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Sensory adaptation is the process whereby brain circuits adjust neuronal activity in response to redundant sensory stimuli. Although sensory adaptation has been extensively studied for individual neurons on timescales of tens of milliseconds to a few seconds, little is known about it over longer timescales or at the population level. We investigated population-level adaptation in the barrel field of the mouse somatosensory cortex (S1BF) using in vivo two-photon calcium imaging and Neuropixels recordings in awake mice. Among stimulus-responsive neurons, we found both adapting and facilitating neurons, which decreased or increased their firing, respectively, with repetitive whisker stimulation. The former outnumbered the latter by 2:1 in layers 2/3 and 4; hence, the overall population response of mouse S1BF was slightly adapting. We also discovered that population adaptation to one stimulus frequency (5 Hz) does not necessarily generalize to a different frequency (12.5 Hz). Moreover, responses of individual neurons to repeated rounds of stimulation over tens of minutes were strikingly heterogeneous and stochastic, such that their adapting or facilitating response profiles were not stable across time. Such representational drift was particularly striking when recording longitudinally across 8-9 days, as adaptation profiles of most whisker-responsive neurons changed drastically from one day to the next. Remarkably, repeated exposure to a familiar stimulus paradoxically shifted the population away from strong adaptation and toward facilitation. Thus, the adapting vs. facilitating response profile of S1BF neurons is not a fixed property of neurons but rather a highly dynamic feature that is shaped by sensory experience across days.

Thermogenetics is a promising neuromodulation technique based on the use of heat-sensitive ion channels. However, on the way to its clinical application, a number of questions have to be addressed. First, to avoid immune response in future human applications, human ion channels should be studied as thermogenetic actuators. Second, heating levels necessary to activate these channels in vivo in brain tissue should be studied and cytotoxicity of these temperatures addressed. Third, the possibility and safety of chronic neuromodulation has to be demonstrated. In this study, we present a comprehensive framework for thermogenetic neuromodulation in vivo using the thermosensitive human ion channel hTRPV1. By targeting hTRPV1 expression to excitatory neurons of the mouse brain and activating them within a non-harmful temperature range with a fiber-coupled infrared laser, we not only induced neuronal firing and stimulated locomotion in mice, but also demonstrated that thermogenetics can be employed for repeated neuromodulation without causing evident brain tissue injury. Our results lay the foundation for the use of thermogenetic neuromodulation in brain research and therapy of neuropathologies. [ABSTRACT FROM AUTHOR]

A comprehensive understanding of physio-pathological processes necessitates non-invasive intravital three-dimensional (3D) imaging over varying spatial and temporal scales. However, huge data throughput, optical heterogeneity, surface irregularity, and phototoxicity pose great challenges, leading to an inevitable trade-off between volume size, resolution, speed, sample health, and system complexity. Here, we introduce a compact real-time, ultra-large-scale, high-resolution 3D mesoscope (RUSH3D), achieving uniform resolutions of 2.6 × 2.6 × 6 μm3 across a volume of 8,000 × 6,000 × 400 μm3 at 20 Hz with low phototoxicity. Through the integration of multiple computational imaging techniques, RUSH3D facilitates a 13-fold improvement in data throughput and an orders-of-magnitude reduction in system size and cost. With these advantages, we observed premovement neural activity and cross-day visual representational drift across the mouse cortex, the formation and progression of multiple germinal centers in mouse inguinal lymph nodes, and heterogeneous immune responses following traumatic brain injury—all at single-cell resolution, opening up a horizon for intravital mesoscale study of large-scale intercellular interactions at the organ level. [Display omitted] • RUSH3D allows long-term, fast 3D imaging at centimeter-scale and single-cell resolution • Compact, high-fidelity mesoscale imaging is achieved in native intravital environments • Intercellular interactions across multiple mouse germinal centers are observed in vivo • Cortex-wide multisensory, locomotion neural activities and immune responses are explored in mice RUSH3D, with the integration of multiple computational imaging methods based on a scanning light-field framework, facilitates long-term, high-speed, centimeter-wide mesoscale 3D imaging at single-cell resolution in a compact system for broad practical applications in understanding large-scale intercellular dynamics at the mammalian organ level. We have demonstrated cortex-wide large-population neural recording with cross-day registration during multisensory input, the formation and progression of multiple germinal centers in mouse inguinal lymph nodes, and cortex-wide heterogeneous immune responses after traumatic brain injury. [ABSTRACT FROM AUTHOR]

In recent years, research has progressively increased the importance of considering sex differences in stress and fear memory studies. Many studies have traditionally focused on male subjects, potentially overlooking critical differences with females. Emerging evidence suggests that males and females can exhibit distinct behavioral and neurophysiological responses to stress and fear conditioning. These differences may be attributable to variations in hormone levels, brain structure, and neural circuitry, particularly in regions such as the prefrontal cortex (PFC). In the present study, we explored sex differences in prelimbic cortex (PL) calcium activity in animals submitted to immobilization stress (IMO), fear conditioning (FC), and fear extinction (FE). While no significant sex differences were found in behavioral responses, we did observe differences in several PL calcium activity parameters. To determine whether these results were related to behaviors beyond stress and fear memory, we conducted correlation studies between the movement of the animals and PL activity during IMO and freezing behavior during FC and FE. Our findings revealed a clear correlation between PL calcium activity with movement during stress exposure and freezing behavior, with no sex differences observed in these correlations. These results suggest a significant role for the PL in movement and locomotion, in addition to its involvement in fear-related processes. The inclusion of both female and male subjects is crucial for studies like this to fully understand the role of the PFC and other brain areas in stress and fear responses. Recognizing sex differences enhances our comprehension of brain function and can lead to more personalized and effective approaches in the study and treatment of stress and fear-related conditions. Plain English summary: In recent years, researchers have started paying more attention to the differences between males and females in how they handle stress and remember fearful events. Traditionally, many studies focused mainly on males, which might have missed important differences in females. New findings seem to suggest that males and females can respond differently to stress and fear due to differences in hormone levels, brain structure, and brain circuits, especially in the prefrontal cortex (PFC). In this study, we looked at how male and female animals' brains reacted to being restrained, experiencing a strong trauma, and then trying to learn a new fearful memory. While their behaviors didn't show significant differences between sexes, their brain activities did. We found that the prelimbic area of the brain shows calcium activity linked to the animals' movements during stress and their freezing behavior during fear-related tests. These results show that the PL is involved in both movement and fear responses. Including both male and female subjects in such studies is vital to fully understand how the prefrontal cortex and other brain areas work in stress and fear situations. Recognizing these differences helps improve our understanding of brain function and can lead to better, more personalized treatments for stress and fear-related conditions. Highlights: • Sex differences were found in the animals' calcium activity in the prelimbic cortex (PL) during IMO. Females showed higher frequency, but lower amplitude of calcium events in early IMO. • No significant sex or stage differences were found in high movement periods during IMO. However, a positive correlation between calcium activity in the PL, and movement was observed in both sexes. • Both sexes showed increased freezing during fear conditioning (FC). Sex differences were noted during the second extinction session (FE2), with males showing significant changes across tones. • A negative correlation between calcium activity in the PL and freezing behavior was identified during FC and both extinction phases (FE1 and FE2). • Significant sex differences in the percentage of excited neurons were observed during FC, but not in inhibited neurons. Differences were also noted in the percentage of excited neurons during FE1, but not during FE2. [ABSTRACT FROM AUTHOR]

