic connections between M1 and layer 5 (L5) intratelencephalic (IT) and pyramidal tract (PT) neurons in S1 of mice (both sexes). We found that M1 synapses onto IT cells displayed modest short-term depression, whereas synapses onto PT neurons showed robust short-term facilitation. Despite M1 inputs to IT cells depressing, their slower kinetics resulted in summation and a response that increased during short trains. In contrast, summation was minimal in PT neurons due to the fast time course of their M1 responses. The functional consequences of this reduced summation, however, were outweighed by the strong facilitation at these M1 synapses, resulting in larger response amplitudes in PT neurons than IT cells during repetitive stimulation. To understand the impact of facilitating M1 inputs on PT output, we paired trains of inputs with single backpropagating action potentials, finding that repetitive M1 activation increased the probability of bursts in PT cells without impacting the time dependence of this coupling. Thus, there are two parallel but dynamically distinct systems of M1 synaptic excitation in L5 of S1, each defined by the short-term dynamics of its synapses, the class of postsynaptic neurons, and how the neurons respond to those inputs.

Visual Corticotectal Neurons in Awake Rabbits: Receptive Fields and Driving Monosynaptic Thalamocortical Inputs.

The superior colliculus receives powerful synaptic inputs from corticotectal neurons in the visual cortex. The function of these corticotectal neurons remains largely unknown due to a limited understanding of their response properties and connectivity. Here, we use antidromic methods to identify corticotectal neurons in awake male and female rabbits, and measure their axonal conduction times, thalamic inputs and receptive field properties. All corticotectal neurons responded to sinusoidal drifting gratings with a nonlinear (nonsinusoidal) increase in mean firing rate but showed pronounced differences in their ON-OFF receptive field structures that we classified into three groups, Cx, S2, and S1. Cx receptive fields had highly overlapping ON and OFF subfields as classical complex cells, S2 had largely separated ON and OFF subfields as classical simple cells, and S1 had a single ON or OFF subfield. Thus, all corticotectal neurons are homogeneous in their nonlinear spatial summation but very heterogeneous in their spatial integration of ON and OFF inputs. The Cx type had the fastest conducting axons, the highest spontaneous activity, and the strongest and fastest visual responses. The S2 type had the highest orientation selectivity, and the S1 type had the slowest conducting axons. Moreover, our cross-correlation analyses found that a subpopulation of corticotectal neurons with very fast conducting axons and high spontaneous firing rates (largely "Cx" type) receives monosynaptic input from retinotopically aligned thalamic neurons. This previously unrecognized fast-conducting thalamic-mediated corticotectal pathway may provide specialized information to superior colliculus and prime recipient neurons for subsequent corticotectal or retinal synaptic input.

Analysis of the effect of the scorpion toxin AaH-II on action potential generation in the axon initial segment.

The toxin AaH-II, from the scorpion Androctonus australis Hector venom, is a 64 amino acid peptide that targets voltage-gated Na+ channels (VGNCs) and slows their inactivation. While at macroscopic cellular level AaH-II prolongs the action potential (AP), a functional analysis of the effect of the toxin in the axon initial segment (AIS), where VGNCs are highly expressed, was never performed so far. Here, we report an original analysis of the effect of AaH-II on the AP generation in the AIS of neocortical layer-5 pyramidal neurons from mouse brain slices. After determining that AaH-II does not discriminate between Nav1.2 and Nav1.6, i.e. between the two VGNC isoforms expressed in this neuron, we established that 7 nM was the smallest toxin concentration producing a minimal detectable deformation of the somatic AP after local delivery of the toxin. Using membrane potential imaging, we found that, at this minimal concentration, AaH-II substantially widened the AP in the AIS. Using ultrafast Na+ imaging, we found that local application of 7 nM AaH-II caused a large increase in the slower component of the Na+ influx in the AIS. Finally, using ultrafast Ca2+ imaging, we observed that 7 nM AaH-II produces a spurious slow Ca2+ influx via Ca2+-permeable VGNCs. Molecules targeting VGNCs, including peptides, are proposed as potential therapeutic tools. Thus, the present analysis in the AIS can be considered a general proof-of-principle on how high-resolution imaging techniques can disclose drug effects that cannot be observed when tested at the macroscopic level.

