Myo-optogenetics: optogenetic stimulation and electrical recording in skeletal muscles.

Complex movements involve highly coordinated control of local muscle elements. Highly controlled perturbations of motor outputs can reveal insights into the neural control of movements. Here we introduce an optogenetic method, compatible with electromyography (EMG) recordings, to perturb muscles in transgenic mice. By expressing channelrhodopsin in muscle fibers, we achieved noninvasive, focal activation of orofacial muscles, enabling detailed examination of the mechanical properties of optogenetically evoked jaw muscle contractions. We demonstrated simultaneous EMG recording and optical stimulation, revealing the electrophysiological characteristics of optogenetically triggered muscle activity. Additionally, we applied optogenetic activation of muscles in physiologically and behaviorally relevant settings, mapping precise muscle actions and perturbing active behaviors. Our findings highlight the potential of muscle optogenetics to precisely manipulate muscle activity, offering a powerful tool for probing neuromuscular control systems and advancing our understanding of motor control.

Inflammation of the Nasal Mucosa is Associated with Susceptibility to Experimental Pneumococcal Challenge in Older Adults.

Streptococcus pneumoniae colonization in the upper respiratory tract is linked to pneumococcal disease development, predominantly affecting young children and older adults. As the global population ages and comorbidities increase, there is a heightened concern about this infection. We investigated the immunological responses of older adults to pneumococcal controlled human infection by analysing the cellular composition and gene expression in the nasal mucosa. Our comparative analysis with data from a concurrent study in younger adults revealed distinct gene expression patterns in older individuals susceptible to colonization, highlighted by neutrophil activation and elevated levels of CXCL9 and CXCL10. Unlike younger adults challenged with pneumococcus, older adults did not show recruitment of monocytes into the nasal mucosa following nasal colonization. However, older adults who were protected from colonization showed increased degranulation of CD8+ T cells, both before and after pneumococcal challenge. These findings suggest age-associated cellular changes, in particular enhanced mucosal inflammation, that may predispose older adults to pneumococcal colonization.

Longitudinal detection of gait alterations associated with hypertension-induced cerebral microhemorrhages in mice: predictive role of stride length and stride time asymmetry and increased gait entropy.

Cerebral microhemorrhages (CMHs) are of paramount importance as they not only signify underlying vascular pathology but also have profound implications for cognitive function and neurological health, serving as a critical indicator for the early detection and management of vascular cognitive impairment (VCI). This study aimed to investigate the effects of hypertension-induced CMHs on gait dynamics in a mouse model, focusing on the utility of advanced gait metrics as sensitive indicators of subclinical neurological alterations associated with CMHs. To induce CMHs, we employed a hypertensive mouse model, using a combination of Angiotensin II and L-NAME to elevate blood pressure, further supplemented with phenylephrine to mimic transient blood pressure fluctuations. Gait dynamics were analyzed using the CatWalk system, with emphasis on symmetry indices for Stride Length (SL), Stride Time (ST), and paw print area, as well as measures of gait entropy and regularity. The study spanned a 30-day experimental period, capturing day-to-day variations in gait parameters to assess the impact of CMHs. Temporary surges in gait asymmetry, detected as deviations from median gait metrics, suggested the occurrence of subclinical neurological signs associated with approximately 50% of all histologically verified CMHs. Our findings also demonstrated that increases in gait entropy correlated with periods of increased gait asymmetry, providing insights into the complexity of gait dynamics in response to CMHs. Significant correlations were found between SL and ST symmetry indices and between these indices and the paw print area symmetry index post-hypertension induction, indicating the interdependence of spatial and temporal aspects of gait affected by CMHs. Collectively, advanced gait metrics revealed sensitive, dynamic alterations in gait regulation associated with CMHs, resembling the temporal characteristics of transient ischemic attacks (TIAs). This underscores their potential as non-invasive indicators of subclinical neurological impacts. This study supports the use of detailed gait analysis as a valuable tool for detecting subtle neurological changes, with implications for the early diagnosis and monitoring of cerebral small vessel disease (CSVD) in clinical settings.