Different populations of hypothalamic kisspeptin (KISS1) neurons located in the rostral periventricular area of the third ventricle (RP3V) and arcuate nucleus (ARC) are thought to generate the sex‐specific patterns of gonadotropin secretion. These neuronal populations integrate gonadal sex steroid feedback with internal and external cues relayed via the actions of neurotransmitters and neuropeptides. The excitatory amino acid neurotransmitter glutamate, the main excitatory neurotransmitter in the brain, plays a role in regulating gonadotropin secretion, at least partially through engaging KISS1 signaling. The expression and function of individual glutamate receptor subtypes in KISS1 neurons, however, are not well characterized. Here, we used GCaMP‐based calcium imaging and patch‐clamp electrophysiology to assess the impact of activating individual ionotropic (iGluR) and group I metabotropic (mGluR) glutamate receptors on KISS1 neuron activity in the mouse RP3V and ARC. Our results indicate that activation of all iGluR subtypes and of group I mGluRs, likely mGluR1, consistently drives activity in the majority of KISS1 neurons within the RP3V and ARC of males and females. Our results also revealed, somewhat unexpectedly, sex‐ and region‐specific differences. Indeed, activating (S)‐α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA) type iGluRs evoked larger responses in female ARCKISS1 neurons than in their male counterparts whereas activating group I mGluRs induced larger responses in RP3VKISS1 neurons than in ARCKISS1 neurons in females. Together, our findings suggest that glutamatergic neurotransmission in KISS1 neurons, and its impact on the activity of these cells, might be sex‐ and region‐dependent in mice. [ABSTRACT FROM AUTHOR]

Improved genetically encoded calcium indicators (GECIs) are essential for capturing intracellular dynamics of both muscle and neurons. A novel set of GECIs with ultrafast kinetics and high sensitivity was recently reported by Zhang et al. (2023). While these indicators, called jGCaMP8, were demonstrated to work in Drosophila and mice, data for Caenorhabditis elegans were not reported. Here, we present an optimized construct for C. elegans and use this to generate several strains expressing GCaMP8f (fast variant of the indicator). Utilizing the myo-2 promoter, we compare pharyngeal muscle activity measured with GCaMP7f and GCaMP8f and find that GCaMP8f is brighter upon binding to calcium, shows faster kinetics, and is not disruptive to the intrinsic contraction dynamics of the pharynx. Additionally, we validate its application for detecting neuronal activity in touch receptor neurons which reveals robust calcium transients even at small stimulus amplitudes. As such, we establish GCaMP8f as a potent tool for C. elegans research which is capable of extracting fast calcium dynamics at very low magnifications across multiple cell types. [ABSTRACT FROM AUTHOR]

Recent literature supports a prominent role for astrocytes in regulation of drug-seeking behaviors. The dorsal striatum, specifically, is known to play a role in reward processing with neuronal activity that can be influenced by astrocyte Ca2+. However, the manner in which Ca2+ in dorsal striatum astrocytes impacts neuronal signaling after exposure to self-administered cocaine remains unclear. We addressed this question following over-expression of the Ca2+ extrusion pump, hPMCA2w/b, in dorsal striatum astrocytes and the Ca2+ indicator, GCaMP6f, in dorsal striatum neurons of rats that were trained to self-administer cocaine. Following extinction of cocaine-seeking behavior, the rats over-expressing hMPCA2w/b showed a significant increase in cue-induced reinstatement of cocaine seeking. Suppression of astrocyte Ca2+ increased the amplitude of neuronal Ca2+ transients in brain slices, but only after cocaine self-administration. This was accompanied by decreased duration of neuronal Ca2+ events in the cocaine group and no changes in Ca2+ event frequency. Acute administration of cocaine to brain slices decreased amplitude of neuronal Ca2+ in both the control and cocaine self-administration groups regardless of hPMCA2w/b expression. These results indicated that astrocyte Ca2+ control over neuronal Ca2+ transients was enhanced by cocaine self-administration experience, although sensitivity to acutely applied cocaine remained comparable across all groups. To explore this further, we found that neither the hMPCA2w/b expression nor the cocaine self-administration experience altered regulation of neuronal Ca2+ events by NPS-2143, a Ca2+ sensing receptor (CaSR) antagonist, suggesting that plasticity of neuronal signaling after hPMCA2w/b over-expression was unlikely to result from elevated extracellular Ca2+. We conclude that astrocyte Ca2+ in the dorsal striatum impacts neurons via cell-intrinsic mechanisms (e.g., gliotransmission, metabolic coupling, etc.) and impacts long-term neuronal plasticity after cocaine self-administration differently from neuronal response to acute cocaine. Overall, astrocyte Ca2+ influences neuronal output in the dorsal striatum to promote resistance to cue-induced reinstatement of cocaine seeking. [ABSTRACT FROM AUTHOR]

Fiber photometry is an optical method to monitor fluorescent signals using a fiber optic cannula. Over the past two decades, together with the development of various genetically encoded biosensors, it has been applied to investigate various types of activity in the central nervous system. This includes not only type-specific neuronal population activity, but also non-neuronal activity and neurotransmitter/neuropeptide signals in awake, freely behaving animals. In this perspective, we summarize the recent development of this technique. After describing common technical pitfalls, we discuss future directions of this powerful approach for investigating brain function and dysfunction. [ABSTRACT FROM AUTHOR]

Kelch‐like family member 17 (KLHL17), an actin‐associated adaptor protein, is linked to neurological disorders, including infantile spasms and autism spectrum disorders. The key morphological feature of Klhl17‐deficient neurons is impaired dendritic spine enlargement, resulting in the amplitude of calcium events being increased. Our previous studies have indicated an involvement of F‐actin and the spine apparatus in KLHL17‐mediated dendritic spine enlargement. Here, we show that KLHL17 further employs different mechanisms to control the expression of two types of glutamate receptors, that is, α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid receptor (AMPAR) and kainate receptors (KARs), to regulate dendritic spine enlargement and calcium influx. We deployed proteomics to reveal that KLHL17 interacts with N‐ethylmaleimide‐sensitive fusion protein (NSF) in neurons, with this interaction of KLHL17 and NSF enhancing NSF protein levels. Consistent with the function of NSF in regulating the surface expression of AMPAR, Klhl17 deficiency limits the surface expression of AMPAR, but not its total protein levels. The NSF pathway also contributes to synaptic F‐actin distribution and the dendritic spine enlargement mediated by KLHL17. KLHL17 is known to act as an adaptor mediating degradation of the KAR subunit GluK2 by the CUL3 ubiquitin ligase complex, and Klhl17 deficiency impairs activity‐dependent degradation of GluK2. Herein, we further demonstrate that GluK2 is critical to the increased amplitude of calcium influx in Klhl17‐deficient neurons. Moreover, GluK2 is also involved in KLHL17‐regulated dendritic spine enlargement. Thus, our study reveals that KLHL17 controls AMPAR and KAR expression via at least two mechanisms, consequently regulating dendritic spine enlargement. The regulatory effects of KLHL17 on these two glutamate receptors likely contribute to neuronal features in patients suffering from certain neurological disorders. [ABSTRACT FROM AUTHOR]

Neurotransmission is critical for brain function, allowing neurons to communicate through neurotransmitters and neuropeptides. RVD-hemopressin (RVD-Hp), a novel peptide identified in noradrenergic neurons, modulates cannabinoid receptors CB1 and CB2. Unlike hemopressin (Hp), which induces anxiogenic behaviors via transient receptor potential vanilloid 1 (TRPV1) activation, RVD-Hp counteracts these effects, suggesting that it may block TRPV1. This study investigates RVD-Hp's role as a TRPV1 channel blocker using HEK293 cells expressing TRPV1-GFP. Calcium imaging and patch-clamp recordings demonstrated that RVD-Hp reduces TRPV1-mediated calcium influx and TRPV1 ion currents. Molecular docking and dynamics simulations indicated that RVD-Hp interacts with TRPV1's selectivity filter, forming stable hydrogen bonds and van der Waals contacts, thus preventing ion permeation. These findings highlight RVD-Hp's potential as a therapeutic agent for conditions involving TRPV1 activation, such as pain and anxiety. [ABSTRACT FROM AUTHOR]

