The Neural Basis for Biased Behavioral Responses Evoked by Galvanic Vestibular Stimulation in Primates.

Noninvasive electrical stimulation of the vestibular system in humans has become an increasingly popular tool with a broad range of research and clinical applications. However, common assumptions regarding the neural mechanisms that underlie the activation of central vestibular pathways through such stimulation, known as galvanic vestibular stimulation (GVS), have not been directly tested. Here, we show that GVS is encoded by VIIIth nerve vestibular afferents with nonlinear dynamics that differ markedly from those predicted by current models. GVS produced asymmetric activation of both semicircular canal and otolith afferents to the onset versus offset and cathode versus anode of applied current, that in turn produced asymmetric eye movement responses in three awake-behaving male monkeys. Additionally, using computational methods, we demonstrate that the experimentally observed nonlinear neural response dynamics lead to an unexpected directional bias in the net population response when the information from both vestibular nerves is centrally integrated. Together our findings reveal the neural basis by which GVS activates the vestibular system, establish that neural response dynamics differ markedly from current predictions, and advance our mechanistic understanding of how asymmetric activation of the peripheral vestibular system alters vestibular function. We suggest that such nonlinear encoding is a general feature of neural processing that will be common across different noninvasive electrical stimulation approaches.SIGNIFICANCE STATEMENT Here, we show that the application of noninvasive electrical currents to the vestibular system (GVS) induces more complex responses than commonly assumed. We recorded vestibular afferent activity in macaque monkeys exposed to GVS using a setup analogous to human studies. GVS evoked notable asymmetries in irregular afferent responses to cathodal versus anodal currents. We developed a nonlinear model explaining these GVS-evoked afferent responses. Our model predicts that GVS induces directional biases in centrally integrated head motion signals and establishes electrical stimuli that recreate physiologically plausible sensations of motion. Altogether, our findings provide new insights into how GVS activates the vestibular system, which will be vital to advancing new clinical and biomedical applications.

Vestibular control of deep and superficial lumbar muscles.

The active control of the lumbar musculature provides a stable platform critical for postures and goal-directed movements. Voluntary and perturbation-evoked motor commands can recruit individual lumbar muscles in a task-specific manner according to their presumed biomechanics. Here, we investigated the vestibular control of the deep and superficial lumbar musculature. Ten healthy participants were exposed to noisy electrical vestibular stimulation while balancing upright with their head facing forward, left, or right to characterize the differential modulation in the vestibular-evoked lumbar extensor responses in generating multidirectional whole body motion. We quantified the activation of the lumbar muscles on the right side using indwelling [deep multifidus, superficial multifidus, caudal longissimus (L4), and cranial longissimus (L1)] and high-density surface recordings. We characterized the vestibular-evoked responses using coherence and peak-to-peak cross-covariance amplitude between the vestibular and electromyographic signals. Participants exhibited responses in all lumbar muscles. The vestibular control of the lumbar musculature exhibited muscle-specific modulations: responses were larger in the longissimus (combined cranio-caudal) compared with the multifidus (combined deep-superficial) when participants faced forward (P < 0.001) and right (P = 0.011) but not when they faced left. The high-density surface recordings partly supported this observation: the location of the responses was more lateral when facing right compared with left (P < 0.001). The vestibular control of muscle subregions within the longissimus or the multifidus was similar. Our results demonstrate muscle-specific vestibular control of the lumbar muscles in response to perturbations of vestibular origin. The lack of differential activation of lumbar muscle subregions suggests the vestibular control of these subregions is co-regulated for standing balance.NEW & NOTEWORTHY We investigated the vestibular control of the deep and superficial lumbar extensor muscles using electrical vestibular stimuli. Vestibular stimuli elicited preferential activation of the longissimus muscle over the multifidus muscle. We did not observe clear regional activation of lumbar muscle subregions in response to the vestibular stimuli. Our findings show that the central nervous system can finely tune the vestibular control of individual lumbar muscles and suggest minimal regional variations in the activation of lumbar muscle subregions.

Cortical facilitation of somatosensory inputs using gravity-related tactile information in humans with vestibular hypofunction.

