Publications using the ES1 & EC1

Adolescence is a developmental period characterized by heightened plasticity. Yet, how ongoing development affects sensory processing and cognitive function is unclear. We investigated how adolescent (postnatal day 20–42) and adult (postnatal day 60–82) mice differ in performance on a pure tone Go/No-Go auditory discrimination task of varying difficulty. Using dense electrophysiological recordings, we measured spiking activity at single neuron resolution in the auditory cortex while mice were engaged in the task. As compared to adults, adolescent mice showed lower auditory discrimination performance in a difficult task. This difference in performance was due to higher response variability and weaker cognitive control expressed as higher lick bias. Adolescent and adult neuronal responses differed only slightly in representations of pure tones when measured outside the context of learning and the task. However, cortical representations after learning within the context of the task were markedly different. We found differences in stimulus- and choice-related activity at the single neuron level representations, as well as lower population-level decoding of the difficult task in adolescents. Overall, cortical decoding in adolescents was lower and slower, especially for difficult sound discrimination, reflecting immature cortical representations of sounds and choices. Notably, we found age-related differences, which were more pronounced after learning, reflecting the combined impact of age and learning. Our findings highlight distinct neurophysiological and behavioral profiles in adolescence, underscoring the ongoing development of cognitive control mechanisms and cortical plasticity during this sensitive developmental period.

Cross-modal responses are common in primary sensory cortex, yet a recent study showed that signals in primary visual cortex (V1) once labeled multisensory were instead more closely related to face movements. To what extent can multisensory processing in other modalities also be explained by face movements? We measured face movements and neural activity in awake mouse primary auditory cortex (A1) and V1 in response to visual and auditory stimulation as well as during spontaneous activity. We find that visual stimuli rarely evoke face movements, A1 responses to visual input remain robust even in the absence of face movement, and optogenetic silencing of V1 reduces A1 visual responses. Taken together, these results confirm that cross-modal responses in the auditory system are primarily sensory, not state driven, in origin.

The neural representations of acoustic features that differ in the location or timbre of the emitter elicit similar perceptions, suggesting the existence of a robust stimulus‐response function between complex sounds and the activity of neural populations at all stages of the auditory system. This hypothesis is tested by decoding a random sound stream, using spike trains from a biophysical model of the auditory nerve and from large‐scale recordings in the inferior colliculus, the auditory thalamus, and the auditory cortex of awake mice. At the level of individual neurons, the reliability of temporal and rate codes is found to decrease along the ascending auditory pathways. Rate coding is progressively favored with increasing independence of neuron frequency tuning. Firing in the auditory cortex is found to be synergistic, whereas that in subcortical areas is more redundant. Finally, combinatorial codes involving neural firing and neural silence within neuron pairs are shown to efficiently encode sound information, particularly in the auditory cortex. Overall, these findings reveal a progressive transformation of the neural code from an individual, redundant, and temporal code at the periphery to a more distributed rate‐based code in the auditory cortex.