Rule-based modulation of a sensorimotor transformation across cortical areas.

Flexible responses to sensory stimuli based on changing rules are critical for adapting to a dynamic environment. However, it remains unclear how the brain encodes and uses rule information to guide behavior. Here, we made single-unit recordings while head-fixed mice performed a cross-modal sensory selection task where they switched between two rules: licking in response to tactile stimuli while rejecting visual stimuli, or vice versa. Along a cortical sensorimotor processing stream including the primary (S1) and secondary (S2) somatosensory areas, and the medial (MM) and anterolateral (ALM) motor areas, single-neuron activity distinguished between the two rules both prior to and in response to the tactile stimulus. We hypothesized that neural populations in these areas would show rule-dependent preparatory states, which would shape the subsequent sensory processing and behavior. This hypothesis was supported for the motor cortical areas (MM and ALM) by findings that (1) the current task rule could be decoded from pre-stimulus population activity; (2) neural subspaces containing the population activity differed between the two rules; and (3) optogenetic disruption of pre-stimulus states impaired task performance. Our findings indicate that flexible action selection in response to sensory input can occur via configuration of preparatory states in the motor cortex.

Imaging of cerebrovascular complications from blunt skull base trauma.

Cerebrovascular complications from blunt trauma to the skull base, though rare, can lead to potentially devastating outcomes, emphasizing the importance of timely diagnosis and management. Due to the insidious clinical presentation, subtle nature of imaging findings, and complex anatomy of the skull base, diagnosing cerebrovascular injuries and their complications poses considerable challenges. This article offers a comprehensive review of skull base anatomy and pathophysiology pertinent to recognizing cerebrovascular injuries and their complications, up-to-date screening criteria and imaging techniques for assessing these injuries, and a case-based review of the spectrum of cerebrovascular complications arising from skull base trauma. This review will enhance understanding of cerebrovascular injuries and their complications from blunt skull base trauma to facilitate diagnosis and timely treatment.

Children with perinatally acquired HIV exhibit distinct immune responses to 4CMenB vaccine.

Children with perinatally acquired HIV (PHIV) have special vaccination needs, as they make suboptimal immune responses. Here, we evaluated safety and immunogenicity of 2 doses of 4-component group B meningococcal vaccine in antiretroviral therapy-treated children with PHIV and healthy controls (HCs). Assessments included the standard human serum bactericidal antibody (hSBA) assay and measurement of IgG titers against capsular group B Neisseria meningitidis antigens (fHbp, NHBA, NadA). The B cell compartment and vaccine-induced antigen-specific (fHbp+) B cells were investigated by flow cytometry, and gene expression was investigated by multiplexed real-time PCR. A good safety and immunogenicity profile was shown in both groups; however, PHIV demonstrated a reduced immunogenicity compared with HCs. Additionally, PHIV showed a reduced frequency of fHbp+ and an altered B cell subset distribution, with higher fHbp+ frequency in activated memory and tissue-like memory B cells. Gene expression analyses on these cells revealed distinct mechanisms between PHIV and HC seroconverters. Overall, these data suggest that PHIV presents a diverse immune signature following vaccination. The impact of such perturbation on long-term maintenance of vaccine-induced immunity should be further evaluated in vulnerable populations, such as people with PHIV.

Exercise-induced ß2-adrenergic Receptor Activation Enhances the Antileukemic Activity of Expanded γδ T-Cells via DNAM-1 Upregulation and PVR/Nectin-2 Recognition.

