Interactions between skin-resident dendritic and Langerhans cells and pain-sensing neurons.

Various immune cells in the skin contribute to its function as a first line of defense against infection and disease, and the skin's dense innervation by pain-sensing sensory neurons protects the host against injury or damage signals. Dendritic cells (DCs) are a heterogeneous population of cells that link the innate immune response to the adaptive response by capturing, processing, and presenting antigens to promote T-cell differentiation and activation. DCs are abundant across peripheral tissues, including the skin, where they are found in the dermis and epidermis. Langerhans cells (LCs) are a DC subset located only in the epidermis; both populations of cells can migrate to lymph nodes to contribute to broad immune responses. Dermal DCs and LCs are found in close apposition with sensory nerve fibers in the skin and express neurotransmitter receptors, allowing them to communicate directly with the peripheral nervous system. Thus, neuroimmune signaling between DCs and/or LCs and sensory neurons can modulate physiologic and pathophysiologic pathways, including immune cell regulation, host defense, allergic response, homeostasis, and wound repair. Here, we summarize the latest discoveries on DC- and LC-neuron interaction with neurons while providing an overview of gaps and areas not previously explored. Understanding the interactions between these 2 defence systems may provide key insight into developing therapeutic targets for treating diseases such as psoriasis, neuropathic pain, and lupus.

Advanced Dynamic Weight Bearing as an Observer-independent Measure of Hyperacute Hypersensitivity in Mice.

Background: Standard methods assessing pain in rodents are often observer dependent, potentially resulting in biased outcomes. Advanced dynamic weight bearing (ADWB) offers an observer-independent approach that can provide objective, reliable data in preclinical pain research. Aims: The aim of this study was to characterize the use of ADWB in assessing murine responses to allyl isothiocyanate (AITC)-induced hyperacute hypersensitivity and identify best practices for use of the device. Methods: Male C57BL/6J mice received intraplantar injections of saline or 0.1% AITC solution and were assessed using the ADWB system; simultaneous observer-dependent durations of paw licking and biting were measured. ADWB data were analyzed using the proprietary software from Bioseb and correlated to observer-dependent results, with parameters assessed to optimize data collected. Results: ADWB detected pain-directed changes in weight and surface area distribution in AITC-treated mice, with paw weight and surface area placement correlating to paw licking and biting. Optimization of adjustable threshold parameters allowed for reduced coefficients of variability and increased duration of validated data. Conclusions: The ADWB assay provides an efficient and unbiased measure of chemical-induced hyperacute hypersensitivity in mice. ADWB detection parameters influence amount of validated data and variability, a consideration for data analysis in future studies.

Contexte: Les méthodes standard d'évaluation de la douleur chez les rongeurs dépendent souvent de l'observateur, ce qui peut fausser les résultats. La mise en charge dynamique avancée offre une approche indépendante de l'observateur qui peut fournir des données objectives et fiables dans la recherche préclinique sur la douleur.Objectifs: L'objectif de cette étude était de caractériser l'utilisation de la mise en charge dynamique avancée dans l'évaluation des réponses murines à l'hypersensibilité hyperaiguë induite par l'isothiocyanate d'allyle et de répertorier les meilleures pratiques d'utilisation de l'appareil.Méthodes: Des souris C57BL/6J mâles ont reçu des injections intraplantaires de solution saline ou de solution d'isothiocyanate d'allyle à 0,1 % et ont été évaluées à l'aide du système de mise en charge dynamique avancée; les durées simultanées de léchage et de morsure des pattes, dépendantes de l'observateur, ont été mesurées. Les données obtenues par la mise en charge dynamique avancée ont été analysées à l'aide du logiciel propriétaire de Bioseb et corrélées aux résultats dépendants de l'observateur, avec des paramètres évalués pour optimiser les données collectées.Résultats: L'essai réalisé à l'aide de la mise en charge dynamique avancée a détecté des changements de poids et de distribution de surface liés à la douleur chez les souris traitées à l'isothiocyanate d'allyle, le poids des pattes et le placement de la surface étant corrélés au léchage et à la morsure des pattes. L'optimisation des paramètres de seuil ajustables a permis de réduire les coefficients de variabilité et d'augmenter la durée des données validées.Conclusion: L'essai réalisé à l'aide de la mise en charge dynamique avancée fournit une mesure efficace et impartiale de l'hypersensibilité hyperaiguë induite par les produits chimiques chez la souris. Les paramètres de détection du système de mise en charge dynamique avancée influencent la quantité de données validées et la variabilité, ce qui doit être pris en compte pour l'analyse des données dans les études futures.