Although olfaction is well known to guide animal behavior, the neural circuits underlying the motor responses elicited by olfactory inputs are poorly understood. In the sea lamprey, anatomical evidence shows that olfactory inputs project to the posterior tuberculum (PT), a structure containing dopaminergic (DA) neurons homologous to the mammalian ventral tegmental area and the substantia nigra pars compacta. Olfactory inputs travel directly from the medial olfactory bulb (medOB) or indirectly through the main olfactory bulb and the lateral pallium (LPal). Here, we characterized the transmission of olfactory inputs to the PT in the sea lamprey, Petromyzon marinus. Abundant projections from the medOB were observed close to DA neurons of the PT. Moreover, electrophysiological experiments revealed that PT neurons are activated by both the medOB and LPal, and calcium imaging indicated that the olfactory signal is then relayed to the mesencephalic locomotor region to initiate locomotion. In semi-intact preparations, stimulation of the medOB and LPal induced locomotion that was tightly associated with neural activity in the PT. Moreover, PT neurons were active throughout spontaneously occurring locomotor bouts. Altogether, our observations suggest that the medOB and LPal convey olfactory inputs to DA neurons of the PT, which in turn activate the brainstem motor command system to elicit locomotion. [ABSTRACT FROM AUTHOR]

Gut motility undergoes a switch from myogenic to neurogenic control in late embryonic development. Here, we report on the electrical events that underlie this transition in the enteric nervous system, using the GCaMP6f reporter in neural crest cell derivatives. We found that spontaneous calcium activity is tetrodotoxin (TTX) resistant at stage E11.5, but not at E18.5. Motility at E18.5 was characterized by periodic, alternating high‐ and low‐frequency contractions of the circular smooth muscle; this frequency modulation was inhibited by TTX. Calcium imaging at the neurogenic‐motility stages E18.5–P3 showed that CaV1.2‐positive neurons exhibited spontaneous calcium activity, which was inhibited by nicardipine and 2‐aminoethoxydiphenyl borate (2‐APB). Our protocol locally prevented muscle tone relaxation, arguing for a direct effect of nicardipine on enteric neurons, rather than indirectly by its relaxing effect on muscle. We demonstrated that the ENS was mechanosensitive from early stages on (E14.5) and that this behaviour was TTX and 2‐APB resistant. We extended our results on L‐type channel‐dependent spontaneous activity and TTX‐resistant mechanosensitivity to the adult colon. Our results shed light on the critical transition from myogenic to neurogenic motility in the developing gut, as well as on the intriguing pathways mediating electro‐mechanical sensitivity in the enteric nervous system. Highlights: What is the central question of this study?What are the first neural electric events underlying the transition from myogenic to neurogenic motility in the developing gut, what channels do they depend on, and does the enteric nervous system already exhibit mechanosensitivity?What is the main finding and its importance?ENS calcium activity is sensitive to tetrodotoxin at stage E18.5 but not E11.5. Spontaneous electric activity at fetal and adult stages is crucially dependent on L‐type calcium channels and IP3R receptors, and the enteric nervous system exhibits a tetrodotoxin‐resistant mechanosensitive response. Abstract figure legend Tetrodotoxin‐resistant Ca2+ rise induced by mechanical stimulation in the E18.5 mouse duodenum. [ABSTRACT FROM AUTHOR]

Abstract In recent years, research has progressively increased the importance of considering sex differences in stress and fear memory studies. Many studies have traditionally focused on male subjects, potentially overlooking critical differences with females. Emerging evidence suggests that males and females can exhibit distinct behavioral and neurophysiological responses to stress and fear conditioning. These differences may be attributable to variations in hormone levels, brain structure, and neural circuitry, particularly in regions such as the prefrontal cortex (PFC). In the present study, we explored sex differences in prelimbic cortex (PL) calcium activity in animals submitted to immobilization stress (IMO), fear conditioning (FC), and fear extinction (FE). While no significant sex differences were found in behavioral responses, we did observe differences in several PL calcium activity parameters. To determine whether these results were related to behaviors beyond stress and fear memory, we conducted correlation studies between the movement of the animals and PL activity during IMO and freezing behavior during FC and FE. Our findings revealed a clear correlation between PL calcium activity with movement during stress exposure and freezing behavior, with no sex differences observed in these correlations. These results suggest a significant role for the PL in movement and locomotion, in addition to its involvement in fear-related processes. The inclusion of both female and male subjects is crucial for studies like this to fully understand the role of the PFC and other brain areas in stress and fear responses. Recognizing sex differences enhances our comprehension of brain function and can lead to more personalized and effective approaches in the study and treatment of stress and fear-related conditions.

Abstract Gut motility undergoes a switch from myogenic to neurogenic control in late embryonic development. Here, we report on the electrical events that underlie this transition in the enteric nervous system, using the GCaMP6f reporter in neural crest cell derivatives. We found that spontaneous calcium activity is tetrodotoxin (TTX) resistant at stage E11.5, but not at E18.5. Motility at E18.5 was characterized by periodic, alternating high‐ and low‐frequency contractions of the circular smooth muscle; this frequency modulation was inhibited by TTX. Calcium imaging at the neurogenic‐motility stages E18.5–P3 showed that CaV1.2‐positive neurons exhibited spontaneous calcium activity, which was inhibited by nicardipine and 2‐aminoethoxydiphenyl borate (2‐APB). Our protocol locally prevented muscle tone relaxation, arguing for a direct effect of nicardipine on enteric neurons, rather than indirectly by its relaxing effect on muscle. We demonstrated that the ENS was mechanosensitive from early stages on (E14.5) and that this behaviour was TTX and 2‐APB resistant. We extended our results on L‐type channel‐dependent spontaneous activity and TTX‐resistant mechanosensitivity to the adult colon. Our results shed light on the critical transition from myogenic to neurogenic motility in the developing gut, as well as on the intriguing pathways mediating electro‐mechanical sensitivity in the enteric nervous system. Highlights What is the central question of this study? What are the first neural electric events underlying the transition from myogenic to neurogenic motility in the developing gut, what channels do they depend on, and does the enteric nervous system already exhibit mechanosensitivity? What is the main finding and its importance? ENS calcium activity is sensitive to tetrodotoxin at stage E18.5 but not E11.5. Spontaneous electric activity at fetal and adult stages is crucially dependent on L‐type calcium channels and IP3R receptors, and the enteric nervous system exhibits a tetrodotoxin‐resistant mechanosensitive response. Abstract figure legend Tetrodotoxin‐resistant Ca2+ rise induced by mechanical stimulation in the E18.5 mouse duodenum.

