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Pyramidal cells and GABAergic interneurons fire together in balanced cortical networks. In contrast to this general rule, we describe a distinct neuron type in mice and rats whose spiking activity is anti-correlated with all principal cells and interneurons in all brain states but, most prevalently, during the down state of non-REM (NREM) sleep. We identify these down state-active (DSA) neurons as deep-layer neocortical neurogliaform cells that express ID2 and Nkx2.1 and are weakly immunoreactive to neuronal nitric oxide synthase. DSA neurons are weakly excited by deep-layer pyramidal cells and strongly inhibited by several other GABAergic cell types. Spiking of DSA neurons modified the sequential firing order of other neurons at down–up transitions. Optogenetic activation of ID2(+)Nkx2.1(+) interneurons in the posterior parietal cortex during NREM sleep, but not during waking, interfered with consolidation of cue discrimination memory. Despite their sparsity, DSA neurons perform critical physiological functions.

The combination of in vivo extracellular recording and genetic-engineering-assisted optical stimulation is a powerful tool for the study of neuronal circuits. Precise analysis of complex neural circuits requires high-density integration of multiple cellular-size light sources and recording electrodes. However, high-density integration inevitably introduces stimulation artifact. We present minimal-stimulation-artifact (miniSTAR) μLED optoelectrodes that enable effective elimination of stimulation artifact. A multi-metal-layer structure with a shielding layer effectively suppresses capacitive coupling of stimulation signals. A heavily boron-doped silicon substrate silences the photovoltaic effect induced from LED illumination. With transient stimulation pulse shaping, we reduced stimulation artifact on miniSTAR μLED optoelectrodes to below 50 μVpp, much smaller than a typical spike detection threshold, at optical stimulation of >50 mW mm–2 irradiance. We demonstrated high-temporal resolution (, Artifact-free opto-electrophysiology is key for precise modulation and monitoring of individual neurons at high spatio-temporal resolution. The authors present a method for eliminating stimulation artifacts in high-density micro-LED optoelectrodes for accurate functional mapping of local circuits.

During non-rapid eye movement (NREM) sleep, neuronal populations in the mammalian forebrain alternate between periods of spiking and inactivity. Termed the slow oscillation in the neocortex and sharp wave-ripples in the hippocampus, these alternations are often considered separately but are both crucial for NREM functions. By directly comparing experimental observations of naturally-sleeping rats with a mean field model of an adapting, recurrent neuronal population, we find that the neocortical alternations reflect a dynamical regime in which a stable active state is interrupted by transient inactive states (slow waves) while the hippocampal alternations reflect a stable inactive state interrupted by transient active states (sharp waves). We propose that during NREM sleep in the rodent, hippocampal and neocortical populations are excitable: each in a stable state from which internal fluctuations or external perturbation can evoke the stereotyped population events that mediate NREM functions., NREM sleep in rodents is characterized by internal dynamics in the form of UP/DOWN states in the neocortex and SWRs in the hippocampus. Here, the authors report that a mean field model with excitable dynamics captures the transition probabilities between these states from rodent sleep data.

High-yield electrophysiological extracellular recording in freely moving rodents provides a unique window into the temporal dynamics of neural circuits. Recording from unrestrained animals is critical to investigate brain activity during natural behaviors. The use and implantation of high-channel-count silicon probes represent the largest cost and experimental complexity associated with such recordings making a recoverable and reusable system desirable. To address this, we have designed and tested a novel 3D printed head-gear system for freely moving mice and rats. The system consists of a recoverable microdrive printed in stainless steel and a plastic head cap system, allowing researchers to reuse the silicon probes with ease, decreasing the effective cost, and the experimental effort and complexity. The cap designs are modular and provide structural protection and electrical shielding to the implanted hardware and electronics. We provide detailed procedural instructions allowing researchers to adapt and flexibly modify the head-gear system.

INTRODUCTION: Learning induces a dynamic reorganization of brain circuits but the neuronal mechanisms underlying this process are not well understood. Interregional gamma-frequency oscillations (~30 to 130 Hz) have been postulated as a mechanism to precisely coordinate upstream and downstream neuronal ensembles, for example, in the hippocampal system. The lateral (LEC) and medial (MEC) entorhinal cortex receive inputs from two distinct streams of cortical hierarchy (the “what” and the “where” paths) and convey these neuronal messages to the hippocampus. However, the mechanisms by which such messages are packaged and integrated or segregated by hippocampal circuits had yet to be explored. RATIONALE: Neuronal assemblies firing within gamma time frames in an upstream region can most effectively discharge their downstream partners. This gamma-time-scale organization appears essential for physiological functions because manipulations that impair precision timing of spikes in the hippocampus often affect behavior. However, direct support for distinct gamma-frequency communication in appropriate behavioral situations is missing. To bring physiological operations closer to behavior, we designed “spatial” and “object” learning tasks and examined the selective engagement of gamma-frequency communication between the MEC and LEC inputs and their target neuronal assemblies in the hippocampal dentate gyrus. We combined these correlational observations with optogenetic perturbation of gamma oscillations in LEC and MEC, respectively, to test their roles in pathway-specific neuronal communication and learning. RESULTS: During spatial learning, fast gamma (100 to 150 Hz) oscillations synchronized MEC and dentate gyrus and entrained predominantly granule cells. During object learning, slow gamma (30 to 50 Hz) oscillations synchronized LEC and dentate gyrus and preferentially recruited mossy cells and CA3 pyramidal neurons, suggesting task-specific routing of MEC and LEC messages in the form of gamma-cycle-spike packets of selected cell types. The low- and high-frequency gamma sub-bands were dominant in the outer and middle third of the dentate molecular layer, respectively, and their amplitude maxima were locked to different phases of theta oscillation. Gamma frequency optogenenetic perturbation of MEC and LEC led to learning impairments in a spatial and object learning task, respectively. In the same animals, the dentate layer–specific low- and high-frequency gamma sub-bands and spike-gamma LFP coupling were selectively reduced, coupled with deterioration of spatial and object-related parameters of dentate neurons. CONCLUSION: These findings demonstrate that distinct gamma-frequency-specific communication between MEC and LEC and the hippocampal cell assemblies are critical for routing task-relevant information, and our selective gamma-band perturbation experiments suggest that they support specific aspects of learning. We hypothesize that sending neuronal messages by segregated gamma-frequency carriers allows a target “reader” area to disambiguate convergent inputs. In general, these results demonstrate that specific projected gamma patterns dynamically engage functionally related cell assemblies across brain regions in a task-specific manner. post-print 2234 KB

