ATP-Binding cassette (ABC) transporters are a family of integral membrane ATPases that transport a large number of structurally unrelated compounds. The physiological role of the orphan transporter Abcc5 remains poorly understood. As previous work demonstrated that the loss of Abcc5 activity leads to elevated levels of NAAG in the brain, the impact of Abcc5 ablation was ascertained using behavioural phenotyping, circadian rhythm analysis and electrophysiological recordings of brain slices from Abcc5-/- mice and compared to wild-type littermates. Behavioural phenotyping of Abcc5-/- mice shows that the loss of murine Abcc5 activity results in profound changes in pre-pulse inhibition (PPI) as well as altered memory consolidation. Circadian measures of activity showed a delay in the timing of Abcc5-/- mice activity rhythm peak. Additionally, activity defined sleep analysis highlighted differences in sleep patterns in Abcc5-/- mice compared to wild-type controls. Patch clamp recording from pyramidal cells in the 2/3 layer of the frontal cortex showed altered synaptic AMPA/NMDA receptor current ratios and increased frequency of spontaneous excitatory postsynaptic currents (sEPSC). This study demonstrates that the loss of functional Abcc5 transporters does have behavioural consequences in mammals and alters NMDA receptor activity. These results highlight a previously unknown role of Abcc5 in the brain.

Cortical layer 6 neurons are the only projection neuron population in the cortical mantle known to electrophysiologically respond to orexin—a neuropeptide involved in cortical arousal and emotive behaviour. These neurons exhibit extensive intercortical and thalamic projections, yet the exact mechanisms underlying these responses are not fully understood. We hypothesize that cortical circuits activated by orexin sensitive L6 neurons in the medial prefrontal cortex (mPFC) are responsible for detecting salient features of sensory stimuli and are therefore involved in regulating emotional states. Here, we show that Drd1a-Cre+ neurons in the mPFC are selectively sensitive to orexin and gate the activation of the prefrontal network in vivo. Moreover, we demonstrated that chronically “silencing” this subpopulation of L6 neurons (Drd1a-Cre+/+:Snap25fl/fl) across the cortical mantle from birth abolishes the orexin-induced prefrontal activation. Consequently, the chronic silencing of these neurons had strong anxiolytic effects on several anxiety-related behavioural paradigms, indicating that orexin-responsive L6 neurons modulate emotional states and may be a substrate for anxiety regulation.

Depolarization of axons is necessary for somatic action potentials to trigger axonal neurotransmitter release. Here we show that striatal cholinergic interneurons (ChIs) and nicotinic receptors (nAChRs) on mouse dopamine axons interrupt this relationship. After nAChR-mediated depolarization, dopamine release by subsequent depolarization events was suppressed for ~100 ms. This suppression was not due to depletion of dopamine or acetylcholine, but to a limited reactivation of dopamine axons after nAChR-mediated depolarization, and is more prominent in dorsal than in ventral striatum. In vivo, nAChRs predominantly depressed dopamine release, as nAChR antagonism in dorsal striatum elevated dopamine detected with optic-fiber photometry of dopamine sensor GRAB DA2m and promoted conditioned place preference. Our findings reveal that ChIs acting via nAChRs transiently limit the reactivation of dopamine axons for subsequent action potentials in dopamine neurons and therefore generate a dynamic inverse scaling of dopamine release according to ChI activity.

Parvalbumin-expressing (PV+) interneurons exert exquisite control over spike output, and plasticity in these inhibitory circuits may be important for maintaining network stability in learning and memory. PV+ interneuron recruitment is primarily mediated by GluA2-lacking Ca 2+ -permeable AMPA receptors (CP-AMPAR), which support anti-Hebbian plasticity. However, the functional significance of CP-AMPAR-mediated plasticity remains unknown. Using a viral approach to artificially express the GluA2 subunit in hippocampal PV+ interneurons, we replaced CP-AMPAR with GluA2-containing receptors, and in doing so reduced synaptically-evoked Ca 2+ transients and anti-Hebbian plasticity. Transfection of hippocampal PV+ interneurons with GluA2 resulted in delay-dependent spatial working memory deficits which increased across trials per session, and impaired reversal learning in the Morris water maze but not initial acquisition. Our data suggest that loss of CP-AMPAR-mediated plasticity in these cells leads to proactive interference, revealing a significant role for dynamic recruitment of PV+ interneurons in the segregation of memories and accurate memory retrieval. Highlights:Viral expression of GluA2 in PV+ interneurons alters synaptic AMPA receptor profile GluA2 overexpression in PV+ cells reduces synaptic Ca 2+ transients and plasticity Upregulating GluA2 in PV+ cells causes delay-dependent deficits in working memoryAcquisition of reference memories is preserved, but reversal learning is impaired.