Kinetics and functional consequences of BK channels activation by N-type Ca2+ channels in the dendrite of mouse neocortical layer-5 pyramidal neurons.

The back-propagation of an action potential (AP) from the axon/soma to the dendrites plays a central role in dendritic integration. This process involves an intricate orchestration of various ion channels, but a comprehensive understanding of the contribution of each channel type remains elusive. In this study, we leverage ultrafast membrane potential recordings (Vm) and Ca2+ imaging techniques to shed light on the involvement of N-type voltage-gated Ca2+ channels (VGCCs) in layer-5 neocortical pyramidal neurons' apical dendrites. We found a selective interaction between N-type VGCCs and large-conductance Ca2+-activated K+ channels (BK CAKCs). Remarkably, we observe that BK CAKCs are activated within a mere 500 µs after the AP peak, preceding the peak of the Ca2+ current triggered by the AP. Consequently, when N-type VGCCs are inhibited, the early broadening of the AP shape amplifies the activity of other VGCCs, leading to an augmented total Ca2+ influx. A NEURON model, constructed to replicate and support these experimental results, reveals the critical coupling between N-type and BK channels. This study not only redefines the conventional role of N-type VGCCs as primarily involved in presynaptic neurotransmitter release but also establishes their distinct and essential function as activators of BK CAKCs in neuronal dendrites. Furthermore, our results provide original functional validation of a physical interaction between Ca2+ and K+ channels, elucidated through ultrafast kinetic reconstruction. This insight enhances our understanding of the intricate mechanisms governing neuronal signaling and may have far-reaching implications in the field.

Cholinergic Activation of Corticofugal Circuits in the Adult Mouse Prefrontal Cortex.

Acetylcholine (ACh) promotes neocortical output to the thalamus and brainstem by preferentially enhancing the postsynaptic excitability of layer 5 pyramidal tract (PT) neurons relative to neighboring intratelencephalic (IT) neurons. Less is known about how ACh regulates the excitatory synaptic drive of IT and PT neurons. To address this question, spontaneous excitatory postsynaptic potentials (sEPSPs) were recorded in dual recordings of IT and PT neurons in slices of prelimbic cortex from adult female and male mice. ACh (20 µM) enhanced sEPSP amplitudes, frequencies, rise-times, and half-widths preferentially in PT neurons. These effects were blocked by the muscarinic receptor antagonist atropine (1 µM). When challenged with pirenzepine (1 µM), an antagonist selective for M1-type muscarinic receptors, ACh instead reduced sEPSP frequencies, suggesting that ACh may generally suppress synaptic transmission in the cortex via non-M1 receptors. Cholinergic enhancement of sEPSPs in PT neurons was not sensitive to antagonism of GABA receptors with gabazine (10 µM) and CGP52432 (2.5 µM) but was blocked by tetrodotoxin (1 µM), suggesting that ACh enhances action-potential-dependent excitatory synaptic transmission in PT neurons. ACh also preferentially promoted the occurrence of synchronous sEPSPs in dual recordings of PT neurons relative to IT-PT and IT-IT parings. Finally, selective chemogenetic silencing of hM4Di-expressing PT, but not commissural IT, neurons blocked cholinergic enhancement of sEPSP amplitudes and frequencies in PT neurons. These data suggest that, in addition to selectively enhancing the postsynaptic excitability of PT neurons, M1 receptor activation promotes corticofugal output by amplifying recurrent excitation within networks of PT neurons.

Hyperpolarization-Activated Cation Channels Shape the Spiking Frequency Preference of Human Cortical Layer 5 Pyramidal Neurons.