A few years after their bilateral vestibular loss, patients usually show a motor repertoire that is almost back to normal. This recovery is thought to involve an upregulation of the visual and proprioceptive information that compensates for the lack of vestibular information. Here, we investigated whether plantar tactile inputs, which provide body information relative to the ground and to the Earth vertical, contribute to this compensation. More specifically, we tested the hypothesis that somatosensory cortex response to electric stimulation of the plantar sole in standing adults will be greater in humans (n = 10) with bilateral vestibular hypofunction (VH) than in an age-matched healthy group (n = 10). Showing significantly greater somatosensory evoked potentials (i.e., P1N1) in VH than in control subjects, the electroencephalographic recordings supported this hypothesis. Furthermore, we found evidence that increasing the differential pressure between both feet, by adding a 1-kg mass at each pendant wrist, enhanced the internal representation of body orientation and motion relative to a gravitational reference frame. The large decrease in alpha power in the right posterior parietal cortex (and not in the left) is in line with this assumption. Finally, behavioral analyses showed that trunk oscillations were smaller than head oscillations in VH and showed a reverse pattern for healthy participants. These findings are consistent with a tactile-based postural control strategy in the absence of vestibular input and a vestibular-based control strategy in healthy participants where the head serves as a reference for balance control.NEW & NOTEWORTHY Somatosensory cortex excitability is greater in participants with bilateral vestibular hypofunction than in age-matched healthy humans. To control balance, healthy humans "locked" the head whereas participants with vestibular hypofunction "locked" their pelvis. For participants with vestibular hypofunction, increasing loading/unloading of the feet enhances the internal representation of body state in the posterior parietal cortex.

Noninvasive prenatal diagnosis of Mendelian disorders for consanguineous couples by relative genotype dosage.

Noninvasive prenatal diagnosis relies on the presence in maternal blood of circulating cell-free fetal DNA released by apoptotic trophoblast cells. Widely used for aneuploidy screening, it can also be applied to monogenic diseases (NIPD-M) in case of known parental mutations. Due to the confounding effect of maternal DNA, detection of maternal or biparental mutations requires relative haplotype dosage (RHDO), a method relying on the presence of SNPs that are heterozygous in one parent and homozygous in the other. Unavoidably, there is a risk of test failure by lack of such informative SNPs, an event particularly likely for consanguineous couples who often share common haplotypes in regions of identity-by-descent. Here we present a novel approach, relative genotype dosage (RGDO) that bypasses this predicament by directly assessing fetal genotype with SNPs that are heterozygous in both parents (frequent in regions of identity-by-descent). We show that RGDO is as sensitive as RHDO and that it performs well over a large range of fetal fractions and DNA amounts, thereby opening NIPD-M to most consanguineous couples. We also report examples of couples, consanguineous or not, where combining RGDO and RHDO allowed a diagnosis that would not have been possible with only one approach.

Atypical diabetes with spontaneous remission associated with systemic lupus erythematosus in an adolescent girl of African ancestry, a case report.

BACKGROUND: New-onset diabetes in youth encompasses type 1 diabetes, type 2 diabetes, monogenic diabetes, and rarer subtypes like Type B insulin resistance syndrome and ketosis-prone atypical diabetes in African populations. Some cases defy classification, posing management challenges. Here, we present a case of a unique, reversible diabetes subtype. CASE PRESENTATION: We describe an adolescent African girl recently diagnosed with systemic lupus erythematosus. At age 15, she presented with ketoacidosis, HbA1c of 108.7 mmol/mol (12.1%), and positive anti-insulin antibodies. Initially diagnosed with type 1 diabetes, insulin was prescribed. Due to the presence of obesity and signs of insulin resistance, we added metformin. Concurrently, she received treatment for lupus with hydroxychloroquine, mycophenolate mofetil, and prednisone. After discharge, she stopped insulin due to cultural beliefs. Five months later, her glycemia and HbA1c normalized (37 mmol/mol or 5.5%) without insulin, despite corticosteroid therapy and weight gain. Autoantibodies normalized, and lupus activity decreased. Genetic testing for monogenic diabetes was negative, and the type 1 genetic risk score was exceptionally low. CONCLUSIONS: We present a complex, reversible diabetes subtype. Features suggest an autoimmune origin, possibly influenced by overlapping HLA risk haplotypes with lupus. Lupus treatment or immunomodulation may have impacted diabetes remission. Ancestry-tailored genetic risk scores are currently designed to improve diagnostic accuracy.

Iron chelation rescues hemolytic anemia and skin photosensitivity in congenital erythropoietic porphyria.