Sound harmonicity is foundational in complex auditory stimuli like music and vocalizations but it remains unclear how such spectrally complex stimuli are processed in the auditory cortex (ACtx). Subregions of the auditory cortex process are thought to process harmonic stimuli differently, and secondary ACtx (A2) layer (L) 2/3 is believed to be the most selective. Selective responses to sound features in ACtx are thought to emerge hierarchically starting from A1 L4. Since the spectral complexity of harmonic stacks can range from two to more than ten components, harmonic selectivity and sensitivity might also arise hierarchically across layers and areas. We studied responses to simple and complex harmonic stacks across A1 L4, A1 L2/3, and A2 L2/3 in adult mice using in vivo two-photon microscopy. We found harmonic-sensitive neurons (HN) responding only to harmonic stacks but not to their pure-tone components in all areas at similar proportions. HNs showed non-linear processing of component tones with onset-responsive HNs showing greater nonlinearity, which decreased with harmonic complexity. Co-tuned HNs in A1 L4 exhibited the highest signal correlation, regardless of harmonic complexity. A1 L4 HNs also showed the lowest noise correlation with other neurons. Moreover, A1 L4 HNs achieve robust spectral integration and harmonic sensitivity by receiving diverse inputs and maintaining high signal correlation, ensuring independent, strong responses to harmonic stimuli. Therefore, harmonic sensitivity is present in A1 L4 and is not a unique feature of A2. Thus, tuning to complex spectral sounds is a fundamental property of ACtx and is already established in A1 L4. Significance statement Harmonics are essential in auditory perception, influencing how we process complex sounds like music and speech. This study reveals that neurons in the primary auditory cortex (A1) and secondary auditory cortex (A2) integrate simple and complex harmonic structures with distinct mechanisms of neuronal recruitment. A1 L4 harmonic-sensitive neurons (HNs) demonstrated strong, independent responses through high signal correlation and minimal noise correlation, suggesting a robust mechanism for spectral integration. Our results show that harmonic relationships are already extracted at the input layers of A1, and that HNs show non-linear facilitative integration. Thus, tuning to sounds of complex spectral contents might be a fundamental processing function of the auditory cortex and is already established in A1 L4, which receives major thalamic inputs.

Maternal defense, a hallmark of motherhood, often involves protecting offspring at personal risk. However, the neural mechanisms underlying the emergence and modulation of this behavior remain not-well understood. Here, we introduce a novel paradigm in which mother mice under threat prioritize offspring protection over self-preservation by approaching, collecting, and retreating pups to safety, a risk-associated behavior that also develops in virgin females following maternal experience. Microendoscopic calcium imaging reveals sequential activation of distinct corticofugal neuronal populations in the medial prefrontal cortex (mPFC) during maternal defense. Specifically, layer (L)6 dopamine receptor D1-expressing (D1) neurons are active during pup approach, while L5 neurons are engaged during pup collection and retreat. Activation of the L6 neurons bidirectionally modulates maternal defense via projections to the medial preoptic area. These findings suggest that mPFC L6 D1 neurons may facilitate the transition to maternal behavior by influencing behavioral selection and initiating maternal defense response under threat.

The insular cortex is involved in a wide range of auditory functions in the mammalian brain. We studied the organization and basic response properties of auditory neurons in the insular cortex and adjacent areas by recording responses to sound stimuli in anesthetized female rats. Auditory neurons were present in an insulo-parietal auditory field that spans the boundary between the posterior insula, particularly in the granular insular cortex, and the ventral part of the adjacent nonprimary somatosensory fields. Confirming previous reports, neurons in this field had narrow tuning and were preferentially tuned to relatively low frequencies (<16 kHz). Intriguingly, auditory units in this field displayed shorter minimal latencies than units in the primary auditory cortex, as well as weaker sensitivity to deviance. These results establish the existence of a rapid auditory information stream with distinct properties through the insulo-parietal cortex that may parallel the pathway through the primary and anterior auditory fields.

Fear retrieval is fundamental for animals to rapidly enter a self-defense state, which enhances their survival rate. Although the cellular mechanisms and neural circuits underlying fear retrieval have been relatively well studied, particularly in classic fear-related nuclei such as the amygdala, the neural circuits and intrinsic microcircuits of some non-classical fear-related nuclei remain poorly understood. A disinhibitory microcircuit in the temporal association cortex (TeA) was identified in this study, demonstrating a critical role in fear retrieval. Additionally, the response of the TeA to fear retrieval is largely driven by acetylcholine (Ach), which then relay information to the tail of striatum (TS) to mediate freezing behavior. Together, these findings demonstrate the essential role of the TeA and its microcircuit in auditory fear retrieval in response to pure tones, providing new insights into the mechanisms of fear retrieval in the cortex.