Discerning the contribution of specific ionic currents to complex neuronal dynamics is a difficult, but important, task. This challenge is exacerbated in the human setting, although the widely characterized uniqueness of the human brain compared with preclinical models necessitates the direct study of human neurons. Neuronal spiking frequency preference is of particular interest given its role in rhythm generation and signal transmission in cortical circuits. Here, we combine the frequency-dependent gain (FDG), a measure of spiking frequency preference, and novel in silico analyses to dissect the contributions of individual ionic currents to the suprathreshold features of human layer 5 (L5) neurons captured by the FDG. We confirm that a contemporary model of such a neuron, primarily constrained to capture subthreshold activity driven by the hyperpolarization-activated cyclic nucleotide gated (h-) current, replicates key features of the in vitro FDG both with and without h-current activity. With the model confirmed as a viable approximation of the biophysical features of interest, we applied new analysis techniques to quantify the activity of each modeled ionic current in the moments before spiking, revealing unique dynamics of the h-current. These findings motivated patch-clamp recordings in analogous rodent neurons to characterize their FDG, which confirmed that a biophysically detailed model of these neurons captures key interspecies differences in the FDG. These differences are correlated with distinct contributions of the h-current to neuronal activity. Together, this interdisciplinary and multispecies study provides new insights directly relating the dynamics of the h-current to suprathreshold spiking frequency preference in human L5 neurons.

Pyramidal neurons form active, transient, multilayered circuits perturbed by autism-associated mutations at the inception of neocortex.

Cortical circuits are composed predominantly of pyramidal-to-pyramidal neuron connections, yet their assembly during embryonic development is not well understood. We show that mouse embryonic Rbp4-Cre cortical neurons, transcriptomically closest to layer 5 pyramidal neurons, display two phases of circuit assembly in vivo. At E14.5, they form a multi-layered circuit motif, composed of only embryonic near-projecting-type neurons. By E17.5, this transitions to a second motif involving all three embryonic types, analogous to the three adult layer 5 types. In vivo patch clamp recordings and two-photon calcium imaging of embryonic Rbp4-Cre neurons reveal active somas and neurites, tetrodotoxin-sensitive voltage-gated conductances, and functional glutamatergic synapses, from E14.5 onwards. Embryonic Rbp4-Cre neurons strongly express autism-associated genes and perturbing these genes interferes with the switch between the two motifs. Hence, pyramidal neurons form active, transient, multi-layered pyramidal-to-pyramidal circuits at the inception of neocortex, and studying these circuits could yield insights into the etiology of autism.

Motor cortex projections to red and pontine nuclei have distinct roles during movement in the mouse.

Motor control largely depends on the deep layer 5 (L5) pyramidal neurons that project to subcortical structures. However, it is largely unknown if these neurons are functionally segregated with distinct roles in movement performance. Here, we analyzed mouse motor cortex L5 pyramidal neurons projecting to the red and pontine nuclei during movement preparation and execution. Using photometry to analyze the calcium activity of L5 pyramidal neurons projecting to the red nucleus and pons, we reveal that both types of neurons activate with different temporal dynamics. Optogenetic inhibition of either kind of projection differentially affects forelimb movement onset and execution in a lever press task, but only the activity of corticopontine neurons is significantly correlated with trial-by-trial variations in reaction time. The results indicate that cortical neurons projecting to the red and pontine nuclei contribute differently to sensorimotor integration, suggesting that L5 output neurons are functionally compartmentalized generating, in parallel, different downstream information.

Nav1.2 and BK channel interaction shapes the action potential in the axon initial segment.

