Neuroprotective role of lactate in a human in vitro model of the ischemic penumbra.

In patients suffering from cerebral ischemic stroke, there is an urgent need for treatments to protect stressed yet viable brain cells. Recently, treatment strategies that induce neuronal activity have been shown to be neuroprotective. Here, we hypothesized that neuronal activation might maintain or trigger the astrocyte-to-neuron lactate shuttle (ANLS), whereby lactate is released from astrocytes to support the energy requirements of ATP-starved hypoxic neurons, and this leads to the observed neuroprotection. We tested this by using a human cell based in vitro model of the ischemic penumbra and investigating whether lactate might be neuroprotective in this setting. We found that lactate transporters are involved in the neuroprotective effect mediated by neuronal activation. Furthermore, we showed that lactate exogenously administered before hypoxia correlated with neuroprotection in our cellular model. In addition, stimulation of astrocyte with consequent endogenous production of lactate resulted in neuroprotection. To conclude, here we presented evidence that lactate transport into neurons contributes to neuroprotection during hypoxia providing a potential basis for therapeutic approaches in ischemic stroke.

Integrating across behaviors and timescales to understand the neural control of movement.

The nervous system evolved to enable navigation throughout the environment in the pursuit of resources. Evolutionarily newer structures allowed increasingly complex adaptations but necessarily added redundancy. A dominant view of movement neuroscientists is that there is a one-to-one mapping between brain region and function. However, recent experimental data is hard to reconcile with the most conservative interpretation of this framework, suggesting a degree of functional redundancy during the performance of well-learned, constrained behaviors. This apparent redundancy likely stems from the bidirectional interactions between the various cortical and subcortical structures involved in motor control. We posit that these bidirectional connections enable flexible interactions across structures that change depending upon behavioral demands, such as during acquisition, execution or adaptation of a skill. Observing the system across both multiple actions and behavioral timescales can help isolate the functional contributions of individual structures, leading to an integrated understanding of the neural control of movement.

SCN1A-deficient excitatory neuronal networks display mutation-specific phenotypes.

Dravet syndrome is a severe epileptic encephalopathy, characterized by (febrile) seizures, behavioural problems and developmental delay. Eighty per cent of patients with Dravet syndrome have a mutation in SCN1A, encoding Nav1.1. Milder clinical phenotypes, such as GEFS+ (generalized epilepsy with febrile seizures plus), can also arise from SCN1A mutations. Predicting the clinical phenotypic outcome based on the type of mutation remains challenging, even when the same mutation is inherited within one family. This clinical and genetic heterogeneity adds to the difficulties of predicting disease progression and tailoring the prescription of anti-seizure medication. Understanding the neuropathology of different SCN1A mutations may help to predict the expected clinical phenotypes and inform the selection of best-fit treatments. Initially, the loss of Na+-current in inhibitory neurons was recognized specifically to result in disinhibition and consequently seizure generation. However, the extent to which excitatory neurons contribute to the pathophysiology is currently debated and might depend on the patient clinical phenotype or the specific SCN1A mutation. To examine the genotype-phenotype correlations of SCN1A mutations in relation to excitatory neurons, we investigated a panel of patient-derived excitatory neuronal networks differentiated on multi-electrode arrays. We included patients with different clinical phenotypes, harbouring various SCN1A mutations, along with a family in which the same mutation led to febrile seizures, GEFS+ or Dravet syndrome. We hitherto describe a previously unidentified functional excitatory neuronal network phenotype in the context of epilepsy, which corresponds to seizurogenic network prediction patterns elicited by proconvulsive compounds. We found that excitatory neuronal networks were affected differently, depending on the type of SCN1A mutation, but did not segregate according to clinical severity. Specifically, loss-of-function mutations could be distinguished from missense mutations, and mutations in the pore domain could be distinguished from mutations in the voltage sensing domain. Furthermore, all patients showed aggravated neuronal network responses at febrile temperatures compared with controls. Finally, retrospective drug screening revealed that anti-seizure medication affected GEFS+ patient- but not Dravet patient-derived neuronal networks in a patient-specific and clinically relevant manner. In conclusion, our results indicate a mutation-specific excitatory neuronal network phenotype, which recapitulates the foremost clinically relevant features, providing future opportunities for precision therapies.