Congenital erythropoietic porphyria (CEP) is an inborn error of heme synthesis resulting from uroporphyrinogen III synthase (UROS) deficiency and the accumulation of nonphysiological porphyrin isomer I metabolites. Clinical features are heterogeneous among patients with CEP but usually combine skin photosensitivity and chronic hemolytic anemia, the severity of which is related to porphyrin overload. Therapeutic options include symptomatic strategies only and are unsatisfactory. One promising approach to treating CEP is to reduce the erythroid production of porphyrins through substrate reduction therapy by inhibiting 5-aminolevulinate synthase 2 (ALAS2), the first and rate-limiting enzyme in the heme biosynthetic pathway. We efficiently reduced porphyrin accumulation after RNA interference-mediated downregulation of ALAS2 in human erythroid cellular models of CEP disease. Taking advantage of the physiological iron-dependent posttranscriptional regulation of ALAS2, we evaluated whether iron chelation with deferiprone could decrease ALAS2 expression and subsequent porphyrin production in vitro and in vivo in a CEP murine model. Treatment with deferiprone of UROS-deficient erythroid cell lines and peripheral blood CD34+-derived erythroid cultures from a patient with CEP inhibited iron-dependent protein ALAS2 and iron-responsive element-binding protein 2 expression and reduced porphyrin production. Furthermore, porphyrin accumulation progressively decreased in red blood cells and urine, and skin photosensitivity in CEP mice treated with deferiprone (1 or 3 mg/mL in drinking water) for 26 weeks was reversed. Hemolysis and iron overload improved upon iron chelation with full correction of anemia in CEP mice treated at the highest dose of deferiprone. Our findings highlight, in both mouse and human models, the therapeutic potential of iron restriction to modulate the phenotype in CEP.

Neural Mechanisms Underlying High-Frequency Vestibulocollic Reflexes In Humans And Monkeys.

The vestibulocollic reflex is a compensatory response that stabilizes the head in space. During everyday activities, this stabilizing response is evoked by head movements that typically span frequencies from 0 to 30 Hz. Transient head impacts, however, can elicit head movements with frequency content up to 300-400 Hz, raising the question whether vestibular pathways contribute to head stabilization at such high frequencies. Here, we first established that electrical vestibular stimulation modulates human neck motor unit (MU) activity at sinusoidal frequencies up to 300 Hz, but that sensitivity increases with frequency up to a low-pass cutoff of ∼70-80 Hz. To examine the neural substrates underlying the low-pass dynamics of vestibulocollic reflexes, we then recorded vestibular afferent responses to the same electrical stimuli in monkeys. Vestibular afferents also responded to electrical stimuli up to 300 Hz, but in contrast to MUs their sensitivity increased with frequency up to the afferent resting firing rate (∼100-150 Hz) and at higher frequencies afferents tended to phase-lock to the vestibular stimulus. This latter nonlinearity, however, was not transmitted to neck motoneurons, which instead showed minimal phase-locking that decreased at frequencies >75 Hz. Similar to human data, we validated that monkey muscle activity also exhibited low-pass filtered vestibulocollic reflex dynamics. Together, our results show that neck MUs are activated by high-frequency signals encoded by primary vestibular afferents, but undergo low-pass filtering at intermediate stages in the vestibulocollic reflex. These high-frequency contributions to vestibular-evoked neck muscle responses could stabilize the head during unexpected head transients.SIGNIFICANCE STATEMENT Vestibular-evoked neck muscle responses rely on accurate encoding and transmission of head movement information to stabilize the head in space. Unexpected transient events, such as head impacts, are likely to push the limits of these neural pathways since their high-frequency features (0-300 Hz) extend beyond the frequency bandwidth of head movements experienced during everyday activities (0-30 Hz). Here, we demonstrate that vestibular primary afferents encode high-frequency stimuli through frequency-dependent increases in sensitivity and phase-locking. When transmitted to neck motoneurons, these signals undergo low-pass filtering that limits neck motoneuron phase-locking in response to stimuli >75 Hz. This study provides insight into the neural dynamics producing vestibulocollic reflexes, which may respond to high-frequency transient events to stabilize the head.

Molecular characterization of pathogenic OTOA gene conversions in hearing loss patients.