Antibiotic-induced socio-sexual behavioral deficits are reversed via cecal microbiota transplantation but not androgen treatment.

Recent evidence has demonstrated a sex-specific role of the gut microbiome on social behavior such as anxiety, possibly driven by a reciprocal relationship between the gut microbiome and gonadal hormones. For instance, gonadal hormones drive sex differences in gut microbiota composition, and certain gut bacteria can produce androgens from glucocorticoids. We thus asked whether the gut microbiome can influence androgen-dependent socio-sexual behaviors. We first treated C57BL/6 mice with broad-spectrum antibiotics (ABX) in drinking water to deplete the gut microbiota either transiently during early development (embryonic day 16-postnatal day [PND] 21) or in adulthood (PND 60-85). We hypothesized that if ABX interferes with androgens, then early ABX would interfere with critical periods for sexual differentiation of brain and thus lead to long-term decreases in males' socio-sexual behavior, while adult ABX would interfere with androgens' activational effects on behavior. We found that in males but not females, early and adult ABX treatment decreased territorial aggression, and adult ABX also decreased sexual odor preference. We then assessed whether testosterone and/or cecal microbiota transplantation (CMT) via oral gavage could prevent ABX-induced socio-sexual behavioral deficits in adult ABX-treated males. Mice were treated with same- or other-sex control cecum contents or with testosterone for two weeks. While testosterone was not effective in rescuing any behavior, we found that male CMT restored both olfactory preference and aggression in adult ABX male mice, while female CMT restored olfactory preference but not aggression. These results suggest sex-specific effects of the gut microbiome on socio-sexual behaviors, independent of androgens.

Circadian rhythms and glial cells of the central nervous system.

Glial cells are the most abundant cells in the central nervous system and play crucial roles in neural development, homeostasis, immunity, and conductivity. Over the past few decades, glial cell activity in mammals has been linked to circadian rhythms, the 24-h chronobiological clocks that regulate many physiological processes. Indeed, glial cells rhythmically express clock genes that cell-autonomously regulate glial function. In addition, recent findings in rodents have revealed that disruption of the glial molecular clock could impact the entire organism. In this review, we discuss the impact of circadian rhythms on the function of the three major glial cell types - astrocytes, microglia, and oligodendrocytes - across different locations within the central nervous system. We also review recent evidence uncovering the impact of glial cells on the body's circadian rhythm. Together, this sheds new light on the involvement of glial clock machinery in various diseases.

Cholinergic modulation is independent of T lymphocytes in a mouse model of neuropathic pain.