Auditory processing in the cerebral cortex is considered to begin with thalamocortical inputs to layer 4 (L4) of the primary auditory cortex (A1). In this canonical model, A1 L4 inputs initiate a hierarchical cascade that propagates to higher-order cortices for slower integration of complex sounds. Here, we identify parallel ascending pathways in mice that bypass A1 and directly reach the secondary auditory cortex (A2), alongside the canonical hierarchical route. We found that layer 6 (L6) of both A1 and A2 receive short-latency (<10 ms) sound inputs via higher-order thalamic nuclei. Additionally, A2 L4 is innervated by a caudal subdivision of the traditionally defined primary thalamus, which we now re-classify as non-primary. Notably, both identified thalamic regions receive projections from distinct subdivisions of the higher-order inferior colliculus, which in turn receive direct projections from cochlear nucleus neurons. Thus, higher-order auditory cortex integrates both slower, pre-processed information and rapid, direct sensory inputs, enabling parallel processing of fast sensory information across cortical areas.

Analyzing the functional connectivity of the brain is an enormous challenge, as deciphering functional connectivity requires knowledge of functional responses and connections. One promising strategy is analyzing the spatial pattern of activity correlations across cell populations. In the primary auditory cortex (A1), cells respond to different sound features. On the large scale, there exists a tonotopic map, which is fractured at the small scale, raising the question of whether functional connections are spatially ordered or disordered. To test whether functional connectivity on a local and a global scale is also disordered, we first designed a robust statistical model to estimate parameters and test for the significance of the estimated correlation maps. We developed an inference method that allows efficient model fitting and statistical testing to project the correlation maps to 2D space. We then performed in vivo two-photon calcium imaging in layer 2/3 of A1 with pure tones (PT) or a combination of two tones (TT; harmonically related or not). We found that the spatial patterns of signal correlations (SCs) depend on the type of sound stimuli that were presented. The functional 2D maps of PT-driven SCs are more restricted to local neurons than TT signal correlations which showed more global textures. 2D SC patterns for harmonic stimuli showed spatially distinct relationships. TT SCs revealed spatially precise functional connectivity between harmonically related neurons. Thus, even though the frequency preference of neighboring neurons in A1 is functionally diverse, the functional connection pattern of these neurons is functionally precise and harmonically related.

Negative feedback of the cochlear efferent system plays a critical role in control of hearing sensitivity and protection from noise trauma. Type II auditory nerves (ANs) innervate outer hair cells (OHCs) in the cochlea and provide an input to the cochlear efferent system to achieve hearing sensitivity controlling and protection; in particular, medial olivocochlear efferent nerves innervate OHCs to control OHC electromotility, which is an active cochlear amplifier in mammals. However, little is known about channel information underlying type II AN activity and consequent function. Here, we report that ATP-gated P2x7 receptor had a predominant expression at type II spiral ganglion (SG) neurons and the synaptic areas under inner hair cells and OHCs with lateral and medial olivocochlear efferent nerves. Knockout (KO) of P2x7 increased hearing sensitivity with enhanced acoustic startle response, auditory brainstem response, and cochlear microphonics by increasing OHC electromotility. P2x7 KO also increased susceptibility to noise and exacerbated ribbon synapse degeneration. Middle-level noise exposure could impair active cochlear mechanics resulting in hearing loss in P2x7 KO mice. These data demonstrate that P2x7 receptors have a critical role in type II SG neuron’s function and the cochlear efferent system to control hearing sensitivity; deficiency of P2x7 receptors can impair type II SG neuron’s function and the cochlear efferent suppression leading to increase of active cochlear amplification and hearing oversensitivity, i.e., hyperacusis, and susceptibility to noise, which may also associate with other hearing disorders, such as tinnitus.