Attention deficit is one of the most prominent and disabling symptoms in Fragile X syndrome (FXS). Hypersensitivity to sensory stimuli contributes to attention difficulties by overwhelming and/or distracting affected individuals, which disrupts activities of daily living at home and learning at school. We find that auditory or visual distractors selectively impair visual discrimination performance in humans and mice with FXS but not in typically developing controls. In both species, males and females were examined. Vasoactive intestinal polypeptide (VIP) neurons were significantly modulated by incorrect responses in the poststimulus period during early distractor trials in WT mice, consistent with their known role as error signals. Strikingly, however, VIP cells from Fmr1 -/- mice showed little modulation in error trials, and this correlated with their poor performance on the distractor task. Thus, VIP interneurons and their reduced modulatory influence on pyramidal cells could be a potential therapeutic target for attentional difficulties in FXS.SIGNIFICANCE STATEMENT Sensory hypersensitivity, impulsivity, and persistent inattention are among the most consistent clinical features of FXS, all of which impede daily functioning and create barriers to learning. However, the neural mechanisms underlying sensory over-reactivity remain elusive. To overcome a significant challenge in translational FXS research we demonstrate a compelling alignment of sensory over-reactivity in both humans with FXS and Fmr1 -/- mice (the principal animal model of FXS) using a novel analogous distractor task. Two-photon microscopy in mice revealed that lack of modulation by VIP cells contributes to susceptibility to distractors. Implementing research efforts we describe here can help identify dysfunctional neural mechanisms associated not only with sensory issues but broader impairments, including those in learning and cognition.

Changes in the function of inhibitory interneurons (INs) during cortical development could contribute to the pathophysiology of neurodevelopmental disorders. Using all-optical in vivo approaches, we find that parvalbumin (PV) INs and their immature precursors are hypoactive and transiently decoupled from excitatory neurons in postnatal mouse somatosensory cortex (S1) of Fmr1 KO mice, a model of fragile X syndrome (FXS). This leads to a loss of parvalbumin INs (PV-INs) in both mice and humans with FXS. Increasing the activity of future PV-INs in neonatal Fmr1 KO mice restores PV-IN density and ameliorates transcriptional dysregulation in S1, but not circuit dysfunction. Critically, administering an allosteric modulator of Kv3.1 channels after the S1 critical period does rescue circuit dynamics and tactile defensiveness. Symptoms in FXS and related disorders could be mitigated by targeting PV-INs.

During learning of a sensory discrimination task, the cortical and subcortical regions display complex spatiotemporal dynamics. During learning, both the amygdala and cortex link stimulus information to its appropriate association, for example, a reward. In addition, both structures are also related to nonsensory parameters such as body movements and licking during the reward period. However, the emergence of the cortico-amygdala relationships during learning is largely unknown. To study this, we combined wide-field cortical imaging with fiber photometry to simultaneously record cortico-amygdala population dynamics as male mice learn a whisker-dependent go/no-go task. We were able to simultaneously record neuronal populations from the posterior cortex and either the basolateral amygdala (BLA) or central/medial amygdala (CEM). Prior to learning, the somatosensory and associative cortex responded during sensation, while amygdala areas did not show significant responses. As mice became experts, amygdala responses emerged early during the sensation period, increasing in the CEM, while decreasing in the BLA. Interestingly, amygdala and cortical responses were associated with task-related body movement, displaying significant responses ∼200 ms before movement initiation which led to licking for the reward. A correlation analysis between the cortex and amygdala revealed negative and positive correlation with the BLA and CEM, respectively, only in the expert case. These results imply that learning induces an involvement of the cortex and amygdala which may aid to link sensory stimuli with appropriate associations. [ABSTRACT FROM AUTHOR]

The ability to perceive and adapt to environmental changes is crucial for the survival of all organisms. Neural functional imaging, particularly in model organisms, such as Caenorhabditis elegans, provides valuable insights into how animals sense and process external cues through their nervous systems. Because of its fully mapped neural anatomy, transparent body, and genetic tractability, C. elegans serves as an ideal model for these studies. This review focuses on advanced methods for neural functional imaging in C. elegans, highlighting calcium imaging techniques, lab-on-a-chip technologies, and their applications in the study of various sensory modalities, including chemosensation, mechanosensation, thermosensation, photosensation, and magnetosensation. We discuss the benefits of these methods in terms of precision, reproducibility, and ability to study dynamic neural processes in real time, ultimately advancing our understanding of the fundamental principles of neural activity and connectivity. [ABSTRACT FROM AUTHOR]

Synaptic plasticity is a key cellular model for learning, memory and chronic pain. Most previous studies were carried out in rats and mice, and less is known about synaptic plasticity in non-human primates. In the present study, we used integrative experimental approaches to study long-term potentiation (LTP) in the anterior cingulate cortex (ACC) of adult tree shrews. We found that glutamate is the major excitatory transmitter and α-amino-3-hydroxy-5-methyl-4-isoxazole-propionicacid (AMPA) receptors mediate postsynaptic responses. LTP in tree shrews was greater than that in adult mice and lasted for at least 5 h. N-methyl-d-aspartic acid (NMDA) receptors, Ca2+ influx and adenylyl cyclase 1 (AC1) contributed to tree shrew LTP. Our results suggest that LTP is a major form of synaptic plasticity in the ACC of primate-like animals. This article is part of a discussion meeting issue 'Long-term potentiation: 50 years on'. [ABSTRACT FROM AUTHOR]

Linking sensory input and its consequences is a fundamental brain operation. During behavior, the neural activity of neocortical and limbic systems often reflects dynamic combinations of sensory and task-dependent variables, and these "mixed representations" are suggested to be important for perception, learning, and plasticity. However, the extent to which such integrative computations might occur outside of the forebrain is less clear. Here, we conduct cellular-resolution two-photon Ca2+ imaging in the superficial "shell" layers of the inferior colliculus (IC), as head-fixed mice of either sex perform a reward-based psychometric auditory task. We find that the activity of individual shell IC neurons jointly reflects auditory cues, mice's actions, and behavioral trial outcomes, such that trajectories of neural population activity diverge depending on mice's behavioral choice. Consequently, simple classifier models trained on shell IC neuron activity can predict trial-by-trial outcomes, even when training data are restricted to neural activity occurring prior to mice's instrumental actions. Thus, in behaving mice, auditory midbrain neurons transmit a population code that reflects a joint representation of sound, actions, and task-dependent variables. [ABSTRACT FROM AUTHOR]

Introduction: Neuroinflammation is a hallmark of multiple neurodegenerative diseases, shared by all pathological processes which primarily impact on neurons, including Central Nervous System (CNS) injuries. In reactive CNS, activated glia releases extracellular vesicles (EVs), nanosized membranous particles known to play a key role in intercellular communication. EVs mediate neuroinflammatory responses and might exacerbate tissue deterioration, ultimately influencing neurodegenerative disease progression. Methods: We treated spinal cord organotypic slices with LPS, a ligand extensively used to induce sEVs release, to mimic mild inflammatory conditions. We combine atomic force microscopy (AFM), nanoparticle tracking (NTA) and western blot (WB) analysis to validate the isolation and characterisation of sEVs. We further use immunofluorescence and confocal microscopy with live calcium imaging by GCaMP6f reporter to compare glial reactivity to treatments with sEVs when isolated from resting and LPS treated organ slices. Results: In our study, we focus on CNS released small EVs (sEVs) and their impact on the biology of inflammatory environment. We address sEVs local signalling within the CNS tissue, in particular their involvement in inflammation spreading mechanism(s). sEVs are harvested from mouse organotypic spinal cord cultures, an in vitro model which features 3D complexity and retains spinal cord resident cells. By confocal microscopy and live calcium imaging we monitor glial responses in naïve spinal slices when exposed to sEVs isolated from resting and LPS treated organ slices. Discussion: We show that sEVs, only when released during LPS neuroinflammation, recruit naïve astrocytes in the neuroinflammation cycle and we propose that such recruitment be mediated by EVs hemichannel (HC) permeability. [ABSTRACT FROM AUTHOR]