T lymphocytes are increasingly implicated in pain signaling. A subset of T lymphocytes, termed TChAT, express the rate-limiting enzyme for acetylcholine (ACh) production, choline acetyltransferase (ChAT), and mediate numerous physiological functions. Given that cholinergic signaling has long been known to modulate pain processing and is the basis for several analgesics used clinically, we asked whether TChAT could be the intersection between T lymphocyte and cholinergic mediation of pain signaling. In this study, we used a mouse gene knockout strategy to ablate ChAT specifically from T lymphocytes and examined the development and expression of mechanical and thermal hypersensitivity in a spared nerve injury (SNI) mouse model of neuropathic pain. We found that mice with ChAT knockout in T cells (floxed Chat plus CD4-Cre recombinase) did not differ from control mice with intact ChAT (floxed Chat, but no Cre recombinase) in their expression of mechanical sensitivity before or after injury. Similarly, thermal sensitivity was unaffected after injury, with control mice expressing similar patterns of thermal preference to mice whose T cells do not express ChAT. Our experiments demonstrate that cholinergic signaling initiated by T lymphocytes neither dampens nor exacerbates the expression of mechanical or thermal sensitivity in neuropathic mice. Thus, while both cholinergic signaling and T lymphocytes have established roles in modulating pain phenotypes, it is not cholinergic signaling initiated by T lymphocytes that drive this. Our findings will help to narrow in on which aspects of T-cell modulation may prove useful as therapies.

Muscle BDNF improves synaptic and contractile muscle strength in Kennedy's disease mice in a muscle-type specific manner.

KEY POINTS: Muscle-derived neurotrophic factors may offer therapeutic promise for treating neuromuscular diseases. We report that a muscle-derived neurotrophic factor, BDNF, rescues synaptic and muscle function in a muscle-type specific manner in mice modelling Kennedy's disease (KD). We also find that BDNF rescues select molecular mechanisms in slow and fast muscle that may underlie the improved cellular function. We also report for the first time that expression of BDNF, but not other members of the neurotrophin family, is perturbed in muscle from patients with KD. Given that muscle BDNF had divergent therapeutic effects that depended on muscle type, a combination of neurotrophic factors may optimally rescue neuromuscular function via effects on both pre- and postsynaptic function, in the face of disease. ABSTRACT: Deficits in muscle brain-derived neurotrophic factor (BDNF) correlate with neuromuscular deficits in mouse models of Kennedy's disease (KD), suggesting that restoring muscle BDNF might restore function. To test this possibility, transgenic mice expressing human BDNF in skeletal muscle were crossed with '97Q' KD mice. We found that muscle BDNF slowed disease, doubling the time between symptom onset and endstage. BDNF also improved expression of genes in muscle known to play key roles in neuromuscular function, including counteracting the expression of neonatal isoforms induced by disease. Intriguingly, BDNF's ameliorative effects differed between muscle types: synaptic strength was rescued only in slow-twitch muscle, while contractile strength was improved only in fast-twitch muscle. In sum, muscle BDNF slows disease progression, rescuing select cellular and molecular mechanisms that depend on fibre type. Muscle BDNF expression was also affected in KD patients, reinforcing its translational and therapeutic potential for treating this disorder.

Sex-Dependent Mechanisms of Chronic Pain: A Focus on Microglia and P2X4R.

For over two decades, purinergic signaling in microglia has persisted in the spotlight as a major pathomechanism of chronic pain. Of the many purinoreceptors, the P2X4R of the ionotropic family, has a well-described causal role underlying chronic neuropathic pain. This review will briefly examine microglial P2X4R signaling in the spinal cord as it relates to chronic pain through a historical lens, followed by a more in-depth examination of recent work, which has revealed major sex differences. We also discuss the generalizability of sex differences in microglial and P2X4R signaling in other pain conditions as well as in nonspinal regions. Finally, we speculate on remaining gaps in the literature as well as what can be done to address them with the ultimate goal of using our collective knowledge to treat chronic pain effectively and in both sexes. SIGNIFICANCE STATEMENT: Effective treatments are lacking for chronic pain sufferers, and this may be explained by the vast sex differences underlying chronic pain mechanisms. In this minireview, we focus on the roles of microglia and P2X4R in chronic pain, with specific attention to the circumstances under which these pathomechanisms differ between males and females. By delineating the ways in which pain occurs differently between the sexes, we can start developing successful therapies for all.

Disease Affects Bdnf Expression in Synaptic and Extrasynaptic Regions of Skeletal Muscle of Three SBMA Mouse Models.

