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The cerebral cortex, basal ganglia and motor thalamus form circuits important for purposeful movement. In Parkinsonism, basal ganglia neurons often exhibit dysrhythmic activity during, and with respect to, the slow (∼1 Hz) and beta-band (15-30 Hz) oscillations that emerge in cortex in a brain state-dependent manner. There remains, however, a pressing need to elucidate the extent to which motor thalamus activity becomes similarly dysrhythmic after dopamine depletion relevant to Parkinsonism. To address this, we recorded single-neuron and ensemble outputs in the basal ganglia-recipient zone (BZ) and cerebellar-recipient zone (CZ) of motor thalamus in anesthetized male dopamine-intact rats and 6-OHDA-lesioned rats during two brain states, respectively defined by cortical slow-wave activity and activation. Two forms of thalamic input zone-selective dysrhythmia manifested after dopamine depletion: (1) BZ neurons, but not CZ neurons, exhibited abnormal phase-shifted firing with respect to cortical slow oscillations prevalent during slow-wave activity; and (2) BZ neurons, but not CZ neurons, inappropriately synchronized their firing and engaged with the exaggerated cortical beta oscillations arising in activated states. These dysrhythmias were not accompanied by the thalamic hypoactivity predicted by canonical firing rate-based models of circuit organization in Parkinsonism. Complementary recordings of neurons in substantia nigra pars reticulata suggested that their altered activity dynamics could underpin the BZ dysrhythmias. Finally, pharmacological perturbations demonstrated that ongoing activity in the motor thalamus bolsters exaggerated beta oscillations in motor cortex. We conclude that BZ neurons are selectively primed to mediate the detrimental influences of abnormal slow and beta-band rhythms on circuit information processing in Parkinsonism.SIGNIFICANCE STATEMENTMotor thalamus neurons mediate the influences of basal ganglia and cerebellum on the cerebral cortex to govern movement. Chronic depletion of dopamine from the basal ganglia causes some symptoms of Parkinson's disease. Here, we elucidate how dopamine depletion alters the ways motor thalamus neurons engage with two distinct oscillations emerging in cortico-basal ganglia circuitsin vivo. We discovered that, after dopamine depletion, neurons in the thalamic zone receiving basal ganglia inputs are particularly prone to becoming dysrhythmic, changing the phases and/or synchronization (but not rate) of their action potential firing. This bolsters cortical dysrhythmia. Our results provide important new insights into how aberrant rhythmicity in select parts of motor thalamus could detrimentally affect neural circuit dynamics and behavior in Parkinsonism.

Loss-of-function mutations in endosomal Na+/H+exchanger 6 (NHE6) cause the X-linked neurologic disorder Christianson syndrome. Patients exhibit symptoms associated with both neurodevelopmental and neurodegenerative abnormalities. While loss of NHE6 has been shown to overacidify the endosome lumen, and is associated with endolysosome neuropathology, NHE6-mediated mechanisms in endosome trafficking and lysosome function have been understudied. Here, we show that NHE6-null mouse neurons demonstrate worsening lysosome function with time in culture, likely as a result of defective endosome trafficking. NHE6-null neurons exhibit overall reduced lysosomal proteolysis despite overacidification of the endosome and lysosome lumen. Akin to Nhx1 mutants inSaccharomyces cerevisiae, we observe decreased endosome-lysosome fusion in NHE6-null neurons. Also, we find premature activation of pH-dependent cathepsin D (CatD) in endosomes. While active CatD is increased in endosomes, CatD activation and CatD protein levels are reduced in the lysosome. Protein levels of another mannose 6-phosphate receptor (M6PR)-dependent enzyme, β-N-acetylglucosaminidase, were also decreased in lysosomes of NHE6-null neurons. M6PRs accumulate in late endosomes, suggesting defective M6PR recycling and retromer function in NHE6-null neurons. Finally, coincident with decreased endosome-lysosome fusion, using total internal reflection fluorescence, we also find a prominent increase in fusion between endosomal multivesicular bodies and the plasma membrane, indicating enhanced exosome secretion from NHE6-null neurons. In summary, in addition to overacidification of endosomes and lysosomes, loss of NHE6 leads to defects in endosome maturation and trafficking, including enhanced exosome release, contributing to lysosome deficiency and potentially leading to neurodegenerative disease.SIGNIFICANCE STATEMENTLoss-of-function mutations in the endosomal Na+/H+exchanger 6 (NHE6) cause Christianson syndrome, an X-linked neurologic disorder. Loss of NHE6 has been shown to overacidify endosomes; however, endosome trafficking mechanisms have been understudied, and the mechanisms leading to neurodegeneration are largely unknown. In NHE6-null mouse neuronsin vitro, we find worsening lysosome function with days in culture. Notably, pH-dependent lysosome enzymes, such as cathepsin D, have reduced activity in lysosomes yet increased, precocious activity in endosomes in NHE6-null neurons. Further, endosomes show reduced fusion to lysosomes, and increased fusion to the plasma membrane with increased exosome release. This study identifies new mechanisms involving defective endosome maturation and trafficking that impair lysosome function in Christianson syndrome, likely contributing to neurodegeneration.

Recent large genome-wide association studies (GWAS) have identified multiple confident risk loci linked to addiction-associated behavioral traits. Genetic variants linked to addiction-associated traits lie largely in non-coding regions of the genome, likely disrupting cis-regulatory element (CRE) function. CREs tend to be highly cell type-specific and may contribute to the functional development of the neural circuits underlying addiction. Yet, a systematic approach for predicting the impact of risk variants on the CREs of specific cell populations is lacking. To dissect the cell types and brain regions underlying addiction-associated traits, we applied LD score regression to compare GWAS to genomic regions collected from human and mouse assays for open chromatin, which is associated with CRE activity. We found enrichment of addiction-associated variants in putativeCREsmarked by open chromatin in neuronal (NeuN+) nuclei collected from multiple prefrontal cortical areas and striatal regions known to play major roles in reward and addiction. To further dissect the cell type-specific basis of addiction-associated traits, we also identified enrichments in human orthologs of open chromatin regions of mouseneuronal subtypes: cortical excitatory, D1, D2, and PV. Lastly, we developed machine learning models from mouse cell type-specific regions of open chromatin to further dissect human NeuN+ open chromatin regions into cortical excitatory or striatal D1 and D2 neurons and predict the functional impact of addiction-associated genetic variants. Our results suggest that differentneuronal subtypeswithin the reward system play distinct roles in the variety of traits that contribute to addiction.Significance StatementWe combine statistical genetic and machine learning techniques to find that the predisposition to for nicotine, alcohol, and cannabis use behaviors can be partially explained by genetic variants in conserved regulatory elements within specific brain regions and neuronal subtypes of the reward system.This computational framework can flexibly integrateopen chromatindata across species to screen for putative causal variants in a cell type-and tissue-specific manner across numerous complex traits.

Reactive oxygen species (ROS) produced by NADPH oxidases (Nox) contribute to the development of different types of sensorineural hearing loss (SNHL), a common impairment in humans with no established treatment. Although the essential role of Nox3 in otoconia biosynthesis and its possible involvement in hearing have been reported in rodents, immunohistological methods targeted at detecting Nox3 expression in inner ear cells reveal ambiguous results. Therefore, the mechanism underlying Nox3-dependent SNHL remains unclear and warrants further investigation. We generatedNox3-Creknock-in mice, in which Nox3 was replaced withCre recombinase(Cre). UsingNox3-Cre;tdTomatomice of either sex, in which tdTomato is expressed under the control of theNox3promoter, we determined Nox3-expressing regions and cell types in the inner ear. Nox3-expressing cells in the cochlea included various types of supporting cells, outer hair cells, inner hair cells, and spiral ganglion neurons. Nox3 expression increased with cisplatin, age, and noise insults. Moreover, increased Nox3 expression in supporting cells and outer hair cells, especially at the basal turn of the cochlea, played essential roles in ROS-related SNHL. The extent of Nox3 involvement in SNHL follows the following order: cisplatin-induced hearing loss > age-related hearing loss > noise-induced hearing loss. Here, on the basis ofNox3-Cre;tdTomato, which can be used as a reporter system (Nox3-Cre+/−;tdTomato+/+andNox3-Cre+/+;tdTomato+/+), andNox3-KO (Nox3-Cre+/+;tdTomato+/+) mice, we demonstrate that Nox3 inhibition in the cochlea is a promising strategy for ROS-related SNHL, such as cisplatin-induced HL, age-related HL, and noise-induced HL.SIGNIFICANCE STATEMENTWe found Nox3-expressing regions and cell types in the inner ear, especially in the cochlea, usingNox3-Cre;tdTomatomice, a reporter system generated in this study. Nox3 expression increased with cisplatin, age, and noise insults in specific cell types in the cochlea and resulted in the loss (apoptosis) of outer hair cells. Thus, Nox3 might serve as a molecular target for the development of therapeutics for sensorineural hearing loss, particularly cisplatin-induced, age-related, and noise-induced hearing loss.

Tau aggregation within neurons is a critical feature of Alzheimer's disease (AD) and related tauopathies. It is believed that soluble pathologic tau species seed the formation of tau aggregates in a prion-like manner and propagate through connected neurons during the progression of disease. Both soluble and aggregated forms of tau are thought to have neurotoxic properties. In addition, different strains of misfolded tau may cause differential neurotoxicity. In this work, we present an accelerated human neuronal model of tau-induced neurotoxicity that incorporates both soluble tau species and tau aggregation. Using patient-derived induced pluripotent stem cell (iPSC) neurons expressing a tau aggregation biosensor, we develop a cell culture system that allows continuous assessment of both induced tau aggregation and neuronal viability at single-cell resolution for periods of >1 week. We show that exogenous tau “seed” uptake, as measured by tau repeat domain (TauRD) reporter aggregation, increases the risk for subsequent neuronal deathin vitro. These results are the first to directly visualize neuronal TauRD aggregation and subsequent cell death in single human iPSC neurons. Specific morphologic strains or patterns of TauRD aggregation are then identified and associated with differing neurotoxicity. Furthermore, we demonstrate that familial AD iPSC neurons expressing the PSEN1 L435F mutation exhibit accelerated TauRD aggregation kinetics and a tau strain propagation bias when compared with control iPSC neurons.SIGNIFICANCE STATEMENTNeuronal intracellular aggregation of the microtubule binding protein tau occurs in Alzheimer's disease and related neurodegenerative tauopathies. Tau aggregates are believed to spread from neuron to neuron via prion-like misfolded tau seeds. Our work develops a human neuronal live-imaging system to visualize seeded tau aggregation and tau-induced neurotoxicity within single neurons. Using an aggregation-sensing tau reporter, we find that neuronal uptake and propagation of tau seeds reduces subsequent survival. In addition, human induced pluripotent stem cell (iPSC) neurons carrying an Alzheimer's disease-causing mutation in presenilin-1 undergo tau seeding more rapidly than control iPSC neurons. However, they do not show subsequent differences in neuronal survival. Finally, specific morphologies of tau aggregates are associated with increased neurotoxicity.