How do firing patterns in a cortical circuit change when inhibitory neurons are excited? We virally expressed an excitatory designer receptor exclusively activated by a designer drug (Gq-DREADD) in all inhibitory interneuron types of the CA1 region of the hippocampus in the rat. While clozapine N-oxide (CNO) activation of interneurons suppressed firing of pyramidal cells, unexpectedly the majority of interneurons also decreased their activity. CNO-induced inhibition decreased over repeated sessions, which we attribute to long-term synaptic plasticity between interneurons and pyramidal cells. Individual interneurons did not display sustained firing but instead transiently enhanced their activity, interleaved with suppression of others. The power of the local fields in the theta band was unaffected, while power at higher frequencies was attenuated, likely reflecting reduced pyramidal neuron spiking. The incidence of sharp wave ripples decreased but the surviving ripples were associated with stronger population firing compared with the control condition. These findings demonstrate that DREADD activation of interneurons brings about both short-term and long-term circuit reorganization, which should be taken into account in the interpretation of chemogenic effects on behavior.

We describe the spatiotemporal course of cortical high-gamma activity (HGA), hippocampal ripple activity and interictal epileptiform discharges (IEDs) during an associative memory task in 15 epilepsy patients undergoing invasive electroencephalography. Successful encoding trials manifested significantly greater HGA in hippocampus and frontal regions. Successful cued recall trials manifested sustained HGA in hippocampus compared to failed responses. Hippocampal ripple rates were greater during successful encoding and retrieval trials. IEDs during encoding were associated with 15% decreased odds of remembering in hippocampus (95% CI 6-23%). Hippocampal IEDs during retrieval predicted 25% decreased odds of remembering (15-33%). Odds of remembering were reduced by 25-52% if IEDs occurred during the 500-2000 ms window of encoding or by 41% during retrieval. During encoding and retrieval, hippocampal IEDs were followed by a transient decrease in ripple rate. We hypothesize that IEDs impair associative memory in a regionally and temporally specific manner by decreasing physiologic hippocampal ripples necessary for effective encoding and recall. Because dynamic memory impairment arises from pathological IED events competing with physiological ripples, IEDs represent a promising therapeutic target for memory remediation in patients with epilepsy.SummaryHippocampal interictal epileptiform discharges in hippocampus acutely impair declarative memory, potentially by hijacking physiological processes essential for encoding and recall.

The hippocampus is thought to guide navigation by forming a cognitive map of space. However, the behavioral demands for such a map can vary depending on particular features of a given environment. For example, an environment rich in cues may require a finer resolution map than an open space. It is unclear how the hippocampal cognitive map adjusts to meet these distinct behavioral demands. To address this issue, we examined the spatial coding characteristics of hippocampal neurons in mice and rats navigating different environments. We found that CA1 place cells located in the superficial sublayer were more active in cue-poor environments, and preferentially used a firing rate code driven by intra-hippocampal inputs. In contrast, place cells located in the deep sublayer were more active in cue-rich environments and expressed a phase code driven by entorhinal inputs. Switching between these two spatial coding modes was supported by the interaction between excitatory gamma inputs and local inhibition.

Summary The mnemonic functions of hippocampal sharp wave ripples (SPW-Rs) have been studied extensively. Because hippocampal outputs affect not only cortical but also subcortical targets, we examined the impact of SPW-Rs on the firing patterns of lateral septal (LS) neurons in behaving rats. A large fraction of SPW-Rs were temporally locked to high-frequency oscillations (HFOs) (120–180 Hz) in LS, with strongest coupling during non-rapid eye movement (NREM) sleep, followed by waking immobility. However, coherence and spike-local field potential (LFP) coupling between the two structures were low, suggesting that HFOs are generated locally within the LS GABAergic population. This hypothesis was supported by optogenetic induction of HFOs in LS. Spiking of LS neurons was largely independent of the sequential order of spiking in SPW-Rs but instead correlated with the magnitude of excitatory synchrony of the hippocampal output. Thus, LS is strongly activated by SPW-Rs and may convey hippocampal population events to its hypothalamic and brainstem targets.

Longer ripples make better memories Sharp wave ripples in the hippocampus are thought to play a role in memory formation and action planning. Fernández-Ruiz et al. used multisite electrophysiological recordings combined with optogenetic activation of hippocampal pyramidal neurons in rats performing learning tasks. Learning and correct recall in spatial memory tasks were associated with extended sharp wave ripples. Artificially prolonging these ripples improved working memory performance, whereas aborting the late part of ripples decreased performance. Science , this issue p. 1082

We describe a technique to semichronically record the cortical extracellular neural activity in the behaving monkey employing commercial high-density electrodes. After the design and construction of low cost microdrives that allow varying the depth of the recording locations after the implantation surgery, we recorded the extracellular unit activity from pools of neurons at different depths in the presupplementary motor cortex (pre-SMA) of a rhesus monkey trained in a tapping task. The collected data were processed to classify cells as putative pyramidal cells or interneurons on the basis of their waveform features. We also demonstrate that short time cross-correlogram occasionally yields unit pairs with high short latency (

Interactions between the hippocampus and the cortex are critical for memory. Interictal epileptiform discharges (IEDs) identify epileptic brain regions and can impair memory, but the mechanisms by which they interact with physiological patterns of network activity are mostly undefined. We show in a rat model of temporal lobe epilepsy that spontaneous hippocampal IEDs correlate with impaired memory consolidation, and that they are precisely coordinated with spindle oscillations in the prefrontal cortex during nonrapid-eye-movement (NREM) sleep. This coordination surpasses the normal physiological ripple-spindle coupling and is accompanied by decreased ripple occurrence. IEDs also induce spindles during rapid-eye movement (REM) sleep and wakefulness-behavioral states that do not naturally express these oscillations-by generating a cortical 'down' state. In a pilot clinical examination of four subjects with focal epilepsy, we confirm a similar correlation of temporofrontal IEDs with spindles over anatomically restricted cortical regions. These findings imply that IEDs may impair memory via the misappropriation of physiological mechanisms for hippocampal-cortical coupling, which suggests a target for the treatment of memory impairment in epilepsy.