Spinal bulbar muscular atrophy (SBMA) is a slowly progressive, androgen-dependent neuromuscular disease in men that is characterized by both muscle and synaptic dysfunction. Because gene expression in muscle is heterogeneous, with synaptic myonuclei expressing genes that regulate synaptic function and extrasynaptic myonuclei expressing genes to regulate contractile function, we used quantitative PCR to compare gene expression in these two domains of muscle from three different mouse models of SBMA: the "97Q" model that ubiquitously expresses mutant human androgen receptor (AR), the 113Q knock-in (KI) model that expresses humanized mouse AR with an expanded glutamine tract, and the "myogenic" model that overexpresses wild-type rat AR only in skeletal muscle. We were particularly interested in neurotrophic factors because of their role in maintaining neuromuscular function via effects on both muscle and synaptic function, and their implicated role in SBMA. We confirmed previous reports of the enriched expression of select genes (e.g., the acetylcholine receptor) in the synaptic region of muscle, and are the first to report the synaptic enrichment of others (e.g., glial cell line-derived neurotrophic factor). Interestingly, all three models displayed comparably dysregulated expression of most genes examined in both the synaptic and extrasynaptic domains of muscle, with only modest differences between regions and models. These findings of comprehensive gene dysregulation in muscle support the emerging view that skeletal muscle may be a prime therapeutic target for restoring function of both muscles and motoneurons in SBMA.

Microglial P2X4R-evoked pain hypersensitivity is sexually dimorphic in rats.

Microglia-neuron signalling in the spinal cord is a key mediator of mechanical allodynia caused by peripheral nerve injury. We recently reported sex differences in microglia in pain signalling in mice: spinal mechanisms underlying nerve injury-induced allodynia are microglial dependent in male but not female mice. Whether this sex difference in pain hypersensitivity mechanisms is conserved in other species is unknown. Here, we show that in rats, the spinal mechanisms of nerve injury-induced hypersensitivity in males differ from those in females, with microglial P2X4 receptors (P2X4Rs) being a key point of divergence. In rats, nerve injury produced comparable allodynia and reactive microgliosis in both sexes. However, inhibiting microglia in the spinal cord reversed allodynia in male rats but not female rats. In addition, pharmacological blockade of P2X4Rs, by an intrathecally administered antagonist, attenuated pain hypersensitivity in male rats only. Consistent with the behavioural findings, nerve injury increased cell surface expression and function of P2X4Rs in acutely isolated spinal microglia from male rats but not from female rats. Moreover, in microglia cultured from male rats, but not in those from female rats, stimulating P2X4Rs drove intracellular signalling through p38 mitogen-activated protein kinase. Furthermore, chromatin immunoprecipitation-qPCR revealed that the transcription factor IRF5 differentially binds to the P2rx4 promoter region in female rats vs male rats. Finally, mechanical allodynia was produced in otherwise naive rats by intrathecally administering P2X4R-stimulated microglia from male rats but not those from female rats. Together, our findings demonstrate the existence of sexually dimorphic pain signalling in rats, suggesting that this sex difference is evolutionarily conserved, at least across rodent species.

Pre-clinical symptoms of SBMA may not be androgen-dependent: implications from two SBMA mouse models.