In neocortical layer-5 pyramidal neurons, the action potential (AP) is generated in the axon initial segment (AIS) when the membrane potential (Vm ) reaches the threshold for activation of the voltage-gated Na+ channels (VGNCs) Nav 1.2 and Nav 1.6. Yet, whereas these VGNCs are known to differ in spatial distribution along the AIS and in biophysical properties, our understanding of the functional differences between the two channels remains elusive. Here, using ultrafast Na+ , Vm and Ca2+ imaging in combination with partial block of Nav 1.2 by the peptide G1 G4 -huwentoxin-IV, we demonstrate an exclusive role of Nav 1.2 in shaping the generating AP. Precisely, we show that selective block of ∼30% of Nav 1.2 widens the AP in the distal part of the AIS and we demonstrate that this effect is due to a loss of activation of BK Ca2+ -activated K+ channels (CAKCs). Indeed, Ca2+ influx via Nav 1.2 activates BK CAKCs, determining the amplitude and the early phase of repolarization of the AP in the AIS. By using control experiments using 4,9-anhydrotetrodotoxin, a moderately selective inhibitor of Nav 1.6, we concluded that the Ca2+ influx shaping the early phase of the AP is exclusive of Nav 1.2. Hence, we mimicked this result with a neuron model in which the role of the different ion channels tested reproduced the experimental evidence. The exclusive role of Nav 1.2 reported here is important for understanding the physiology and pathology of neuronal excitability. KEY POINTS: We optically analysed the action potential generated in the axon initial segment of mouse layer-5 neocortical pyramidal neurons and its associated Na+ and Ca2+ currents using ultrafast imaging techniques. We found that partial selective block of the voltage-gated Na+ channel Nav 1.2, produced by a recently developed peptide, widens the shape of the action potential in the distal part of the axon initial segment. We demonstrate that this effect is due to a reduction of the Ca2+ influx through Nav 1.2 that activates BK Ca2+ -activated K+ channels. To validate our conclusions, we generated a neuron model that reproduces the ensemble of our experimental results. The present results indicate a specific role of Nav 1.2 in the axon initial segment for shaping of the action potential during its generation.

Altered integration of excitatory inputs onto the basal dendrites of layer 5 pyramidal neurons in a mouse model of Fragile X syndrome.

Layer 5 (L5) pyramidal neurons receive predictive and sensory inputs in a compartmentalized manner at their apical and basal dendrites, respectively. To uncover how integration of sensory inputs is affected in autism spectrum disorders (ASD), we used two-photon glutamate uncaging to activate spines in the basal dendrites of L5 pyramidal neurons from a mouse model of Fragile X syndrome (FXS), the most common genetic cause of ASD. While subthreshold excitatory inputs integrate linearly in wild-type animals, surprisingly those with FXS summate sublinearly, contradicting what would be expected of sensory hypersensitivity classically associated with ASD. We next investigated the mechanism underlying this sublinearity by performing knockdown of the regulatory ß4 subunit of BK channels, which rescued the synaptic integration, a result that was corroborated with numerical simulations. Taken together, these findings suggest that there is a differential impairment in the integration of feedforward sensory and feedback predictive inputs in L5 pyramidal neurons in FXS and potentially other forms of ASD, as a result of specifically localized subcellular channelopathies. These results challenge the traditional view that FXS and other ASD are characterized by sensory hypersensitivity, proposing instead a hyposensitivity of sensory inputs and hypersensitivity of predictive inputs onto cortical neurons.

Micropopulation mapping of the mouse parafascicular nucleus connections reveals diverse input-output motifs.

Introduction: In primates, including humans, the centromedian/parafascicular (CM-Pf) complex is a key thalamic node of the basal ganglia system. Deep brain stimulation in CM-Pf has been applied for the treatment of motor disorders such as Parkinson's disease or Tourette syndrome. Rodents have become widely used models for the study of the cellular and genetic mechanisms of these and other motor disorders. However, the equivalence between the primate CM-Pf and the nucleus regarded as analogous in rodents (Parafascicular, Pf) remains unclear. Methods: Here, we analyzed the neurochemical architecture and carried out a brain-wide mapping of the input-output motifs in the mouse Pf at micropopulation level using anterograde and retrograde labeling methods. Specifically, we mapped and quantified the sources of cortical and subcortical input to different Pf subregions, and mapped and compared the distribution and terminal structure of their axons. Results: We found that projections to Pf arise predominantly (>75%) from the cerebral cortex, with an unusually strong (>45%) Layer 5b component, which is, in part, contralateral. The intermediate layers of the superior colliculus are the main subcortical input source to Pf. On its output side, Pf neuron axons predominantly innervate the striatum. In a sparser fashion, they innervate other basal ganglia nuclei, including the subthalamic nucleus (STN), and the cerebral cortex. Differences are evident between the lateral and medial portions of Pf, both in chemoarchitecture and in connectivity. Lateral Pf axons innervate territories of the striatum, STN and cortex involved in the sensorimotor control of different parts of the contralateral hemibody. In contrast, the mediodorsal portion of Pf innervates oculomotor-limbic territories in the above three structures. Discussion: Our data thus indicate that the mouse Pf consists of several neurochemically and connectively distinct domains whose global organization bears a marked similarity to that described in the primate CM-Pf complex.