Spinocerebellar ataxias (SCAs) are diseases characterized by cerebellar atrophy and loss of Purkinje neurons caused by mutations in diverse genes. In SCA14, the disease is caused by point mutations or small deletions in protein kinase C γ (PKCγ), a crucial signaling protein in Purkinje cells. It is still unclear whether increased or decreased PKCγ activity may be involved in the SCA14 pathogenesis. In this study, we present a new knock-in mouse model related to SCA14 with a point mutation in the pseudosubstrate domain, PKCγ-A24E, known to induce a constitutive PKCγ activation. In this protein conformation, the kinase domain of PKCγ is activated, but at the same time the protein is subject to dephosphorylation and protein degradation. As a result, we find a dramatic reduction of PKCγ protein expression inPKCγ-A24Emice of either sex. Despite this reduction, there is clear evidence for an increased PKC activity in Purkinje cells fromPKCγ-A24Emice. Purkinje cells derived from PKCγ-A24E have short thickened dendrites typical for PKC activation. These mice also develop a marked ataxia and signs of Purkinje cell dysfunction making them an interesting new mouse model related to SCA. Recently, a similar mutation in a human patient was discovered and found to be associated with overt SCA14. RNA profiling ofPKCγ-A24Emice showed a dysregulation of related signaling pathways, such as mGluR1 or mTOR. Our results show that the induction of PKCγ activation in Purkinje cells results in the SCA-like phenotype indicating PKC activation as one pathogenetic avenue leading to a SCA.SIGNIFICANCE STATEMENTSpinocerebellar ataxias (SCAs) are hereditary diseases affecting cerebellar Purkinje cells and are a one of neurodegenerative diseases. While mutation in several genes have been identified as causing SCAs, it is unclear how these mutations cause the disease phenotype. Mutations in PKCγ cause one subtype of SCAs, SCA14. In this study, we have generated a knock-in mouse with a mutation in the pseudosubstrate domain of PKCγ, which keeps PKCγ in the constitutive active open conformation. We show that this mutation leading to a constant activation of PKCγ results in a SCA-like phenotype in these mice. Our findings establish the constant activation of PKC signaling as one pathogenetic avenue leading to an SCA phenotype and a mechanism causing a neurodegenerative disease.

Alzheimer's disease is a devastating neurodegenerative disease with a dramatically increasing prevalence and no disease-modifying treatment. Inflammatory lifestyle factors increase the risk of developing Alzheimer's disease. Zinc deficiency is the most prevalent malnutrition in the world and may be a risk factor for Alzheimer's disease potentially through enhanced inflammation, although evidence for this is limited. Here we provide epidemiological evidence suggesting that zinc supplementation was associated with reduced risk and slower cognitive decline, in people with Alzheimer's disease and mild cognitive impairment. Using the APP/PS1 mouse model of Alzheimer's disease fed a control (35 mg/kg zinc) or diet deficient in zinc (3 mg/kg zinc), we determined that zinc deficiency accelerated Alzheimer's-like memory deficits without modifying amyloid β plaque burden in the brains of male mice. The NLRP3-inflammasome complex is one of the most important regulators of inflammation, and we show here that zinc deficiency in immune cells, including microglia, potentiated NLRP3 responses to inflammatory stimuliin vitro, including amyloid oligomers, while zinc supplementation inhibited NLRP3 activation.APP/PS1mice deficient in NLRP3 were protected against the accelerated cognitive decline with zinc deficiency. Collectively, this research suggests that zinc status is linked to inflammatory reactivity and may be modified in people to reduce the risk and slow the progression of Alzheimer's disease.SIGNIFICANCE STATEMENTAlzheimer's disease is a common condition mostly affecting the elderly. Zinc deficiency is also a global problem, especially in the elderly and also in people with Alzheimer's disease. Zinc deficiency contributes to many clinical disorders, including immune dysfunction. Inflammation is known to contribute to the risk and progression of Alzheimer's disease; thus, we hypothesized that zinc status would affect Alzheimer's disease progression. Here we show that zinc supplementation reduced the prevalence and symptomatic decline in people with Alzheimer's disease. In an animal model of Alzheimer's disease, zinc deficiency worsened cognitive decline because of an enhancement in NLRP3-driven inflammation. Overall, our data suggest that zinc status affects Alzheimer's disease progression, and that zinc supplementation could slow the rate of cognitive decline.

Preclinical studies show a link between subthalamic nucleus (STN) deep brain stimulation (DBS) and neuroprotection of nigrostriatal dopamine (DA) neurons, potentially through brain-derived neurotrophic factor (BDNF) signaling. However, the question of whether DBS of the STN can be disease-modifying in Parkinson's disease (PD) remains unanswered. In particular, the impact of STN DBS on α-synuclein (α-syn) aggregation, inclusion-associated neuroinflammation, and BDNF levels has yet to be examined in the context of synucleinopathy. To address this, we examined the effects of STN DBS on BDNF using the α-syn preformed fibril (PFF) model in male rats. While PFF injection resulted in accumulation of phosphorylated α-syn (pSyn) inclusions in the substantia nigra pars compacta (SNpc) and cortical areas, STN DBS did not impact PFF-induced accumulation of pSyn inclusions in the SNpc. In addition, nigral pSyn inclusions were associated with increased microgliosis and astrogliosis; however, the magnitude of these processes was not altered by STN DBS. Total BDNF protein was not impacted by pSyn inclusions, but the normally positive association of nigrostriatal and corticostriatal BDNF was reversed in rats with PFF-induced nigrostriatal and corticostriatal inclusions. Despite this, rats receiving both STN DBS and PFF injection showed increased BDNF protein in the striatum, which partially restored the normal corticostriatal relationship. Our results suggest that pathologic α-syn inclusions disrupt anterograde BDNF transport within nigrostriatal and corticostriatal circuitry. Further, STN DBS has the potential to exert protective effects by modifying the long-term neurodegenerative consequences of synucleinopathy.SIGNIFICANCE STATEMENTAn increase in brain-derived neurotrophic factor (BDNF) has been linked to the neuroprotection elicited by subthalamic nucleus (STN) deep brain stimulation (DBS) in neurotoxicant models of Parkinson's disease (PD). However, whether STN DBS can similarly increase BDNF in nigrostriatal and corticostriatal circuitry in the presence of α-synuclein (α-syn) inclusions has not been examined. We examined the impact of STN DBS on rats in which accumulation of α-syn inclusions is induced by injection of α-syn preformed fibrils (PFFs). STN DBS significantly increased striatal BDNF protein in rats seeded with α-syn inclusions and partially restored the normal corticostriatal BDNF relationship. These findings suggest that STN DBS can drive BDNF in the parkinsonian brain and retains the potential for neuroprotection in PD.

Misophonia is a common disorder characterized by the experience of strong negative emotions of anger and anxiety in response to certain everyday sounds, such as those generated by other people eating, drinking, and breathing. The commonplace nature of these “trigger” sounds makes misophonia a devastating disorder for sufferers and their families. How such innocuous sounds trigger this response is unknown. Since most trigger sounds are generated by orofacial movements (e.g., chewing) in others, we hypothesized that the mirror neuron system related to orofacial movements could underlie misophonia. We analyzed resting state fMRI (rs-fMRI) connectivity (N = 33, 16 females) and sound-evoked fMRI responses (N = 42, 29 females) in misophonia sufferers and controls. We demonstrate that, compared with controls, the misophonia group show no difference in auditory cortex responses to trigger sounds, but do show: (1) stronger rs-fMRI connectivity between both auditory and visual cortex and the ventral premotor cortex responsible for orofacial movements; (2) stronger functional connectivity between the auditory cortex and orofacial motor area during sound perception in general; and (3) stronger activation of the orofacial motor area, specifically, in response to trigger sounds. Our results support a model of misophonia based on “hyper-mirroring” of the orofacial actions of others with sounds being the “medium” via which action of others is excessively mirrored. Misophonia is therefore not an abreaction to sounds, per se, but a manifestation of activity in parts of the motor system involved in producing those sounds. This new framework to understand misophonia can explain behavioral and emotional responses and has important consequences for devising effective therapies. SIGNIFICANCE STATEMENT Conventionally, misophonia, literally “hatred of sounds” has been considered as a disorder of sound emotion processing, in which “simple” eating and chewing sounds produced by others cause negative emotional responses. Our data provide an alternative but complementary perspective on misophonia that emphasizes the action of the trigger-person rather than the sounds which are a byproduct of that action. Sounds, in this new perspective, are only a “medium” via which action of the triggering-person is mirrored onto the listener. This change in perspective has important consequences for devising therapies and treatment methods for misophonia. It suggests that, instead of focusing on sounds, which many existing therapies do, effective therapies should target the brain representation of movement.

Alzheimer's disease (AD) is characterized by progressive neurodegeneration in the cerebral cortex, histopathologically hallmarked by amyloid β (Aβ) extracellular plaques and intracellular neurofibrillary tangles, constituted by hyperphosphorylated tau protein. Correlation between these pathologic features and dementia has been challenged by the emergence of “nondemented with Alzheimer's neuropathology” (NDAN) individuals, cognitively intact despite displaying pathologic features of AD. The existence of these subjects suggests that some unknown mechanisms are triggered to resist Aβ-mediated detrimental events. Aβ accumulation affects mitochondrial redox balance, increasing oxidative stress status, which in turn is proposed as a primary culprit in AD pathogenesis. To clarify the relationship linking Aβ, oxidative stress, and cognitive impairment, we performed a comparative study on AD, NDAN, and aged-matched human postmortem frontal cortices of either sex. We quantitatively analyzed immunofluorescence distribution of oxidative damage markers, and of SOD2 (superoxide dismutase 2), PGC1α [peroxisome proliferator-activated receptor (PPAR) γ-coactivator 1α], PPARα, and catalase as key factors in antioxidant response, as well as the expression of miRNA-485, as a PGC1α upstream regulator. Our results confirm dramatic redox imbalance, associated with impaired antioxidant defenses in AD brain. By contrast, NDAN individuals display low oxidative damage, which is associated with high levels of scavenging systems, possibly resulting from a lack of PGC1α miRNA-485-related inhibition. Comparative analyses in neurons and astrocytes further highlighted cell-specific mechanisms to counteract redox imbalance. Overall, our data emphasize the importance of transcriptional and post-transcriptional regulation of antioxidant response in AD. This suggests that an efficient PGC1α-dependent “safety mechanism” may prevent Aβ-mediated oxidative stress, supporting neuroprotective therapies aimed at ameliorating defects in antioxidant response pathways in AD patients.

A single stressful event can cause morphologic and functional changes in neurons and even malfunction of vascular systems, which can lead to acute stress disorder or post-traumatic stress disorder. However, there is a lack of evidence regarding how acute stress impacts neuronal activity, the concurrent vascular response, and the relationship between these two factors, which is defined as neurovascular coupling. Here, usingin vivotwo-photon imaging, we found that NMDA-evoked calcium transients of excitatory neurons were impaired and that vasodilation of penetrating arterioles was concomitantly disrupted in acutely stressed male mice. Furthermore, acute stress altered the relationship between excitatory neuronal calcium coherence and vascular responses. By measuring NMDA-evoked excitatory and inhibitory neuronal calcium activity in acute brain slices, we confirmed that neuronal coherence both between excitatory neurons and between excitatory and inhibitory neurons was reduced by acute stress but restored by blockade of glucocorticoid receptor signaling. Furthermore, the ratio of sEPSCs to sIPSCs was altered by acute stress, suggesting that the excitation-inhibition balance was disrupted by acute stress. In summary,in vivo,ex vivo, and whole-cell recording studies demonstrate that acute stress modifies excitatory-inhibitory neuronal coherence, disrupts the excitation-inhibition balance, and causes consequent neurovascular coupling changes, providing critical insights into the neural mechanism of stress-induced disorders.SIGNIFICANCE STATEMENTAcute stress can cause pathologic conditions, such as acute stress disorder and post-traumatic stress disorder, by affecting the functions of neurons and blood vessels. However, investigations into the impacts of acute stress on neurovascular coupling, the tight connection between local neural activity and subsequent blood flow changes, are lacking. Through investigations at thein vivo,ex vivo, and whole-cell recording levels, we found that acute stress alters the NMDA-evoked vascular response, impairs the function and coherence of excitatory and inhibitory neurons, and disrupts the excitatory and inhibitory balance. These novel findings provide insights into the relevance of the excitatory-inhibitory balance, neuronal coherence, and neurovascular coupling to stress-induced disorders.