Electroanalytical methods are used to understand, modify, and control bionic devices. Bionic devices can record or stimulate cells to understand and/or control normal or abnormal biological processes. These devices contain electrodes that transduce electrical current within the electrical circuit into ionic current within a tissue. Despite the similarity between electroanalysis and electrophysiology, there remains a poor understanding of the relationship between the two techniques, including their methodology and theory. This paper investigates the electrochemical and acute electrophysiological recording performance of neural electrodes. A range of behaviors is achieved by modifying electrodes with the conducting polymer poly(3,4‐ethylenedioxythiophene) (PEDOT) doped with chondroitin sulfate, dextran sulfate, or para ‐toluene sulfonate. The results support previous studies showing that increased electrode area reduced total impedance below the Maxwell–Wagner relaxation frequency and thermal noise while increasing the signal‐to‐noise ratio and neural spike count. The results allowed novel investigation of relative contributions of biological and electrode properties to electrophysiological performance, with increased electrode area having a larger impact on neural population within recording range rather than reducing thermal noise. The utility of measuring electrode impedance for predicting electrophysiological performance is mainly for an indirect measure of electrode area. The results provide insight into noise sources from electrophysiological recordings and limitations in cable theory in neuroscience.

Socially effective vocal communication requires brain regions that encode expressive and receptive aspects of vocal communication in a social context-dependent manner. Here, we combined a novel behavioral assay with microendoscopic calcium imaging to interrogate neuronal activity (regions of interest [ROIs]) in the posterior insula (pIns) in socially interacting mice as they switched rapidly between states of vocal expression and reception. We found that largely distinct subsets of pIns ROIs were active during vocal expression and reception. Notably, pIns activity during vocal expression increased prior to vocal onset and was also detected in congenitally deaf mice, pointing to a motor signal. Furthermore, receptive pIns activity was modulated strongly by social context. Lastly, tracing experiments reveal that deep-layer neurons in the pIns directly bridge the auditory thalamus to a midbrain vocal gating region. Therefore, the pIns is a site that encodes vocal expression and reception in a manner that depends on social context.

Aminoglycoside antibiotics remain indispensable despite their ototoxicity. Like other sensorineural forms, aminoglycoside-induced hearing loss (AGIHL) has no effective pharmacotherapy. Oxidative stress, apoptosis, excitotoxicity and inflammation are key pathological factors of the disease. We hypothesised that selegiline, an irreversible monoamine oxidase-B (MAO-B) inhibitor used in Parkinson’s disease, could be repurposed as an otoprotective agent against AGIHL and its effect on dopamine (DA) release from lateral olivocochlear (LOC) fibres, the efferent division of a protective feedback loop plays a major role in the protection against excitotoxicity. Selegiline mitigated AGIHL in BALB/c mice in a dose-dependent manner at different auditory brainstem response frequencies, including 16 kHz, the hearing sensitivity optimum of the animals. It also enhanced the action potential-evoked DA release from LOC efferents in mouse cochlear preparation dose-dependently. Inhibition of DA reuptake contributed to its basic effect of saving DA from metabolism. Among four selegiline analogues tested, the one that increased LOC DA release also provided otoprotection. In contrast, neither safinamide (a reversible MAO-B inhibitor) nor LJP-1207 (a selective semicarbazide-sensitive amine oxidase/vascular-adhesion protein 1 (SSAO/VAP1) inhibitor) prevented AGIHL, despite their antioxidant and anti-inflammatory properties. The reversibility or lack of MAO-B inhibition in safinamide and LJP-1207, respectively, as well as the absence of the propargylamine moiety with known intrinsic neuroprotective activity in both molecules, may explain their ineffectiveness. Selegiline, or certain propargylamine analogues of it, offer a promising therapy against AGIHL by addressing its multifactorial pathology through antioxidant, antiapoptotic, neuroprotective, and anti-inflammatory actions, while enhancing endogenous DAergic protective mechanisms.