SummaryWe report a scalable method to monolithically integrate microscopic light emitting diodes (μLEDs) and recording sites onto silicon neural probes for optogenetic applications in neuroscience. Each μLED and recording site has dimensions similar to a pyramidal neuron soma, providing confined emission and electrophysiological recording of action potentials and local field activity. We fabricated and implanted the four-shank probes, each integrated with 12 μLEDs and 32 recording sites, into the CA1 pyramidal layer of anesthetized and freely moving mice. Spikes were robustly induced by 60 nW light power, and fast population oscillations were induced at the microwatt range. To demonstrate the spatiotemporal precision of parallel stimulation and recording, we achieved independent control of distinct cells ∼50 μm apart and of differential somato-dendritic compartments of single neurons. The scalability and spatiotemporal resolution of this monolithic optogenetic tool provides versatility and precision for cellular-level circuit analysis in deep structures of intact, freely moving animals.

Hippocampal theta oscillations coordinate neuronal firing to support memory and spatial navigation. The medial septum (MS) is critical in theta generation by two possible mechanisms: either a unitary "pacemaker" timing signal is imposed on the hippocampal system, or it may assist in organizing target subcircuits within the phase space of theta oscillations. We used temperature manipulation of the MS to test these models. Cooling of the MS reduced both theta frequency and power and was associated with an enhanced incidence of errors in a spatial navigation task, but it did not affect spatial correlates of neurons. MS cooling decreased theta frequency oscillations of place cells and reduced distance-time compression but preserved distance-phase compression of place field sequences within the theta cycle. Thus, the septum is critical for sustaining precise theta phase coordination of cell assemblies in the hippocampal system, a mechanism needed for spatial memory.

The relationship between mesoscopic local field potentials (LFPs) and single-neuron firing in the multi-layered neocortex is poorly understood. Simultaneous recordings from all layers in the primary visual cortex (V1) of the behaving mouse revealed functionally defined layers in V1. The depth of maximum spike power and sink-source distributions of LFPs provided consistent laminar landmarks across animals. Coherence of gamma oscillations (30-100 Hz) and spike-LFP coupling identified six physiological layers and further sublayers. Firing rates, burstiness, and other electrophysiological features of neurons displayed unique layer and brain state dependence. Spike transmission strength from layer 2/3 cells to layer 5 pyramidal cells and interneurons was stronger during waking compared with non-REM sleep but stronger during non-REM sleep among deep-layer excitatory neurons. A subset of deep-layer neurons was active exclusively in the DOWN state of non-REM sleep. These results bridge mesoscopic LFPs and single-neuron interactions with laminar structure in V1.

SUMMARY Hippocampal sharp-wave ripples (SPW-Rs) support consolidation of recently acquired episodic memories and planning future actions by generating ordered neuronal sequences of previous or future experiences. SPW-Rs are characterized by several spectral components: a slow (5–15 Hz) sharp-wave, a high-frequency “ripple” oscillation (150–200 Hz), and a slow “gamma” oscillation (20–40 Hz). Using laminar hippocampal recordings and optogenetic manipulations, we dissected the origin of these spectral components. We show that increased power in the 20–40 Hz band does not reflect an entrainment of CA1 and CA3 neurons at gamma frequency but the power envelope of overlapping ripples. Spike-local field potential coupling between unit firing in CA1 and CA3 regions during SPW-Rs is lowest in the gamma band. Longer SPW-Rs are preceded by increased firing in the entorhinal cortex. Thus, fusion of SPW-Rs leads to lengthening of their duration associated with increased power in the slow gamma band without the presence of true oscillation., Graphical Abstract, In Brief SWRs are thought to support memory consolidation and planning. They are characterized by several spectral components: a slow-frequency wave (sharp-wave), a high-frequency oscillation (ripple), and a slow “gamma” band. Here, we report a mechanism, involving the concatenation of several SWRs, which explains the generation of the “gamma” frequency band.

The local field potential (LFP) is an aggregate measure of group neuronal activity and is often correlated with the action potentials of single neurons. In recent years investigators have found that action potential firing rates increase during elevations in power high-frequency band oscillations (50-200 Hz range). However action potentials also contribute to the LFP signal itself, making the spike–LFP relationship complex. Here we examine the relationship between spike rates and LFPs in varying frequency bands in rat neocortical recordings. We find that 50-180Hz oscillations correlate most consistently with high firing rates, but that other LFPs bands also carry information relating to spiking, including in some cases anti-correlations. Relatedly, we find that spiking itself and electromyographic activity contribute to LFP power in these bands. The relationship between spike rates and LFP power varies between brain states and between individual cells. Finally, we create an improved oscillation-based predictor of action potential activity by specifically utilizing information from across the entire recorded frequency spectrum of LFP. The findings illustrate both caveats and improvements to be taken into account in attempts to infer spiking activity from LFP.

Transcranial electric stimulation is a non-invasive tool that can influence brain activity; however, the parameters necessary to affect local circuits in vivo remain to be explored. Here, we report that in rodents and human cadaver brains, ~75% of scalp-applied currents are attenuated by soft tissue and skull. Using intracellular and extracellular recordings in rats, we find that at least 1 mV/mm voltage gradient is necessary to affect neuronal spiking and subthreshold currents. We designed an ‘intersectional short pulse’ stimulation method to inject sufficiently high current intensities into the brain, while keeping the charge density and sensation on the scalp surface relatively low. We verify the regional specificity of this novel method in rodents; in humans, we demonstrate how it affects the amplitude of simultaneously recorded EEG alpha waves. Our combined results establish that neuronal circuits are instantaneously affected by intensity currents that are higher than those used in conventional protocols., Though transcranial electric stimulation has been used to influence brain activity, it is debated whether neuronal spiking activity is directly affected by commonly-used protocols. Here, the authors quantify the voltage gradients necessary to instantaneously affect neuronal spiking and show that they are higher than commonly-used protocols.

Sequential activity of multineuronal spiking can be observed during theta and high-frequency ripple oscillations in the hippocampal CA1 region and is linked to experience, but the mechanisms underlying such sequences are unknown. We compared multineuronal spiking during theta oscillations, spontaneous ripples, and focal optically induced high-frequency oscillations ("synthetic" ripples) in freely moving mice. Firing rates and rate modulations of individual neurons, and multineuronal sequences of pyramidal cell and interneuron spiking, were correlated during theta oscillations, spontaneous ripples, and synthetic ripples. Interneuron spiking was crucial for sequence consistency. These results suggest that participation of single neurons and their sequential order in population events are not strictly determined by extrinsic inputs but also influenced by local-circuit properties, including synapses between local neurons and single-neuron biophysics.