Porewater constituents inhibit microbially mediated greenhouse gas production (GHG) and regulate the response of soil organic matter decomposition to warming in anoxic peat from a Sphagnum-dominated bog.

Northern peatlands store approximately one-third of terrestrial soil carbon. Climate warming is expected to stimulate the microbially mediated degradation of peat soil organic matter (SOM), leading to increasing greenhouse gas (GHG; carbon dioxide, CO2; methane, CH4) production and emission. Porewater dissolved organic matter (DOM) plays a key role in SOM decomposition; however, the mechanisms controlling SOM decomposition and its response to warming remain unclear. The temperature dependence of GHG production and microbial community dynamics were investigated in anoxic peat from a Sphagnum-dominated peatland. In this study, peat decomposition, which was quantified by GHG production and carbon substrate utilization is limited by terminal electron acceptors (TEA) and DOM, and these controls of microbially mediated SOM degradation are temperature-dependent. Elevated temperature led to a slight decrease in microbial diversity, and stimulated the growth of specific methanotrophic and syntrophic taxa. These results confirm that DOM is a major driver of decomposition in peatland soils contains inhibitory compounds, but the inhibitory effect is alleviated by warming.

Peat loss collocates with a threshold in plant-mycorrhizal associations in drained peatlands encroached by trees.

Drainage-induced encroachment by trees may have major effects on the carbon balance of northern peatlands, and responses of microbial communities are likely to play a central mechanistic role. We profiled the soil fungal community and estimated its genetic potential for the decay of lignin and phenolics (class II peroxidase potential) along peatland drainage gradients stretching from interior locations (undrained, open) to ditched locations (drained, forested). Mycorrhizal fungi dominated the community across the gradients. When moving towards ditches, the dominant type of mycorrhizal association abruptly shifted from ericoid mycorrhiza to ectomycorrhiza at c. 120 m from the ditches. This distance corresponded with increased peat loss, from which more than half may be attributed to oxidation. The ectomycorrhizal genus Cortinarius dominated at the drained end of the gradients and its relatively higher genetic potential to produce class II peroxidases (together with Mycena) was positively associated with peat humification and negatively with carbon-to-nitrogen ratio. Our study is consistent with a plant-soil feedback mechanism, driven by a shift in the mycorrhizal type of vegetation, that potentially mediates changes in aerobic decomposition during postdrainage succession. Such feedback may have long-term legacy effects upon postdrainage restoration efforts and implication for tree encroachment onto carbon-rich soils globally.

Cadherin-13 is a critical regulator of GABAergic modulation in human stem-cell-derived neuronal networks.

Activity in the healthy brain relies on a concerted interplay of excitation (E) and inhibition (I) via balanced synaptic communication between glutamatergic and GABAergic neurons. A growing number of studies imply that disruption of this E/I balance is a commonality in many brain disorders; however, obtaining mechanistic insight into these disruptions, with translational value for the patient, has typically been hampered by methodological limitations. Cadherin-13 (CDH13) has been associated with autism and attention-deficit/hyperactivity disorder. CDH13 localizes at inhibitory presynapses, specifically of parvalbumin (PV) and somatostatin (SST) expressing GABAergic neurons. However, the mechanism by which CDH13 regulates the function of inhibitory synapses in human neurons remains unknown. Starting from human-induced pluripotent stem cells, we established a robust method to generate a homogenous population of SST and MEF2C (PV-precursor marker protein) expressing GABAergic neurons (iGABA) in vitro, and co-cultured these with glutamatergic neurons at defined E/I ratios on micro-electrode arrays. We identified functional network parameters that are most reliably affected by GABAergic modulation as such, and through alterations of E/I balance by reduced expression of CDH13 in iGABAs. We found that CDH13 deficiency in iGABAs decreased E/I balance by means of increased inhibition. Moreover, CDH13 interacts with Integrin-ß1 and Integrin-ß3, which play opposite roles in the regulation of inhibitory synaptic strength via this interaction. Taken together, this model allows for standardized investigation of the E/I balance in a human neuronal background and can be deployed to dissect the cell-type-specific contribution of disease genes to the E/I balance.