Many animal training paradigms rely on head-fixation. Head-fixation training is typically laborious and can benefit from automation to relieve the workload and reduce the variability in the training outcome. Several groups have reported successful implementations of such systems, but throughput varied greatly across groups. In addition, most studies relied on brief periods of head-fixation sessions (≤ 1 minute) to reduce the potential stress on the animal. Here, we report the design of a new system that could achieve head-fixation sessions on the order of minutes with a high participation rate from the animal (100%). Throughout the training period, each mouse performed a total of close to 40 minutes of head-fixation training on average each day and learned common psychoacoustic tasks, i.e., tone detection and tone discrimination. Our system can achieve highly efficient training with minimum idling time, allowing combinations with high-end neural recording equipment to achieve maximum training and data collection efficiency.

Foraging theory predicts animal behavior in many contexts. In patch-based foraging behaviors, the marginal value theorem (MVT) gives the optimal strategy for deterministic environments whose parameters are fully known to the forager. In natural settings, environmental parameters exhibit variability and are only partially known to the animal based on its experience, creating uncertainty. Models of uncertainty in foraging are well established. However, natural environments also exhibit unpredicted changes in their statistics. As a result, animals must ascertain whether the currently observed quality of the environment is consistent with their internal models, or whether something has changed, creating meta-uncertainty. Behavioral strategies for optimizing foraging behavior under meta-uncertainty, and their neural underpinnings, are largely unknown. Here, we developed a novel behavioral task and computational framework for studying patch-leaving decisions in head-fixed and freely moving mice in conditions of meta-uncertainty. We stochastically varied between-patch travel time, as well as within-patch reward depletion rate. We find that, when uncertainty is minimal, mice adopt patch residence times in a manner consistent with the MVT and not explainable by simple ethologically motivated heuristic strategies. However, behavior in highly variable environments was best explained by modeling both first- and second-order uncertainty in environmental parameters, wherein local variability and global statistics are captured by a Bayesian estimator and dynamic prior, respectively. Thus, mice forage under meta-uncertainty by employing a hierarchical Bayesian strategy, which is essential for efficiently foraging in volatile environments. The results provide a foundation for understanding the neural basis of decision-making that exhibits naturalistic meta-uncertainty.

While evolutionary cortical expansion is thought to underlie the evolution of human cognitive capabilities, excessive developmental expansion can lead to megalencephaly, often found in neurodevelopmental disorders. Still, little is known about how the overproduction of cortical neurons during development affects cortical processing and behavior in later life. Here we show that developmental overproduction of cortical superficial neurons impairs auditory processing in adult mice. We applied XAV939 to overproduce cortical superficial excitatory neurons during development. XAV939-treated adult mice exhibited auditory behavioral deficits and abnormal auditory cortical processing. Furthermore, we found fewer functional monosynaptic connections between cortical putative excitatory neurons. Altogether, our results suggest that abnormal auditory cortical processing contributes to the atypical auditory detectability in XAV939-treated mice. Although the expansion of cortical size is evolutionarily advantageous, an abnormal expansion during development can result in detrimental effects on cortical processing and perceptual behavior in adulthood.

Molecular profiling of inner ear neurons has broadened the classification of the primary afferents that support neural coding for hearing and balance. To extend spatiotemporal characterization of auditory and vestibular neuron diversity, we established a transgenic reporter mouse model (Prphp-mCherry), where elements of the peripherin promoter (Prphp) drive expression of the mCherry fluorescent reporter. Type III intermediate filament protein peripherin expression is a marker for type II spiral ganglion neurons (SGN) that innervate the cochlear outer hair cells, and the small diameter 'bouton' vestibular ganglion neurons (VGN) innervating the type II vestibular hair cells. Using Nanopore genome sequencing, the integration site of the transgene construct was identified within the class III metabotropic glutamate receptor 8 gene (Grm8, chromosome 6). Use of CUBIC / PEGASOS clearing of early postnatal to adult inner ear tissues enabled in situ 3D spatial localization of a dispersed population of cochlear mCherry + ve SGN, with highest expression and density in the hook (high frequency encoding) basal region. Of these mCherry + ve SGN, type II SGN (peripherin-immunopositive) were all co-labeled in the basal region, but the majority of the overall mCherry-delineated SGN auditory subpopulation were type I SGN innervating inner hair cells. In the VGN, mCherry + ve neurons represented ~ 15% of the adult population, dispersed as a small diameter subpopulation throughout both the inferior and superior VGN regions. These findings resolve heterogeneous type I and type II cochlear SGN subpopulations, particularly in the structurally complex hook region, and further differentiate vestibular primary afferents across postnatal development.