The head direction (HD) system functions as a compass with member neurons robustly increasing their firing rates when the animal’s head points in a specific direction. HD neurons may be driven by peripheral sensors or, as computational models postulate, internally-generated (‘attractor’) mechanisms. We addressed the contributions of stimulus-driven and internally-generated activity by recording ensembles of HD neurons in the antero-dorsal thalamic nucleus and the postsubiculum of mice by comparing their activity in various brain states. The temporal correlation structure of HD neurons is preserved during sleep, characterized by a 60°-wide correlated neuronal firing (‘activity packet’), both within as well as across these two brain structures. During REM, the spontaneous drift of the activity packet was similar to that observed during waking and accelerated tenfold during slow wave sleep. These findings demonstrate that peripheral inputs impinge upon an internally-organized network, which provides amplification and enhanced precision of the head-direction signal.

International audience; Cognitive representation of the environment requires a stable hippocampal map, but the mechanisms maintaining a given map are unknown. Because sharp wave-ripples (SPW-R) orchestrate both retrospective and prospective spatial information, we hypothesized that disrupting neuronal activity during SPW-Rs affects spatial representation. Mice learned new sets of three goal locations daily in a multiwell maze. We used closed-loop SPW-R detection at goal locations to trigger optogenetic silencing of a subset of CA1 pyramidal neurons. Control place cells (nonsilenced or silenced outside SPW-Rs) largely maintained the location of their place fields after learning and showed increased spatial information content. In contrast, the place fields of SPW-Rsilenced place cells remapped, and their spatial information remained unaltered. SPW-R silencing did not impact the firing rates or proportions of place cells. These results suggest that interference with SPW-R-associated activity during learning prevents stabilization and refinement of hippocampal maps.

Memory transfer for long-term storage Explicit memory formation involves the transfer of rapidly encoded information from the hippocampus to long-term storage sites in the association cortex. Khodagholy et al. developed a microelectrode system for large-scale simultaneous electrophysiological monitoring of multiple sites in the rat neocortex. They observed discrete high-frequency neocortical oscillations called ripples only in the association cortex. These cortical ripples shared many properties with hippocampal ripples. Hippocampal ripples were coupled with cortical ripples in the posterior parietal cortex, an association cortical area linked to navigational planning. This coupling was increased during sleep after the induction of long-term hippocampal-dependent spatial memory. Science , this issue p. 369

Summary Excitatory control of inhibitory neurons is poorly understood due to the difficulty of studying synaptic connectivity in vivo . We inferred such connectivity through analysis of spike timing and validated this inference using juxtacellular and optogenetic control of presynaptic spikes in behaving mice. We observed that neighboring CA1 neurons had stronger connections and that superficial pyramidal cells projected more to deep interneurons. Connection probability and strength were skewed, with a minority of highly connected hubs. Divergent presynaptic connections led to synchrony between interneurons. Synchrony of convergent presynaptic inputs boosted postsynaptic drive. Presynaptic firing frequency was read out by postsynaptic neurons through short-term depression and facilitation, with individual pyramidal cells and interneurons displaying a diversity of spike transmission filters. Additionally, spike transmission was strongly modulated by prior spike timing of the postsynaptic cell. These results bridge anatomical structure with physiological function.

Fiberless optoelectrodes are an emerging tool to enable brain circuit mapping by providing precise optical modulation and electrical monitoring of many neurons. While optoelectrodes having an on-board light source offer compact and optically efficient device solutions, many of them fail to provide robust thermal and electrical design to fully exploit the recording capabilities of the device. In this work, we present a novel fiberless multicolor optoelectrode solution, which meets the optical and thermal design requirements of an in vivo neural optoelectrode and offers potential for low-noise neural recording. The total optical loss measured for 405 nm and 635 nm wavelengths through the waveguide is 11.7±1.1 dB and 9.9±0.7 dB, corresponding to respective irradiances of 1928 mW/mm2 and 2905 mW/mm2 at the waveguide tip from 6 mW laser diode chips. The efficient thermal packaging enables continuous device operation for up to 190 seconds at 10% duty cycle. We validated the fully packaged device in the intact brain of anesthetized mice co-expressing Channelrhodopsin-2 and Archaerhodopsin in the hippocampal CA1 region and achieved activation and silencing of the same neurons. We discuss improvements made to reduce the stimulation artifact induced by applying currents to the laser diode chips.

The consolidation of context-dependent emotional memory requires communication between the hippocampus and the basolateral amygdala (BLA), but the mechanisms of this process are unknown. We recorded neuronal ensembles in the hippocampus and BLA while rats learned the location of an aversive air puff on a linear track, as well as during sleep before and after training. We found coordinated reactivations between the hippocampus and the BLA during non-REM sleep following training. These reactivations peaked during hippocampal sharp wave-ripples (SPW-Rs) and involved a subgroup of BLA cells positively modulated during hippocampal SPW-Rs. Notably, reactivation was stronger for the hippocampus-BLA correlation patterns representing the run direction that involved the air puff than for the 'safe' direction. These findings suggest that consolidation of contextual emotional memory occurs during ripple-reactivation of hippocampus-amygdala circuits.

Slow oscillations and spindle activity during non-rapid eye movement sleep have been implicated in memory consolidation. Closed-loop acoustic stimulation has previously been shown to enhance slow oscillations and spindle activity during sleep and improve verbal associative memory. We assessed the effect of closed-loop acoustic stimulation during a daytime nap on a virtual reality spatial navigation task in 12 healthy human subjects in a randomized within-subject crossover design. We show robust enhancement of slow oscillation and spindle activity during sleep. However, no effects on behavioral performance were observed when comparing real versus sham stimulation. To explore whether memory enhancement effects were task specific and dependent on nocturnal sleep, in a second experiment with 19 healthy subjects, we aimed to replicate a previous study that used closed-loop acoustic stimulation to enhance memory for word pairs. The methods used were as close as possible to those used in the original study, except that we used a double-blind protocol, in which both subject and experimenter were unaware of the test condition. Again, we successfully enhanced slow oscillation and spindle power, but again did not strengthen associative memory performance with stimulation. We conclude that enhancement of sleep oscillations may be insufficient to enhance memory performance in spatial navigation or verbal association tasks, and provide possible explanations for lack of behavioral replication.