Bi-allelic loss-of-function variants of OTOA are a well-known cause of moderate-to-severe hearing loss. Whereas non-allelic homologous recombination-mediated deletions of the gene are well known, gene conversions to pseudogene OTOAP1 have been reported in the literature but never fully described nor their pathogenicity assessed. Here, we report two unrelated patients with moderate hearing-loss, who were compound heterozygotes for a converted allele and a deletion of OTOA. The conversions were initially detected through sequencing depths anomalies at the OTOA locus after exome sequencing, then confirmed with long range polymerase chain reactions. Both conversions lead to loss-of-function by introducing a premature stop codon in exon 22 (p.Glu787*). Using genomic alignments and long read nanopore sequencing, we found that the two probands carry stretches of converted DNA of widely different lengths (at least 9 kbp and around 900 bp, respectively).

A task-relevant experimental pain model to target motor adaptation.

KEY POINTS: Motor adaptation is thought to be a strategy to avoid pain. Current experimental pain models do not allow for consistent modulation of pain perception depending on movement. We showed that low-frequency sinusoidal stimuli delivered at painful intensity result in minimal habituation of pain perception (over 60 s) and minimal stimulation artefacts on electromyographic signals. When the amplitude of the low-frequency sinusoidal stimuli was modulated based on the vertical force participants applied to the ground with their right leg while standing upright, we demonstrated a strong association between perceived pain and motor adaptation. By enabling task-relevant modulation of perceived pain intensity and the recording electromyographic signals during electrical painful stimulation, our novel pain model will permit direct experimental testing of the relationship between pain and motor adaptation. ABSTRACT: Contemporary pain adaptation theories predict that motor adaptation occurs to limit pain. Current experimental pain models, however, do not allow for pain intensity modulation according to one's posture or movements. We developed a task-relevant experimental pain model using low-frequency sinusoidal electrical stimuli applied over the infrapatellar fat pad. In fourteen participants, we compared perceived pain habituation and stimulation-induced artefacts in vastus medialis electromyographic recordings elicited by sinusoidal (4, 10, 20 and 50 Hz) and square electrical waveforms delivered at constant peak stimulation amplitude. Next, we simulated a clinical condition where perceived knee pain intensity is proportional to the load applied on the leg by controlling sinusoidal current amplitude (4 Hz) according to the vertical force the participants applied with their right leg to the ground while standing upright. Pain ratings habituated over a 60 s period for 50 Hz sinusoidal and square waveforms but not for low-frequency sinusoidal stimuli (P < 0.001). EMG filters removed most stimulation artefacts for low-frequency sinusoidal stimuli (4 Hz). While balancing upright, participants' pain ratings were correlated with the force applied by the right leg (R2  = 0.65), demonstrating task-relevant changes in perceived pain intensity. Low-frequency sinusoidal stimuli can induce knee pain of constant intensity for 60 s with minimal EMG artefacts while enabling task-relevant pain modulation when controlling current amplitude. By enabling task-dependent modulation of perceived pain intensity, our novel experimental model replicates key temporal aspects of clinical musculoskeletal pain while allowing quantification of neuromuscular activation during painful electrical stimulation. This approach will enable researchers to test the predicted relationship between movement strategies and pain.

Soleus responses to Achilles tendon stimuli are suppressed by heel and enhanced by metatarsal cutaneous stimuli during standing.

KEY POINTS: We examined the influence of cutaneous feedback from the heel and metatarsal regions of the foot sole on the soleus stretch reflex pathway during standing. We found that heel electrical stimuli suppressed and metatarsal stimuli enhanced the soleus vibration response. Follow-up experiments indicated that the interaction between foot sole cutaneous feedback and the soleus vibration response was likely not mediated by presynaptic inhibition and was contingent upon a modulation at the ⍺-motoneuron pool level. The spatially organized interaction between cutaneous feedback from the foot sole and the soleus vibration response provides information about how somatosensory information is combined to appropriately respond to perturbations during standing. ABSTRACT: Cutaneous feedback from the foot sole provides balance-relevant information and has the potential to interact with spinal reflex pathways. In this study, we examined how cutaneous feedback from the foot sole (heel and metatarsals) influenced the soleus response to proprioceptive stimuli during standing. We delivered noisy vibration (10-115 Hz) to the right Achilles tendon while we intermittently applied electrical pulse trains (five 1-ms pulses at 200 Hz, every 0.8-1.0 s) to the skin under either the heel or the metatarsals of the ipsilateral foot sole. We analysed time-dependent (referenced to cutaneous stimuli) coherence and cross-correlations between the vibration acceleration and rectified soleus EMG. Vibration-EMG coherence was observed across a bandwidth of ∼10-80 Hz, and coherence was suppressed by heel but enhanced by metatarsal cutaneous stimuli. Cross-correlations showed soleus EMG was correlated with the vibration (∼40 ms lag) and cross-correlations were also suppressed by heel (from 104-155 ms) but enhanced by metatarsal (from 76-128 ms) stimuli. To examine the neural mechanisms mediating this reflex interaction, we conducted two further experiments to probe potential contributions from (1) presynaptic inhibition, and (2) modulations at the ⍺- and γ-motoneuron pools. Results suggest the cutaneous interactions with the stretch reflex pathway required a modulation at the ⍺-motoneuron pool and were likely not mediated by presynaptic inhibition. These findings demonstrate that foot sole cutaneous information functionally tunes the stretch reflex pathway during the control of upright posture and balance.