A distinguishing aspect of spinal and bulbar muscular atrophy (SBMA) is its androgen-dependence, possibly explaining why only males are clinically affected. This disease, which impairs neuromuscular function, is linked to a polyglutamine expansion mutation in the androgen receptor (AR). In mouse models of SBMA, motor dysfunction is associated with pronounced defects in neuromuscular transmission, including defects in evoked transmitter release (quantal content, QC) and fiber membrane excitability (based on the resting membrane potential, RMP). However, whether such defects are androgen-dependent is unknown. Thus, we recorded synaptic potentials intracellularly from adult muscle fibers of transgenic (Tg) AR97Q male mice castrated pre-symptomatically. Although castration largely protects both QC and the RMP of fibers, correlating with the protective effect of castration on motor function, significant deficits in QC and RMP remained. Surprisingly, comparable defects in QC and RMP were also observed in pre-symptomatic AR97Q males, indicating that such defects emerge early and are pre-clinical. Exposing asymptomatic Tg females to androgens also induces both motor dysfunction and comparable defects in QC and RMP. Notably, asymptomatic Tg females also showed significant deficits in QC and RMP, albeit less severe, supporting their pre-clinical nature, but also raising questions about the androgen-dependence of pre-clinical symptoms. In summary, current evidence indicates that disease progression depends on androgens, but early pathogenic events may be triggered by the mutant AR allele independent of androgens. Such early, androgen-independent disease mechanisms may also be relevant to females carrying the SBMA allele.

Muscle Contraction Regulates BDNF/TrkB Signaling to Modulate Synaptic Function through Presynaptic cPKCα and cPKCßI.

The neurotrophin brain-derived neurotrophic factor (BDNF) acts via tropomyosin-related kinase B receptor (TrkB) to regulate synapse maintenance and function in the neuromuscular system. The potentiation of acetylcholine (ACh) release by BDNF requires TrkB phosphorylation and Protein Kinase C (PKC) activation. BDNF is secreted in an activity-dependent manner but it is not known if pre- and/or postsynaptic activities enhance BDNF expression in vivo at the neuromuscular junction (NMJ). Here, we investigated whether nerve and muscle cell activities regulate presynaptic conventional PKC (cPKCα and ßI) via BDNF/TrkB signaling to modulate synaptic strength at the NMJ. To differentiate the effects of presynaptic activity from that of muscle contraction, we stimulated the phrenic nerve of rat diaphragms (1 Hz, 30 min) with or without contraction (abolished by µ-conotoxin GIIIB). Then, we performed ELISA, Western blotting, qRT-PCR, immunofluorescence and electrophysiological techniques. We found that nerve-induced muscle contraction: (1) increases the levels of mature BDNF protein without affecting pro-BDNF protein or BDNF mRNA levels; (2) downregulates TrkB.T1 without affecting TrkB.FL or p75 neurotrophin receptor (p75) levels; (3) increases presynaptic cPKCα and cPKCßI protein level through TrkB signaling; and (4) enhances phosphorylation of cPKCα and cPKCßI. Furthermore, we demonstrate that cPKCßI, which is exclusively located in the motor nerve terminals, increases activity-induced acetylcholine release. Together, these results show that nerve-induced muscle contraction is a key regulator of BDNF/TrkB signaling pathway, retrogradely activating presynaptic cPKC isoforms (in particular cPKCßI) to modulate synaptic function. These results indicate that a decrease in neuromuscular activity, as occurs in several neuromuscular disorders, could affect the BDNF/TrkB/PKC pathway that links pre- and postsynaptic activity to maintain neuromuscular function.

Decreased Peak Expiratory Flow Associated with Muscle Fiber-Type Switching in Spinal and Bulbar Muscular Atrophy.

The aim of this study was to characterize the respiratory function profile of subjects with spinal and bulbar muscular atrophy (SBMA), and to explore the underlying pathological mechanism by comparing the clinical and biochemical indices of this disease with those of amyotrophic lateral sclerosis (ALS). We enrolled male subjects with SBMA (n = 40) and ALS (n = 25) along with 15 healthy control subjects, and assessed their respiratory function, motor function, and muscle strength. Predicted values of peak expiratory flow (%PEF) and forced vital capacity were decreased in subjects with SBMA compared with controls. In SBMA, both values were strongly correlated with the trunk subscores of the motor function tests and showed deterioration relative to disease duration. Compared with activities of daily living (ADL)-matched ALS subjects, %PEF, tongue pressure, and grip power were substantially decreased in subjects with SBMA. Both immunofluorescence and RT-PCR demonstrated a selective decrease in the expression levels of the genes encoding the myosin heavy chains specific to fast-twitch fibers in SBMA subjects. The mRNA levels of peroxisome proliferator-activated receptor gamma coactivator 1-alpha and peroxisome proliferator-activated receptor delta were up-regulated in SBMA compared with ALS and controls. In conclusion, %PEF is a disease-specific respiratory marker for the severity and progression of SBMA. Explosive muscle strength, including %PEF, was selectively affected in subjects with SBMA and was associated with activation of the mitochondrial biogenesis-related molecular pathway in skeletal muscles.