Air pollution and the pandemic: Long-term PM2.5 exposure and disease severity in COVID-19 patients.

BACKGROUND AND OBJECTIVE: Ecological studies have suggested an association between exposure to particulate matter ≤2.5 µm (PM2.5 ) and coronavirus disease 2019 (COVID-19) severity. However, these findings are yet to be validated in individual-level studies. We aimed to determine the association of long-term PM2.5 exposure with hospitalization among individual patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). METHODS: We estimated the 10-year (2009-2018) PM2.5 exposure at the residential zip code of COVID-19 patients diagnosed at the University of Cincinnati healthcare system between 13 March 2020 and 30 September 2020. Logistic regression was used to determine the odds ratio (OR) and 95% CI for COVID-19 hospitalizations associated with PM2.5 , adjusting for socioeconomic characteristics and comorbidities. RESULTS: Among the 14,783 COVID-19 patients included in our study, 13.6% were hospitalized; the geometric mean (SD) PM2.5 was 10.48 (1.12) µg/m3 . In adjusted analysis, 1 µg/m3 increase in 10-year annual average PM2.5 was associated with 18% higher hospitalization (OR: 1.18, 95% CI: 1.11-1.26). Likewise, 1 µg/m3 increase in PM2.5 estimated for the year 2018 was associated with 14% higher hospitalization (OR: 1.14, 95% CI: 1.08-1.21). CONCLUSION: Long-term PM2.5 exposure is associated with increased hospitalization in COVID-19. Therefore, more stringent COVID-19 prevention measures may be needed in areas with higher PM2.5 exposure to reduce the disease morbidity and healthcare burden.

Soil metabolome response to whole-ecosystem warming at the Spruce and Peatland Responses under Changing Environments experiment.

In this study, a suite of complementary environmental geochemical analyses, including NMR and gas chromatography-mass spectrometry (GC-MS) analyses of central metabolites, Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) of secondary metabolites, and lipidomics, was used to investigate the influence of organic matter (OM) quality on the heterotrophic microbial mechanisms controlling peatland CO2, CH4, and CO2:CH4 porewater production ratios in response to climate warming. Our investigations leverage the Spruce and Peatland Responses under Changing Environments (SPRUCE) experiment, where air and peat warming were combined in a whole-ecosystem warming treatment. We hypothesized that warming would enhance the production of plant-derived metabolites, resulting in increased labile OM inputs to the surface peat, thereby enhancing microbial activity and greenhouse gas production. Because shallow peat is most susceptible to enhanced warming, increases in labile OM inputs to the surface, in particular, are likely to result in significant changes to CO2 and CH4 dynamics and methanogenic pathways. In support of this hypothesis, significant correlations were observed between metabolites and temperature consistent with increased availability of labile substrates, which may stimulate more rapid turnover of microbial proteins. An increase in the abundance of methanogenic genes in response to the increase in the abundance of labile substrates was accompanied by a shift toward acetoclastic and methylotrophic methanogenesis. Our results suggest that as peatland vegetation trends toward increasing vascular plant cover with warming, we can expect a concomitant shift toward increasingly methanogenic conditions and amplified climate-peatland feedbacks.

Flexible scaling and persistence of social vocal communication.

Innate vocal sounds such as laughing, screaming or crying convey one's feelings to others. In many species, including humans, scaling the amplitude and duration of vocalizations is essential for effective social communication1-3. In mice, female scent triggers male mice to emit innate courtship ultrasonic vocalizations (USVs)4,5. However, whether mice flexibly scale their vocalizations and how neural circuits are structured to generate flexibility remain largely unknown. Here we identify mouse neurons from the lateral preoptic area (LPOA) that express oestrogen receptor 1 (LPOAESR1 neurons) and, when activated, elicit the complete repertoire of USV syllables emitted during natural courtship. Neural anatomy and functional data reveal a two-step, di-synaptic circuit motif in which primary long-range inhibitory LPOAESR1 neurons relieve a clamp of local periaqueductal grey (PAG) inhibition, enabling excitatory PAG USV-gating neurons to trigger vocalizations. We find that social context shapes a wide range of USV amplitudes and bout durations. This variability is absent when PAG neurons are stimulated directly; PAG-evoked vocalizations are time-locked to neural activity and stereotypically loud. By contrast, increasing the activity of LPOAESR1 neurons scales the amplitude of vocalizations, and delaying the recovery of the inhibition clamp prolongs USV bouts. Thus, the LPOA disinhibition motif contributes to flexible loudness and the duration and persistence of bouts, which are key aspects of effective vocal social communication.