Neocortical layer 5 subclasses: From cellular properties to roles in behavior.

Layer 5 (L5) serves as the main output layer of cortical structures, where long-range projecting pyramidal neurons broadcast the columnar output to other cortical and extracortical regions of the brain. L5 pyramidal neurons are grouped into two subclasses based on their projection targets; while intratelencephalic (IT) neurons project to cortical areas and the striatum, extratelencephalic (ET) neurons project to subcortical areas such as the thalamus, midbrain, and brainstem. Each L5 subclass possesses distinct morphological and electrophysiological properties and is incorporated into a unique synaptic network. Thanks to recent advances in genetic tools and methodologies, it has now become possible to distinguish between the two subclasses in the living brain. There is increasing evidence indicating that each subclass plays a unique role in sensory processing, decision-making, and learning. This review first summarizes the anatomical and physiological properties as well as the neuromodulation of IT and ET neurons in the rodent neocortex, and then reviews recent literature on their roles in sensory processing and rodent behavior. Our ultimate goal is to provide a comprehensive understanding of the role of each subclass in cortical function by examining their operational regimes based on their cellular properties.

Layer 5 of cortex innervates the thalamic reticular nucleus in mice.

Neurons in the thalamic reticular nucleus (TRN) are a primary source of inhibition to the dorsal thalamus and, as they are innervated in part by the cortex, are a means of corticothalamic regulation. Previously, cortical inputs to the TRN were thought to originate solely from layer 6 (L6), but we recently reported the presence of putative synaptic terminals from layer 5 (L5) neurons in multiple cortical areas in the TRN [J. A. Prasad, B. J. Carroll, S. M. Sherman, J. Neurosci. 40, 5785-5796 (2020)]. Here, we demonstrate with electron microscopy that L5 terminals from multiple cortical regions make bona fide synapses in the TRN. We further use light microscopy to localize these synapses relative to recently described TRN subdivisions and show that L5 terminals target the edges of the somatosensory TRN, where neurons reciprocally connect to higher-order thalamus, and that L5 terminals are scarce in the core of the TRN, where neurons reciprocally connect to first-order thalamus. In contrast, L6 terminals densely innervate both edge and core subregions and are smaller than those from L5. These data suggest that a sparse but potent input from L5 neurons of multiple cortical regions to the TRN may yield transreticular inhibition targeted to higher-order thalamus.

Cell-type-specific integration of feedforward and feedback synaptic inputs in the posterior parietal cortex.

The integration of feedforward (sensory) and feedback (top-down) neuronal signals is a principal function of the neocortex. Yet, we have limited insight into how these information streams are combined by individual neurons. Using a two-color optogenetic strategy, we found that layer 5 pyramidal neurons in the posterior parietal cortex receive monosynaptic dual innervation, combining sensory inputs with top-down signals. Subclasses of layer 5 pyramidal neurons integrated these synapses with distinct temporal dynamics. Specifically, regular spiking cells exhibited supralinear enhancement of delayed-but not coincident-inputs, while intrinsic burst-firing neurons selectively boosted coincident synaptic events. These subthreshold integration characteristics translated to a nonlinear summation of action potential firing. Complementing electrophysiology with computational modeling, we found that distinct integration profiles arose from a cell-type-specific interaction of ionic mechanisms and feedforward inhibition. These data provide insight into the cellular properties that guide the nonlinear interaction of distinct long-range afferents in the neocortex.

Diversity in spatial frequency, temporal frequency, and speed tuning across mouse visual cortical areas and layers.