Here, we characterized the dynamics of sleep spindles, focusing on their damping, which we estimated using a metric called oscillatory-Quality (o-Quality), derived by fitting an autoregressive model to electrophysiological signals, recorded from the cortex in mice. The o-Quality of sleep spindles correlates weakly with their amplitude, shows marked laminar differences and regional topography across cortical regions, reflects the level of synchrony within and between cortical networks, is strongly modulated by sleep-wake history, reflects the degree of sensory disconnection, and correlates with the strength of coupling between spindles and slow waves. As most spindle events are highly localized and not detectable with conventional low-density recording approaches, o-Quality thus emerges as a valuable metric that allows us to infer the spread and dynamics of spindle activity across the brain and directly links their spatiotemporal dynamics with local and global regulation of brain states, sleep regulation, and function.

Bidirectional communication between neurons and glial cells is crucial to establishing and maintaining normal brain function. Some of these interactions are activity-dependent, yet it remains largely unexplored how acute changes in neuronal activity affect glial-to-neuron and neuron-to-glial dynamics. Here, we use excitatory and inhibitory designer receptors exclusively activated by designer drugs (DREADD) to study the effects of acute chemogenetic manipulations of a subpopulation of layer 5 cortical projection and dentate gyrus neurons in adult (Rbp4Cre) mouse brains. We show that acute chemogenetic neuronal activation reduces synaptic density, and increases microglia and astrocyte reactivity, but does not affect parvalbumin (PV+) neurons, only perineuronal nets (PNN). Conversely, acute silencing increases synaptic density and decreases glial reactivity. We show fast glial response upon clozapine-N-oxide (CNO) administration in cortical and subcortical regions. Together, our work provides evidence of fast, activity-dependent, bidirectional interactions between neurons and glial cells. Acute manipulations of neuronal activity of cortical projection neurons using DREADDs affects synaptic connectivity and leads to rapid response and changes in glial cell dynamics in cortical and subcortical regions.

Depolarisation of distal axons is necessary for neurons to translate somatic action potentials into neurotransmitter release. Studies have shown that striatal cholinergic interneurons (ChIs) can directly drive ectopic action potentials in dopamine (DA) axons and trigger DA release. However, here we show that this action occurs within a broader context of axonal signal integration whereby activation of ChIs and depolarisation of DA axons by nicotinic receptors (nAChRs) limits the subsequent depolarisation and release of DA in response to ensuing activity. We demonstrate that activation of ChIs and nAChRs in ex vivo mouse striatum, even when it does not trigger DA release that is detectable by fast-scan cyclic voltammetry, limits DA release for ∼100 ms by depressing subsequent axonal depolarisation and calcium summation. This axonal brake on DA release is stronger in dorsal than ventral striatum, and is unrelated to DA depletion. In vivo , antagonism of nAChRs in dorsal striatum elevated extracellular DA levels and promoted conditioned place-preference, underscoring its physiological relevance. Our findings reveal that under physiological conditions in vivo, ChIs acting via nAChRs dynamically attenuate DA output driven by DA neuron activity, leading to a predominantly inverse relationship between ACh and DA signalling that varies continuously with ChI activity. ### Competing Interest Statement Y.-F.Z. is the owner of patent (WO2023175357A1).

Sleep spindles are traditionally defined as 10-15Hz thalamo-cortical oscillations typical of NREM sleep. While substantial heterogeneity in the appearance or spatio-temporal dynamics of spindle events is well recognised, the physiological relevance of the underlying fundamental property - the oscillatory strength - has not been studied. Here we introduce a novel metric called oscillatory Quality ( o-Quality ), which is derived by fitting an auto-regressive model to short segments of electrophysiological signals, recorded from the cortex in mice, to identify and calculate the damping of spindle oscillations. We find that the o-Quality of spindles varies markedly across cortical layers and regions and reflects the level of synchrony within and between cortical networks. Furthermore, the o-Quality of spindles varies as a function of sleep-wake history, determines the strength of coupling between spindles and slow waves, and influences the responsiveness to sensory stimulation during sleep. Thus, the o-Quality emerges as a metric that, for the first time, directly links the spatio-temporal dynamics of sleep spindles with their functional role.