17β-Estradiol (E2) is produced from androgens via the action of the enzyme aromatase. E2 is known to be made in neurons in the brain, but the functions of neuron-derived E2 in the ischemic brain are unclear. Here, we used a forebrain neuron-specific aromatase KO (FBN-ARO-KO) mouse model to deplete neuron-derived E2 in the forebrain and determine its roles after global cerebral ischemia. We demonstrated that ovariectomized female FBN-ARO-KO mice exhibited significantly attenuated astrocyte activation, astrocytic aromatization, and decreased hippocampal E2 levels compared with FLOX mice. Furthermore, FBN-ARO-KO mice had exacerbated neuronal damage and worse cognitive dysfunction after global cerebral ischemia. Similar results were observed in intact male mice. RNA-seq analysis revealed alterations in pathways and genes associated with astrocyte activation, neuroinflammation, and oxidative stress in FBN-ARO-KO mice. The compromised astrocyte activation in FBN-ARO-KO mice was associated with robust downregulation of the astrocyte-derived neurotrophic factors, BDNF and IGF-1, as well as the astrocytic glutamate transporter, GLT-1. Νeuronal FGF2, which acts in a paracrine manner to suppress astrocyte activation, was increased in FBN-ARO-KO neurons. Interestingly, blocking FGF2 signaling by central injection of FGFR3-neutralizing antibody was able to reverse the diminishment in neuroprotective astrocyte reactivity, and attenuate neuronal damage in FBN-ARO-KO mice. Moreover, in vivo E2 replacement suppressed FGF2 signaling and rescued the compromised reactive astrogliosis and cognitive deficits. Collectively, our data provide novel genetic evidence for a beneficial role of neuron-derived E2 in astrocyte activation, neuroprotection, and cognitive preservation following ischemic injury to the brain. SIGNIFICANCE STATEMENT Following cerebral ischemia, astrocytes become highly reactive and can exert neuroprotection through the release of neurotrophic factors and clearance of neurotoxic glutamate. The current study advances our understanding of this process by demonstrating that neuron-derived 17β-estradiol (E2) is neuroprotective and critical for induction of reactive astrocytes and their ability to produce astrocyte-derived neurotrophic factors, BDNF and IGF-1, and the glutamate transporter, GLT-1 after ischemic brain damage. These beneficial effects of neuron-derived E2 appear to be due, at least in part, to suppression of neuronal FGF2 signaling, which is a known suppressor of astrocyte activation. These findings suggest that neuron-derived E2 is neuroprotective after ischemic brain injury via a mechanism that involves suppression of neuronal FGF2 signaling, thereby facilitating astrocyte activation.

In vivo functional and structural brain imaging of synucleinopathies in humans have provided a rich new understanding of the affected networks across the cortex and subcortex. Despite this progress, the temporal relationship between α-synuclein (α-syn) pathology and the functional and structural changes occurring in the brain is not well understood., In vivo functional and structural brain imaging of synucleinopathies in humans have provided a rich new understanding of the affected networks across the cortex and subcortex. Despite this progress, the temporal relationship between α-synuclein (α-syn) pathology and the functional and structural changes occurring in the brain is not well understood. Here, we examine the temporal relationship between locomotor ability, brain microstructure, functional brain activity, and α-syn pathology by longitudinally conducting rotarod, diffusion magnetic resonance imaging (MRI), resting-state functional MRI (fMRI), and sensory-evoked fMRI on 20 mice injected with α-syn fibrils and 20 PBS-injected mice at three timepoints (10 males and 10 females per group). Intramuscular injection of α-syn fibrils in the hindlimb of M83+/− mice leads to progressive α-syn pathology along the spinal cord, brainstem, and midbrain by 16 weeks post-injection. Our results suggest that peripheral injection of α-syn has acute systemic effects on the central nervous system such that structural and resting-state functional activity changes occur in the brain by four weeks post-injection, well before α-syn pathology reaches the brain. At 12 weeks post-injection, a separate and distinct pattern of structural and sensory-evoked functional brain activity changes was observed that are co-localized with previously reported regions of α-syn pathology and immune activation. Microstructural changes in the pons at 12 weeks post-injection were found to predict survival time and preceded measurable locomotor deficits. This study provides preliminary evidence for diffusion and fMRI markers linked to the progression of synuclein pathology and has translational importance for understanding synucleinopathies in humans. SIGNIFICANCE STATEMENT α-Synuclein (α-syn) pathology plays a critical role in neurodegenerative diseases such as Parkinson's disease, dementia with Lewy bodies, and multiple system atrophy. The longitudinal effects of α-syn pathology on locomotion, brain microstructure, and functional brain activity are not well understood. Using high field imaging, we show preliminary evidence that peripheral injection of α-syn fibrils induces unique patterns of functional and structural changes that occur at different temporal stages of α-syn pathology progression. Our results challenge existing assumptions that α-syn pathology must precede changes in brain structure and function. Additionally, we show preliminary evidence that diffusion and functional magnetic resonance imaging (fMRI) are capable of resolving such changes and thus should be explored further as markers of disease progression.

To investigate the role of neuronal NF-κB activity in pathogenesis of amyotrophic lateral sclerosis (ALS), we generated transgenic mice with neuron-specific expression of a super-repressor form of the NF-κB inhibitor (IκBα-SR), which were then crossed with mice of both sexes, expressing ALS-linked gene mutants for TAR DNA-binding protein (TDP-43) and superoxide dismutase 1 (SOD1). Remarkably, neuronal expression of IκBα-SR transgene in mice expressing TDP-43A315T or TDP-43G348C mice led to a decrease in cytoplasmic to nuclear ratio of human TDP-43. The mitigation of TDP-43 neuropathology by IκBα-SR, which is likely due to an induction of autophagy, was associated with amelioration of cognitive and motor deficits as well as reduction of motor neuron loss and gliosis. Neuronal suppression of NF-κB activity in SOD1G93A mice also resulted in neuroprotection with reduction of misfolded SOD1 levels and significant extension of life span. The results suggest that neuronal NF-κB signaling constitutes a novel therapeutic target for ALS disease and related disorders with TDP-43 proteinopathy. SIGNIFICANCE STATEMENT This study reports that neuron-specific expression of IκB super-repressor mitigated behavioral and pathologic changes in transgenic mouse models of amyotrophic lateral sclerosis expressing mutant forms of either Tar DNA-binding protein 43 or superoxide dismutase. The results suggest that neuronal NF-κB signaling constitutes a novel therapeutic target for amyotrophic lateral sclerosis and related disorders with Tar DNA-binding protein 43 proteinopathy.

Krabbe's disease is an infantile neurodegenerative disease, which is affected by mutations in the lysosomal enzyme galactocerebrosidase, leading to the accumulation of its metabolite psychosine. We have shown previously that the S1P receptor agonist fingolimod (FTY720) attenuates psychosine-induced glial cell death and demyelination both in vitro and ex vivo models., Krabbe's disease is an infantile neurodegenerative disease, which is affected by mutations in the lysosomal enzyme galactocerebrosidase, leading to the accumulation of its metabolite psychosine. We have shown previously that the S1P receptor agonist fingolimod (FTY720) attenuates psychosine-induced glial cell death and demyelination both in vitro and ex vivo models. These data, together with a lack of therapies for Krabbe's disease, prompted the current preclinical study examining the effects of fingolimod in twitcher mice, a murine model of Krabbe's disease. Twitcher mice, both male and female, carrying a natural mutation in the galc gene were given fingolimod via drinking water (1 mg/kg/d). The direct impact of fingolimod administration was assessed via histochemical and biochemical analysis using markers of myelin, astrocytes, microglia, neurons, globoid cells, and immune cells. The effects of fingolimod on twitching behavior and life span were also demonstrated. Our results show that treatment of twitcher mice with fingolimod significantly rescued myelin levels compared with vehicle-treated animals and also regulated astrocyte and microglial reactivity. Furthermore, nonphosphorylated neurofilament levels were decreased, indicating neuroprotective and neurorestorative processes. These protective effects of fingolimod on twitcher mice brain pathology was reflected by an increased life span of fingolimod-treated twitcher mice. These in vivo findings corroborate initial in vitro studies and highlight the potential use of S1P receptors as drug targets for treatment of Krabbe's disease. SIGNIFICANCE STATEMENT This study demonstrates that the administration of the therapy known as fingolimod in a mouse model of Krabbe's disease (namely, the twitcher mouse model) significantly rescues myelin levels. Further, the drug fingolimod also regulates the reactivity of glial cells, astrocytes and microglia, in this mouse model. These protective effects of fingolimod result in an increased life span of twitcher mice.

Neurofibrillary tangles likely cause neurodegeneration in Alzheimer's disease (AD). We demonstrate that the CX3CL1 C-terminal domain can upregulate neurogenesis, which may ameliorate neurodegeneration. Here we generated transgenic (Tg-CX3CL1) mice by overexpressing CX3CL1 in neurons. Tg-CX3CL1 mice exhibit enhanced neurogenesis in both subgranular and subventricular zones. This enhanced neurogenesis correlates well with elevated expression of TGF-β2 and TGF-β3, and activation of their downstream signaling molecule Smad2. Intriguingly, the enhanced adult neurogenesis was mitigated when Smad2 expression was deleted in neurons, supporting a role for the CX3CL1–TGF-β2/3–Smad2 pathway in the control of adult neurogenesis. When Tg-CX3CL1 mice were crossed with Alzheimer's PS19 mice, which overexpress a tau P301S mutation and exhibit age-dependent neurofibrillary tangles and neurodegeneration, overexpressed CX3CL1 in both male and female mice was sufficient to rescue the neurodegeneration, increase survival time, and improve cognitive function. Hence, we providein vivoevidence that CX3CL1 is a strong activator of adult neurogenesis, and that it reduces neuronal loss and improves cognitive function in AD.SIGNIFICANCE STATEMENTThis study will be the first to demonstrate that enhanced neurogenesis by overexpressed CX3CL1 is mitigated by disruption of Smad2 signaling and is independent of its interaction with CX3CR1. Overexpression of CX3CL1 lengthens the life span of PS19 tau mice by enhancing adult neurogenesis while having minimal effect on tau pathology. Enhancing neuronal CX3CL1, mainly the C-terminal fragment, is a therapeutic strategy for blocking or reversing neuronal loss in Alzheimer's disease or related neurodegenerative disease patients.

Memory deficits are common in epilepsy patients. In these patients, the interictal EEG commonly shows interictal epileptiform discharges (IEDs). While IEDs are associated with transient cognitive impairments, it remains poorly understood why this is. We investigated the effects of human (male and female) hippocampal IEDs on single-neuron activity during a memory task in patients with medically refractory epilepsy undergoing depth electrode monitoring. We quantified the effects of hippocampal IEDs on single-neuron activity and the impact of this modulation on subjectively declared memory strength. Across all recorded neurons, the activity of 50 of 728 neurons were significantly modulated by IEDs, with the strongest modulation in the medial temporal lobe (33 of 416) and in particular the right hippocampus (12 of 58). Putative inhibitory neurons, as identified by their extracellular signature, were more likely to be modulated by IEDs than putative excitatory neurons (19 of 157 vs 31 of 571). Behaviorally, the occurrence of hippocampal IEDs was accompanied by a disruption of recognition of familiar images only if they occurred up to 2 s before stimulus onset. In contrast, IEDs did not impair encoding or recognition of novel images, indicating high temporal and task specificity of the effects of IEDs. The degree of modulation of individual neurons by an IED correlated with the declared confidence of a retrieval trial, with higher firing rates indicative of reduced confidence. Together, these data link the transient modulation of individual neurons by IEDs to specific declarative memory deficits in specific cell types, thereby revealing a mechanism by which IEDs disrupt medial temporal lobe-dependent declarative memory retrieval processes.SIGNIFICANCE STATEMENTInterictal epileptiform discharges (IEDs) are thought to be a cause of memory deficits in chronic epilepsy patients, but the underlying mechanisms are not understood. Utilizing single-neuron recordings in epilepsy patients, we found that hippocampal IEDs transiently change firing of hippocampal neurons and disrupted selectively the retrieval, but not encoding, of declarative memories. The extent of the modulation of the individual firing of hippocampal neurons by an IED predicted the extent of reduction of subjective retrieval confidence. Together, these data reveal a specific kind of transient cognitive impairment caused by IEDs and link this impairment to the modulation of the activity of individual neurons. Understanding the mechanisms by which IEDs impact memory is critical for understanding memory impairments in epilepsy patients.