Exercise mobilizes cytotoxic lymphocytes to blood which may allow superior cell products to be harvested and manufactured for cancer therapy. Gamma-Delta (γδ) T-cells have shown promise for treating solid tumors, but there is a need to increase their potency against hematologic malignancies. Here, we show that human γδ T-cells mobilized to blood in response to just 20 minutes of graded exercise have surface phenotypes and transcriptomic profiles associated with cytotoxicity, adhesion, migration, and cytokine signaling. Following 14 days ex vivo expansion with zoledronic acid and IL2, exercise mobilized γδ T-cells had surface phenotypes and transcriptomic profiles associated with enhanced effector functions and demonstrated superior cytotoxic activity against multiple hematologic tumors in vitro and in vivo in leukemia-bearing xenogeneic mice. Infusing humans with the ß1+ß2-agonist isoproterenol and administering ß1 or ß1+ß2 antagonists prior to exercise revealed these effects to be ß2-adrenergic receptor (AR) dependent. Antibody blocking of DNAM-1 on expanded γδ T-cells, as well as the DNAM-1 ligands PVR and Nectin-2 on leukemic targets, abolished the enhanced antileukemic effects of exercise. These findings provide a mechanistic link between exercise, ß2-AR activation, and the manufacture of superior γδ T-cell products for adoptive cell therapy against hematologic malignancies. SIGNIFICANCE: Exercise mobilizes effector γδ T-cells to blood via ß2-adrenergic signaling which allows for generation of a potent expanded γδ T-cell product that is highly cytotoxic against hematologic malignancies.

Tactile processing in mouse cortex depends on action context.

The brain receives constant tactile input, but only a subset guides ongoing behavior. Actions associated with tactile stimuli thus endow them with behavioral relevance. It remains unclear how the relevance of tactile stimuli affects processing in the somatosensory (S1) cortex. We developed a cross-modal selection task in which head-fixed mice switched between responding to tactile stimuli in the presence of visual distractors or to visual stimuli in the presence of tactile distractors using licking movements to the left or right side in different blocks of trials. S1 spiking encoded tactile stimuli, licking actions, and direction of licking in response to tactile but not visual stimuli. Bidirectional optogenetic manipulations showed that sensory-motor activity in S1 guided behavior when touch but not vision was relevant. Our results show that S1 activity and its impact on behavior depend on the actions associated with a tactile stimulus.

Natural History of Opioid Use in Naive and Tolerant Patients in Revision Total Hip Arthroplasty.

BACKGROUND: Opioid use after revision total hip arthroplasty (rTHA) has not been well characterized. The purpose of this study was to characterize preoperative, perioperative, and postoperative opioid use during rTHA. METHODS: Patients undergoing revision THA from 2010 to 2018 were screened for opioid use 3 months before revision surgery and tracked 24 months postoperatively. Patients were categorized as naïve or tolerant. Opioid prescriptions and average morphine milligram equivalents (MME) were compared between the two groups. RESULTS: One hundred twenty-four of 247 patients (50%) in the tolerant group averaged a preoperative MME of 23.7 mg/day. Postoperatively, tolerant patients received significantly higher daily MME at all time points, including at 3 months 31.4 versus 18.1 mg/day (P < 0.001), 6 months 19.9 versus 2.95 mg/day (P < 0.001), 12 months 14.3 versus 3.5 mg/day (P < 0.001), and 24 months 10.7 versus 2.17 mg/day (P < 0.001). Tolerant patients were more likely to have a prescription at 6 months (44% versus 22%), 12 months (41.4% versus 24%), and 24 months (38% versus 19.3%) (P < 0.001, P = 0.002, P < 0.001, respectively). DISCUSSION: Opioid-tolerant patients had higher postoperative MME requirements for longer recovery duration. Both groups reduced opioid use at 3 months and plateaued at 6 months. These findings can help the revision surgeon counsel patients and expectations.

Multi-omics analysis reveals COVID-19 vaccine induced attenuation of inflammatory responses during breakthrough disease.