Bi-allelic loss of ERGIC1 causes relatively mild arthrogryposis.

Arthrogryposis describes the presence of multiple joint-contractures. Clinical severity of this phenotype is variable, and more than 400 causative genes have been proposed. Among these, ERGIC1 is a recently reported candidate encoding a putative transmembrane protein of the ER-Golgi interface. Two homozygous missense variants have been reported in patients with relatively mild non-syndromic arthrogryposis. In a consanguineous family with two affected siblings presenting congenital arthrogryposis and some facial dysmorphism we performed prenatal array-CGH, postnatal targeted exome and genome sequencing. Genome sequencing identified a homozygous 22.6 Kb deletion encompassing the promoter and first exon of ERGIC1. mRNA quantification showed the complete absence of ERGIC1 expression in the two affected siblings and a decrease in heterozygous parents. Our observations validate the pathogenic role of ERGIC1 in congenital arthrogryposis and demonstrate that complete loss of function causes a relatively mild phenotype. These findings will contribute to improve genetic counseling of ERGIC1 mutations.

Immune deficiency, autoimmune disease and intellectual disability: A pleiotropic disorder caused by biallelic variants in the TPP2 gene.

Four individuals from two families presented with a multisystemic condition of suspected genetic origin that was diagnosed only after genome analysis. The main phenotypic features were immune system dysregulation (severe immunodeficiency with autoimmunity) and intellectual disability. The four individuals were found to be homozygous for a 4.4 Kb deletion removing exons 20-23 (NM_003291.4) of the TPP2 gene, predicting a frameshift with premature termination of the protein. The deletion was located on a shared chromosome 13 haplotype indicating a Swiss founder mutation. Tripeptidyl peptidase 2 (TPP2) is a protease involved in HLA/antigen complex processing and amino acid homeostasis. Biallelic variants in TPP2 have been described in 10 individuals with variable features including immune deficiency, autoimmune cytopenias, and intellectual disability or chronic sterile brain inflammation mimicking multiple sclerosis. Our observations further delineate this severe condition not yet included in the OMIM catalog. Timely recognition of TPP2 deficiency is crucial since (1) immune surveillance is needed and hematopoietic stem cell transplantation may be necessary, and (2) for provision of genetic counselling. Additionally, enzyme replacement therapy, as already established for TPP1 deficiency, might be an option in the future.

Regional activation in the human longissimus thoracis pars lumborum muscle.

KEY POINTS: •Longissimus activity in the lumbar region was measured using indwelling electromyography to characterize the territory of its motor units. •The distribution of motor units in the longissimus pars lumborum muscle was mainly grouped into two distinct regions. •Regional activation of the longissimus pars lumborum was also observed during functional tasks involving trunk movements. •The regional activation of the longissimus pars lumborum muscle may play a role in segmental stabilization of the lumbar spine. ABSTRACT: The longissimus pars lumborum contributes to lumbar postural control and movement. While animal studies suggest a segmental control of this muscle, the territory of motor units constituting the human longissimus pars lumborum remains unknown. The aims of this study were to identify the localization of motor unit territories in the longissimus and assess the activation of this muscle during functional tasks. Eight healthy participants were recruited. During isometric back extension contractions, single motor-unit (at L1, L2, L3 and L4) and multi-unit indwelling recordings (at L1, L1-L2, L2, L2-L3, L3, L3-L4 and L4) were used to estimate motor unit territories in the longissimus pars lumborum based on the motor-unit spike-triggered averages from fine-wire electrodes. A series of functional tasks involving trunk and arm movements were also performed. A total of 73 distinct motor units were identified along the length of the longissimus: only two motor units spanned all recording sites. The majority of the recorded motor units had muscle fibres located in two main rostro-caudal territories (32 motor units spanned L1 to L3 and 30 spanned ∼L3 to L4) and 11 had muscle fibres outside these two main territories. We also observed distinct muscle activation between the rostral and caudal regions of the longissimus pars lumborum during a trunk rotation task. Our results show clear rostral and caudal motor unit territories in the longissimus pars lumborum muscle and suggest that the central nervous system can selectively activate regions of the superficial lumbar muscles to provide local stabilization of the spine.