TREM2 is an Alzheimer's disease (AD) risk gene expressed in microglia. To study the role of Trem2 in a mouse model of β-amyloidosis, we compared PS2APP transgenic mice versus PS2APP mice lacking Trem2 (PS2APP;Trem2ko) at ages ranging from 4 to 22 months. Microgliosis was impaired in PS2APP;Trem2ko mice, with Trem2-deficient microglia showing compromised expression of proliferation/Wnt-related genes and marked accumulation of ApoE., TREM2 is an Alzheimer's disease (AD) risk gene expressed in microglia. To study the role of Trem2 in a mouse model of β-amyloidosis, we compared PS2APP transgenic mice versus PS2APP mice lacking Trem2 (PS2APP;Trem2ko) at ages ranging from 4 to 22 months. Microgliosis was impaired in PS2APP;Trem2ko mice, with Trem2-deficient microglia showing compromised expression of proliferation/Wnt-related genes and marked accumulation of ApoE. Plaque abundance was elevated in PS2APP;Trem2ko females at 6–7 months; but by 12 or 19–22 months of age, it was notably diminished in female and male PS2APP;Trem2ko mice, respectively. Across all ages, plaque morphology was more diffuse in PS2APP;Trem2ko brains, and the Aβ42:Aβ40 ratio was elevated. The amount of soluble, fibrillar Aβ oligomers also increased in PS2APP;Trem2ko hippocampi. Associated with these changes, axonal dystrophy was exacerbated from 6 to 7 months onward in PS2APP;Trem2ko mice, notwithstanding the reduced plaque load at later ages. PS2APP;Trem2ko mice also exhibited more dendritic spine loss around plaque and more neurofilament light chain in CSF. Thus, aggravated neuritic dystrophy is a more consistent outcome of Trem2 deficiency than amyloid plaque load, suggesting that the microglial packing of Aβ into dense plaque is an important neuroprotective activity. SIGNIFICANCE STATEMENT Genetic studies indicate that TREM2 gene mutations confer increased Alzheimer's disease (AD) risk. We studied the effects of Trem2 deletion in the PS2APP mouse AD model, in which overproduction of Aβ peptide leads to amyloid plaque formation and associated neuritic dystrophy. Interestingly, neuritic dystrophies were intensified in the brains of Trem2-deficient mice, despite these mice displaying reduced plaque accumulation at later ages (12–22 months). Microglial clustering around plaques was impaired, plaques were more diffuse, and the Aβ42:Aβ40 ratio and amount of soluble, fibrillar Aβ oligomers were elevated in Trem2-deficient brains. These results suggest that the Trem2-dependent compaction of Aβ into dense plaques is a protective microglial activity, limiting the exposure of neurons to toxic Aβ species.

Systemic infection triggers a spectrum of metabolic and behavioral changes, collectively termed sickness behavior, which while adaptive, can affect mood and cognition. In vulnerable individuals, acute illness can also produce profound, maladaptive, cognitive dysfunction including delirium, but our understanding of delirium pathophysiology remains limited. Here, we used bacterial lipopolysaccharide (LPS) in female C57BL/6J mice and acute hip fracture in humans to address whether disrupted energy metabolism contributes to inflammation-induced behavioral and cognitive changes. LPS (250 µg/kg) induced hypoglycemia, which was mimicked by interleukin (IL)-1β (25 µg/kg) but not prevented in IL-1RI−/− mice, nor by IL-1 receptor antagonist (IL-1RA; 10 mg/kg). LPS suppression of locomotor activity correlated with blood glucose concentrations, was mitigated by exogenous glucose (2 g/kg), and was exacerbated by 2-deoxyglucose (2-DG) glycolytic inhibition, despite preventing IL-1β synthesis. Using the ME7 model of chronic neurodegeneration in female mice, to examine vulnerability of the diseased brain to acute stressors, we showed that LPS (100 µg/kg) produced acute cognitive dysfunction, selectively in those animals. These acute cognitive impairments were mimicked by insulin (11.5 IU/kg) and mitigated by glucose, demonstrating that acutely reduced glucose metabolism impairs cognition selectively in the vulnerable brain. To test whether these acute changes might predict altered carbohydrate metabolism during delirium, we assessed glycolytic metabolite levels in CSF in humans during inflammatory trauma-induced delirium. Hip fracture patients showed elevated CSF lactate and pyruvate during delirium, consistent with acutely altered brain energy metabolism. Collectively, the data suggest that disruption of energy metabolism drives behavioral and cognitive consequences of acute systemic inflammation. SIGNIFICANCE STATEMENT Acute systemic inflammation alters behavior and produces disproportionate effects, such as delirium, in vulnerable individuals. Delirium has serious short and long-term sequelae but mechanisms remain unclear. Here, we show that both LPS and interleukin (IL)-1β trigger hypoglycemia, reduce CSF glucose, and suppress spontaneous activity. Exogenous glucose mitigates these outcomes. Equivalent hypoglycemia, induced by lipopolysaccharide (LPS) or insulin, was sufficient to trigger cognitive impairment selectively in animals with existing neurodegeneration and glucose also mitigated those impairments. Patient CSF from inflammatory trauma-induced delirium also shows altered brain carbohydrate metabolism. The data suggest that the degenerating brain is exquisitely sensitive to acute behavioral and cognitive consequences of disrupted energy metabolism. Thus “bioenergetic stress” drives systemic inflammation-induced dysfunction. Elucidating this may offer routes to mitigating delirium.

Neurovascular coupling (NVC), the interaction between neural activity and vascular response, ensures normal brain function by maintaining brain homeostasis. We previously reported altered cerebrovascular responses during functional hyperemia in chronically stressed animals. However, the underlying neuronal-level changes associated with those hemodynamic changes remained unclear. Here, usingin vivoandex vivoexperiments, we investigate the neuronal origins of altered NVC dynamics under chronic stress conditions in adult male mice. Stimulus-evoked hemodynamic and neural responses, especially beta and gamma-band local field potential activity, were significantly lower in chronically stressed animals, and the NVC relationship, itself, had changed. Further, using acute brain slices, we discovered that the underlying cause of this change was dysfunction of neuronal nitric oxide synthase (nNOS)-mediated vascular responses. Using FISH to check the mRNA expression of several GABAergic subtypes, we confirmed that only nNOS mRNA was significantly decreased in chronically stressed mice. Ultimately, chronic stress impairs NVC by diminishing nNOS-mediated vasodilation responses to local neural activity. Overall, these findings provide useful information in understanding NVC dynamics in the healthy brain. More importantly, this study reveals that impaired nNOS-mediated NVC function may be a contributory factor in the progression of stress-related diseases.SIGNIFICANCE STATEMENTThe correlation between neuronal activity and cerebral vascular dynamics is defined as neurovascular coupling (NVC), which plays an important role for meeting the metabolic demands of the brain. However, the impact of chronic stress, which is a contributory factor of many cerebrovascular diseases, on NVC is poorly understood. We therefore investigated the effects of chronic stress on impaired neurovascular response to sensory stimulation and their underlying mechanisms. Multimodal approaches, fromin vivohemodynamic imaging and electrophysiology toex vivovascular imaging with pharmacological treatment, patch-clamp recording, FISH, and immunohistochemistry revealed that chronic stress-induced dysfunction of nNOS-expressing interneurons contributes to NVC impairment. These findings will provide useful information to understand the role of nNOS interneurons in NVC in normal and pathological conditions.

Microglia are resident myeloid cells of the CNS. Recently, single-cell RNA sequencing (scRNAseq) has enabled description of a disease-associated microglia (DAM) with a role in neurodegeneration and demyelination. In this study, we use scRNAseq to investigate the temporal dynamics of immune cells harvested from the epicenter of traumatic spinal cord injury (SCI) induced in female mice. We find that as a consequence of SCI, baseline microglia undergo permanent transcriptional reprogramming into a previously uncharacterized subtype of microglia with striking similarities to previously reported DAM as well as a distinct microglial state found during development. Using a microglia depletion model we showed that DAM in SCI are derived from baseline microglia and strongly enhance recovery of hindlimb locomotor function following injury. SIGNIFICANCE STATEMENT Although disease-associated microglia (DAM) have been the subject of strong research interest during recent years (Keren-Shaul, 2017; Jordão, 2019), their cellular origin and their role in “normal” acute injury processes is not well understood. Our work directly addresses the origin and the role of DAM in traumatic injury response. Further, we use a microglia depletion model to prove that DAM in spinal cord injury (SCI) are indeed derived from homeostatic microglia, and that they strongly enhance recovery. Thus, in this work we significantly expand the knowledge of immune response to traumatic injury, demonstrate the applicability to human injury via our unique access to injured human spinal cord tissue, and provide the community with a comprehensive dataset for further exploration.

The expansion of a polyglutamine tract in the ubiquitously expressed huntingtin protein causes Huntington's disease (HD), a dominantly inherited neurodegenerative disease. We show that the activity of the cholesterol biosynthetic pathway is altered in HD. In particular, the transcription of key genes of the cholesterol biosynthetic pathway is severely affected in vivo in brain tissue from HD mice and in human postmortem striatal and cortical tissue; this molecular dysfunction is biologically relevant because cholesterol biosynthesis is reduced in cultured human HD cells, and total cholesterol mass is significantly decreased in the CNS of HD mice and in brain-derived ST14A cells in which the expression of mutant huntingtin has been turned on. The transcription of the genes of the cholesterol biosynthetic pathway is regulated via the activity of sterol regulatory element-binding proteins (SREBPs), and we found an ∼50% reduction in the amount of the active nuclear form of SREBP in HD cells and mouse brain tissue. As a consequence, mutant huntingtin reduces the transactivation of an SRE-luciferase construct even under conditions of SREBP overexpression or in the presence of an exogenous N-terminal active form of SREBP. Finally, the addition of exogenous cholesterol to striatal neurons expressing mutant huntingtin prevents their death in a dose-dependent manner. We conclude that the cholesterol biosynthetic pathway is impaired in HD cells, mice, and human subjects, and that the search for HD therapies should also consider cholesterol levels as both a potential target and disease biomarker.

Patients with advanced Parkinson's can be treated by deep brain stimulation (DBS) of the subthalamic nucleus (STN). This affords a unique opportunity to record from this nucleus and stimulate it in a controlled manner. Previous work has shown that activity in the STN is modulated in a rhythmic pattern when Parkinson's patients perform stepping movements, raising the question whether the STN is involved in the dynamic control of stepping., Patients with advanced Parkinson's can be treated by deep brain stimulation (DBS) of the subthalamic nucleus (STN). This affords a unique opportunity to record from this nucleus and stimulate it in a controlled manner. Previous work has shown that activity in the STN is modulated in a rhythmic pattern when Parkinson's patients perform stepping movements, raising the question whether the STN is involved in the dynamic control of stepping. To answer this question, we tested whether an alternating stimulation pattern resembling the stepping-related modulation of activity in the STN could entrain patients' stepping movements as evidence of the STN's involvement in stepping control. Group analyses of 10 Parkinson's patients (one female) showed that alternating stimulation significantly entrained stepping rhythms. We found a remarkably consistent alignment between the stepping and stimulation cycle when the stimulation speed was close to the stepping speed in the five patients that demonstrated significant individual entrainment to the stimulation cycle. Our study suggests that the STN is causally involved in dynamic control of step timing and motivates further exploration of this biomimetic stimulation pattern as a potential basis for the development of DBS strategies to ameliorate gait impairments. SIGNIFICANCE STATEMENT We tested whether the subthalamic nucleus (STN) in humans is causally involved in controlling stepping movements. To this end, we studied patients with Parkinson's disease who have undergone therapeutic deep brain stimulation (DBS), as in these individuals we can stimulate the STNs in a controlled manner. We developed an alternating pattern of stimulation that mimics the pattern of activity modulation recorded in this nucleus during stepping. The alternating DBS (altDBS) could entrain patients' stepping rhythm, suggesting a causal role of the STN in dynamic gait control. This type of stimulation may potentially form the basis for improved DBS strategies for gait.