Non-Cell-Autonomous Regulation of Retrograde Motoneuronal Axonal Transport in an SBMA Mouse Model.

Defects in axonal transport are seen in motoneuronal diseases, but how that impairment comes about is not well understood. In spinal bulbar muscular atrophy (SBMA), a disorder linked to a CAG/polyglutamine repeat expansion in the androgen receptor (AR) gene, the disease-causing AR disrupts axonal transport by acting in both a cell-autonomous fashion in the motoneurons themselves, and in a non-cell-autonomous fashion in muscle. The non-cell-autonomous mechanism is suggested by data from a unique "myogenic" transgenic (TG) mouse model in which an AR transgene expressed exclusively in skeletal muscle fibers triggers an androgen-dependent SBMA phenotype, including defects in retrograde transport. However, motoneurons in this TG model retain the endogenous AR gene, leaving open the possibility that impairments in transport in this model also depend on ARs in the motoneurons themselves. To test whether non-cell-autonomous mechanisms alone can perturb retrograde transport, we generated male TG mice in which the endogenous AR allele has the testicular feminization mutation (Tfm) and, consequently, is nonfunctional. Males carrying the Tfm allele alone show no deficits in motor function or axonal transport, with or without testosterone treatment. However, when Tfm males carrying the myogenic transgene (Tfm/TG) are treated with testosterone, they develop impaired motor function and defects in retrograde transport, having fewer retrogradely labeled motoneurons and deficits in endosomal flux based on time-lapse video microscopy of living axons. These findings demonstrate that non-cell-autonomous disease mechanisms originating in muscle are sufficient to induce defects in retrograde transport in motoneurons.

Defects in Neuromuscular Transmission May Underlie Motor Dysfunction in Spinal and Bulbar Muscular Atrophy.

UNLABELLED: Spinal and bulbar muscular atrophy (SBMA) in men is an androgen-dependent neuromuscular disease caused by expanded CAG repeats in the androgen receptor (AR). Whether muscle or motor neuron dysfunction or both underlies motor impairment in SBMA is unknown. Muscles of SBMA mice show significant contractile dysfunction, implicating them as a likely source of motor dysfunction, but whether disease also impairs neuromuscular transmission is an open question. Thus, we examined synaptic function in three well-studied SBMA mouse models-the AR97Q, knock-in (KI), and myogenic141 models-by recording in vitro miniature and evoked end-plate potentials (MEPPs and EPPs, respectively) intracellularly from adult muscle fibers. We found striking defects in neuromuscular transmission suggesting that toxic AR in SBMA impairs both presynaptic and postsynaptic mechanisms. Notably, SBMA causes neuromuscular synapses to become weak and muscles to become hyperexcitable in all three models. Presynaptic defects included deficits in quantal content, reduced size of the readily releasable pool, and impaired short-term facilitation. Postsynaptic defects included prolonged decay times for both MEPPs and EPPs, marked resistance to µ-conotoxin (a sodium channel blocker), and enhanced membrane excitability. Quantitative PCR revealed robust upregulation of mRNAs encoding neonatal isoforms of the AChR (γ-subunit) and the voltage-gated sodium channel (NaV1.5) in diseased adult muscles of all three models, consistent with the observed slowing of synaptic potentials and resistance to µ-conotoxin. These findings suggest that muscles of SBMA patients regress to an immature state that impairs neuromuscular function. SIGNIFICANCE STATEMENT: We have discovered that SBMA is accompanied by marked defects in neuromuscular synaptic transmission involving both presynaptic and postsynaptic mechanisms. For three different mouse models, we find that diseased synapses are weak, having reduced quantal content due to reductions in the size of the readily releasable pool and/or probability of release. Synaptic potentials in diseased adult fibers are slowed, explained by an aberrant upregulation of the neonatal isoform of the acetylcholine receptor. Diseased fibers also show marked resistance to µ-conotoxin, explained by an aberrant upregulation in the neonatal isoform of the sodium channel, and are hyperexcitable, reminiscent of myotonic dystrophy, showing anode-break action potentials. This work identifies several new molecular targets for recovering function in SBMA.