Influence of age on the frequency characteristics of the soleus muscle response to Achilles tendon vibration during standing.

KEY POINTS: Proprioceptive sensory information from the ankle joint is critical for the control of upright posture and balance. We examined the influence of age (n = 54 healthy adults, 20-82 years old) on lower limb muscle responses to proprioceptive perturbations evoked by Achilles tendon vibration during standing. The frequency bandwidth of the muscle response became narrower, and the gain (the muscle response relative to the stimulus) and scaling (increases in response amplitude with increases in stimulus amplitude) decreased with age. Mechanics of the muscle-tendon unit (mechanical admittance) did not differ with age during standing, and thus probably did not mediate the age-related changes observed in soleus muscle responses to vibration. These findings add to our understanding of how altered proprioceptive responses may contribute to impaired mobility and falls with ageing. ABSTRACT: Proprioceptive information from the ankle joint plays an important role in the control of upright posture and balance. Ageing influences many components of the sensorimotor system, which leads to poor mobility and falls. However, little is known about the influence of age on the characteristics of short latency muscle responses to proprioceptive stimuli during standing across frequencies that are encoded by muscle spindles. We examined the frequency characteristics of the soleus muscle response to noisy (10-115 Hz) Achilles tendon vibration during standing in 54 healthy adults across a broad age range (20-82 years). The results showed the frequency bandwidth of the soleus response (vibration-electromyography coherence) became progressively narrower with ageing. Coherence was significantly lower in middle-aged relative to young adults between ∼7-11 and 28-62 Hz, lower in older relative to middle-aged adults between ∼30-50 Hz and lower in older relative to young adults between ∼7-64 Hz. Muscle response gain was similar between age groups at low frequencies, although gain was lower in older relative to young adults between ∼28-54 Hz. Across the age range, the response amplitude (peak-to-peak cross-covariance) and the scaling of the response with stimulus amplitude were both negatively correlated with age. Muscle-tendon mechanics (admittance) did not differ with age, suggesting this did not mediate differences in soleus responses. Our findings suggest there is a progressive change in the soleus response to proprioceptive stimuli with ageing during standing, which could contribute to poorer mobility and falls.

Independent Early and Late Sensory Processes for Proprioceptive Integration When Planning a Step.

Somatosensory inputs to the cortex undergo an early and a later stage of processing which are characterized by an early and a late somatosensory evoked potentials (SEP). The early response is highly representative of the stimulus characteristics whereas the late response reflects a more integrative, task specific, stage of sensory processing. We hypothesized that the later processing stage is independent of the early processing stage. We tested the prediction that a reduction of the first volley of input to the cortex should not prevent the increase of the late SEP. Using the sensory interference phenomenon, we halved the amplitude of the early response to somatosensory input of the ankle joints (evoked by vibration) when participants either planned a step forward or remained still. Despite the initial cortical response to the vibration being massively decreased in both tasks, the late response was still enhanced during step planning. Source localization indicated the posterior parietal cortex (PPC) as the likely origin of the late response modulation. Overall these results support the dissociation between the processes underlying the early and late SEP. The later processing stage could involve both direct and indirect thalamic connections to PPC which bypass the postcentral somatosensory cortex.

Virtual signals of head rotation induce gravity-dependent inferences of linear acceleration.