The slow oscillation (SO) is a central neuronal dynamic during sleep and is generated by alternating periods of high and low neuronal activity (ON and OFF states). Mounting evidence causally links the SO to sleep’s functions, and it has recently become possible to manipulate the SO non-invasively and phase-specifically. These developments represent promising clinical avenues, but they also highlight the importance of improving our understanding of how ON/OFF states affect incoming stimuli and what role they play in neuronal plasticity. Most studies using closed-loop stimulation rely on the electroencephalogram (EEG) and local field potential (LFP) signals, which reflect neuronal ON and OFF states only indirectly. Here we develop an online detection algorithm based on spiking activity recorded from laminar arrays in mouse motor cortex. We find that online detection of ON and OFF states reflects specific phases of spontaneous LFP SO. Our neuronal-spiking-based closed-loop procedure offers a novel opportunity for testing the functional role of SO in sleep-related restorative processes and neural plasticity.

The spatiotemporal distribution of mitochondria is crucial for precise ATP provision and calcium buffering required to support neuronal signaling. Fast-spiking GABAergic interneurons expressing parvalbumin (PV+) have a high mitochondrial content reflecting their large energy utilization. The importance for correct trafficking and precise mitochondrial positioning remains poorly elucidated in inhibitory neurons. Miro1 is a Ca²+-sensing adaptor protein that links mitochondria to the trafficking apparatus, for their microtubule-dependent transport along axons and dendrites, in order to meet the metabolic and Ca2+-buffering requirements of the cell. Here, we explore the role of Miro1 in PV+ interneurons and how changes in mitochondrial trafficking could alter network activity in the mouse brain. By employing live and fixed imaging, we found that the impairments in Miro1-directed trafficking in PV+ interneurons altered their mitochondrial distribution and axonal arborization, while PV+ interneuron-mediated inhibition remained intact. These changes were accompanied by an increase in the ex vivo hippocampal γ-oscillation (30–80 Hz) frequency and promoted anxiolysis. Our findings show that precise regulation of mitochondrial dynamics in PV+ interneurons is crucial for proper neuronal signaling and network synchronization.

Sleep and wakefulness are not simple, homogenous all-or-none states but represent a spectrum of substates, distinguished by behavior, levels of arousal, and brain activity at the local and global levels. Until now, the role of the hypothalamic circuitry in sleep-wake control was studied primarily with respect to its contribution to rapid state transitions. In contrast, whether the hypothalamus modulates within-state dynamics (state "quality") and the functional significance thereof remains unexplored. Here, we show that photoactivation of inhibitory neurons in the lateral preoptic area (LPO) of the hypothalamus of adult male and female laboratory mice does not merely trigger awakening from sleep, but the resulting awake state is also characterized by an activated electroencephalogram (EEG) pattern, suggesting increased levels of arousal. This was associated with a faster build-up of sleep pressure, as reflected in higher EEG slow-wave activity (SWA) during subsequent sleep. In contrast, photoinhibition of inhibitory LPO neurons did not result in changes in vigilance states but was associated with persistently increased EEG SWA during spontaneous sleep. These findings suggest a role of the LPO in regulating arousal levels, which we propose as a key variable shaping the daily architecture of sleep-wake states.

We have shown previously that prebiotic (Bimuno galacto-oligosacharides, B-GOS®) administration to neonatal rats increased hippocampal NMDAR proteins. The present study has investigated the effects of postnatal B-GOS® supplementation on hippocampus-dependent behavior in young, adolescent, and adult rats and applied electrophysiological, metabolomic and metagenomic analyses to explore potential underlying mechanisms. The administration of B-GOS® to suckling, but not post-weaned, rats reduced anxious behavior until adulthood. Neonatal prebiotic intake also reduced the fast decay component of hippocampal NMDAR currents, altered age-specific trajectories of the brain, intestinal, and liver metabolomes, and reduced abundance of fecal Enterococcus and Dorea bacteria. Our data are the first to show that prebiotic administration to rats during a specific postnatal period has long-term effects on behavior and hippocampal physiology. The study also suggests that early-life prebiotic intake may affect host brain function through the reduction of stress-related gut bacteria rather than increasing the proliferation of beneficial microbes.