To navigate real-world listening conditions, the auditory system relies on the integration of multiple sources of information. However, to avoid inappropriate cross-talk between inputs, highly connected neural systems need to strike a balance between integration and segregation. Here, we develop a novel approach to examine how repeated neurochemical modules in the mouse inferior colliculus lateral cortex (LC) allow controlled integration of its multimodal inputs. The LC had been impossible to study via imaging because it is buried in a sulcus. Therefore, we coupled two-photon microscopy with the use of a microprism to reveal the first-ever sagittal views of the LC to examine neuronal responses with respect to its neurochemical motifs under anesthetized and awake conditions. This approach revealed marked differences in the acoustic response properties of LC and neighboring non-lemniscal portions of the inferior colliculus. In addition, we observed that the module and matrix cellular motifs of the LC displayed distinct somatosensory and auditory responses. Specifically, neurons in modules demonstrated primarily offset responses to acoustic stimuli with enhancement in responses to bimodal stimuli, whereas matrix neurons showed onset response to acoustic stimuli and suppressed responses to bimodal stimulation. Thus, this new approach revealed that the repeated structural motifs of the LC permit functional integration of multimodal inputs while retaining distinct response properties.

Auditory neuropathies affect the spiral ganglion neurons of the auditory nerve or their synapses with the sensory hair cells, distorting the sound information transmitted from the ear to the brain. Deciphering the underlying pathophysiological mechanisms remains challenging owing to the diversity of spiral ganglion neuron subtypes and associated central auditory circuits. An auditory neuropathy mechanism is unraveled by investigating the origin of auditory hyperexcitability in a mouse model for hereditary congenital deafness. Otogl encodes the large Otogelin‐like protein, which is related to secreted epithelial mucins and is implicated in the mechanical stimulation of cochlear outer hair cells. Heterozygous Otogl +/− mutant mice display auditory hyperexcitability, highlighted by their susceptibility to audiogenic seizures induced by loud sounds. It is shown that Otogl is transiently expressed in a subpopulation of spiral ganglion neurons during cochlear development. Despite their apparently normal hearing, Otogl +/− mice display poor activation of the spiral ganglion neurons processing loud sounds and an elevation of the activation threshold of the middle the ear muscle reflex that attenuates loud sounds. The findings reveal how a neuropathy affecting spiral ganglion neurons specialized in loud sound processing and associated with the middle the ear muscle reflex can manifest itself as auditory hyperexcitability.

Transgenic mice provide unprecedented access to manipulate and visualize neural circuits; however, those on a C57BL/6 background develop progressive hearing loss, significantly confounding systems-level and behavioral analysis. While outbreeding can limit hearing loss, it introduces strain variability and complicates the generation of complex genotypes. Here, we propose an approach to preserve hearing by crossing transgenic mice with congenic B6.CAST- Cdh23 Ahl + mice, which maintain low-threshold hearing into adulthood. Widefield and two-photon imaging of the auditory cortex revealed that 2.5-month-old C57BL/6 mice exhibit elevated thresholds to high-frequency tones and widespread cortical reorganization, with most neurons responding best to lower frequencies. In contrast, Ahl+ C57BL/6 mice exhibited robust neural responses across tested frequencies and sound levels (4–64 kHz, 30–90 dB SPL) and retained low thresholds into adulthood. Our approach offers a cost-effective solution for generating complex genotypes and facilitates more interpretable systems neuroscience research by eliminating confounding effects from hearing loss.