KEY POINTS: Considerable debate exists regarding whether electrical vestibular stimuli encoded by vestibular afferents induce a net signal of linear acceleration, rotation or a combination of the two. This debate exists because an isolated signal of head rotation encoded by the vestibular afferents can cause perceptions of both linear and angular motion. We recorded participants' perceptions in different orientations relative to gravity and predicted their responses by modelling the effect of electrical vestibular stimuli on vestibular afferents and a current model of central vestibular processing. We show that, even if electrical vestibular stimuli are encoded as a net signal of head rotation, participants perceive both linear acceleration and rotation motions, provided the electrical stimulation-induced rotational vector has a component orthogonal to gravity. The emergence of a perception of linear acceleration from a single rotational input signal clarifies the origins of the neural mechanisms underlying electrical vestibular stimulation. ABSTRACT: Electrical vestibular stimulation (EVS) is an increasingly popular biomedical tool for generating sensations of virtual motion in humans, for which the mechanism of action is a topic of considerable debate. Contention surrounds whether the evoked vestibular afferent activity encodes a signal of net rotation and/or linear acceleration. Central processing of vestibular self-motion signals occurs through an internal representation of gravity that can lead to inferred linear accelerations in absence of a true inertial acceleration. Applying this model to virtual signals of rotation evoked by EVS, we predict that EVS will induce behaviours attributed to both angular and linear motion, depending on the head orientation relative to gravity. To demonstrate this, 18 subjects indicated their perceived motion during sinusoidal EVS when in one of four head/body positions orienting the gravitational vector parallel or orthogonal to the EVS rotation vector. During stimulation, participants selected one simulated movement from seven that corresponded best to what they perceived. Participants' responses in each orientation were predicted by a model combining the influence of EVS on vestibular afferents with known mechanisms of vestibular processing. When the EVS rotation vector had a component orthogonal to gravity, human perceptual responses were consistent with a non-zero central estimate of interaural or superior-inferior linear acceleration. The emergence of a perception of linear acceleration from a single rotational input signal clarifies the origins of the neural mechanisms underlying EVS, which has important implications for its use in human biomedical or sensory augmentation applications.

Soleus single motor units show stronger coherence with Achilles tendon vibration across a broad bandwidth relative to medial gastrocnemius units while standing.

To probe the frequency characteristics of somatosensory responses in the triceps surae muscles, we previously applied suprathreshold noisy vibration to the Achilles tendon and correlated it with ongoing triceps surae muscle activity (recorded via surface EMG) during standing. Stronger responses to tendon stimuli were observed in soleus (Sol) relative to medial gastrocnemius (MGas) surface EMG; however, it is unknown whether differences in motor unit activity or limitations of surface EMG could have influenced this finding. Here, we inserted indwelling EMG into Sol and MGas to record the activity of single motor units while we applied noisy vibration (10-115 Hz) to the right Achilles tendon of standing participants. We analyzed the relationship between vibration acceleration and the spike activity of active single motor units through estimates of coherence, gain, phase, and cross-covariance. We also applied sinusoidal vibration at frequencies from 10 to 100 Hz (in 5-Hz increments) to examine whether motor units demonstrate nonlinear synchronization or phase locking at higher frequencies. Relative to MGas single motor units, Sol units demonstrated stronger coherence and higher gain with noisy vibration across a bandwidth of 7-68 Hz, and larger peak-to-peak cross-covariance at all four stimulus amplitudes examined. Sol and MGas motor unit activity was modulated over the time course of the sinusoidal stimuli across all frequencies, but their phase-locking behavior was minimal. These findings suggest Sol plays a prominent role in responding to disturbances transmitted through the Achilles tendon across a broad frequency band during standing.NEW & NOTEWORTHY We examined the relationship between Achilles tendon stimuli and spike times of single soleus (Sol) and medial gastrocnemius (MGas) motor units during standing. Relative to MGas, Sol units demonstrated stronger coherence and higher gain with noisy stimuli across a bandwidth of 7-68 Hz. Sol and MGas units demonstrated minimal nonlinear phase locking with sinusoidal stimuli. These findings indicate Sol plays a prominent role in responding to tendon stimuli across a broad frequency band.

Missense UROS mutations causing congenital erythropoietic porphyria reduce UROS homeostasis that can be rescued by proteasome inhibition.