Autoantibodies Targeting Intracellular and Extracellular Proteins in Autoimmunity.

Detecting autoantibodies provides foundational information for the diagnosis of most autoimmune diseases. An important pathophysiological distinction is whether autoantibodies are directed against extracellular or intracellular proteins. Autoantibodies targeting extracellular domains of proteins, such as membrane receptors, channels or secreted molecules are often directly pathogenic, whereby autoantibody binding to the autoantigen disrupts the normal function of a critical protein or pathway, and/or triggers antibody-dependent cell surface complement killing. By comparison, autoantibodies directed against intracellular proteins are recognized as useful diagnostic biomarkers of abnormal autoimmune activity, but the link between antigenicity and pathogenicity is less straightforward. Because intracellular autoantigens are generally inaccessible to autoantibody binding, for the most part, they do not directly contribute to pathogenesis. In a few diseases, autoantibodies to intracellular targets cause damage indirectly by immune complex formation, immune activation, and other processes. In this review, the general features of and differences between autoimmune diseases segregated on the basis of intracellular or extracellular autoantigens are explored using over twenty examples. Expression profiles of autoantigens in relation to the tissues targeted by autoimmune disease and the temporal appearance of autoantibodies before clinical diagnosis often correlate with whether the respective autoantibodies mostly recognize either intracellular or extracellular autoantigens. In addition, current therapeutic strategies are discussed from this vantage point. One drug, rituximab, depletes CD20+ B-cells and is highly effective for autoimmune disorders associated with autoantibodies against extracellular autoantigens. In contrast, diseases associated with autoantibodies directed predominately against intracellular autoantigens show much more complex immune cell involvement, such as T-cell mediated tissue damage, and require different strategies for optimal therapeutic benefit. Understanding the clinical ramifications of autoimmunity derived by autoantibodies against either intracellular or extracellular autoantigens, or a spectrum of both, has practical implications for guiding drug development, generating monitoring tools, stratification of patient interventions, and designing trials based on predictive autoantibody profiles for autoimmune diseases.

Long-term exposure to fine particulate matter and hospitalization in COVID-19 patients.

BACKGROUND: Ecological evidence suggests that exposure to air pollution affects coronavirus disease 2019 (COVID-19) outcomes. However, no individual-level study has confirmed the association to date. METHODS: We identified COVID-19 patients diagnosed at the University of Cincinnati hospitals and clinics and estimated particulate matter ≤2.5 µm (PM2.5) exposure over a 10-year period (2008-2017) at their residential zip codes. We used logistic regression to evaluate the association between PM2.5 exposure and hospitalizations for COVID-19, adjusting for socioeconomic characteristics and comorbidities. RESULTS: Among the 1128 patients included in our study, the mean (standard deviation) PM2.5 was 11.34 (0.70) µg/m3 for the 10-year average exposure and 13.83 (1.03) µg/m3 for the 10-year maximal exposures. The association between long-term PM2.5 exposure and hospitalization for COVID-19 was contingent upon having pre-existing asthma or chronic obstructive pulmonary (COPD) (Pinteraction = 0.030 for average PM2.5 and Pinteraction = 0.001 for maximal PM2.5). In COVID-19 patients with asthma or COPD, the odds of hospitalization were 62% higher with 1 µg/m3 increment in 10-year average PM2.5 (odds ratio [OR]: 1.62, 95% confidence interval [CI]: 1.00-2.64) and 65% higher with 1 µg/m3 increase in 10-year maximal PM2.5 levels (OR: 1.65, 95% CI: 1.16-2.35). However, among COVID-19 patients without asthma or COPD, PM2.5 exposure was not associated with higher hospitalizations (OR: 0.84, 95% CI: 0.65-1.09 for average PM2.5 and OR: 0.78, 95% CI: 0.65-0.95 for maximal PM2.5). CONCLUSIONS: Long-term exposure to PM2.5 is associated with higher odds of hospitalization in COVID-19 patients with pre-existing asthma or COPD.