Preclinical assessments of pain have often relied upon behavioral measurements and anesthetized neurophysiological recordings. Current technologies enabling large-scale neural recordings, however, have the potential to unveil quantifiable pain signals in conscious animals for preclinical studies. Although pain processing is distributed across many brain regions, the anterior cingulate cortex (ACC) is of particular interest in isolating these signals given its suggested role in the affective (“unpleasant”) component of pain. Here, we explored the utility of the ACC toward preclinical pain research using head-mounted miniaturized microscopes to record calcium transients in freely moving male mice expressing genetically encoded calcium indicator 6f (GCaMP6f) under the Thy1 promoter. We verified the expression of GCaMP6f in excitatory neurons and found no intrinsic behavioral differences in this model. Using a multimodal stimulation paradigm across naive, pain, and analgesic conditions, we found that while ACC population activity roughly scaled with stimulus intensity, single-cell representations were highly flexible. We found only low-magnitude increases in population activity after complete Freund’s adjuvant (CFA) and insufficient evidence for the existence of a robust nociceptive ensemble in the ACC. However, we found a temporal sharpening of response durations and generalized increases in pairwise neural correlations in the presence of the mechanistically distinct analgesics gabapentin or ibuprofen after (but not before) CFA-induced inflammatory pain. This increase was not explainable by changes in locomotion alone. Taken together, these results highlight challenges in isolating distinct pain signals among flexible representations in the ACC but suggest a neurophysiological hallmark of analgesia after pain that generalizes to at least two analgesics. [ABSTRACT FROM AUTHOR]

Optical and electrophysiological methods are widely used in neuroscience to study brain activity. Each technique has its own strengths and weaknesses that complement each other to provide a more comprehensive understanding of neuronal activity. To this end, we developed an implantable CMOS image sensor device with an integrated carbon electrode for the simultaneous measurement of fluorescence and extracellular signals. The device is composed of a 450 × 1660 µm CMOS chip and micro-LED secured to a flexible printed circuit substrate. The chip has an imaging surface measuring 900 × 300 µm2, comprising 120 × 90 pixels recording at 10 frames per second. An absorption filter film was placed on top of the imaging surface to block excitation light. The device was coated with parylene-C for waterproofing and, finally, the coating film was readily turned into carbon electrodes by laser carbonization. We characterized the lasercarbonized electrodes by cyclic voltammetry, electrochemical impedance spectroscopy, and X-ray photoelectron spectroscopy to confirm carbonization. We then demonstrated simultaneous in vivo recording of fluorescence and extracellular signals in hippocampus CA1 using the device. [ABSTRACT FROM AUTHOR]

Genetically encoded optical indicators and actuators of neural activity allow for all-optical investigations of signaling in the nervous system. But commonly used indicators, actuators, and expression strategies are poorly suited for systematic measurements of signal propagation at brain scale and cellular resolution. Large-scale measurements of the brain require indicators and actuators with compatible excitation spectra to avoid optical crosstalk. They must be highly expressed in every neuron but at the same time avoid lethality and permit the animal to reach adulthood. Their expression must also be compatible with additional fluorescent labels to locate and identify neurons, such as those in the NeuroPAL cell identification system. We present TWISP, a transgenic worm for interrogating signal propagation, that addresses these needs and enables optical measurements of evoked calcium activity at brain scale and cellular resolution in the nervous system of the nematode Caenorhabditis elegans. In every neuron we express a nonconventional optical actuator, the gustatory receptor homolog GUR-3 + PRDX-2 , under the control of a drug-inducible system QF + hGR, and a calcium indicator GCAMP6s, in a background with additional fluorophores from the NeuroPAL cell ID system. We show that this combination, but not others tested, avoids optical crosstalk, creates strong expression in the adult, and generates stable transgenic lines for systematic measurements of signal propagation in the worm brain. [ABSTRACT FROM AUTHOR]

The miniature single-photon fluorescent microscope (miniscope) enables the visualization of calcium activity in vivo in freely moving laboratory animals, providing the capability to track cellular activity during the investigation of memory formation, learning, sleep, and social interactions. However, the use of calcium sensors for in vivo imaging is limited by their relatively slow (millisecond-scale) kinetics, which complicates the recording of high-frequency spike activity. The integration of methods from single-photon miniature fluorescent microscopy with electrophysiological recording, which possesses microsecond resolution, represents a potential solution to this issue. Such a combination of techniques allows for the simultaneous recording of optical and electrophysiological activity in a single animal in vivo. In this study, a flexible polyimide microelectrode was developed and integrated with the gradient lens of the miniscope. The in vivo tests conducted in this research confirmed that the microelectrode combined with the gradient lens facilitates simultaneous single-photon calcium imaging and local field potential recording in the hippocampus of an adult mouse. [ABSTRACT FROM AUTHOR]

Objective: The increased amplitude of ictal activity is a common feature of epileptic seizures, but the determinants of this amplitude have not been identified. Clinically, ictal amplitudes are measured electrographically (using, e.g., electroencephalography, electrocorticography, and depth electrodes), but these methods do not enable the assessment of the activity of individual neurons. Population signal may increase from three potential sources: (1) increased synchrony (i.e., more coactive neurons); (2) altered active state, from bursts of action potentials and/or paroxysmal depolarizing shifts in membrane potential; and (3) altered subthreshold state, which includes all lower levels of activity. Here, we quantify the fraction of ictal signal from each source. Methods: To identify the cellular determinants of the ictal signal, we measured single cell and population electrical activity and neuronal calcium levels via optical imaging of the genetically encoded calcium indicator (GECI) GCaMP. Spontaneous seizure activity was assessed with microendoscopy in an APP/PS1 mouse with focal cortical injury and via widefield imaging in the organotypic hippocampal slice cultures (OHSCs) model of posttraumatic epilepsy. Single cell calcium signals were linked to a range of electrical activities by performing simultaneous GECI‐based calcium imaging and whole‐cell patch‐clamp recordings in spontaneously seizing OHSCs. Neuronal resolution calcium imaging of spontaneous seizures was then used to quantify the cellular contributions to population‐level ictal signal. Results: The seizure onset signal was primarily driven by increased subthreshold activity, consistent with either barrages of excitatory postsynaptic potentials or sustained membrane depolarization. Unsurprisingly, more neurons entered the active state as seizure activity progressed. However, the increasing fraction of active cells was primarily driven by synchronous reactivation and not from continued recruitment of new populations of neurons into the seizure. Significance: This work provides a critical link between single neuron activity and population measures of seizure activity. [ABSTRACT FROM AUTHOR]

Our understanding of human brain function can be greatly aided by studying analogous brain structures in other organisms. One brain structure with neurochemical and anatomical homology throughout vertebrate species is the locus coeruleus (LC), a small collection of norepinephrine (NE)-containing neurons in the brainstem that project throughout the central nervous system. The LC is involved in nearly every aspect of brain function, including arousal and learning, which has been extensively examined in rats and nonhuman primates using single-unit recordings. Recent work has expanded into putative LC single-unit electrophysiological recordings in a nonmodel species, the zebra finch. Given the importance of correctly identifying analogous structures as research efforts expand to other vertebrates, we suggest adoption of consensus anatomical and electrophysiological guidelines for identifying LC neurons across species when evaluating brainstem single-unit spiking or calcium imaging. Such consensus criteria will allow for confident cross-species understanding of the roles of the LC in brain function and behavior. [ABSTRACT FROM AUTHOR]