Conducting polymer–based electrodes can acquire population and individual neural activity from the surface of the human brain., Localizing neuronal patterns that generate pathological brain signals may assist with tissue resection and intervention strategies in patients with neurological diseases. Precise localization requires high spatiotemporal recording from populations of neurons while minimizing invasiveness and adverse events. We describe a large-scale, high-density, organic material–based, conformable neural interface device (“NeuroGrid”) capable of simultaneously recording local field potentials (LFPs) and action potentials from the cortical surface. We demonstrate the feasibility and safety of intraoperative recording with NeuroGrids in anesthetized and awake subjects. Highly localized and propagating physiological and pathological LFP patterns were recorded, and correlated neural firing provided evidence about their local generation. Application of NeuroGrids to brain disorders, such as epilepsy, may improve diagnostic precision and therapeutic outcomes while reducing complications associated with invasive electrodes conventionally used to acquire high-resolution and spiking data.

Sleep exerts many effects on mammalian forebrain networks, including homeostatic effects on both synaptic strengths and firing rates. We used large-scale recordings to examine the activity of neurons in the frontal cortex of rats and firstly observed that the distribution of pyramidal cell firing rates was wide and strongly skewed towards high firing rates. Moreover, neurons from different parts of that distribution were differentially modulated by sleep sub-states. Periods of nonREM sleep reduced the activity of high firing rate neurons and tended to upregulate firing of slow firing neurons. By contrast, the effect of REM was to reduce firing rates across the entire rate spectrum. Microarousals, interspersed within nonREM epochs, increased firing rates of slow firing neurons. The net result of sleep was to homogenize the firing rate distribution. These findings are at variance with current homeostatic models and provide a novel view of sleep in adjusting network excitability.

Coding what is known and what is newDo neural activity patterns during sleep reflect the replay of a novel experience or an invariant preexisting dynamic? Grosmark and Buzsáki observed that both familiar and novel aspects of learned information are replayed during synchronous bursts of activity in the hippocampus. Familiarity was encoded by fast-firing less-modifiable neurons that showed rate and sequence correlations that persisted into postlearning sleep. The novel features of an experience were represented by a different set of slowly firing and highly plastic cells.Science, this issue p.1440

Developments in microfabrication technology have enabled the production of neural electrode arrays with hundreds of closely-spaced recording sites, and electrodes with thousands of sites are currently under development. These probes will in principle allow the simultaneous recording of very large numbers of neurons. However, use of this technology requires the development of techniques for decoding the spike times of the recorded neurons, from the raw data captured from the probes. There currently exists no practical solution to this problem of ?spike sorting? for large, dense electrode arrays. Here, we present a set of novel tools to solve this problem, implemented in a suite of practical, user-friendly, open-source software. We validate these methods on data from rat cortex, demonstrating error rates as low as 5%. Fil: Rossant, Cyrille . Colegio Universitario de Londres; Reino Unido Fil: Kadir, Shabnam N. . Colegio Universitario de Londres; Reino Unido Fil: Goodman, Dan F. M. . Imperial College London; Reino Unido Fil: Schulman, John. University Of California Berkeley; Estados Unidos Fil: Hunter, Maximilian L. D.. Colegio Universitario de Londres; Reino Unido Fil: Saleem, Aman B.. Colegio Universitario de Londres; Reino Unido Fil: Grosmark, Andres. University Of New York; Estados Unidos Fil: Belluscio, Mariano Andres. University Of New York; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Fisiología y Biofísica Bernardo Houssay; Argentina Fil: Denfield, George H.. Baylor College of Medicine. Department of Neuroscience; Estados Unidos Fil: Ecker, Alexander S.. Baylor College of Medicine. Department of Neuroscience; Estados Unidos Fil: Tolias, Andreas S.. Baylor College of Medicine. Department of Neuroscience; Estados Unidos Fil: Solomon, Samuel. Colegio Universitario de Londres; Reino Unido Fil: Buzsáki, György . University Of New York; Estados Unidos Fil: Carandini, Matteo. Colegio Universitario de Londres; Reino Unido Fil: Harris, Kenneth D.. Colegio Universitario de Londres; Reino Unido

Neuronal control with high temporal precision is possible with optogenetics, yet currently available methods do not enable to control independently multiple locations in the brains of freely moving animals. Here, we describe a diode-probe system that allows real-time and location-specific control of neuronal activity at multiple sites. Manipulation of neuronal activity in arbitrary spatiotemporal patterns is achieved by means of an optoelectronic array, manufactured by attaching multiple diode-fiber assemblies to high-density silicon probes or wire tetrodes and implanted into the brains of animals that are expressing light-responsive opsins. Each diode can be controlled separately, allowing localized light stimulation of neuronal activators and silencers in any temporal configuration and concurrent recording of the stimulated neurons. Because the only connections to the animals are via a highly flexible wire cable, unimpeded behavior is allowed for circuit monitoring and multisite perturbations in the intact brain. The capacity of the system to generate unique neural activity patterns facilitates multisite manipulation of neural circuits in a closed-loop manner and opens the door to addressing novel questions.

Karoly Schaffer (1864–1939) was a Hungarian neurologist who distinguished himself through original discoveries in human neuropathology. At the beginning of his scientific carrier, he described the cellular and fiber structure of the hippocampus, earning him a high reputation in neuroscience. Schaffer (1892) described the so-called “collateral fiber system” that connects the CA3 and CA1 regions of the hippocampus, known today as Schaffer collaterals. To decipher the history of this well-known eponym, we review Schaffer's original German publication and follow the impact of his research in the contemporary literature. © 2012 Wiley Periodicals Inc.

Neuronal oscillations allow for temporal segmentation of neuronal spikes. Interdependent oscillators can integrate multiple layers of information. We examined phase–phase coupling of theta and gamma oscillators in the CA1 region of rat hippocampus during maze exploration and rapid eye movement sleep. Hippocampal theta waves were asymmetric, and estimation of the spatial position of the animal was improved by identifying the waveform-based phase of spiking, compared to traditional methods used for phase estimation. Using the waveform-based theta phase, three distinct gamma bands were identified: slow gammaS(gammaS; 30–50 Hz), midfrequency gammaM(gammaM; 50–90 Hz), and fast gammaF(gammaF; 90–150 Hz or epsilon band). The amplitude of each sub-band was modulated by the theta phase. In addition, we found reliable phase–phase coupling between theta and both gammaSand gammaMbut not gammaFoscillators. We suggest that cross-frequency phase coupling can support multiple time-scale control of neuronal spikes within and across structures.