Androgen-dependent loss of muscle BDNF mRNA in two mouse models of SBMA.

Transgenic expression of neurotrophic factors in skeletal muscle has been found to protect mice from neuromuscular disease, including spinal bulbar muscular atrophy (SBMA), triggering renewed interest in neurotrophic factors as therapeutic agents for treating neuromuscular disease. Because SBMA is an androgen-dependent disease, and brain-derived neurotrophic factor (BDNF) mediates effects of androgens on neuromuscular systems, we asked whether BDNF expression is impaired in two different transgenic (Tg) mouse models of SBMA, the so called "97Q" and "myogenic" SBMA models. The 97Q model globally overexpresses a full length human AR with 97 glutamine repeats whereas the myogenic model of SBMA overexpresses a wild-type rat androgen receptor (AR) only in skeletal muscle fibers. Using quantitative PCR, we find that muscle BDNF mRNA declines in an androgen-dependent manner in both models, paralleling changes in motor function, with robust deficits (6-8 fold) in both fast and slow twitch muscles of impaired Tg males. Castration rescues or reverses disease-related deficits in muscle BDNF mRNA in both models, paralleling its effect on motor function. Moreover, when disease is acutely induced in Tg females, both motor function and muscle BDNF mRNA expression plummet, with the deficit in muscle BDNF emerging before overt motor dysfunction. That androgen-dependent motor dysfunction is tightly associated with a robust and early down-regulation of muscle BDNF mRNA suggests that BDNF delivered to skeletal muscle may have therapeutic value for SBMA.

Distinct Etiological Roles for Myocytes and Motor Neurons in a Mouse Model of Kennedy's Disease/Spinobulbar Muscular Atrophy.

Polyglutamine (polyQ) expansion of the androgen receptor (AR) causes Kennedy's disease/spinobulbar muscular atrophy (KD/SBMA) through poorly defined cellular mechanisms. Although KD/SBMA has been thought of as a motor neuron disease, recent evidence indicates a key role for skeletal muscle. To resolve which early aspects of the disease can be caused by neurogenic or myogenic mechanisms, we made use of the tet-On and Cre-loxP genetic systems to selectively and acutely express polyQ AR in either motor neurons (NeuroAR) or myocytes (MyoAR) of transgenic mice. After 4 weeks of transgene induction in adulthood, deficits in gross motor function were seen in NeuroAR mice, but not MyoAR mice. Conversely, reduced size of fast glycolytic fibers and alterations in expression of candidate genes were observed only in MyoAR mice. Both NeuroAR and MyoAR mice exhibited reduced oxidative capacity in skeletal muscles, as well as a shift in fast fibers from oxidative to glycolytic. Markers of oxidative stress were increased in the muscle of NeuroAR mice and were reduced in motor neurons of both NeuroAR and MyoAR mice. Despite secondary pathology in skeletal muscle and behavioral deficits, no pathological signs were observed in motor neurons of NeuroAR mice, possibly due to relatively low levels of polyQ AR expression. These results indicate that polyQ AR in motor neurons can produce secondary pathology in muscle. Results also support both neurogenic and myogenic contributions of polyQ AR to several acute aspects of pathology and provide further evidence for disordered cellular respiration in KD/SBMA skeletal muscle.