Congenital erythropoietic porphyria (CEP) is an inborn error of heme biosynthesis characterized by uroporphyrinogen III synthase (UROS) deficiency resulting in deleterious porphyrin accumulation in blood cells responsible for hemolytic anemia and cutaneous photosensitivity. We analyzed here the molecular basis of UROS impairment associated with twenty nine UROS missense mutations actually described in CEP patients. Using a computational and biophysical joint approach we predicted that most disease-causing mutations would affect UROS folding and stability. Through the analysis of enhanced green fluorescent protein-tagged versions of UROS enzyme we experimentally confirmed these data and showed that thermodynamic instability and premature protein degradation is a major mechanism accounting for the enzymatic deficiency associated with twenty UROS mutants in human cells. Since the intracellular loss in protein homeostasis is in excellent agreement with the in vitro destabilization, we used molecular dynamic simulation to rely structural 3D modification with UROS disability. We found that destabilizing mutations could be clustered within three types of mechanism according to side chain rearrangements or contact alterations within the pathogenic UROS enzyme so that the severity degree correlated with cellular protein instability. Furthermore, proteasome inhibition using bortezomib, a clinically available drug, significantly enhanced proteostasis of each unstable UROS mutant. Finally, we show evidence that abnormal protein homeostasis is a prevalent mechanism responsible for UROS deficiency and that modulators of UROS proteolysis such as proteasome inhibitors or chemical chaperones may represent an attractive therapeutic option to reduce porphyrin accumulation and prevent skin photosensitivity in CEP patients when the genotype includes a missense variant.

Targeted gene therapy in human-induced pluripotent stem cells from a patient with primary hyperoxaluria type 1 using CRISPR/Cas9 technology.

Primary hyperoxaluria type 1 (PH1) is an inherited metabolic disorder caused by a deficiency of the peroxisomal enzyme alanine-glyoxylate aminotransferase (AGT), which leads to overproduction of oxalate by the liver and results in urolithiasis, nephrocalcinosis and renal failure. The only curative treatment for PH1 is combined liver and kidney transplantation, which is limited by the lack of suitable organs, significant complications, and the life-long requirement for immunosuppressive agents to maintain organ tolerance. Hepatocyte-like cells (HLCs) generated from CRISPR/Cas9 genome-edited human-induced pluripotent stem cells would offer an attractive unlimited source of autologous gene-corrected liver cells as an alternative to orthotopic liver transplantation (OLT). Here we report the CRISPR/Cas9 nuclease-mediated gene targeting of a single-copy AGXT therapeutic minigene into the safe harbour AAVS1 locus in PH1-induced pluripotent stem cells (PH1-iPSCs) without off-target inserts. We obtained a robust expression of a codon-optimized AGT in HLCs derived from AAVS1 locus-edited PH1-iPSCs. Our study provides the proof of concept that CRISPR/Cas9-mediated integration of an AGXT minigene into the AAVS1 safe harbour locus in patient-specific iPSCs is an efficient strategy to generate functionally corrected hepatocytes, which in the future may serve as a source for an autologous cell-based gene therapy for the treatment of PH1.

Genetic background influences hepcidin response to iron imbalance in a mouse model of hemolytic anemia (Congenital erythropoietic porphyria).

Clinical severity is heterogeneous among patients suffering from congenital erythropoietic porphyria (CEP) suggesting a modulation of the disease (UROS deficiency) by environmental factors and modifier genes. A KI model of CEP due to a missense mutation of UROS gene present in human has been developed on 3 congenic mouse strains (BALB/c, C57BL/6, and 129/Sv) in order to study the impact of genetic background on disease severity. To detect putative modifiers of disease expression in congenic mice, hematologic data, iron parameters, porphyrin content and tissue samples were collected. Regenerative hemolytic anemia, a consequence of porphyrin excess in RBCs, had various expressions: 129/Sv mice were more hemolytic, BALB/c had more regenerative response to anemia, C57BL/6 were less affected. Iron status and hemolysis level were directly related: C57BL/6 and BALB/c had moderate hemolysis and active erythropoiesis able to reduce iron overload in the liver, while, 129/Sv showed an imbalance between iron release due to hemolysis and erythroid use. The negative control of hepcidin on the ferroportin iron exporter appeared strain specific in the CEP mice models tested. Full repression of hepcidin was observed in BALB/c and 129/Sv mice, favoring parenchymal iron overload in the liver. Unchanged hepcidin levels in C57BL/6 resulted in retention of iron predominantly in reticuloendothelial tissues. These findings open the field for potential therapeutic applications in the human disease, of hepcidin agonists and iron depletion in chronic hemolytic anemia.