CADENCE is an open Python 3-written neuroinformatics tool with Qt6 graphic user interface for supervised calcium events detection. In neuronal ensembles recording during calcium imaging experiments, the output of instruments such as Celena X, Zeiss LSM 5 Live confocal microscope and Miniscope is a movie showing flashing cells somata. There are few pipelines to convert video to relative fluorescence ΔF/F, from simplest ImageJ plugins to sophisticated tools like MiniAn (Dong et al. in Elife 11, https://doi.org/10.7554/eLife.70661, 2022). Minian, an open-source miniscope analysis pipeline. Elife, 11.). While in some areas of study relative fluorescence ΔF/F may be the desired result in itself, researchers of neuronal ensembles are typically interested in a more detailed analysis of calcium events as indirect proxy of neuronal electrical activity. For such analyses, researchers need a tool to infer calcium events from the continuous ΔF/F curve in order to create a raster representation of calcium events for later use in analysis software, such as Elephant (Denker, M., Yegenoglu, A., & Grün, S. (2018). Collaborative HPC-enabled workflows on the HBP Collaboratory using the Elephant framework. Neuroinformatics, 19.). Here we present such an open tool with supervised calcium events detection. [ABSTRACT FROM AUTHOR]

Kilohertz high-frequency spinal cord stimulation (kHF-SCS) is a rapidly advancing neuromodulatory technique in the clinical management of chronic pain. However, the precise cellular mechanisms underlying kHF-SCS-induced paresthesia-free pain relief, as well as the neural responses within spinal pain circuits, remain largely unexplored. In this study, using a novel preparation, we investigated the impact of varying kilohertz frequency SCS on dorsal horn neuron activation. Employing calcium imaging on isolated spinal cord slices, we found that extracellular electric fields at kilohertz frequencies (1, 3, 5, 8, and 10 kHz) induce distinct patterns of activation in dorsal horn neurons. Notably, as the frequency of extracellular electric fields increased, there was a clear and significant monotonic escalation in neuronal activity. This phenomenon was observed not only in superficial dorsal horn neurons, but also in those located deeper within the dorsal horn. Our study demonstrates the unique patterns of dorsal horn neuron activation in response to varying kilohertz frequencies of extracellular electric fields, and we contribute to a deeper understanding of how kHF-SCS induces paresthesia-free pain relief. Furthermore, our study highlights the potential for kHF-SCS to modulate sensory information processing within spinal pain circuits. These insights pave the way for future research aimed at optimizing kHF-SCS parameters and refining its therapeutic applications in the clinical management of chronic pain. [ABSTRACT FROM AUTHOR]

Cortical responses to visual stimuli are believed to rely on the geniculo-striate pathway. However, recent work has challenged this notion by showing that responses in the postrhinal cortex (POR), a visual cortical area, instead depend on the tecto-thalamic pathway, which conveys visual information to the cortex via the superior colliculus (SC). Does PORs SC-dependence point to a wider system of tecto-thalamic cortical visual areas? What information might this system extract from the visual world? We discovered multiple mouse cortical areas whose visual responses rely on SC, with the most lateral showing the strongest SC-dependence. This system is driven by a genetically defined cell type that connects the SC to the pulvinar thalamic nucleus. Finally, we show that SC-dependent cortices distinguish self-generated from externally generated visual motion. Hence, lateral visual areas comprise a system that relies on the tecto-thalamic pathway and contributes to processing visual motion as animals move through the environment.

Aversive stimuli can cause hippocampal place cells to remap their firing fields, but it is not known whether remapping plays a role in storing memories of aversive experiences. Here, we addressed this question by performing in vivo calcium imaging of CA1 place cells in freely behaving rats (n = 14). Rats were first trained to prefer a short path over a long path for obtaining food reward, then trained to avoid the short path by delivering a mild footshock. Remapping was assessed by comparing place cell population vector similarity before acquisition versus after extinction of avoidance. Some rats received shock after systemic injections of the amnestic drug scopolamine at a dose (1 mg/kg) that impaired avoidance learning but spared spatial tuning and shock-evoked responses of CA1 neurons. Place cells remapped significantly more following remembered than forgotten shocks (drug-free versus scopolamine conditions); shock-induced remapping did not cause place fields to migrate toward or away from the shocked location and was similarly prevalent in cells that were responsive versus non-responsive to shocks. When rats were exposed to a neutral barrier rather than aversive shock, place cells remapped significantly less in response to the barrier. We conclude that place cell remapping occurs in response to events that are remembered rather than merely perceived and forgotten, suggesting that reorganization of hippocampal population codes may play a role in storing memories for aversive events.

Memory deficits are a debilitating symptom of epilepsy, but little is known about mechanisms underlying cognitive deficits. Here, we describe a Na+ channel-dependent mechanism underlying altered hippocampal dendritic integration, degraded place coding and deficits in spatial memory. Two-photon glutamate uncaging experiments revealed a marked increase in the fraction of hippocampal first-order CA1 pyramidal cell dendrites capable of generating dendritic spikes in the kainate model of chronic epilepsy. Moreover, in epileptic mice dendritic spikes were generated with lower input synchrony, and with a lower threshold. The Nav1.3/1.1 selective Na+ channel blocker ICA-121431 reversed dendritic hyperexcitability in epileptic mice, while the Nav1.2/1.6 preferring anticonvulsant S-Lic did not. We used in vivo two-photon imaging to determine if aberrant dendritic excitability is associated with altered place-related firing of CA1 neurons. We show that ICA-121431 improves degraded hippocampal spatial representations in epileptic mice. Finally, behavioural experiments show that reversing aberrant dendritic excitability with ICA-121431 reverses hippocampal memory deficits. Thus, a dendritic channelopathy may underlie cognitive deficits in epilepsy and targeting it pharmacologically may constitute a new avenue to enhance cognition.

Recent advances in calcium imaging, including the development of fast and sensitive genetically encoded indicators, high-resolution camera chips for wide-field imaging, and resonant scanning mirrors in laser scanning microscopy, have notably improved the temporal and spatial resolution of functional imaging analysis. Nonetheless, the variability of imaging approaches and brain structures challenges the development of versatile and reliable segmentation methods. Standard techniques, such as manual selection of regions of interest or machine learning solutions, often fall short due to either user bias, non-transferability among systems, or computational demand. To overcome these issues, we developed CalciSeg, a data-driven and reproducible approach for unsupervised functional calcium imaging data segmentation. CalciSeg addresses the challenges associated with brain structure variability and user bias by offering a computationally efficient solution for automatic image segmentation based on two parameters: regions’ size limits and number of refinement iterations. We evaluated CalciSeg efficacy on datasets of varied complexity, different insect species (locusts, bees, and cockroaches), and imaging systems (wide-field, confocal, and multiphoton), showing the robustness and generality of our approach. Finally, the user-friendly nature and open-source availability of CalciSeg facilitate the integration of this algorithm into existing analysis pipelines.

The ventrolateral preoptic area (VLPO) contains GABAergic sleep-active neurons. However, the extent to which these neurons are involved in expressing spontaneous sleep and homeostatic sleep regulatory demands is not fully understood. We used calcium (Ca2+) imaging to characterize the activity dynamics of VLPO neurons, especially those expressing the vesicular GABA transporter (VGAT) across spontaneous sleep-waking and in response to homeostatic sleep demands. The VLPOs of wild-type and VGAT-Cre mice were transfected with GCaMP6, and the Ca2+ fluorescence of unidentified (UNID) and VGAT cells was recorded during spontaneous sleep-waking and 3 h of sleep deprivation (SD) followed by 1 h of recovery sleep. Although both VGAT and UNID neurons exhibited heterogeneous Ca2+ fluorescence across sleep-waking, the majority of VLPO neurons displayed increased activity during nonREM/REM (VGAT, 120/303; UNID, 39/106) and REM sleep (VGAT, 32/303; UNID, 19/106). Compared to the baseline waking, VLPO sleep-active neurons (n = 91) exhibited higher activity with increasing SD that remained elevated during the recovery period. These neurons also exhibited increased Ca2+ fluorescence during nonREM sleep, marked by increased slow-wave activity and REM sleep during recovery after SD. These findings support the notion that VLPO sleep-active neurons, including GABAergic neurons, are components of neuronal circuitry that mediate spontaneous sleep and homeostatic responses to sustained wakefulness.