Pathologic tau modifications are characteristic of Alzheimer's disease and related dementias, but mechanisms of tau toxicity continue to be debated. Inherited mutations in tau cause early onset frontotemporal lobar dementias (FTLD-tau) and are commonly used to model mechanisms of tau toxicity in tauopathies. Previous work in the isolated squid axoplasm model demonstrated that several pathogenic forms of tau inhibit axonal transport through a mechanism involving activation of protein phosphatase 1 (PP1). Here, we determined that P301L and R5L FTLD mutant tau proteins elicit a toxic effect on axonal transport as monomeric proteins. We evaluated interactions of wild-type or mutant tau with specific PP1 isoforms (α, β, and γ) to examine how the interaction contributes to this toxic effect using primary rat hippocampal neurons from both sexes. Pull-down and bioluminescence resonance energy transfer experiments revealed selective interactions of wild-type tau with PP1α and PP1γ isoforms, but not PP1β, which were significantly increased by the P301L tau mutation. The results from proximity ligation assays confirmed the interaction in primary hippocampal neurons. Moreover, expression of FTLD-linked mutant tau in these neurons enhanced levels of active PP1, also increasing the pausing frequency of fluorescently labeled vesicles in both anterograde and retrograde directions. Knockdown of PP1γ, but not PP1α, rescued the cargo-pausing effects of P301L and R5L tau, a result replicated by deleting a phosphatase-activating domain in the amino terminus of P301L tau. These findings support a model of tau toxicity involving aberrant activation of a specific PP1γ-dependent pathway that disrupts axonal transport in neurons.SIGNIFICANCE STATEMENTTau pathology is closely associated with neurodegeneration in Alzheimer's disease and other tauopathies, but the toxic mechanisms remain a debated topic. We previously proposed that pathologic tau forms induce dysfunction and degeneration through aberrant activation of a PP1-dependent pathway that disrupts axonal transport. Here, we show that tau directly interacts with specific PP1 isoforms, increasing levels of active PP1. Pathogenic tau mutations enhance this interaction, further increasing active PP1 levels and impairing axonal transport in isolated squid axoplasm and primary hippocampal neurons. Mutant-tau-mediated impairment of axonal transport was mediated by PP1γ and a phosphatase-activating domain located at the amino terminus of tau. This work has important implications for understanding and potentially mitigating tau-mediated neurotoxicity in tauopathies.

We tested the popular, unproven theory that tinnitus is caused by resetting of auditory predictions toward a persistent low-intensity sound. Electroencephalographic mismatch negativity responses, which quantify the violation of sensory predictions, to unattended tinnitus-like sounds were greater in response to upward than downward intensity deviants in 26 unselected chronic tinnitus subjects with normal to severely impaired hearing, and in 15 acute tinnitus subjects, but not in 26 hearing and age-matched controls (p< 0.001, receiver operator characteristic, area under the curve, 0.77), or in 20 healthy and hearing-impaired controls presented with simulated tinnitus. The findings support a prediction resetting model of tinnitus generation, and may form the basis of a convenient tinnitus biomarker, which we name Intensity Mismatch Asymmetry, which is usable across species, is quick and tolerable, and requires no training.SIGNIFICANCE STATEMENTIn current models, perception is based around the generation of internal predictions of the environment, which are tested and updated using evidence from the senses. Here, we test the theory that auditory phantom perception (tinnitus) occurs when a default auditory prediction is formed to explain spontaneous activity in the subcortical pathway, rather than ignoring it as noise. We find that chronic tinnitus patients show an abnormal pattern of evoked responses to unexpectedly loud and quiet sounds that both supports this hypothesis and provides fairly accurate classification of tinnitus status at the individual subject level. This approach to objectively demonstrating the predictions underlying pathological perceptual states may also have a much wider utility, for instance, in chronic pain.

Brainstem locus ceruleus neurons (LCn) are among the first neurons across the lifespan to evidence tau pathology, and LCn are implicated in tau propagation throughout the cortices. Yet, events influencing LCn tau are poorly understood. Activated persistently across wakefulness, LCn experience significant metabolic stress in response to chronic short sleep (CSS). Here we explored whether CSS influences LCn tau and the biochemical, neuroanatomical, and/or behavioral progression of tauopathy in male and female P301S mice. CSS in early adult life advanced the temporal progression of neurobehavioral impairments and resulted in a lasting increase in soluble tau oligomers. Intriguingly, CSS resulted in an early increase in AT8 and MC1 tau pathology in the LC. Over time tau pathology, including tangles, was evident in forebrain tau-vulnerable regions. Sustained microglial and astrocytic activation was observed as well. Remarkably, CSS resulted in significant loss of neurons in the two regions examined: the basolateral amygdala and LC. A second, distinct form of chronic sleep disruption, fragmentation of sleep, during early adult life also increased tau deposition and imparted early neurobehavioral impairment. Collectively, the findings demonstrate that early life sleep disruption has important lasting effects on the temporal progression in P301S mice, influencing tau pathology and hastening neurodegeneration, neuroinflammation, and neurobehavioral impairments.SIGNIFICANCE STATEMENTChronic short sleep (CSS) is pervasive in modern society. Here, we found that early life CSS influences behavioral, biochemical, and neuroanatomic aspects of the temporal progression of tauopathy in a mouse model of the P301S tau mutation. Specifically, CSS hastened the onset of motor impairment and resulted in a greater loss of neurons in both the locus ceruleus and basolateral/lateral amygdala. Importantly, despite a protracted recovery opportunity after CSS, mice evidenced a sustained increase in pathogenic tau oligomers, and increased pathogenic tau in the locus ceruleus and limbic system nuclei. These findings unveil early life sleep habits as an important determinant in the progression of tauopathy.

Traumatic brain injury (TBI) patients often exhibit slowed information processing speed that can underlie diverse symptoms. Processing speed depends on neural circuit function at synapses, in the soma, and along axons. Long axons in white matter (WM) tracts are particularly vulnerable to TBI. We hypothesized that disrupted axon–myelin interactions that slow or block action potential conduction in WM tracts may contribute to slowed processing speed after TBI. Concussive TBI in male/female mice was used to produce traumatic axonal injury in the corpus callosum (CC), similar to WM pathology in human TBI cases. Compound action potential velocity was slowed along myelinated axons at 3 d after TBI with partial recovery by 2 weeks, suggesting early demyelination followed by remyelination. Ultrastructurally, dispersed demyelinated axons and disorganized myelin attachment to axons at paranodes were apparent within CC regions exhibiting traumatic axonal injury. Action potential conduction is exquisitely sensitive to paranode abnormalities. Molecular identification of paranodes and nodes of Ranvier detected asymmetrical paranode pairs and abnormal heminodes after TBI. Fluorescent labeling of oligodendrocyte progenitors inNG2CreER;mTmGmice showed increased synthesis of new membranes extended along axons to paranodes, indicating remyelination after TBI. At later times after TBI, an overall loss of conducting axons was observed at 6 weeks followed by CC atrophy at 8 weeks. These studies identify a progression of both myelinated axon conduction deficits and axon–myelin pathology in the CC, implicating WM injury in impaired information processing at early and late phases after TBI. Furthermore, the intervening recovery reveals a potential therapeutic window.SIGNIFICANCE STATEMENTTraumatic brain injury (TBI) is a major global health concern. Across the spectrum of TBI severities, impaired information processing can contribute to diverse functional deficits that underlie persistent symptoms. We used experimental TBI to exploit technical advantages in mice while modeling traumatic axonal injury in white matter tracts, which is a key pathological feature of human TBI. A combination of approaches revealed slowed and failed signal conduction along with damage to the structure and molecular composition of myelinated axons in the white matter after TBI. An early regenerative response was not sustained yet reveals a potential time window for intervention. These insights into white matter abnormalities underlying axon conduction deficits can inform strategies to improve treatment options for TBI patients.

Alterations in motivated behavior are a hallmark of attention-deficit/hyperactivity disorder (ADHD), one of the most common psychiatric disorders in children and adolescents. The orbitofrontal cortex (OFC) plays a key role in controlling goal-directed behavior, but the link between OFC dysfunction and behavioral deficits in ADHD, particularly in adolescence, remains poorly understood. Here we used advanced high-resolution functional magnetic resonance imaging (fMRI) of the human OFC in adolescents with ADHD and typically developing (TD) controls (N = 39, age 12–16, all male except for one female per group) to study reward-related OFC responses and how they relate to behavioral dysfunction in ADHD. During fMRI data acquisition, participants performed a simple decision-making task, allowing us to image expectation-related responses to small and large monetary outcomes. Across all participants, we observed significant signal increases to large versus small expected rewards in the OFC. These responses were significantly enhanced in ADHD relative to TD participants. Moreover, stronger reward-related activity was correlated with individual differences in hyperactive/impulsive symptoms in the ADHD group, whereas high cognitive ability was associated with normalized OFC responses. These results provide evidence for the importance of OFC dysfunctions in the neuropathology of ADHD, highlighting the role of OFC-dependent goal-directed control mechanisms in this disorder. SIGNIFICANCE STATEMENT Attention-deficit/hyperactivity disorder (ADHD) is characterized by alterations in motivated behavior which can be understood as diminished goal-directed control. The orbitofrontal cortex (OFC) plays a key role in controlling goal-directed behavior, but its potential contribution to ADHD symptomatology remains poorly understood. Using high-resolution fMRI, we show that adolescent ADHD patients display enhanced OFC signaling of future rewards and that these increased reward-related responses are correlated with the severity of hyperactivity/impulsivity. These findings suggest that an inability to adequately evaluate future outcomes may translate into maladaptive behavior in ADHD patients. They also challenge the idea that dysfunctions in dopaminergic brain areas are the sole contributor to reward-related symptoms in ADHD and point to a central contribution of goal-directed control circuits in hyperactivity.

Gait disturbances in Parkinson's disease are commonly refractory to current treatment options and majorly impair patient's quality of life. Auditory cues facilitate gait and prevent motor blocks. We investigated how neural dynamics in the human subthalamic nucleus of Parkinsons's disease patients (14 male, 2 female) vary during stepping and whether rhythmic auditory cues enhance the observed modulation. Oscillations in the beta band were suppressed after ipsilateral heel strikes, when the contralateral foot had to be raised, and reappeared after contralateral heel strikes, when the contralateral foot rested on the floor. The timing of this 20–30 Hz beta modulation was clearly distinct between the left and right subthalamic nucleus, and was alternating within each stepping cycle. This modulation was similar, whether stepping movements were made while sitting, standing, or during gait, confirming the utility of the stepping in place paradigm. During stepping in place, beta modulation increased with auditory cues that assisted patients in timing their steps more regularly. Our results suggest a link between the degree of power modulation within high beta frequency bands and stepping performance. These findings raise the possibility that alternating deep brain stimulation patterns may be superior to constant stimulation for improving parkinsonian gait.SIGNIFICANCE STATEMENTGait disturbances in Parkinson's disease majorly reduce patients' quality of life and are often refractory to current treatment options. We investigated how neural activity in the subthalamic nucleus of patients who received deep brain stimulation surgery covaries with the stepping cycle. 20–30 Hz beta activity was modulated relative to each step, alternating between the left and right STN. The stepping performance of patients improved when auditory cues were provided, which went along with enhanced beta modulation. This raises the possibility that alternating stimulation patterns may also enhance beta modulation and may be more beneficial for gait control than continuous stimulation, which needs to be tested in future studies.