When monitoring neural activity using intracranial electrical recordings, researchers typically consider the signals to have two primary components: fast action potentials (APs) from neurons near the electrode, and the slower local field potential (LFP), thought to be dominated by postsynaptic currents integrated over a larger volume of tissue. In general, a decrease in signal power with increasing frequency is observed for most brain rhythms. The 100–200 Hz oscillations in the rat hippocampus, including “fast gamma” or “epsilon” oscillations and sharp wave-ripples (SPW-Rs), are one exception, showing an increase in power with frequency within this band. We have used detailed biophysical modeling to investigate the composition of extracellular potentials during fast oscillations in rat CA1. We find that postsynaptic currents exhibit a decreasing ability to generate large-amplitude oscillatory signals at high frequencies, whereas phase-modulated spiking shows the opposite trend. Our estimates indicate that APs and postsynaptic currents contribute similar proportions of the power contained in 140–200 Hz ripples, and the two combined generate a signal that closely resemblesin vivoSPW-Rs. Much of the AP-generated signal originates from neurons further than 100 μm from the recording site, consistent with ripples appearing similarly strong regardless of whether or not they contain recognizable APs. Additionally, substantial power can be generated in the 90–150 Hz epsilon band by the APs of rhythmically firing pyramidal neurons. Thus, high-frequency LFPs may generally contain signatures of local cell assembly activation.

SummaryNetwork oscillations support transient communication across brain structures. We show here, in rats, that task-related neuronal activity in the medial prefrontal cortex (PFC), the hippocampus, and the ventral tegmental area (VTA), regions critical for working memory, is coordinated by a 4 Hz oscillation. A prominent increase of power and coherence of the 4 Hz oscillation in the PFC and the VTA and its phase modulation of gamma power in both structures was present in the working memory part of the task. Subsets of both PFC and hippocampal neurons predicted the turn choices of the rat. The goal-predicting PFC pyramidal neurons were more strongly phase locked to both 4 Hz and hippocampal theta oscillations than nonpredicting cells. The 4 Hz and theta oscillations were phase coupled and jointly modulated both gamma waves and neuronal spikes in the PFC, the VTA, and the hippocampus. Thus, multiplexed timing mechanisms in the PFC-VTA-hippocampus axis may support processing of information, including working memory.

Summary Hippocampal CA1 pyramidal neurons have frequently been regarded as a homogeneous cell population in biophysical, pharmacological and modeling studies. Here we report robust differences between pyramidal neurons residing in the deep and superficial CA1 sublayers in the rat. Compared to their superficial peers, deep pyramidal cells fired at higher rates, burst more frequently, were more likely to have place fields and were more strongly modulated by slow oscillations of sleep. Both deep and superficial pyramidal cells fired preferentially at the trough of theta oscillations during maze exploration, yet during Rapid eye movement (REM) sleep, deep pyramidal cells shifted their preferred phase of firing to the peak of theta. Furthermore, whereas in waking, the majority of REM theta phase-shifting cells fired at the ascending phase of gamma oscillations, non-shifting cells preferred the trough. Thus, CA1 pyramidal cells in adjacent sublayers can address their targets jointly or differentially, depending on brain states.

Hippocampal sharp waves (SPW) and associated fast (‘ripple’) oscillations in the CA1 region are among the most synchronous physiological patterns in the mammalian brain. Using two-dimensional arrays of electrodes for recording local field potentials and unit discharges in freely moving rats, we studied the emergence of ripple oscillations (140–220 Hz) and compared their origin and cellular-synaptic mechanisms with fast gamma oscillations (90–140 Hz). We show that (a) hippocampal SPW-Rs and fast gamma oscillations are quantitatively distinct patterns but involve the same networks and share similar mechanisms, (b) both the frequency and magnitude of fast oscillations is positively correlated with the magnitude of SPWs, (c) during both ripples and fast gamma oscillations the frequency of network oscillation is higher in CA1 than in CA3, (d) SPWs and associated firing of neurons are synchronous in the dorsal hippocampus and dorso-medial entorhinal cortex but ripples are confined to the CA1 pyramidal layer and its downstream targets and (e) the emergence of CA3 population bursts, a prerequisite for SPW-ripples, is biased by activity patterns in the dentate gyrus and entorhinal cortex, with highest probability of ripples associated with an ‘optimum’ level of dentate gamma power. We hypothesize that each hippocampal subnetwork possesses distinct resonant properties, tuned by the magnitude of the excitatory drive.

Glucose is the primary energetic substrate of the brain, and measurements of its metabolism are the basis of major functional cerebral imaging methods. Contrary to the general view that neurons are fueled solely by glucose in proportion to their energetic needs, recentin vitroandex vivoanalyses suggest that glucose preferentially feeds astrocytes. However, the cellular fate of glucose in the intact brain has not yet been directly observed. We have used a real-time method for measuring glucose uptake in astrocytes and neuronsin vivoin male rats by imaging the trafficking of the nonmetabolizable glucose analog 6-deoxy-N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-aminoglucose (6-NBDG) using two-photon microscopy. During resting conditions we found that astrocytes and neurons both take up 6-NBDG at the same rate in the barrel cortex of the rat. However, during intense neuronal activity triggered by whisker stimulation, astrocytes rapidly accelerated their uptake, whereas neuronal uptake remained almost unchanged. After the stimulation period, astrocytes returned to their preactivation rates of uptake paralleling the neuronal rate of uptake. These observations suggest that glucose is taken up primarily by astrocytes, supporting the view that functional imaging experiments based on glucose analogs extraction may predominantly reflect the metabolic activity of the astrocytic network.