Visual processing in the retina depends on the collective activity of large ensembles of neurons organized in different layers. Current techniques for measuring activity of layer-specific neural ensembles rely on expensive pulsed infrared lasers to drive 2-photon activation of calcium-dependent fluorescent reporters. We present a 1-photon light-sheet imaging system that can measure the activity in hundreds of neurons in the ex vivo retina over a large field of view while presenting visual stimuli. This allows for a reliable functional classification of different retinal cell types. We also demonstrate that the system has sufficient resolution to image calcium entry at individual synaptic release sites across the axon terminals of dozens of simultaneously imaged bipolar cells. The simple design, large field of view, and fast image acquisition make this a powerful system for high-throughput and high-resolution measurements of retinal processing at a fraction of the cost of alternative approaches.

Some γ-glutamyl peptides including glutathione (γ-Glu-Cys-Gly) and γ-glutamyl-valyl-glycine (γ-Glu-Val-Gly= γ-EVG) are reported to increase the intensity of basic tastes, such as salty, sweet, and umami, although they have no taste themselves at tested concentrations. The mechanism of action of γ-glutamyl peptides is not clearly understood, but the calcium sensing receptor (CaSR) may be involved. Glutathione and γ-EVG enhance the pungency of some spices, and the present study investigated the effects of γ-EVG on the responses of trigeminal ganglion (TG) cells to thermosensitiveTRP channel agonists. Single-cell RT-PCR revealed that most CaSR-expressing cells co-expressed TRPV1 (sensitive to capsaicin) and TRPA1 (sensitive to allyl isothiocyanate= AITC). Intracellular Ca2+ imaging showed that pretreatment with γ-EVG excited 7% of trigeminal ganglion (TG) cells and increased the amplitude of their responses to AITC, but not to capsaicin or menthol. The enhancing effect of γ-EVG was prevented by a CaSR inhibitor. The results indicate that γ-EVG increases AITC pungency by activating a subset of trigeminal ganglion cells that co-express CaSR and TRPA1.

Mechanosensory neurons that innervate the tongue provide essential information to guide feeding, speech, and social grooming. We use in vivo calcium imaging of mouse trigeminal ganglion neurons to identify functional groups of mechanosensory neurons innervating the anterior tongue. These sensory neurons respond to thermal and mechanical stimulation. Analysis of neuronal activity patterns reveal that most mechanosensory trigeminal neurons are tuned to detect moving stimuli across the tongue. Using an unbiased, multilayer hierarchical clustering approach to classify pressure-evoked activity based on temporal response dynamics, we identify five functional classes of mechanosensory neurons with distinct force-response relations and adaptation profiles. These populations are tuned to detect different features of touch. Molecular markers of functionally distinct clusters are identified by analyzing cluster representation in genetically marked neuronal subsets. Collectively, these studies provide a platform for defining the contributions of functionally distinct mechanosensory neurons to oral behaviors crucial for survival in mammals.

Coordinated peri-ripple activity in the hippocampal-neocortical network is essential for mnemonic information processing in the brain. Hippocampal ripples likely serve different functions in sleep and awake states. Thus, the corresponding neocortical activity patterns may differ in important ways. We addressed this possibility by conducting voltage and glutamate wide-field imaging of the neocortex with concurrent hippocampal electrophysiology in awake mice. Contrary to our previously published sleep results, deactivation and activation were dominant in post-ripple neocortical voltage and glutamate activity, respectively, especially in the agranular retrosplenial cortex (aRSC). Additionally, the spiking activity of aRSC neurons, estimated by two-photon calcium imaging, revealed the existence of two subpopulations of excitatory neurons with opposite peri-ripple modulation patterns: one increases and the other decreases firing rate. These differences in peri-ripple spatiotemporal patterns of neocortical activity in sleep versus awake states might underlie the reported differences in the function of sleep versus awake ripples.

Spontaneous activity is a hallmark of developing neural systems. In the retina, spontaneous activity comes in the form of retinal waves, comprised of three stages persisting from embryonic day 16 (E16) to eye opening at postnatal day 14 (P14). Though postnatal retinal waves have been well characterized, little is known about the spatiotemporal properties or the mechanisms mediating embryonic retinal waves, designated stage 1 waves. Using a custom-built macroscope to record spontaneous calcium transients from whole embryonic retinas, we show that stage 1 waves are initiated at several locations across the retina and propagate across a broad range of areas. Blocking gap junctions reduced the frequency and size of stage 1 waves, nearly abolishing them. Global blockade of nAChRs similarly nearly abolished stage 1 waves. Thus, stage 1 waves are mediated by a complex circuitry involving subtypes of nAChRs and gap junctions. Stage 1 waves in mice lacking the β2 subunit of the nAChRs (β2-nAChR-KO) persisted with altered propagation properties and were abolished by a gap junction blocker. To assay the impact of stage 1 waves on retinal development, we compared the spatial distribution of a subtype of retinal ganglion cells, intrinsically photosensitive retinal ganglion cells (ipRGCs), which undergo a significant amount of cell death, in WT and β2-nAChR-KO mice. We found that the developmental decrease in ipRGC density is preserved between WT and β2-nAChR-KO mice, indicating that processes regulating ipRGC numbers and distributions are not influenced by spontaneous activity.

Epifluorescence miniature microscopes ('miniscopes') are widely used for in vivo calcium imaging of neural population activity. Imaging data are typically collected during a behavioral task and stored for later offline analysis, but emerging techniques for online imaging can support novel closed-loop experiments in which neural population activity is decoded in real time to trigger neurostimulation or sensory feedback. To achieve short feedback latencies, online imaging systems must be optimally designed to maximize computational speed and efficiency while minimizing errors in population decoding. Here we introduce DeCalciOn, an open-source device for real-time imaging and population decoding of in vivo calcium signals that is hardware compatible with all miniscopes that use the UCLA Data Acquisition (DAQ) interface. DeCalciOn performs online motion stabilization, neural enhancement, calcium trace extraction, and decoding of up to 1024 traces per frame at latencies of

Background: Retinal prostheses aim to restore vision by electrically stimulating the remaining viable retinal cells in Retinal Degeneration (RD) cases. Research in this field necessitates a comprehensive analysis of retinal ganglion cells' (RGCs) responses to assess the obtained visual acuity and quality. Here we present a novel animal model which facilitates the optical recording of RGCs activity in an RD rat. This model can significantly enhance the functional evaluation of vision restoration treatments. Methods: The development of the novel rat model is based on crossbreeding a retinal degenerated Royal College of Surgeons (RCS) rat with a transgenic line expressing the genetic calcium indicator GCaMP6f in the RGCs. Characterization of the model was achieved using Optical Coherence Tomography (OCT) imaging, histology, and electroretinography (ERG) at the ages of 4, 8, and 12 weeks. Additionally, optical recordings of RGCs function in response to ex-vivo subretinal electrical stimulations were performed. Results: Histological investigations confirmed the high expression of GCaMP6f in the RGCs and minimal expression in the inner nuclear layer (INL). OCT imaging and histological studies revealed the expected gradual retinal degeneration, as evident by the decrease in retinal thickness with age and the formation of subretinal debris. This degeneration was further confirmed by ERG recordings, which demonstrated a significant decrease in the b-wave amplitude throughout the degeneration process, culminating in its absence at 12 weeks in the GCaMP6f-RCS rat. Importantly, the feasibility of investigating subretinal stimulation was demonstrated, revealing a consistent increase in activation threshold throughout degeneration. Furthermore, an increase in the diameter of the activated area with increasing currents was observed. The spatial spread of the activation area in the GCaMP6f-RCS rat was found to be smaller and exhibited faster activation dynamics compared with the GCaMP6f-LE strain. Conclusion: This novel animal model offers an opportunity to deepen our understanding of prosthetically induced retinal responses, potentially leading to significant advancements in prosthetic interventions in visual impairments. [ABSTRACT FROM AUTHOR]