Hippocampal overexpression of FK506-binding protein 12.6/1b (FKBP1b), a negative regulator of ryanodine receptor Ca2+release, reverses aging-induced memory impairment and neuronal Ca2+dysregulation. Here, we tested the hypothesis thatFKBP1balso can protect downstream transcriptional networks from aging-induced dysregulation. We gave hippocampal microinjections ofFKBP1b-expressing viral vector to male rats at either 13 months of age (long-term, LT) or 19 months of age (short-term, ST) and tested memory performance in the Morris water maze at 21 months of age. Aged rats treated ST or LT withFKBP1bsubstantially outperformed age-matched vector controls and performed similarly to each other and young controls (YCs). Transcriptional profiling in the same animals identified 2342 genes with hippocampal expression that was upregulated/downregulated in aged controls (ACs) compared with YCs (the aging effect). Of these aging-dependent genes, 876 (37%) also showed altered expression in agedFKBP1b-treated rats compared with ACs, withFKBP1brestoring expression of essentially all such genes (872/876, 99.5%) in the direction opposite the aging effect and closer to levels in YCs. This inverse relationship between the aging andFKBP1beffects suggests that the aging effects arise fromFKBP1bdeficiency. Functional category analysis revealed that genes downregulated with aging and restored byFKBP1bwere associated predominantly with diverse brain structure categories, including cytoskeleton, membrane channels, and extracellular region. Conversely, genes upregulated with aging but not restored byFKBP1bassociated primarily with glial–neuroinflammatory, ribosomal, and lysosomal categories. Immunohistochemistry confirmed aging-induced rarefaction andFKBP1b-mediated restoration of neuronal microtubular structure. Therefore, a previously unrecognized genomic network modulating diverse brain structural processes is dysregulated by aging and restored byFKBP1boverexpression.SIGNIFICANCE STATEMENTPreviously, we found that hippocampal overexpression of FK506-binding protein 12.6/1b (FKBP1b), a negative regulator of intracellular Ca2+responses, reverses both aging-related Ca2+dysregulation and cognitive impairment. Here, we tested whether hippocampalFKBP1boverexpression also counteracts aging changes in gene transcriptional networks. In addition to reducing memory deficits in aged rats,FKBP1bselectively counteracted aging-induced expression changes in 37% of aging-dependent genes, with cytoskeletal and extracellular structure categories highly associated with theFKBP1b-rescued genes. Our results indicate that, in parallel with cognitive processes, a novel transcriptional network coordinating brain structural organization is dysregulated with aging and restored byFKBP1b.

Dendritic spine loss is recognized as an early feature of Alzheimer's disease (AD), but the underlying mechanisms are poorly understood. Dendritic spine structure is defined by filamentous actin (F-actin) and we observed depolymerization of synaptosomal F-actin accompanied by increased globular-actin (G-actin) at as early as 1 month of age in a mouse model of AD (APPswe/PS1ΔE9, male mice). This led to recall deficit after contextual fear conditioning (cFC) at 2 months of age in APPswe/PS1ΔE9 male mice, which could be reversed by the actin-polymerizing agent jasplakinolide. Further, the F-actin-depolymerizing agent latrunculin induced recall deficit after cFC in WT mice, indicating the importance of maintaining F-/G-actin equilibrium for optimal behavioral response. Using direct stochastic optical reconstruction microscopy (dSTORM), we show that F-actin depolymerization in spines leads to a breakdown of the nano-organization of outwardly radiating F-actin rods in cortical neurons from APPswe/PS1ΔE9 mice. Our results demonstrate that synaptic dysfunction seen as F-actin disassembly occurs very early, before onset of pathological hallmarks in AD mice, and contributes to behavioral dysfunction, indicating that depolymerization of F-actin is causal and not consequent to decreased spine density. Further, we observed decreased synaptosomal F-actin levels in postmortem brain from mild cognitive impairment and AD patients compared with subjects with normal cognition. F-actin decrease correlated inversely with increasing AD pathology (Braak score, Aβ load, and tangle density) and directly with performance in episodic and working memory tasks, suggesting its role in human disease pathogenesis and progression.SIGNIFICANCE STATEMENTSynaptic dysfunction underlies cognitive deficits in Alzheimer's disease (AD). The cytoskeletal protein actin plays a critical role in maintaining structure and function of synapses. Using cultured neurons and an AD mouse model, we show for the first time that filamentous actin (F-actin) is lost selectively from synapses early in the disease process, long before the onset of classical AD pathology. We also demonstrate that loss of synaptic F-actin contributes directly to memory deficits. Loss of synaptosomal F-actin in human postmortem tissue correlates directly with decreased performance in memory test and inversely with AD pathology. Our data highlight that synaptic cytoarchitectural changes occur early in AD and they may be targeted for the development of therapeutics.

Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels and the Ih current they generate contribute to the pathophysiological mechanisms of absence seizures (ASs), but their precise role in neocortical and thalamic neuronal populations, the main components of the network underlying AS generation, remains controversial. In diverse genetic AS models, Ih amplitude is smaller in neocortical neurons and either larger or unchanged in thalamocortical (TC) neurons compared with nonepileptic strains. A lower expression of neocortical HCN subtype 1 channels is present in genetic AS-prone rats, and HCN subtype 2 knock-out mice exhibit ASs. Furthermore, whereas many studies have characterized Ih contribution to “absence-like” paroxysmal activity in vitro, no data are available on the specific role of cortical and thalamic HCN channels in behavioral seizures. Here, we show that the pharmacological block of HCN channels with the antagonist ZD7288 applied via reverse microdialysis in the ventrobasal thalamus (VB) of freely moving male Genetic Absence Epilepsy Rats from Strasbourg decreases TC neuron firing and abolishes spontaneous ASs. A similar effect is observed on γ-hydroxybutyric acid-elicited ASs in normal male Wistar rats. Moreover, thalamic knockdown of HCN channels via virally delivered shRNA into the VB of male Stargazer mice, another genetic AS model, decreases spontaneous ASs and Ih-dependent electrophysiological properties of VB TC neurons. These findings provide the first evidence that block of TC neuron HCN channels prevents ASs and suggest that any potential anti-absence therapy that targets HCN channels should carefully consider the opposite role for cortical and thalamic Ih in the modulation of absence seizures. SIGNIFICANCE STATEMENT Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels play critical roles in the fine-tuning of cellular and network excitability and have been suggested to be a key element of the pathophysiological mechanism underlying absence seizures. However, the precise contribution of HCN channels in neocortical and thalamic neuronal populations to these nonconvulsive seizures is still controversial. In the present study, pharmacological block and genetic suppression of HCN channels in thalamocortical neurons in the ventrobasal thalamic nucleus leads to a marked reduction in absence seizures in one pharmacological and two genetic rodent models of absence seizures. These results provide the first evidence that block of TC neuron HCN channels prevents absence seizures.

Acoustic overexposure, such as listening to loud music too often, results in noise-induced hearing loss. The pathologies of this prevalent sensory disorder begin within the ear at synapses of the primary auditory receptors, their postsynaptic partners and their supporting cells. The extent of noise-induced damage, however, is determined by overstimulation of primary auditory receptors, upstream of where the pathologies manifest., Acoustic overexposure, such as listening to loud music too often, results in noise-induced hearing loss. The pathologies of this prevalent sensory disorder begin within the ear at synapses of the primary auditory receptors, their postsynaptic partners and their supporting cells. The extent of noise-induced damage, however, is determined by overstimulation of primary auditory receptors, upstream of where the pathologies manifest. A systematic characterization of the electrophysiological function of the upstream primary auditory receptors is warranted to understand how noise exposure impacts on downstream targets, where the pathologies of hearing loss begin. Here, we used the experimentally-accessible locust ear (male, Schistocerca gregaria) to characterize a decrease in the auditory receptor's ability to respond to sound after noise exposure. Surprisingly, after noise exposure, the electrophysiological properties of the auditory receptors remain unchanged, despite a decrease in the ability to transduce sound. This auditory deficit stems from changes in a specialized receptor lymph that bathes the auditory receptors, revealing striking parallels with the mammalian auditory system. SIGNIFICANCE STATEMENT Noise exposure is the largest preventable cause of hearing loss. It is the auditory receptors that bear the initial brunt of excessive acoustic stimulation, because they must convert excessive sound-induced movements into electrical signals, but remain functional afterward. Here we use the accessible ear of an invertebrate to, for the first time in any animal, characterize changes in auditory receptors after noise overexposure. We find that their decreased ability to transduce sound into electrical signals is, most probably, due to changes in supporting (scolopale) cells that maintain the ionic composition of the ear. An emerging doctrine in hearing research is that vertebrate primary auditory receptors are surprisingly robust, something that we show rings true for invertebrate ears too.

International audience; The genotype-phenotype relationship and the physiopathology of Down Syndrome (DS) have been explored in the last 20 years with more and more relevant mouse models. From the early age of transgenesis to the new CRISPR/CAS9-derived chromosomal engineering and the transchromosomic technologies, mouse models have been key to identify homolo-gous genes or entire regions homologous to the human chromosome 21 that are necessary or sufficient to induce DS features, to investigate the complexity of the genetic interactions that are involved in DS and to explore therapeutic strategies. In this review we report the new developments made, how genomic data and new genetic tools have deeply changed our way of making models, extended our panel of animal models, and increased our understanding of the neurobiology of the disease. But even if we have made an incredible progress which promises to make DS a curable condition, we are facing new research challenges to nurture our knowledge of DS pathophysiology as a neurodevelopmental disorder with many comorbidities during ageing.

Refractory focal epilepsy is a devastating disease for which there is frequently no effective treatment. Gene therapy represents a promising alternative, but treating epilepsy in this way involves irreversible changes to brain tissue, so vector design must be carefully optimized to guarantee safety without compromising efficacy. We set out to develop an epilepsy gene therapy vector optimized for clinical translation. The gene encoding the voltage-gated potassium channel Kv1.1, KCNA1, was codon optimized for human expression and mutated to accelerate the recovery of the channels from inactivation. For improved safety, this engineered potassium channel (EKC) gene was packaged into a nonintegrating lentiviral vector under the control of a cell type-specific CAMK2A promoter. In a blinded, randomized, placebo-controlled preclinical trial, the EKC lentivector robustly reduced seizure frequency in a male rat model of focal neocortical epilepsy characterized by discrete spontaneous seizures. When packaged into an adeno-associated viral vector (AAV2/9), the EKC gene was also effective at suppressing seizures in a male rat model of temporal lobe epilepsy. This demonstration of efficacy in a clinically relevant setting, combined with the improved safety conferred by cell type-specific expression and integration-deficient delivery, identify EKC gene therapy as being ready for clinical translation in the treatment of refractory focal epilepsy.SIGNIFICANCE STATEMENT Pharmacoresistant epilepsy affects up to 0.3% of the population. Although epilepsy surgery can be effective, it is limited by risks to normal brain function. We have developed a gene therapy that builds on a mechanistic understanding of altered neuronal and circuit excitability in cortical epilepsy. The potassium channel gene KCNA1 was mutated to bypass post-transcriptional editing and was packaged in a nonintegrating lentivector to reduce the risk of insertional mutagenesis. A randomized, blinded preclinical study demonstrated therapeutic effectiveness in a rodent model of focal neocortical epilepsy. Adeno-associated viral delivery of the channel to both hippocampi was also effective in a model of temporal lobe epilepsy. These results support clinical translation to address a major unmet need.

Imaging biomarkers are frequently proposed as endpoints for clinical trials targeting brain amyloidosis in Alzheimer's disease (AD); however, the specific impact of amyloid-β (Aβ) aggregation on biomarker abnormalities remains elusive in AD. Using the McGill-R-Thy1-APP transgenic rat as a model of selective Aβ pathology, we characterized the longitudinal progression of abnormalities in biomarkers commonly used in AD research. Middle-aged (9–11 months) transgenic animals (both male and female) displayed mild spatial memory impairments and disrupted cingulate network connectivity measured by resting-state fMRI, even in the absence of hypometabolism (measured with PET [18F]FDG) or detectable fibrillary amyloidosis (measured with PET [18F]NAV4694). At more advanced ages (16–19 months), cognitive deficits progressed in conjunction with resting connectivity abnormalities; furthermore, hypometabolism, Aβ plaque accumulation, reduction of CSF Aβ1-42concentrations, and hippocampal atrophy (structural MRI) were detectable at this stage. The present results emphasize the early impact of Aβ on brain connectivity and support a framework in which persistent Aβ aggregation itself is sufficient to impose memory circuits dysfunction, which propagates to adjacent brain networks at later stages.SIGNIFICANCE STATEMENTThe present study proposes a “back translation” of the Alzheimer pathological cascade concept from human to animals. We used the same set of Alzheimer imaging biomarkers typically used in large human cohorts and assessed their progression over time in a transgenic rat model, which allows for a finer spatial resolution not attainable with mice. Using this translational platform, we demonstrated that amyloid-β pathology recapitulates an Alzheimer-like profile of biomarker abnormalities even in the absence of other hallmarks of the disease such as neurofibrillary tangles and widespread neuronal losses.