The mouse visual system consists of several visual cortical areas thought to be specialized for different visual features and/or tasks. Previous studies have revealed differences between primary visual cortex (V1) and other higher visual areas, namely, anterolateral (AL) and posteromedial (PM), and their tuning preferences for spatial and temporal frequency. However, these differences have primarily been characterized using methods that are biased toward superficial layers of cortex, such as two-photon calcium imaging. Fewer studies have investigated cell types in deeper layers of these areas and their tuning preferences. Because superficial versus deep-layer neurons and different types of deep-layer neurons are known to have different feedforward and feedback inputs and outputs, comparing the tuning preferences of these groups is important for understanding cortical visual information processing. In this study, we used extracellular electrophysiology and two-photon calcium imaging targeted toward two different layer 5 cell classes to characterize their tuning properties in V1, AL, and PM. We find that deep-layer neurons, similar to superficial layer neurons, are also specialized for different spatial and temporal frequencies, with the strongest differences between AL and V1, and AL and PM, but not V1 and PM. However, we note that the deep-layer neuron populations preferred a larger range of SFs and TFs compared to previous studies. We also find that extratelencephalically projecting layer 5 neurons are more direction selective than intratelencephalically projecting layer 5 neurons.

Enhanced motor cortex output and disinhibition in asymptomatic female mice with C9orf72 genetic expansion.

Information and action coding by cortical circuits relies on a balanced dialogue between excitation and inhibition. Circuit hyperexcitability is considered a potential pathophysiological mechanism in various brain disorders, but the underlying deficits, especially at early disease stages, remain largely unknown. We report that asymptomatic female mice carrying the chromosome 9 open reading frame 72 (C9orf72) repeat expansion, which represents a high-prevalence genetic abnormality for human amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD) spectrum disorder, exhibit abnormal motor cortex output. The number of primary motor cortex (M1) layer 5 pyramidal neurons is reduced in asymptomatic mice, with the surviving neurons receiving a decreased inhibitory drive that results in a higher M1 output, specifically during high-speed animal locomotion. Importantly, using deep-learning algorithms revealed that speed-dependent M1 output predicts the likelihood of C9orf72 genetic expansion. Our data link early circuit abnormalities with a gene mutation in asymptomatic ALS/FTLD carriers.

General anesthesia and the cortical stranglehold on consciousness.

In this issue of Neuron, Bharioke et al. (2022) demonstrate that diverse general anesthetic regimens all reversibly and selectively synchronize spontaneous activity of pyramidal neurons in layer 5 of mouse cortex. We discuss the implications of these findings for the mechanism of consciousness and anesthetic-induced unconsciousness.

Movement-specific signaling is differentially distributed across motor cortex layer 5 projection neuron classes.

Motor cortex generates descending output necessary for executing a wide range of limb movements. Although movement-related activity has been described throughout motor cortex, the spatiotemporal organization of movement-specific signaling in deep layers remains largely unknown. Here we record layer 5B population dynamics in the caudal forelimb area of motor cortex while mice perform a forelimb push/pull task and find that most neurons show movement-invariant responses, with a minority displaying movement specificity. Using cell-type-specific imaging, we identify that invariant responses dominate pyramidal tract (PT) neuron activity, with a small subpopulation representing movement type, whereas a larger proportion of intratelencephalic (IT) neurons display movement-type-specific signaling. The proportion of IT neurons decoding movement-type peaks prior to movement initiation, whereas for PT neurons, this occurs during movement execution. Our data suggest that layer 5B population dynamics largely reflect movement-invariant signaling, with information related to movement-type being routed through relatively small, distributed subpopulations of projection neurons.

General anesthesia globally synchronizes activity selectively in layer 5 cortical pyramidal neurons.

General anesthetics induce loss of consciousness, a global change in behavior. However, a corresponding global change in activity in the context of defined cortical cell types has not been identified. Here, we show that spontaneous activity of mouse layer 5 pyramidal neurons, but of no other cortical cell type, becomes consistently synchronized in vivo by different general anesthetics. This heightened neuronal synchrony is aperiodic, present across large distances, and absent in cortical neurons presynaptic to layer 5 pyramidal neurons. During the transition to and from anesthesia, changes in synchrony in layer 5 coincide with the loss and recovery of consciousness. Activity within both apical and basal dendrites is synchronous, but only basal dendrites' activity is temporally locked to somatic activity. Given that layer 5 is a major cortical output, our results suggest that brain-wide synchrony in layer 5 pyramidal neurons may contribute to the loss of consciousness during general anesthesia.