Central serotonin (5-HT) orchestrates myriad cognitive processes and lies at the core of many stress-related psychiatric illnesses. However, the basic relationship between its brain-wide axonal projections and functional dynamics is not known. Here we combine optogenetics and fMRI to produce a brain-wide 5-HT evoked functional map. We find that DRN photostimulation leads to an increase in the hemodynamic response in the DRN itself, while projection areas predominately exhibit a reduction of cerebral blood volume mirrored by suppression of cortical delta oscillations. We find that the regional distribution of post-synaptically expressed 5-HT receptors better correlates with DRN 5-HT functional connectivity than anatomical projections. Our work suggests that neuroarchitecture is not the primary determinant of function for the DRN 5-HT. With respect to two 5-HT elevating stimuli, we find that acute stress leads to circuit-wide blunting of the DRN output, while the SSRI fluoxetine noticeably enhances DRN functional connectivity. These data provide fundamental insight into the brain-wide functional dynamics of the 5-HT projection system.

Feeding neonatal rats with a prebiotic (Bimuno galacto-oligosaccharides, B-GOS®), increased hippocampal NMDAR proteins. Here, we tested the effects of postnatal B-GOS® supplementation on anxious behavior, hippocampal electrophysiology, metabolomics, and faecal metagenomics in young, adolescent and adult rats. Postnatal, but not post-wean, B-GOS® administration reduced anxiety-like behaviour in adulthood, decreased the fast decay component of Τ (tau) of hippocampus CA1 NMDA currents and changed the age-specific modifications of the metabolomes of the brain, liver and intestines. Age, but not diet, affected the composition of the fecal microbiome overall on taxonomic genus level. However, B-GOS® supplementation reduced the abundance of Enterococcus and Dorea genera. In conclusion, the long-term anxiolytic effect of early-life B-GOS® intake, may be linked to altered hippocampal NMDA receptors kinetics. The prebiotic mediated changes in gut Enterococcus and Dorea and age-specific metabolic trajectories of central and peripheral tissues, may have contributed to the prolonged behavioural effect.

Contrast gain control is the systematic adjustment of neuronal gain in response to the contrast of sensory input. It is widely observed in sensory cortical areas and has been proposed to be a canonical neuronal computation. Here, we investigated whether shunting inhibition from parvalbumin-positive interneurons-a mechanism involved in gain control in visual cortex-also underlies contrast gain control in auditory cortex. First, we performed extracellular recordings in the auditory cortex of anesthetized male mice and optogenetically manipulated the activity of parvalbumin-positive interneurons while varying the contrast of the sensory input. We found that both activation and suppression of parvalbumin interneuron activity altered the overall gain of cortical neurons. However, despite these changes in overall gain, we found that manipulating parvalbumin interneuron activity did not alter the strength of contrast gain control in auditory cortex. Furthermore, parvalbumin-positive interneurons did not show increases in activity in response to high-contrast stimulation, which would be expected if they drive contrast gain control. Finally, we performed in vivo whole-cell recordings in auditory cortical neurons during high- and low-contrast stimulation and found that no increase in membrane conductance was observed during high-contrast stimulation. Taken together, these findings indicate that while parvalbumin-positive interneuron activity modulates the overall gain of auditory cortical responses, other mechanisms are primarily responsible for contrast gain control in this cortical area.NEW & NOTEWORTHY We investigated whether contrast gain control is mediated by shunting inhibition from parvalbumin-positive interneurons in auditory cortex. We performed extracellular and intracellular recordings in mouse auditory cortex while presenting sensory stimuli with varying contrasts and manipulated parvalbumin-positive interneuron activity using optogenetics. We show that while parvalbumin-positive interneuron activity modulates the gain of cortical responses, this activity is not the primary mechanism for contrast gain control in auditory cortex.

AbstractSleep and wakefulness are not simple homogenous all-or-none states, but instead are characterized by rich dynamics of brain activity across many temporal and spatial scales. Rapid global state transitions between waking and sleeping are believed to be controlled by hypothalamic circuits, but the contribution of the hypothalamus to within-state changes of sleep and wake “intensity” remains largely unexplored. Here we show that stimulation of inhibitory neurons in the preoptic hypothalamus does not merely trigger awakening from sleep, but the resulting awake state is also characterized by increased cortical activity. This activation is associated with a faster build-up of sleep pressure, proportional to the arousal level. These findings show that hypothalamic systems thought to exclusively control global state switching, also regulate within-state “intensity”, which we propose as a key intrinsic variable in shaping the architecture of sleep/wake states across the 24h day.