Neuroinflammation is a key part of the etio-pathogenesis of Alzheimer’s disease. We test the relationship between neuroinflammation and the disruption of functional connectivity in large-scale networks, and their joint influence on cognitive impairment.We combined [11C]PK11195 positron emission tomography (PET) and resting-state functional magnetic resonance imaging (rs-fMRI) in 28 humans (13 females/15 males) with clinical diagnosis of probable Alzheimer’s disease or mild cognitive impairment with positive PET biomarker for amyloid, and 14 age-, sex-, and education-matched healthy humans (8 females/6 males). Source-based ‘inflammetry’ was used to extract principal components of [11C]PK11195 PET signal variance across all participants. rs-fMRI data were pre-processed via independent component analyses to classify neuronal and non-neuronal signals. Multiple linear regression models identified sources of signal co-variance between neuroinflammation and brain connectivity profiles, in relation to group and cognitive status.Patients showed significantly higher [11C]PK11195 binding relative to controls, in a distributed spatial pattern including the hippocampus, medial, and inferior temporal cortex. Patients with enhanced loading on this [11C]PK11195 binding distribution displayed diffuse abnormal functional connectivity. The expression of a stronger association between such abnormal connectivity and higher levels of neuroinflammation correlated with worse cognitive deficits.Our study suggests that neuroinflammation relates to the pathophysiological changes in network function that underlie cognitive deficits in Alzheimer’s disease. Neuroinflammation, and its association with functionally-relevant reorganisation of brain networks, is proposed as a target for emerging immuno-therapeutic strategies aimed at preventing or slowing the emergence of dementia.Significance StatementNeuroinflammation is an important aspect of Alzheimer’s disease (AD), but it was not known whether the influence of neuroinflammation on brain network function in humans was important for cognitive deficit.Our study provides clear evidence that in vivo neuroinflammation in AD impairs large-scale network connectivity; and that the link between inflammation and functional network connectivity is relevant to cognitive impairment.We suggest that future studies should address how neuroinflammation relates to network function as AD progresses; and whether the neuroinflammation in AD is reversible, as the basis of immunotherapeutic strategies to slow the progression of AD.

Mnemonic discrimination, the ability to distinguish similar events in memory, relies on subregions in the human medial temporal lobes (MTLs). Tau pathology is frequently found within the MTL of older adults and therefore likely to affect mnemonic discrimination, even in healthy older individuals. The MTL subregions that are known to be affected early by tau pathology, the perirhinal-transentorhinal region (area 35) and the anterior-lateral entorhinal cortex (alEC), have recently been implicated in the mnemonic discrimination of objects rather than scenes. Here we used an object-scene mnemonic discrimination task in combination with fMRI recordings and analyzed the relationship between subregional MTL activity, memory performance, and levels of total and phosphorylated tau as well as Aβ42/40 ratio in CSF. We show that activity in alEC was associated with mnemonic discrimination of similar objects but not scenes in male and female cognitively unimpaired older adults. Importantly, CSF tau levels were associated with increased fMRI activity in the hippocampus, and both increased hippocampal activity as well as tau levels were associated with mnemonic discrimination of objects, but again not scenes. This suggests that dysfunction of the alEC-hippocampus object mnemonic discrimination network might be a marker for tau-related cognitive decline.SIGNIFICANCE STATEMENTSubregions in the human medial temporal lobe are critically involved in episodic memory and, at the same time, affected by tau pathology. Impaired object mnemonic discrimination performance as well as aberrant activity within the entorhinal-hippocampal circuitry have been reported in earlier studies involving older individuals, but it has thus far remained elusive whether and how tau pathology is implicated in this specific impairment. Using task-related fMRI in combination with measures of tau pathology in CSF, we show that measures of tau pathology are associated with increased hippocampal activity and reduced mnemonic discrimination of similar objects but not scenes. This suggests that object mnemonic discrimination tasks could be promising markers for tau-related cognitive decline.

Synchronized oscillations within and between brain areas facilitate normal processing, but are often amplified in disease. A prominent example is the abnormally sustained beta-frequency (∼20 Hz) oscillations recorded from the cortex and subthalamic nucleus of Parkinson's disease patients. Computational modeling suggests that the amplitude of such oscillations could be modulated by applying stimulation at a specific phase. Such a strategy would allow selective targeting of the oscillation, with relatively little effect on other activity parameters. Here, activity was recorded from 10 awake, parkinsonian patients (6 male, 4 female human subjects) undergoing functional neurosurgery. We demonstrate that stimulation arriving on a particular patient-specific phase of the beta oscillation over consecutive cycles could suppress the amplitude of this pathophysiological activity by up to 40%, while amplification effects were relatively weak. Suppressive effects were accompanied by a reduction in the rhythmic output of subthalamic nucleus (STN) neurons and synchronization with the mesial cortex. While stimulation could alter the spiking pattern of STN neurons, there was no net effect on firing rate, suggesting that reduced beta synchrony was a result of alterations to the relative timing of spiking activity, rather than an overall change in excitability. Together, these results identify a novel intrinsic property of cortico-basal ganglia synchrony that suggests the phase of ongoing neural oscillations could be a viable and effective control signal for the treatment of Parkinson's disease. This work has potential implications for other brain diseases with exaggerated neuronal synchronization and for probing the function of rhythmic activity in the healthy brain.SIGNIFICANCE STATEMENTIn Parkinson's disease (PD), movement impairment is correlated with exaggerated beta frequency oscillations in the cerebral cortex and subthalamic nucleus (STN). Using a novel method of stimulation in PD patients undergoing neurosurgery, we demonstrate that STN beta oscillations can be suppressed when consecutive electrical pulses arrive at a specific phase of the oscillation. This effect is likely because of interrupting the timing of neuronal activity rather than excitability, as stimulation altered the firing pattern of STN spiking without changing overall rate. These findings show the potential of oscillation phase as an input for “closed-loop” stimulation, which could provide a valuable neuromodulation strategy for the treatment of brain disorders and for elucidating the role of neuronal oscillations in the healthy brain.

Fragile X syndrome (FXS) is the most common inherited form of intellectual disability, resulted from the silencing of the Fmr1 gene and the subsequent loss of fragile X mental retardation protein (FMRP). Spine dysgenesis and cognitive impairment have been extensively characterized in FXS; however, the underlying mechanism remains poorly understood., Fragile X syndrome (FXS) is the most common inherited form of intellectual disability, resulted from the silencing of the Fmr1 gene and the subsequent loss of fragile X mental retardation protein (FMRP). Spine dysgenesis and cognitive impairment have been extensively characterized in FXS; however, the underlying mechanism remains poorly understood. As an important regulator of spine maturation, intercellular adhesion molecule 5 (ICAM5) mRNA may be one of the targets of FMRP and involved in cognitive impairment in FXS. Here we show that in Fmr1 KO male mice, ICAM5 was excessively expressed during the late developmental stage, and its expression was negatively correlated with the expression of FMRP and positively related with the morphological abnormalities of dendritic spines. While in vitro reduction of ICAM5 normalized dendritic spine abnormalities in Fmr1 KO neurons, and in vivo knockdown of ICAM5 in the dentate gyrus rescued the impaired spatial and fear memory and anxiety-like behaviors in Fmr1 KO mice, through both granule cell and mossy cell with a relative rate of 1.32 ± 0.15. Furthermore, biochemical analyses showed direct binding of FMRP with ICAM5 mRNA, to the coding sequence of ICAM5 mRNA. Together, our study suggests that ICAM5 is one of the targets of FMRP and is implicated in the molecular pathogenesis of FXS. ICAM5 could be a therapeutic target for treating cognitive impairment in FXS. SIGNIFICANCE STATEMENT Fragile X syndrome (FXS) is characterized by dendritic spine dysgenesis and cognitive dysfunctions, while one of the FMRP latent targets, ICAM5, is well established for contributing both spine maturation and learning performance. In this study, we examined the potential link between ICAM5 mRNA and FMRP in FXS, and further investigated the molecular details and pathological consequences of ICAM5 overexpression. Our results indicate a critical role of ICAM5 in spine maturation and cognitive impairment in FXS and suggest that ICAM5 is a potential molecular target for the development of medication against FXS.

Classical schemes of basal ganglia organization posit that parkinsonian movement difficulties presenting after striatal dopamine depletion stem from the disproportionate firing rates of spiny projection neurons (SPNs) therein. There remains, however, a pressing need to elucidate striatal SPN firing in the context of the synchronized network oscillations that are abnormally exaggerated in cortical–basal ganglia circuits in parkinsonism. To address this, we recorded unit activities in the dorsal striatum of dopamine-intact and dopamine-depleted rats during two brain states, respectively defined by cortical slow-wave activity (SWA) and activation. Dopamine depletion escalated striatal net output but had contrasting effects on “direct pathway” SPNs (dSPNs) and “indirect pathway” SPNs (iSPNs); their firing rates became imbalanced, and they disparately engaged in network oscillations. Disturbed striatal activity dynamics relating to the slow (∼1 Hz) oscillations prevalent during SWA partly generalized to the exaggerated beta-frequency (15–30 Hz) oscillations arising during cortical activation. In both cases, SPNs exhibited higher incidences of phase-locked firing to ongoing cortical oscillations, and SPN ensembles showed higher levels of rhythmic correlated firing, after dopamine depletion. Importantly, in dopamine-depleted striatum, a widespread population of iSPNs, which often displayed excessive firing rates and aberrant phase-locked firing to cortical beta oscillations, preferentially and excessively synchronized their firing at beta frequencies. Conversely, dSPNs were neither hyperactive nor synchronized to a large extent during cortical activation. These data collectively demonstrate a cell type-selective entrainment of SPN firing to parkinsonian beta oscillations. We conclude that a population of overactive, excessively synchronized iSPNs could orchestrate these pathological rhythms in basal ganglia circuits. SIGNIFICANCE STATEMENT Chronic depletion of dopamine from the striatum, a part of the basal ganglia, causes some symptoms of Parkinson's disease. Here, we elucidate how dopamine depletion alters striatal neuron firing in vivo, with an emphasis on defining whether and how spiny projection neurons (SPNs) engage in the synchronized beta-frequency (15–30 Hz) oscillations that become pathologically exaggerated throughout basal ganglia circuits in parkinsonism. We discovered that a select population of so-called “indirect pathway” SPNs not only fire at abnormally high rates, but are also particularly prone to being recruited to exaggerated beta oscillations. Our results provide an important link between two complementary theories that explain the presentation of disease symptoms on the basis of changes in firing rate or firing synchronization/rhythmicity.

Many lines of evidence suggest that the Parkinson's disease (PD)-related protein α-synuclein (α-SYN) can propagate from cell to cell in a prion-like manner. However, the cellular mechanisms behind the spreading remain elusive. Here, we show that human astrocytes derived from embryonic stem cells actively transfer aggregated α-SYN to nearby astrocytes via direct contact and tunneling nanotubes (TNTs). Failure in the astrocytes' lysosomal digestion of excess α-SYN oligomers results in α-SYN deposits in the trans-Golgi network followed by endoplasmic reticulum swelling and mitochondrial disturbances. The stressed astrocytes respond by conspicuously sending out TNTs, enabling intercellular transfer of α-SYN to healthy astrocytes, which in return deliver mitochondria, indicating a TNT-mediated rescue mechanism. Using a pharmacological approach to inhibit TNT formation, we abolished the transfer of both α-SYN and mitochondria. Together, our results highlight the role of astrocytes in α-SYN cell-to-cell transfer, identifying possible pathophysiological events in the PD brain that could be of therapeutic relevance. SIGNIFICANCE STATEMENT Astrocytes are the major cell type in the brain, yet their role in Parkinson's disease progression remains elusive. Here, we show that human astrocytes actively transfer aggregated α-synuclein (α-SYN) to healthy astrocytes via direct contact and tunneling nanotubes (TNTs), rather than degrade it. The astrocytes engulf large amounts of oligomeric α-SYN that are subsequently stored in the trans-Golgi network region. The accumulation of α-SYN in the astrocytes affects their lysosomal machinery and induces mitochondrial damage. The stressed astrocytes respond by sending out TNTs, enabling intercellular transfer of α-SYN to healthy astrocytes. Our findings highlight an unexpected role of astrocytes in the propagation of α-SYN pathology via TNTs, revealing astrocytes as a potential target for therapeutic intervention.