PIEZO2 in sensory neurons and urothelial cells coordinates urination.

Henry Miller stated that "to relieve a full bladder is one of the great human joys". Urination is critically important in health and ailments of the lower urinary tract cause high pathological burden. Although there have been advances in understanding the central circuitry in the brain that facilitates urination1-3, there is a lack of in-depth mechanistic insight into the process. In addition to central control, micturition reflexes that govern urination are all initiated by peripheral mechanical stimuli such as bladder stretch and urethral flow4. The mechanotransduction molecules and cell types that function as the primary stretch and pressure detectors in the urinary tract mostly remain unknown. Here we identify expression of the mechanosensitive ion channel PIEZO2 in lower urinary tract tissues, where it is required for low-threshold bladder-stretch sensing and urethral micturition reflexes. We show that PIEZO2 acts as a sensor in both the bladder urothelium and innervating sensory neurons. Humans and mice lacking functional PIEZO2 have impaired bladder control, and humans lacking functional PIEZO2 report deficient bladder-filling sensation. This study identifies PIEZO2 as a key mechanosensor in urinary function. These findings set the foundation for future work to identify the interactions between urothelial cells and sensory neurons that control urination.

Is Blood Type Associated with COVID-19 Severity?

Blood type purportedly influences susceptibility to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection, but whether it affects severity of coronavirus disease 2019 (COVID-19) is unclear. Therefore, we examined the association of blood type and rhesus with hospitalization and disease severity among 428 COVID-19 patients diagnosed at the University of Cincinnati health system. In the sample, 50.2% of participants had the blood type O, 38.7% had the blood type A, 17.5% had the blood type B, and 3.5% had the blood type AB. In analysis adjusted for sociodemographic characteristics and comorbidities, the blood types A (OR: 0.90, 95% CI: 0.54, 1.50), B (OR: 0.93, 95% CI: 0.51, 1.69), AB (OR: 0.69, 95% CI: 0.20, 2.41), and O (OR: 1.18, 95%: 0.74, 1.98) were not associated with hospitalization for COVID-19. Similarly, the blood types A (OR: 0.93, 95% CI: 0.52, 1.65), B (OR: 0.92, 95% CI: 0.46, 1.84), AB (OR: 0.30, 95% CI: 0.04, 2.44), and O (OR: 1.25, 95%: 0.73, 2.14) were not associated with admission to intensive care unit or death in COVID-19. In conclusion, blood type is not associated with hospitalization or disease severity in COVID-19; therefore, it may not be useful marker for identifying patients at risk for severe COVID-19.

m.3243A > G-Induced Mitochondrial Dysfunction Impairs Human Neuronal Development and Reduces Neuronal Network Activity and Synchronicity.

Epilepsy, intellectual and cortical sensory deficits, and psychiatric manifestations are the most frequent manifestations of mitochondrial diseases. How mitochondrial dysfunction affects neural structure and function remains elusive, mostly because of a lack of proper in vitro neuronal model systems with mitochondrial dysfunction. Leveraging induced pluripotent stem cell technology, we differentiated excitatory cortical neurons (iNeurons) with normal (low heteroplasmy) and impaired (high heteroplasmy) mitochondrial function on an isogenic nuclear DNA background from patients with the common pathogenic m.3243A > G variant of mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS). iNeurons with high heteroplasmy exhibited mitochondrial dysfunction, delayed neural maturation, reduced dendritic complexity, and fewer excitatory synapses. Micro-electrode array recordings of neuronal networks displayed reduced network activity and decreased synchronous network bursting. Impaired neuronal energy metabolism and compromised structural and functional integrity of neurons and neural networks could be the primary drivers of increased susceptibility to neuropsychiatric manifestations of mitochondrial disease.