Cardiac stimulation via sympathetic neurons can potentially trigger arrhythmias. We present approaches to study neuron-cardiomyocyte interactions involving optogenetic selective probing and all-optical electrophysiology to measure activity in an automated fashion. Here we demonstrate the utility of optical interrogation of sympathetic neurons and their effects on macroscopic cardiomyocyte network dynamics to address research targets such as the effects of adrenergic stimulation via the release of neurotransmitters, the effect of neuronal numbers on cardiac behavior, and the applicability of optogenetics in mechanistic in vitro studies. As arrhythmias are emergent behaviors that involve the coordinated activity of millions of cells, we image at macroscopic scales to capture complex dynamics. We show that neurons can both decrease and increase wave stability and re-entrant activity in culture depending on their induced activity-a finding that may help us understand the often conflicting results seen in experimental and clinical studies.

γ-frequency oscillations (30-120 Hz) in cortical networks influence neuronal encoding and information transfer, and are disrupted in multiple brain disorders. While synaptic inhibition is important for synchronization across the γ-frequency range, the role of distinct interneuronal subtypes in slow (<60 Hz) and fast γ states remains unclear. Here, we used optogenetics to examine the involvement of parvalbumin-expressing (PV + ) and somatostatin-expressing (SST + ) interneurons in γ oscillations in the mouse hippocampal CA3 ex vivo , using animals of either sex. Disrupting either PV + or SST + interneuron activity, via either photoinhibition or photoexcitation, led to a decrease in the power of cholinergically induced slow γ oscillations. Furthermore, photoexcitation of SST + interneurons induced fast γ oscillations, which depended on both synaptic excitation and inhibition. Our findings support a critical role for both PV + and SST + interneurons in slow hippocampal γ oscillations, and further suggest that intense activation of SST + interneurons can enable the CA3 circuit to generate fast γ oscillations. SIGNIFICANCE STATEMENT The generation of hippocampal γ oscillations depends on synchronized inhibition provided by GABAergic interneurons. Parvalbumin-expressing (PV + ) interneurons are thought to play the key role in coordinating the spike timing of excitatory pyramidal neurons, but the role distinct inhibitory circuits in network synchronization remains unresolved. Here, we show, for the first time, that causal disruption of either PV + or somatostatin-expressing (SST + ) interneuron activity impairs the generation of slow γ oscillations in the ventral hippocampus ex vivo . We further show that SST + interneuron activation along with general network excitation is sufficient to generate high-frequency γ oscillations in the same preparation. These results affirm a crucial role for both PV + and SST + interneurons in hippocampal γ oscillation generation.

The brain has a remarkable capacity to adapt to changes in sensory inputs and to learn from experience. However, the neural circuits responsible for this flexible processing remain poorly understood. Using optogenetic silencing of ArchT-expressing neurons in adult ferrets, we show that within-trial activity in primary auditory cortex (A1) is required for training-dependent recovery in sound-localization accuracy following monaural deprivation. Because localization accuracy under normal-hearing conditions was unaffected, this highlights a specific role for cortical activity in learning. A1-dependent plasticity appears to leave a memory trace that can be retrieved, facilitating adaptation during a second period of monaural deprivation. However, in ferrets in which learning was initially disrupted by perturbing A1 activity, subsequent optogenetic suppression during training no longer affected localization accuracy when one ear was occluded. After the initial learning phase, the reweighting of spatial cues that primarily underpins this plasticity may therefore occur in A1 target neurons.

Mesostriatal dopaminergic neurons possess extensively branched axonal arbours. Whether action potentials are converted to dopamine output in the striatum will be influenced dynamically and critically by axonal properties and mechanisms that are poorly understood. Here, we address the roles for mechanisms governing release probability and axonal activity in determining short‐term plasticity of dopamine release, using fast‐scan cyclic voltammetry in the ex vivo mouse striatum. We show that brief short‐term facilitation and longer short term depression are only weakly dependent on the level of initial release, i.e. are release insensitive. Rather, short-term plasticity is strongly determined by mechanisms which govern axonal activation, including K + ‐gated excitability and the dopamine transporter, particularly in the dorsal striatum. We identify the dopamine transporter as a master regulator of dopamine short‐term plasticity, governing the balance between release‐dependent and independent mechanisms that also show region‐specific gating.