The MAPK/ERK (mitogen-activated protein kinases/extracellular signal-regulated kinase) pathway is a cardinal regulator of synaptic plasticity, learning, and memory in the hippocampus. One of major endpoints of this signaling cascade is the 5′ mRNA cap binding protein eIF4E (eukaryotic Initiation Factor 4E), which is phosphorylated on Ser 209 by MNK (MAPK-interacting protein kinases) and controls mRNA translation. The precise role of phospho-eIF4E in the brain is yet to be determined. Herein, we demonstrate that ablation of eIF4E phosphorylation in male mice (4Eki mice) does not impair long-term spatial or contextual fear memory, or the late phase of LTP. Using unbiased translational profiling in mouse brain, we show that phospho-eIF4E differentially regulates the translation of a subset of mRNAs linked to inflammation, the extracellular matrix, pituitary hormones, and the serotonin pathway. Consequently, 4Eki male mice display exaggerated inflammatory responses and reduced levels of serotonin, concomitant with depression and anxiety-like behaviors. Remarkably, eIF4E phosphorylation is required for the chronic antidepressant action of the selective serotonin reuptake inhibitor fluoxetine. Finally, we propose a novel phospho-eIF4E-dependent translational control mechanism in the brain, via the GAIT complex (gamma IFN activated inhibitor of translation). In summary, our work proposes a novel translational control mechanism involved in the regulation of inflammation and depression, which could be exploited to design novel therapeutics. SIGNIFICANCE STATEMENT We demonstrate that downstream of the MAPK (mitogen-activated protein kinase) pathway, eukaryotic Initiation Factor 4E (eIF4E) Ser209 phosphorylation is not required for classical forms of hippocampal LTP and memory. We reveal a novel role for eIF4E phosphorylation in inflammatory responses and depression-like behaviors. eIF4E phosphorylation is required for the chronic action of antidepressants, such as fluoxetine in mice. These phenotypes are accompanied by selective translation of extracellular matrix, pituitary hormones, and serotonin pathway genes, in eIF4E phospho-mutant mice. We also describe a previously unidentified translational control mechanism in the brain, whereby eIF4E phosphorylation is required for inhibiting the translation of gamma IFN activated inhibitor of translation element-containing mRNAs. These findings can be used to design novel therapeutics for depression.

Healthy aging is associated with marked effects on sleep, including its daily amount and architecture, as well as the specific EEG oscillations. Neither the neurophysiological underpinnings nor the biological significance of these changes are understood, and crucially the question remains whether aging is associated with reduced sleep need or a diminished capacity to generate sufficient sleep. Here we tested the hypothesis that aging may affect local cortical networks, disrupting the capacity to generate and sustain sleep oscillations, and with it the local homeostatic response to sleep loss. We performed chronic recordings of cortical neural activity and local field potentials from the motor cortex in young and older male C57BL/6J mice, during spontaneous waking and sleep, as well as during sleep after sleep deprivation. In older animals, we observed an increase in the incidence of non-rapid eye movement sleep local field potential slow waves and their associated neuronal silent (OFF) periods, whereas the overall pattern of state-dependent cortical neuronal firing was generally similar between ages. Furthermore, we observed that the response to sleep deprivation at the level of local cortical network activity was not affected by aging. Our data thus suggest that the local cortical neural dynamics and local sleep homeostatic mechanisms, at least in the motor cortex, are not impaired during healthy senescence in mice. This indicates that powerful protective or compensatory mechanisms may exist to maintain neuronal function stable across the life span, counteracting global changes in sleep amount and architecture. SIGNIFICANCE STATEMENT The biological significance of age-dependent changes in sleep is unknown but may reflect either a diminished sleep need or a reduced capacity to generate deep sleep stages. As aging has been linked to profound disruptions in cortical sleep oscillations and because sleep need is reflected in specific patterns of cortical activity, we performed chronic electrophysiological recordings of cortical neural activity during waking, sleep, and after sleep deprivation from young and older mice. We found that all main hallmarks of cortical activity during spontaneous sleep and recovery sleep after sleep deprivation were largely intact in older mice, suggesting that the well-described age-related changes in global sleep are unlikely to arise from a disruption of local network dynamics within the neocortex.

Neuron subtype dysfunction is a key contributor to neurologic disease circuits, but identifying associated gene regulatory pathways is complicated by the molecular complexity of the brain. For example, parvalbumin-expressing (PV+) neurons in the external globus pallidus (GPe) are critically involved in the motor deficits of dopamine-depleted mouse models of Parkinson's disease, where cell type-specific optogenetic stimulation of PV+neurons over other neuron populations rescues locomotion. Despite the distinct roles these cell types play in the neural circuit, the molecular correlates remain unknown because of the difficulty of isolating rare neuron subtypes. To address this issue, we developed a new viral affinity purification strategy, Cre-Specific Nuclear Anchored Independent Labeling, to isolate Cre recombinase-expressing (Cre+) nuclei from the adult mouse brain. Applying this technology, we performed targeted assessments of the cell type-specific transcriptomic and epigenetic effects of dopamine depletion on PV+and PV–cells within three brain regions of male and female mice: GPe, striatum, and cortex. We found GPe PV+neuron-specific gene expression changes that suggested increased hypoxia-inducible factor 2α signaling. Consistent with transcriptomic data, regions of open chromatin affected by dopamine depletion within GPe PV+neurons were enriched for hypoxia-inducible factor family binding motifs. The gene expression and epigenomic experiments performed on PV+neurons isolated by Cre-Specific Nuclear Anchored Independent Labeling identified a transcriptional regulatory network mediated by the neuroprotective factor Hif2a as underlying neural circuit differences in response to dopamine depletion.SIGNIFICANCE STATEMENTCre-Specific Nuclear Anchored Independent Labeling is an enhanced, virus-based approach to isolate nuclei of a specific cell type for transcriptome and epigenome interrogation that decreases dependency on transgenic animals. Applying this technology to GPe parvalbumin-expressing neurons in a mouse model of Parkinson's disease, we discovered evidence for an upregulation of the oxygen homeostasis maintaining pathway involving Hypoxia-inducible factor 2α. These results provide new insight into how neuron subtypes outside the substantia nigra pars compacta may be compensating at a molecular level for differences in the motor production neural circuit during the progression of Parkinson's disease. Furthermore, they emphasize the utility of cell type-specific technologies, such as Cre-Specific Nuclear Anchored Independent Labeling, for isolated assessment of specific neuron subtypes in complex systems.

Status Epilepticus Increases the Intracellular Accumulation of GABAA Receptors Goodkin HP, Yeh JL, Kapur J J Neurosci 2005;25:5511–5520 Status epilepticus is a neurological emergency that results in mortality and neurological morbidity. It has been postulated that the reduction of inhibitory transmission during status epilepticus results from a rapid modification of GABAA receptors. However, the mechanism(s) that contributes to this modification has not been elucidated. We report, using an in vitro model of status epilepticus combined with electrophysiological and cellular imaging techniques, that prolonged epileptiform bursting results in a reduction of GABA-mediated synaptic inhibition. Furthermore, we found that constitutive internalization of GABAA receptors is rapid and accelerated by the increased neuronal activity associated with seizures. Inhibition of neuronal activity reduced the rate of internalization. These findings suggest that the rate of GABAA receptor internalization is regulated by neuronal activity and its acceleration contributes to the reduction of inhibitory transmission observed during prolonged seizures. Trafficking of GABAA Receptors, Loss of Inhibition, and a Mechanism for Pharmacoresistance in Status Epilepticus Naylor DE, Liu H, Wasterlain CG J Neurosci 2005;25:7724–7733 During status epilepticus (SE), GABAergic mechanisms fail and seizures become self-sustaining and pharmacoresistant. During lithiumpilocarpine-induced SE, our studies of postsynaptic GABAA receptors in dentate gyrus granule cells show a reduction in the amplitude of miniature IPSCs (mIPSCs). Anatomical studies show a reduction in the colocalization of the β2/ β3 and γ2 subunits of GABAA receptors with the presynaptic marker synaptophysin and an increase in the proportion of those subunits in the interior of dentate granule cells and other hippocampal neurons with SE. Unlike synaptic mIPSCs, the amplitude of extrasynaptic GABAA tonic currents is augmented during SE. Mathematical modeling suggests that the change of mIPSCs with SE reflects a decrease in the number of functional postsynaptic GABAA receptors. It also suggests that increases in extracellular [GABA] during SE can account for the tonic current changes and can affect postsynaptic receptor kinetics with a loss of paired-pulse inhibition. GABA exposure mimics the effects of SE on mIPSC and tonic GABAA current amplitudes in granule cells, consistent with the model predictions. These results provide a potential mechanism for the inhibitory loss that characterizes initiation of SE and for the pharmacoresistance to benzodiazepines, as a reduction of available functional GABAA postsynaptic receptors. Novel therapies for SE might be directed toward prevention or reversal of these losses. Kainic Acid-Induced Status Epilepticus Alters GABA Receptor Subunit mRNA and Protein Expression in the Developing Rat Hippocampus Lauren HB, Lopez-Picon FR, Korpi ER, Holopainen IE J Neurochem 2005;94:1384–1394 Kainic acid-induced status epilepticus leads to structural and functional changes in inhibitory GABAA receptors in the adult rat hippocampus, but whether similar changes occur in the developing rat is not known. We have used in situ hybridization to study status epilepticus-induced changes in the GABAA α1– α5, β1– β3, γ1, and γ2 subunit mRNA expression in the hippocampus of 9-day-old rats during 1 week after the treatment. Immunocytochemistry was applied to detect the α1, α2, and β3 subunit proteins in the control and treated rats. In the saline-injected control rats, the α1 and α4 subunit mRNA expression significantly increased between the postnatal days 9 to 16, whereas those of α2, β3, and γ2 subunits decreased. The normal developmental changes in the expression of α1, α2, β3, and γ2 subunit mRNAs were altered after the treatment. The immunostainings with antibodies to α1, α2, and β3 subunits confirmed the in situ hybridization findings. No neuronal death was detected in any hippocampal subregion in the treated rats. Our results show that status epilepticus disturbs the normal developmental expression pattern of GABAA receptor subunit in the rat hippocampus during the sensitive postnatal period of brain development. These perturbations could result in altered functional and pharmacological properties of GABAA receptors.

Nerve cells that re-enter a cell cycle will die rather than divide, a fact that likely underlies the neurodegeneration in Alzheimer's disease (AD). Several mouse models of familial AD have been created, and although many display amyloid plaques in their brains, none captures the extensive pattern of nerve cell death found in the human disease. Using both immunocytochemistry and fluorescentin situhybridization, we show that neurons in three different mouse models reproduce the ectopic cell cycling found in human AD. The temporal and spatial appearance of the cell cycle events in the mouse closely mimics the human disease progression. The cell-cycle events are evident 6 months before the first amyloid deposits and significantly precede the appearance of the first CD45+ microglia. These data suggest that the ectopic initiation of cell-cycle processes in neurons is an early sign of neuronal distress in both human and mouse AD. The close phenotypic correspondence indicates a previously unsuspected level of fidelity of the mouse model to the human disease. Finally, the relative timing suggests that neither the activated microglia nor the amyloid plaques themselves are necessary to initiate the pathogenic events in AD.