In mammals, especially rodents, social behaviours, such as parenting, territoriality or mate attraction, are largely based on olfactory communication through chemosignals. These behaviours are mediated by species-specific chemosignals, including small organic molecules and proteins that are secreted in the urine or in various fluids from exocrine glands. Chemosignal detection is mainly ensured by olfactory neurons in two specific sensory organs, the vomeronasal organ (VNO) and the main olfactory epithelium (MOE). This study aimed to characterise the olfactory communication in the fossorial ecotype of the water voles, Arvicola terrestris. We first measured the olfactory investigation of urine and lateral scent gland secretions from conspecifics. Our results showed that water voles can discriminate the sex of conspecifics based on the smell of urine, and that urinary male odour is attractive for female voles. Then, we demonstrated the ability of the VNO and MOE to detect volatile organic compounds (VOCs) found in water vole secretions using live-cell calcium imaging in dissociated cells. Finally, we evaluated the attractiveness of two mixtures of VOCs from urine or lateral scent glands in the field during a cyclical outbreak of vole populations. [ABSTRACT FROM AUTHOR]

Cell therapy has proven to be a promising treatment for a range of neurological disorders, including Parkinson Disease, drug-resistant epilepsy, and stroke, by restoring function after brain damage. Nevertheless, evaluating the true effectiveness of these therapeutic interventions requires a deep understanding of the functional integration of grafted cells into existing neural networks. This review explores a powerful arsenal of molecular techniques revolutionizing our ability to unveil functional integration of grafted cells within the host brain. From precise manipulation of neuronal activity to pinpoint the functional contribution of transplanted cells by using opto- and chemo-genetics, to real-time monitoring of neuronal dynamics shedding light on functional connectivity within the reconstructed circuits by using genetically encoded (calcium) indicators in vivo. Finally, structural reconstruction and mapping communication pathways between grafted and host neurons can be achieved by monosynaptic tracing with viral vectors. The cutting-edge toolbox presented here holds immense promise for elucidating the impact of cell therapy on neural circuitry and guiding the development of more effective treatments for neurological disorders. [ABSTRACT FROM AUTHOR]

The inferior colliculus (IC) of the midbrain is important for complex sound processing, such as discriminating conspecific vocalizations and human speech. The IC's nonlemniscal, dorsal "shell" region is likely important for this process, as neurons in these layers project to higher-order thalamic nuclei that subsequently funnel acoustic signals to the amygdala and nonprimary auditory cortices, forebrain circuits important for vocalization coding in a variety of mammals, including humans. However, the extent to which shell IC neurons transmit acoustic features necessary to discern vocalizations is less clear, owing to the technical difficulty of recording from neurons in the IC's superficial layers via traditional approaches. Here, we use two-photon Ca2+ imaging in mice of either sex to test how shell IC neuron populations encode the rate and depth of amplitude modulation, important sound cues for speech perception. Most shell IC neurons were broadly tuned, with a low neurometric discrimination of amplitude modulation rate; only a subset was highly selective to specific modulation rates. Nevertheless, neural network classifier trained on fluorescence data from shell IC neuron populations accurately classified amplitude modulation rate, and decoding accuracy was only marginally reduced when highly tuned neurons were omitted from training data. Rather, classifier accuracy increased monotonically with the modulation depth of the training data, such that classifiers trained on full-depth modulated sounds had median decoding errors of ∼0.2 octaves. Thus, shell IC neurons may transmit time-varying signals via a population code, with perhaps limited reliance on the discriminative capacity of any individual neuron. NEW & NOTEWORTHY: The IC's shell layers originate a "nonlemniscal" pathway important for perceiving vocalization sounds. However, prior studies suggest that individual shell IC neurons are broadly tuned and have high response thresholds, implying a limited reliability of efferent signals. Using Ca2+ imaging, we show that amplitude modulation is accurately represented in the population activity of shell IC neurons. Thus, downstream targets can read out sounds' temporal envelopes from distributed rate codes transmitted by populations of broadly tuned neurons. [ABSTRACT FROM AUTHOR]

Introduction: Peripheral sensory neurons serve as the initial responders to the external environment. How these neurons react to different sensory stimuli, such as mechanical or thermal forces applied to the skin, remains unclear. Methods: Using in vivo two-photon Ca2+ imaging in the lumbar 4 dorsal root ganglion (DRG) of awake Thy1.2-GCaMP6s mice, we assessed neuronal responses to various mechanical (punctate or dynamic) and thermal forces (heat or cold) sequentially applied to the paw plantar surface. Results: Our data indicate that in normal awake male mice, approximately 14 and 38% of DRG neurons respond to either single or multiple modalities of stimulation. Anesthesia substantially reduces the number of responsive neurons but does not alter the ratio of cells exhibiting single-modal responses versus multi-modal responses. Following peripheral nerve injury, DRG cells exhibit a more than 5.1-fold increase in spontaneous neuronal activity and a 1.5-fold increase in sensory stimulus-evoked activity. As neuropathic pain resulting from nerve injury progresses, the polymodal nature of sensory neurons intensifies. The polymodal population increases from 39.1 to 56.9%, while the modalityspecific population decreases from 14.7 to 5.0% within a period of 5 days. Discussion: Our study underscores polymodality as a significant characteristic of primary sensory neurons, which becomes more pronounced during the development of neuropathic pain. [ABSTRACT FROM AUTHOR]

It has been known for a long time that epileptic seizures provoke brain neuroinflammation involving the activation of microglial cells. However, the role of these cells in this disease context and the consequences of their inflammatory activation on subsequent neuron network activity remain poorly understood so far. To fill this gap of knowledge and gain a better understanding of the role of microglia in the pathophysiology of epilepsy, we used an established zebrafish Dravet syndrome epilepsy model based on Scn1Lab sodium channel loss-of-function, combined with live microglia and neuronal Ca2+ imaging, local field potential (LFP) recording, and genetic microglia ablation. Data showed that microglial cells in scn1Lab-deficient larvae experiencing epileptiform seizures displayed morphological and biochemical changes characteristic of M1-like pro-inflammatory activation; i.e., reduced branching, amoeboid-like morphology, and marked increase in the number of microglia expressing pro-inflammatory cytokine Il1β. More importantly, LFP recording, Ca2+ imaging, and swimming behavior analysis showed that microglia-depleted scn1Lab-KD larvae displayed an increase in epileptiform seizure-like neuron activation when compared to that seen in scn1Lab-KD individuals with microglia. These findings strongly suggest that despite microglia activation and the synthesis of pro-inflammatory cytokines, these cells provide neuroprotective activities to epileptic neuronal networks, making these cells a promising therapeutic target in epilepsy. [ABSTRACT FROM AUTHOR]