The immune mechanisms mediating COVID-19 vaccine attenuation of COVID-19 remain undescribed. We conducted comprehensive analyses detailing immune responses to SARS-CoV-2 virus in blood post-vaccination with ChAdOx1 nCoV-19 or a placebo. Samples from randomised placebo-controlled trials (NCT04324606 and NCT04400838) were taken at baseline, onset of COVID-19-like symptoms, and 7 days later, confirming COVID-19 using nucleic amplification test (NAAT test) via real-time PCR (RT-PCR). Serum cytokines were measured with multiplexed immunoassays. The transcriptome was analysed with long, short and small RNA sequencing. We found attenuation of RNA inflammatory signatures in ChAdOx1 nCoV-19 compared with placebo vaccinees and reduced levels of serum proteins associated with COVID-19 severity. KREMEN1, a putative alternative SARS-CoV-2 receptor, was downregulated in placebo compared with ChAdOx1 nCoV-19 vaccinees. Vaccination ameliorates reductions in cell counts across leukocyte populations and platelets noted at COVID-19 onset, without inducing potentially deleterious Th2-skewed immune responses. Multi-omics integration links a global reduction in miRNA expression at COVID-19 onset to increased pro-inflammatory responses at the mRNA level. This study reveals insights into the role of COVID-19 vaccines in mitigating disease severity by abrogating pro-inflammatory responses associated with severe COVID-19, affirming vaccine-mediated benefit in breakthrough infection, and highlighting the importance of clinically relevant endpoints in vaccine evaluation.

The Many Uses of the Humble Alcohol Swab.

Readily available and comparatively inexpensive, the common alcohol swab can aid dermatologists in everything from diagnosis to preoperative and postoperative care. The 70% isopropyl alcohol swab can aid in the accurate diagnosis of lesions and skin conditions, identification of biopsy sites, and disinfection.

Targeted metagenomics reveals association between severity and pathogen co-detection in infants with respiratory syncytial virus.

Respiratory syncytial virus (RSV) is the leading cause of hospitalisation for respiratory infection in young children. RSV disease severity is known to be age-dependent and highest in young infants, but other correlates of severity, particularly the presence of additional respiratory pathogens, are less well understood. In this study, nasopharyngeal swabs were collected from two cohorts of RSV-positive infants <12 months in Spain, the UK, and the Netherlands during 2017-20. We show, using targeted metagenomic sequencing of >100 pathogens, including all common respiratory viruses and bacteria, from samples collected from 433 infants, that burden of additional viruses is common (111/433, 26%) but only modestly correlates with RSV disease severity. In contrast, there is strong evidence in both cohorts and across age groups that presence of Haemophilus bacteria (194/433, 45%) is associated with higher severity, including much higher rates of hospitalisation (odds ratio 4.25, 95% CI 2.03-9.31). There is no evidence for association between higher severity and other detected bacteria, and no difference in severity between RSV genotypes. Our findings reveal the genomic diversity of additional pathogens during RSV infection in infants, and provide an evidence base for future causal investigations of the impact of co-infection on RSV disease severity.

Bilateral Whisker Representations in the Primary Somatosensory Cortex in Robo3cKO Mice Are Reflected in the Primary Motor Cortex.

In Robo3cKO mice, midline crossing defects of the trigeminothalamic projections from the trigeminal principal sensory nucleus result in bilateral whisker maps in the somatosensory thalamus and consequently in the face representation area of the primary somatosensory (S1) cortex (Renier et al., 2017; Tsytsarev et al., 2017). We investigated whether this bilateral sensory representation in the whisker-barrel cortex is also reflected in the downstream projections from the S1 to the primary motor (M1) cortex. To label these projections, we injected anterograde viral axonal tracer in S1 cortex. Corticocortical projections from the S1 distribute to similar areas across the ipsilateral hemisphere in control and Robo3cKO mice. Namely, in both genotypes they extend to the M1, premotor/prefrontal cortex (PMPF), secondary somatosensory (S2) cortex. Next, we performed voltage-sensitive dye imaging (VSDi) in the left hemisphere following ipsilateral and contralateral single whisker stimulation. While controls showed only activation in the contralateral whisker barrel cortex and M1 cortex, the Robo3cKO mouse left hemisphere was activated bilaterally in both the barrel cortex and the M1 cortex. We conclude that the midline crossing defect of the trigeminothalamic projections leads to bilateral whisker representations not only in the thalamus and the S1 cortex but also downstream from the S1, in the M1 cortex.

