Hippocampal representation during collective spatial behaviour in bats.

Social animals live and move through spaces shaped by the presence, motion and sensory cues of multiple other individuals1-6. Neural activity in the hippocampus is known to reflect spatial behaviour7-9 yet its study is lacking in such dynamic group settings, which are ubiquitous in natural environments. Here we studied hippocampal activity in groups of bats engaged in collective spatial behaviour. We find that, under spontaneous conditions, a robust spatial structure emerges at the group level whereby behaviour is anchored to specific locations, movement patterns and individual social preferences. Using wireless electrophysiological recordings from both stationary and flying bats, we find that many hippocampal neurons are tuned to key features of group dynamics. These include the presence or absence of a conspecific, but not typically of an object, at landing sites, shared spatial locations, individual identities and sensory signals that are broadcasted in the group setting. Finally, using wireless calcium imaging, we find that social responses are anatomically distributed and robustly represented at the population level. Combined, our findings reveal that hippocampal activity contains a rich representation of naturally emerging spatial behaviours in animal groups that could in turn support the complex feat of collective behaviour.

Enhanced Feedback Inhibition Due to Increased Recruitment of Somatostatin-Expressing Interneurons and Enhanced Cortical Recurrent Excitation in a Genetic Mouse Model of Migraine.

Migraine is a complex brain disorder, characterized by attacks of unilateral headache and global dysfunction in multisensory information processing, whose underlying cellular and circuit mechanisms remain unknown. The finding of enhanced excitatory, but unaltered inhibitory, neurotransmission at cortical synapses between pyramidal cells (PCs) and fast-spiking interneurons (FS INs) in mouse models of familial hemiplegic migraine (FHM) suggested the hypothesis that dysregulation of the excitatory-inhibitory (E/I) balance in specific circuits is a key pathogenic mechanism. Here, we investigated the cortical layer 2/3 (L2/3) feedback inhibition microcircuit involving somatostatin-expressing (SOM) INs in FHM1 mice of both sexes carrying a gain-of-function mutation in CaV2.1. Unitary inhibitory neurotransmission at SOM IN-PC synapses was unaltered while excitatory neurotransmission at both PC-SOM IN and PC-PC synapses was enhanced, because of increased probability of glutamate release, in FHM1 mice. Short-term synaptic depression was enhanced at PC-PC synapses while short-term synaptic facilitation was unaltered at PC-SOM IN synapses during 25-Hz repetitive activity. The frequency-dependent disynaptic inhibition (FDDI) mediated by SOM INs was enhanced, lasted longer and required shorter high-frequency bursts to be initiated in FHM1 mice. These findings, together with previous evidence of enhanced disynaptic feedforward inhibition by FS INs, suggest that the increased inhibition may effectively counteract the increased recurrent excitation in FHM1 mice and may even prevail in certain conditions. Considering the involvement of SOM INs in γ oscillations, surround suppression and context-dependent sensory perception, the facilitated recruitment of SOM INs, together with the enhanced recurrent excitation, may contribute to dysfunctional sensory processing in FHM1 and possibly migraine.SIGNIFICANCE STATEMENTMigraine is a complex brain disorder, characterized by attacks of unilateral headache and global dysfunction in multisensory information processing, whose underlying cellular and circuit mechanisms remain unknown, although dysregulation of the excitatory-inhibitory (E/I) balance in specific circuits could be a key pathogenic mechanism. Here, we provide insights into these mechanisms by investigating the cortical feedback inhibition microcircuit involving somatostatin-expressing interneurons (SOM INs) in a mouse model of a rare monogenic migraine. Despite unaltered inhibitory synaptic transmission, the disynaptic feedback inhibition mediated by SOM INs was enhanced in the migraine model because of enhanced recruitment of the INs. Recurrent cortical excitation was also enhanced. These alterations may contribute to context-dependent sensory processing dysfunctions in migraine.

A stable hippocampal code in freely flying bats.

Neural activity in the hippocampus is known to reflect how animals move through an environment1,2. Although navigational behaviour may show considerable stability3-6, the tuning stability of individual hippocampal neurons remains unclear7-12. Here we used wireless calcium imaging to longitudinally monitor the activity of dorsal CA1 hippocampal neurons in freely flying bats performing highly reproducible flights in a familiar environment. We find that both the participation and the spatial selectivity of most neurons remain stable over days and weeks. We also find that apparent changes in tuning can be largely attributed to variations in the flight behaviour of the bats. Finally, we show that bats navigating in the same environment under different room lighting conditions (lights on versus lights off) exhibit substantial changes in flight behaviour that can give the illusion of neuronal instability. However, when similar flight paths are compared across conditions, the stability of the hippocampal code persists. Taken together, we show that the underlying hippocampal code is highly stable over days and across contexts if behaviour is taken into account.