International audience; A thorough knowledge of the intrinsic circuit properties of the entorhinal cortex (EC) and the temporal dynamics these circuits support is essential for understanding how information is exchanged between the hippocampus and neocortex. Using intracellular and extracellular recordings in the anesthetized rat and anatomical reconstruction of single cells, we found that EC5 and EC2 principal neurons form large axonal networks mainly within their layers, interconnected by the more vertically organized axon trees of EC3 pyramidal cells. Principal cells showed layer-specific unique membrane properties and contributed differentially to theta and gamma oscillations. EC2 principal cells were most strongly phase modulated by EC theta. The multiple gamma oscillators, present in the various EC layers, were temporally coordinated by the phase of theta waves. Putative interneurons in all EC layers fired relatively synchronously within the theta cycle, coinciding with the maximum power of gamma oscillation. The special wiring architecture and unique membrane properties of EC neurons may underlie their behaviorally distinct firing patterns in the waking animal.

Although anatomical, lesion, and imaging studies of the hippocampus indicate qualitatively different information processing along its septo-temporal axis, physiological mechanisms supporting such distinction are missing. We found fundamental differences between the dorsal (dCA3) and the ventral-most parts (vCA3) of the hippocampus in both environmental representation and temporal dynamics. Discrete place fields of dCA3 neurons evenly covered all parts of the testing environments. In contrast, vCA3 neurons (1) rarely showed continuous two-dimensional place fields, (2) differentiated open and closed arms of a radial maze, and (3) discharged similar firing patterns with respect to the goals, both on multiple arms of a radial maze and during opposite journeys in a zigzag maze. In addition, theta power and the fraction of theta-rhythmic neurons were substantially reduced in the ventral compared with dorsal hippocampus. We hypothesize that the spatial representation in the septo-temporal axis of the hippocampus is progressively decreased. This change is paralleled with a reduction of theta rhythm and an increased representation of nonspatial information.

Theta (4–10 Hz) oscillations in the hippocampus are thought to be important for plasticity, temporal coding, learning, and memory. The hippocampal system has been postulated to have two (or more) rhythmic sources of theta oscillations, but little is known about the behavior-dependent interplay of theta oscillations in different subregions and layers of the hippocampus. We tested rats in a hippocampus-dependent delayed spatial alternation task on a modified T-maze while simultaneously recording local field potentials from dendritic and somatic layers of the dentate gyrus, CA3, and CA1 regions using high-density, 96-site silicon probes. We found that while theta oscillations were generally coherent throughout the hippocampus, the power, coherence, and phase of theta oscillations fluctuated in a layer-specific manner, confirming the presence of multiple interdependent dipoles. Layer-dependent changes in the power and coherence of theta oscillations varied with aspects of both the memory and control (non-mnemonic) tasks, but only a small fraction of the variance could be explained by running speed or acceleration. Furthermore, the phase lag between theta oscillations in the CA3 and CA1 pyramidal layers was significantly smaller on the maze arm approaching the T-junction than on other arms of the alternation task or on comparable segments of control tasks. Overall, our findings reveal a consortium of layer-specific theta dipoles (current sinks and sources) generated by the rhythmic flow of ions into and out of hippocampal cells. Moreover, our data suggest that these different theta generators flexibly coordinate hippocampal regions and layers to support behavioral task performance.

A long-standing conjecture in neuroscience is that aspects of cognition depend on the brain's ability to self-generate sequential neuronal activity. We found that reliably and continually changing cell assemblies in the rat hippocampus appeared not only during spatial navigation but also in the absence of changing environmental or body-derived inputs. During the delay period of a memory task, each moment in time was characterized by the activity of a particular assembly of neurons. Identical initial conditions triggered a similar assembly sequence, whereas different conditions gave rise to different sequences, thereby predicting behavioral choices, including errors. Such sequences were not formed in control (nonmemory) tasks. We hypothesize that neuronal representations, evolved for encoding distance in spatial navigation, also support episodic recall and the planning of action sequences.

Although short-term plasticity is believed to play a fundamental role in cortical computation, empirical evidence bearing on its role during behavior is scarce. Here we looked for the signature of short-term plasticity in the fine-timescale spiking relationships of a simultaneously recorded population of physiologically identified pyramidal cells and interneurons, in the medial prefrontal cortex of the rat, in a working memory task. On broader timescales, sequentially organized and transiently active neurons reliably differentiated between different trajectories of the rat in the maze. On finer timescales, putative monosynaptic interactions reflected short-term plasticity in their dynamic and predictable modulation across various aspects of the task, beyond a statistical accounting for the effect of the neurons’ co-varying firing rates. Seeking potential mechanisms for such effects, we found evidence for both firing pattern–dependent facilitation and depression, as well as for a supralinear effect of presynaptic coincidence on the firing of postsynaptic targets.

The hippocampal formation is believed to be critical for the encoding, consolidation, and retrieval of episodic memories. Yet, how these processes are supported by the anatomically diverse hippocampal networks is still unknown. To examine this issue, we tested rats in a hippocampus-dependent delayed spatial alternation task on a modified T maze while simultaneously recording local field potentials from dendritic and somatic layers of the dentate gyrus, CA3, and CA1 regions by using high-density, 96-site silicon probes. Both the power and coherence of gamma oscillations exhibited layer-specific changes during task performance. Peak increases in the gamma power and coherence were found in the CA3–CA1 interface on the maze segment approaching the T junction, independent of motor aspects of task performance. These results show that hippocampal networks can be dynamically coupled by gamma oscillations according to specific behavioral demands. Based on these findings, we propose that gamma oscillations may serve as a physiological mechanism by which CA3 output can coordinate CA1 activity to support retrieval of hippocampus-dependent memories.

Cytochrome c (CC) immunoreactivity was quantified in functionally distinct rat hippocampal inhibitory neuron populations using double immunocytochemistry and laser scanning confocal microscopy to measure the CC expression level as well as the amount of mitochondria within the cells, which is a sign of neuronal activity. The CC signal showed a similar distribution to cytochrome c oxidase histochemical staining. Strongly stained somata, dendrites and axon terminal clouds were dispersed over the low intensity neuropil staining. The staining was granular and electron microscopic investigation confirmed that the signal was localized in mitochondria. Intensively labeled neurons, showing the morphological features of inhibitory cells, were most frequently found in the principal cell layers, stratum oriens of the CA1-3 areas, stratum lucidum and hilus. These neurons contained not only a higher number of mitochondria than the principal cells but the intensity of the mitochondrial staining was evidently stronger. Among the examined interneuronal populations, parvalbumin-immunoreactive neurons were intensively labeled for CC. Calbindin D28k- (CB), somatostatin- and cholecystokinin-labeled cells showed heterogeneous CC levels, whereas calretinin-immunoreactive cells never showed a strong CC signal. CB cells in stratum oriens and alveus layers, lucidum and the hilus were strongly labeled for CC. CB cells in such regions are known to project to the medial septum and contain somatostatin. We have demonstrated that the CA1 interneurons that project to the medial septum (hippocampo-septal neurons) express a high level of CC. Thus, similar to the parvalbumin-containing basket and axo-axonic cells, the hippocampo-septal neurons potentially have a high average activity level.