Drug repurposing for rare: progress and opportunities for the rare disease community.

Repurposing is one of the key opportunities to address the unmet rare diseases therapeutic need. Based on cases of drug repurposing in small population conditions, and previous work in drug repurposing, we analyzed the most important lessons learned, such as the sharing of clinical observations, reaching out to regulatory scientific advice at an early stage, and public-private collaboration. In addition, current upcoming trends in the field of drug repurposing in rare diseases were analyzed, including the role these trends could play in the rare diseases' ecosystem. Specifically, we cover the opportunities of innovation platforms, the use of real-world data, the use of artificial intelligence, regulatory initiatives in repurposing, and patient engagement throughout the repurposing project. The outcomes from these emerging activities will help progress the field of drug repurposing for the benefit of patients, public health and medicines development.

A Genome-Wide Association Study of Respiratory Syncytial Virus Infection Severity in Infants.

BACKGROUND: Respiratory syncytial virus (RSV) is a significant cause of infant morbidity and mortality worldwide. Most children experience at least one 1 RSV infection by the age of two 2 years, but not all develop severe disease. However, the understanding of genetic risk factors for severe RSV is incomplete. Consequently, we conducted a genome-wide association study of RSV severity. METHODS: Disease severity was assessed by the ReSVinet scale, in a cohort of 251 infants aged 1 week to 1 year. Genotyping data were collected from multiple European study sites as part of the RESCEU Consortium. Linear regression models were used to assess the impact of genotype on RSV severity and gene expression as measured by microarray. RESULTS: While no SNPs reached the genome-wide statistical significance threshold (P < 5 × 10-8), we identified 816 candidate SNPs with a P-value of <1 × 10-4. Functional annotation of candidate SNPs highlighted genes relevant to neutrophil trafficking and cytoskeletal functions, including LSP1 and RAB27A. Moreover, SNPs within the RAB27A locus significantly altered gene expression (false discovery rate, FDR P < .05). CONCLUSIONS: These findings may provide insights into genetic mechanisms driving severe RSV infection, offering biologically relevant information for future investigations.

Association of mobile health (mHealth) use with health status and COVID-19-related concerns by people with mobility impairments.

PURPOSE: Mobile health (mHealth) technology has increased dramatically in the wake of the pandemic. Less research has focused on people with mobility impairing (PMI) disabilities. This study determined the prevalence of mHealth use among PMI adults during the COVID-19 escalation and examines demographic, health and COVID-19 concerns correlates. METHODS: PMI adults (N = 304) completed an online survey investigating mHealth use and COVID-19 concerns related to food access in June of 2020. Smartphone and mHealth use were measured with an adapted version of the survey used in the Pew Internet & American Life project. Descriptive and multivariable analyses were conducted to determine associations of demographics, health status, and COVID-19 concerns with mHealth use. About two-thirds (N = 201) of the sample were mHealth users (owned a smartphone and engaged in health-promoting behaviors with the smartphone; e.g., sought online information, tracked health behaviors, used patient portals). RESULTS: Having hypertension was associated with higher mHealth use, and having higher COVID-19 concerns about food access was associated with higher mHealth use. Those who used mHealth were also more engaged with smartphone apps for communication, services, and entertainment. Only the association between educational attainment and mHealth use remained significant after adjusting for other covariates in multivariable logistic regression models. DISCUSSION: PMIs continue to need support in the use of mHealth technology to help maximize access to potentially important tools for rehabilitation and health management. There is a need to continue to investigate mHealth and its applications for people with disabilities.Implications for RehabilitationMany people with mobility impairing disabilities may be missing opportunities for mHealth rehabilitation and healthcare.COVID-19 has widened existing gaps in access and use of mHealth technology among people with mobility impairing disabilities.Focused education is needed to help people with disabilities exploit the full range of services of their smartphones to increase access to care, social connectivity, and other important goods and services to enhance rehabilitation and health management.