Stimulus Feature-Specific Control of Layer 2/3 Subthreshold Whisker Responses by Layer 4 in the Mouse Primary Somatosensory Cortex.

In the barrel field of the rodent primary somatosensory cortex (S1bf), excitatory cells in layer 2/3 (L2/3) display sparse firing but reliable subthreshold response during whisker stimulation. Subthreshold responses encode specific features of the sensory stimulus, for example, the direction of whisker deflection. According to the canonical model for the flow of sensory information across cortical layers, activity in L2/3 is driven by layer 4 (L4). However, L2/3 cells receive excitatory inputs from other regions, raising the possibility that L4 partially drives L2/3 during whisker stimulation. To test this hypothesis, we combined patch-clamp recordings from L2/3 pyramidal neurons in S1bf with selective optogenetic inhibition of L4 during passive whisker stimulation in both anesthetized and awake head-restrained mice. We found that L4 optogenetic inhibition did not abolish the subthreshold whisker-evoked response nor it affected spontaneous membrane potential fluctuations of L2/3 neurons. However, L4 optogenetic inhibition decreased L2/3 subthreshold responses to whisker deflections in the preferred direction, and it increased L2/3 responses to stimuli in the nonpreferred direction, leading to a change in the direction tuning. Our results contribute to reveal the circuit mechanisms underlying the processing of sensory information in the rodent S1bf.

Optogenetic strategies for high-efficiency all-optical interrogation using blue-light-sensitive opsins.

All-optical methods for imaging and manipulating brain networks with high spatial resolution are fundamental to study how neuronal ensembles drive behavior. Stimulation of neuronal ensembles using two-photon holographic techniques requires high-sensitivity actuators to avoid photodamage and heating. Moreover, two-photon-excitable opsins should be insensitive to light at wavelengths used for imaging. To achieve this goal, we developed a novel soma-targeted variant of the large-conductance blue-light-sensitive opsin CoChR (stCoChR). In the mouse cortex in vivo, we combined holographic two-photon stimulation of stCoChR with an amplified laser tuned at the opsin absorption peak and two-photon imaging of the red-shifted indicator jRCaMP1a. Compared to previously characterized blue-light-sensitive soma-targeted opsins in vivo, stCoChR allowed neuronal stimulation with more than 10-fold lower average power and no spectral crosstalk. The combination of stCoChR, tuned amplified laser stimulation, and red-shifted functional indicators promises to be a powerful tool for large-scale interrogation of neural networks in the intact brain.

Extended field-of-view ultrathin microendoscopes for high-resolution two-photon imaging with minimal invasiveness.

Imaging neuronal activity with high and homogeneous spatial resolution across the field-of-view (FOV) and limited invasiveness in deep brain regions is fundamental for the progress of neuroscience, yet is a major technical challenge. We achieved this goal by correcting optical aberrations in gradient index lens-based ultrathin (≤500 µm) microendoscopes using aspheric microlenses generated through 3D-microprinting. Corrected microendoscopes had extended FOV (eFOV) with homogeneous spatial resolution for two-photon fluorescence imaging and required no modification of the optical set-up. Synthetic calcium imaging data showed that, compared to uncorrected endoscopes, eFOV-microendoscopes led to improved signal-to-noise ratio and more precise evaluation of correlated neuronal activity. We experimentally validated these predictions in awake head-fixed mice. Moreover, using eFOV-microendoscopes we demonstrated cell-specific encoding of behavioral state-dependent information in distributed functional subnetworks in a primary somatosensory thalamic nucleus. eFOV-microendoscopes are, therefore, small-cross-section ready-to-use tools for deep two-photon functional imaging with unprecedentedly high and homogeneous spatial resolution.

Temporal Sharpening of Sensory Responses by Layer V in the Mouse Primary Somatosensory Cortex.