Transcriptional profile of muscle following acute induction of symptoms in a mouse model of Kennedy's disease/spinobulbar muscular atrophy.

BACKGROUND: Kennedy's disease/Spinobulbar muscular atrophy (KD/SBMA) is a degenerative neuromuscular disease affecting males. This disease is caused by polyglutamine expansion mutations of the androgen receptor (AR) gene. Although KD/SBMA has been traditionally considered a motor neuron disease, emerging evidence points to a central etiological role of muscle. We previously reported a microarray study of genes differentially expressed in muscle of three genetically unique mouse models of KD/SBMA but were unable to detect those which are androgen-dependent or are associated with onset of symptoms. METHODOLOGY/PRINCIPAL FINDINGS: In the current study we examined the time course and androgen-dependence of transcriptional changes in the HSA-AR transgenic (Tg) mouse model, in which females have a severe phenotype after acute testosterone treatment. Using microarray analysis we identified differentially expressed genes at the onset and peak of muscle weakness in testosterone-treated Tg females. We found both transient and persistent groups of differentially expressed genes and analysis of gene function indicated functional groups such as mitochondrion, ion and nucleotide binding, muscle development, and sarcomere maintenance. CONCLUSIONS/SIGNIFICANCE: By comparing the current results with those from the three previously reported models we were able to identify KD/SBMA candidate genes that are androgen dependent, and occur early in the disease process, properties which are promising for targeted therapeutics.

Contractile dysfunction in muscle may underlie androgen-dependent motor dysfunction in spinal bulbar muscular atrophy.

Spinal and bulbar muscular atrophy (SBMA) is characterized by progressive muscle weakness linked to a polyglutamine expansion in the androgen receptor (AR). Current evidence indicates that mutant AR causes SBMA by acting in muscle to perturb its function. However, information about how muscle function is impaired is scant. One fundamental question is whether the intrinsic strength of muscles, an attribute of muscle independent of its mass, is affected. In the current study, we assess the contractile properties of hindlimb muscles in vitro from chronically diseased males of three different SBMA mouse models: a transgenic (Tg) model that broadly expresses a full-length human AR with 97 CAGs (97Q), a knock-in (KI) model that expresses a humanized AR containing a CAG expansion in the first exon, and a Tg myogenic model that overexpresses wild-type AR only in skeletal muscle fibers. We found that hindlimb muscles in the two Tg models (97Q and myogenic) showed marked losses in their intrinsic strength and resistance to fatigue, but were minimally affected in KI males. However, diseased muscles of all three models showed symptoms consistent with myotonic dystrophy type 1, namely, reduced resting membrane potential and deficits in chloride channel mRNA. These data indicate that muscle dysfunction is a core feature of SBMA caused by at least some of the same pathogenic mechanisms as myotonic dystrophy. Thus mechanisms controlling muscle function per se independent of mass are prime targets for SBMA therapeutics.

The effects of vitamin D3 during pregnancy and lactation on offspring physiology and behavior in sprague-dawley rats.

Recent findings show that developmental vitamin D deficiency leads to altered brain morphology and behavioral development in the rat offspring. We examined the effects of different dietary vitamin D levels in rat dams on behavior and biochemistry of the offspring. Females were divided into five conditions and received diets containing 0, 1,5, 3.3, 6.0, or 10.0 IU/g of vitamin D3 from mating to weaning. Offspring were tested as juveniles and as adults for anxiety, social learning and behavior, and locomotion. Results show that both deficient and excessive levels of vitamin D3 in juveniles lead to altered physiology and behavior. In juveniles but not adults, variations in vitamin D were related to variations in measures of anxiety and marginally, activity levels. For social behaviors, both juveniles and adults were affected by mothers' diets. In general, offspring of animals receiving abnormal concentrations of vitamin D showed the most deficits.