SummaryBoth episodic memory and spatial navigation require temporal encoding of the relationships between events or locations. In a linear maze, ordered spatial distances between sequential locations were represented by the temporal relations of hippocampal place cell pairs within cycles of theta oscillation in a compressed manner. Such correlations could arise due to spike “phase precession” of independent neurons driven by common theta pacemaker or as a result of temporal coordination among specific hippocampal cell assemblies. We found that temporal correlation between place cell pairs was stronger than predicted by a pacemaker drive of independent neurons, indicating a critical role for synaptic interactions and precise timing within and across cell assemblies in place sequence representation. CA1 and CA3 ensembles, identifying spatial locations, were active preferentially on opposite phases of theta cycles. These observations suggest that interleaving CA3 neuronal sequences bind CA1 assemblies representing overlapping past, present, and future locations into single episodes.

International audience; High-frequency ripple oscillations, observed most prominently in the hippocampal CA1 pyramidal layer, are associated with memory consolidation. The cellular and network mechanisms underlying the generation of the rhythm and the recruitment of spikes from pyramidal neurons are still poorly understood. Using intracellular, sharp electrode recordings in freely moving, drug-free mice, we observed consistent large depolarizations in CA1 pyramidal cells during sharp wave ripples, which are associated with ripple frequency fluctuation of the membrane potential ("intracellular ripple"). Despite consistent depolarization, often exceeding pre-ripple spike threshold values, current pulse-induced spikes were strongly suppressed, indicating that spiking was under the control of concurrent shunting inhibition. Ripple events were followed by a prominent afterhyperpolarization and spike suppression. Action potentials during and outside ripples were orthodromic, arguing against ectopic spike generation, which has been postulated by computational models of ripple generation. These findings indicate that dendritic excitation of pyramidal neurons during ripples is countered by shunting of the membrane and postripple silence is mediated by hyperpolarizing inhibition.

Sharp wave and associated fast oscillatory ripples (140-200 Hz) in the cornu ammonis 1 region are the most synchronous hippocampal patterns in the adult rat. Spike sequences associated with sharp waves are believed to play a critical role in transferring transient memories from the hippocampus to the neocortex and the emergence of superfast ripples is pathognostic in temporal lobe epilepsy. Sharp waves in cornu ammonis 1 stratum radiatum are induced by a strong depolarization by the cornu ammonis 3 Schaffer collaterals, due to the synchronous discharge of cornu ammonis 3 pyramidal cells. Although during the first postnatal week, sharp-wave events are associated with hippocampal unit bursts in the pyramidal layer, ripple oscillations are absent. We investigated the emergence of fast-field oscillations in rat pups ranging from postnatal day 12-20 by recording with wire tetrodes in freely behaving pups and with 16-site linear silicon probes in head fixed animals. Cornu ammonis 1 pyramidal cell layer was determined by the presence of multiple unit activity and a reversal of the field potential in the deeper electrode sites. On-line verification of the recording sites was determined via an evoked response to commissural stimulation, showing a clear reversal in the field potential. Sharp wave-associated fast-field oscillations did not begin to emerge until the end of the second postnatal week and showed a gradual increase until day 18. Once ripples emerged, the intra-ripple frequency assumed adult values. The developmental time course of the ripple parallels the switch in the GABA(A) receptor-mediated signaling from excitation to inhibition. The time course may also reflect hitherto unidentified emergence of neuronal gap junctions.

Local hemodynamics of the cerebral cortex is the basis of modern functional imaging techniques, such as fMRIand PET. Despite the importance of local regulation of the blood flow, capillary level quantification of cerebral blood flow has been limited by the spatial resolution of functional imaging techniques and the depth penetration of conventional optical microscopy. Two-photon laser scanning microscopic imaging technique has the necessary spatial resolution and can image capillaries in the depth of the cortex. We have loaded the serum with fluorescein isothiocyanate dextran and quantified the flow of red blood cells (RBCs) in capillaries in layers 2/3 of the mouse somatosensory cortex in vivo. Basal capillary flux was quantified as approximately 28.9+/-13.6 RBCs/s (n=50, mean+/-S.D.) under ketamine-xylazine anesthesia and 26.7+/-16.0 RBCs/s (n=31) under urethane anesthesia. Focal interictal (epileptiform) activity was induced by local infusion of bicuculline methochloride in the cortex. We have observed that capillary blood flow increased as the cortical local field events developed into epileptiform in the vicinity of GABA receptor blockade (300 microm from the administration site). Local blood flow in the interictal focus increased significantly (42.5+/-18.5RBCs/s, n=52) relative to the control conditions or to blood flow measured in capillaries at distant (1mm from the focus) sites from the epileptic focus (27.8+/-12.9 RBCs/s, n=30). These results show that hyper-synchronized neural activity is associated with increased capillary perfusion in a localized cortical area. This volume is significantly smaller than the currently available resolution of the fMRI signal.

Neural-network oscillations at distinct frequencies have been implicated in the encoding, consolidation and retrieval of information in the hippocampus. Some GABA (gamma-aminobutyric acid)-containing interneurons fire phase-locked to theta oscillations (4-8 Hz) or to sharp-wave-associated ripple oscillations (120-200 Hz), which represent different behavioural states. Interneurons also entrain pyramidal cells in vitro. The large diversity of interneurons poses the question of whether they have specific roles in shaping distinct network activities in vivo. Here we report that three distinct interneuron types--basket, axo-axonic and oriens-lacunosum-moleculare cells--visualized and defined by synaptic connectivity as well as by neurochemical markers, contribute differentially to theta and ripple oscillations in anaesthetized rats. The firing patterns of individual cells of the same class are remarkably stereotyped and provide unique signatures for each class. We conclude that the diversity of interneurons, innervating distinct domains of pyramidal cells, emerged to coordinate the activity of pyramidal cells in a temporally distinct and brain-state-dependent manner.