The timing of stimulus-evoked spikes encodes information about sensory stimuli. Here we studied the neural circuits controlling this process in the mouse primary somatosensory cortex. We found that brief optogenetic activation of layer V pyramidal cells just after whisker deflection modulated the membrane potential of neurons and interrupted their long-latency whisker responses, increasing their accuracy in encoding whisker deflection time. In contrast, optogenetic inhibition of layer V during either passive whisker deflection or active whisking decreased accuracy in encoding stimulus or touch time, respectively. Suppression of layer V pyramidal cells increased reaction times in a texture discrimination task. Moreover, two-color optogenetic experiments revealed that cortical inhibition was efficiently recruited by layer V stimulation and that it mainly involved activation of parvalbumin-positive rather than somatostatin-positive interneurons. Layer V thus performs behaviorally relevant temporal sharpening of sensory responses through circuit-specific recruitment of cortical inhibition.

Two-Photon Bidirectional Control and Imaging of Neuronal Excitability with High Spatial Resolution In Vivo.

Sensory information is encoded within the brain in distributed spatiotemporal patterns of neuronal activity. Understanding how these patterns influence behavior requires a method to measure and to bidirectionally perturb with high spatial resolution the activity of the multiple neuronal cell types engaged in sensory processing. Here, we combined two-photon holography to stimulate neurons expressing blue light-sensitive opsins (ChR2 and GtACR2) with two-photon imaging of the red-shifted indicator jRCaMP1a in the mouse neocortex in vivo. We demonstrate efficient control of neural excitability across cell types and layers with holographic stimulation and improved spatial resolution by opsin somatic targeting. Moreover, we performed simultaneous two-photon imaging of jRCaMP1a and bidirectional two-photon manipulation of cellular activity with negligible effect of the imaging beam on opsin excitation. This all-optical approach represents a powerful tool to causally dissect how activity patterns in specified ensembles of neurons determine brain function and animal behavior.

Interneuron-specific signaling evokes distinctive somatostatin-mediated responses in adult cortical astrocytes.

The signaling diversity of GABAergic interneurons to post-synaptic neurons is crucial to generate the functional heterogeneity that characterizes brain circuits. Whether this diversity applies to other brain cells, such as the glial cells astrocytes, remains unexplored. Using optogenetics and two-photon functional imaging in the adult mouse neocortex, we here reveal that parvalbumin- and somatostatin-expressing interneurons, two key interneuron classes in the brain, differentially signal to astrocytes inducing weak and robust GABAB receptor-mediated Ca2+ elevations, respectively. Furthermore, the astrocyte response depresses upon parvalbumin interneuron repetitive stimulations and potentiates upon somatostatin interneuron repetitive stimulations, revealing a distinguished astrocyte plasticity. Remarkably, the potentiated response crucially depends on the neuropeptide somatostatin, released by somatostatin interneurons, which activates somatostatin receptors at astrocytic processes. Our study unveils, in the living brain, a hitherto unidentified signaling specificity between interneuron subtypes and astrocytes opening a new perspective into the role of astrocytes as non-neuronal components of inhibitory circuits.

Unaltered Network Activity and Interneuronal Firing During Spontaneous Cortical Dynamics In Vivo in a Mouse Model of Severe Myoclonic Epilepsy of Infancy.

Severe myoclonic epilepsy of infancy (SMEI) is associated with loss of function of the SCN1A gene encoding the NaV1.1 sodium channel isoform. Previous studies in Scn1a(-/+) mice during the pre-epileptic period reported selective reduction in interneuron excitability and proposed this as the main pathological mechanism underlying SMEI. Yet, the functional consequences of this interneuronal dysfunction at the circuit level in vivo are unknown. Here, we investigated whether Scn1a(-/+) mice showed alterations in cortical network function. We found that various forms of spontaneous network activity were similar in Scn1a(-/+) during the pre-epileptic period compared with wild-type (WT) in vivo. Importantly, in brain slices from Scn1a(-/+) mice, the excitability of parvalbumin (PV) and somatostatin (SST) interneurons was reduced, epileptiform activity propagated more rapidly, and complex synaptic changes were observed. However, in vivo, optogenetic reduction of firing in PV or SST cells in WT mice modified ongoing network activities, and juxtasomal recordings from identified PV and SST interneurons showed unaffected interneuronal firing during spontaneous cortical dynamics in Scn1a(-/+) compared with WT. These results demonstrate that interneuronal hypoexcitability is not observed in Scn1a(-/+) mice during spontaneous activities in vivo and suggest that additional mechanisms may contribute to homeostatic rearrangements and the pathogenesis of SMEI.