Exploring Factors Associated With Accelerometer Validity Among Ethnically Diverse Toddlers.

PURPOSE: Studying physical activity in toddlers using accelerometers is challenging due to noncompliance with wear time (WT) and activity log (AL) instructions. The aims of this study are to examine relationships between WT and AL completion and (1) demographic and socioeconomic variables, (2) parenting style, and (3) whether sedentary time differs by AL completion. METHODS: Secondary analysis was performed using baseline data from a community wellness program randomized controlled trial for parents with toddlers (12-35 mo). Parents had toddlers wear ActiGraph wGT3x accelerometers and completed ALs. Valid days included ≥600-minute WT. Analysis of variance and chi-square analyses were used. RESULTS: The sample (n = 50) comprised racial and ethnically diverse toddlers (mean age = 27 mo, 58% male) and parents (mean age = 31.7 y, 84% female). Twenty-eight families (56%) returned valid accelerometer data with ALs. Participants in relationships were more likely to complete ALs (P < .05). Toddler sedentary time did not differ between those with ALs and those without. CONCLUSIONS: We found varied compliance with WT instructions and AL completion. Returned AL quality was poor, presenting challenges in correctly characterizing low-activity counts to improve internal validity of WT and physical activity measures. Support from marital partners may be important for adherence to study protocols.

Rule-based modulation of a sensorimotor transformation across cortical areas.

Flexible responses to sensory stimuli based on changing rules are critical for adapting to a dynamic environment. However, it remains unclear how the brain encodes rule information and uses this information to guide behavioral responses to sensory stimuli. Here, we made single-unit recordings while head-fixed mice performed a cross-modal sensory selection task in which they switched between two rules in different blocks of trials: licking in response to tactile stimuli applied to a whisker while rejecting visual stimuli, or licking to visual stimuli while rejecting the tactile stimuli. Along a cortical sensorimotor processing stream including the primary (S1) and secondary (S2) somatosensory areas, and the medial (MM) and anterolateral (ALM) motor areas, the single-trial activity of individual neurons distinguished between the two rules both prior to and in response to the tactile stimulus. Variable rule-dependent responses to identical stimuli could in principle occur via appropriate configuration of pre-stimulus preparatory states of a neural population, which would shape the subsequent response. We hypothesized that neural populations in S1, S2, MM and ALM would show preparatory activity states that were set in a rule-dependent manner to cause processing of sensory information according to the current rule. This hypothesis was supported for the motor cortical areas by findings that (1) the current task rule could be decoded from pre-stimulus population activity in ALM and MM; (2) neural subspaces containing the population activity differed between the two rules; and (3) optogenetic disruption of pre-stimulus states within ALM and MM impaired task performance. Our findings indicate that flexible selection of an appropriate action in response to a sensory input can occur via configuration of preparatory states in the motor cortex.

Simultaneous recordings of pupil size variation and locus coeruleus activity in mice.

An extensive literature describes how pupil size reflects neuromodulatory activity, including the noradrenergic system. Here, we present a protocol for the simultaneous recording of optogenetically identified locus coeruleus (LC) units and pupil diameter in mice under different conditions. We describe steps for building an optrode, performing surgery to implant the optrode and headpost, searching for opto-tagged LC units, and performing dual LC-pupil recording. We then detail procedures for data processing and analysis. For complete details on the use and execution of this protocol, please refer to Megemont et al.1.