Distinct Pathogenic Genes Causing Intellectual Disability and Autism Exhibit a Common Neuronal Network Hyperactivity Phenotype.

Pathogenic mutations in either one of the epigenetic modifiers EHMT1, MBD5, MLL3, or SMARCB1 have been identified to be causative for Kleefstra syndrome spectrum (KSS), a neurodevelopmental disorder with clinical features of both intellectual disability (ID) and autism spectrum disorder (ASD). To understand how these variants lead to the phenotypic convergence in KSS, we employ a loss-of-function approach to assess neuronal network development at the molecular, single-cell, and network activity level. KSS-gene-deficient neuronal networks all develop into hyperactive networks with altered network organization and excitatory-inhibitory balance. Interestingly, even though transcriptional data reveal distinct regulatory mechanisms, KSS target genes share similar functions in regulating neuronal excitability and synaptic function, several of which are associated with ID and ASD. Our results show that KSS genes mainly converge at the level of neuronal network communication, providing insights into the pathophysiology of KSS and phenotypically congruent disorders.

Neuronal network dysfunction in a model for Kleefstra syndrome mediated by enhanced NMDAR signaling.

Kleefstra syndrome (KS) is a neurodevelopmental disorder caused by mutations in the histone methyltransferase EHMT1. To study the impact of decreased EHMT1 function in human cells, we generated excitatory cortical neurons from induced pluripotent stem (iPS) cells derived from KS patients. Neuronal networks of patient-derived cells exhibit network bursting with a reduced rate, longer duration, and increased temporal irregularity compared to control networks. We show that these changes are mediated by upregulation of NMDA receptor (NMDAR) subunit 1 correlating with reduced deposition of the repressive H3K9me2 mark, the catalytic product of EHMT1, at the GRIN1 promoter. In mice EHMT1 deficiency leads to similar neuronal network impairments with increased NMDAR function. Finally, we rescue the KS patient-derived neuronal network phenotypes by pharmacological inhibition of NMDARs. Summarized, we demonstrate a direct link between EHMT1 deficiency and NMDAR hyperfunction in human neurons, providing a potential basis for more targeted therapeutic approaches for KS.

Past, Present, and Future of Neuronal Models In Vitro.

Over the past century, robust methods were developed that enable the isolation, culture, and dynamic observation of mammalian neuronal networks in vitro. But even if neuronal culture cannot yet fully recapitulate the normal brain, the knowledge that has been acquired from these surrogate in vitro models is invaluable. Indeed, neuronal culture has continued to propel basic neuroscience research, proving that in vitro systems have legitimacy when it comes to studying either the healthy or diseased human brain. Furthermore, scientific advancement typically parallels technical refinements in the field. A pertinent example is that a collective drive in the field of neuroscience to better understand the development, organization, and emergent properties of neuronal networks is being facilitated by progressive advances in micro-electrode array (MEA) technology. In this chapter, we briefly review the emergence of neuronal cell culture as a technique, the current trends in human stem cell-based modeling, and the technologies used to monitor neuronal communication. We conclude by highlighting future prospects that are evolving specifically out of the combination of human neuronal models and MEA technology.

Voluntary urination control by brainstem neurons that relax the urethral sphincter.

Voluntary urination ensures that waste is eliminated when safe and socially appropriate, even without a pressing urge. Uncontrolled urination, or incontinence, is a common problem with few treatment options. Normal urine release requires a small region in the brainstem known as Barrington's nucleus (Bar), but specific neurons that relax the urethral sphincter and enable urine flow are unknown. Here we identify a small subset of Bar neurons that control the urethral sphincter in mice. These excitatory neurons express estrogen receptor 1 (BarESR1), project to sphincter-relaxing interneurons in the spinal cord and are active during natural urination. Optogenetic stimulation of BarESR1 neurons rapidly initiates sphincter bursting and efficient voiding in anesthetized and behaving animals. Conversely, optogenetic and chemogenetic inhibition reveals their necessity in motivated urination behavior. The identification of these cells provides an expanded model for the control of urination and its dysfunction.