The Journal of Physiology

Motoneurons adapt to both resistance and endurance training in reduced animal preparations, with adaptations seemingly more apparent in higher threshold neurons, but similar evidence in humans is lacking. Here, we compared the identified motor unit (MU) discharge patterns from decomposed electromyography signals acquired during triangular dorsiflexion contractions up to 70% of maximal voluntary force (MVF) between resistance-trained, endurance-trained, and untrained individuals (n=23 in each group). We then estimated intrinsic motoneuron properties and garnered insight about the proportion of excitatory, inhibitory, and neuromodulatory inputs contributing to motor commands across contraction intensities in each group. Participants also performed a task where a triangular contraction was superimposed onto a sustained one designed to challenge inhibitory control of dendritic persistent inward currents (PICs). Both trained groups demonstrated greater MU discharge rates with greater ascending discharge rate modulation during higher contraction forces ([≥]50% MVF), which were accompanied by more linear MU discharge patterns and greater post-acceleration attenuation slopes of the ascending discharge rates. No differences in discharge rate hysteresis or the discharge rate characteristics during the sombrero tasks between groups, suggesting no differences in neuromodulatory input. Conversely, resistance-compared to endurance-trained individuals exhibited greater acceleration slopes during lower contractions forces ([≤]50% MVF), indicating the possibility of enhanced initial activation of PICs. Collectively, the greater and more linear MU discharge patterns in the trained groups either suggests a more reciprocal (i.e., push-pull) excitation-inhibition coupling during higher contraction forces or enhanced excitatory synaptic input to the motor pool, which might underpin greater force production of trained individuals.

The intrinsic properties of spinal motoneurons support flexible movement, including the maintenance of postural tone. Motoneurons can produce sustained action potential output that outlasts synaptic input, a phenomenon traditionally attributed to persistent inward currents (PICs) mediated by sodium and calcium channels. Using whole-cell patch clamp electrophysiology, we examined how specific ion channels contribute to PIC maturation and non-linear firing dynamics that allow motoneurons to sustain their output in fast and slow lumbar motoneurons across postnatal development in mice. PIC amplitude and non-linear firing dynamics increased after weight bearing in fast but not slow motoneurons. Blocking Nav1.6 channels reduced PIC amplitude at both pre- and post-weight-bearing stages, whereas L-type calcium channel blockade only reduced PICs after weight bearing emerged. However, reducing PIC amplitude--either individually or in combination--did not abolish sustained firing hysteresis. Unexpectedly, activation of muscarinic receptors increased PIC amplitude while promoting adaptive firing dynamics, suggesting that PICs alone do not drive this behavior. Instead, pharmacological manipulation of potassium currents mediated by KCNQ and Kv1.2 channels, which oppose PICs, produced substantial changes in firing dynamics. Strikingly, blocking HCN channels promoted sustained firing dynamics and led to the emergence of self-sustained firing in fast motoneurons. These results indicate that while PICs and non-linear firing dynamics mature together, sustained firing relies on mechanisms beyond PICs, with potassium and HCN channels playing key modulatory roles. Key PointsO_LIPersistent inward currents (PICs) and recruitment-derecruitment hysteresis increase in parallel in fast, but not slow, motoneurons following the onset of hindlimb weight bearing. C_LIO_LIIncreased expression or function of L-type calcium channels may contribute to enhanced PICs in fast motoneurons after weight bearing emerges. C_LIO_LINeither Nav1.6 nor L-type calcium channels are required for sustained firing hysteresis in fast motoneurons. C_LIO_LIKCNQ channels attenuate PICs and, together with Kv1.2 channels, shape recruitment-derecruitment asymmetry, thereby modulating firing hysteresis in fast motoneurons. C_LIO_LIHCN channels generate a resting H-current that delays recruitment, modulates firing hysteresis, and prevents the emergence of self-sustained firing in fast motoneurons. C_LI

1The energetic cost of skeletal muscle contraction is a fundamental driver in the selection of locomotor strategies. Muscle energy consumption depends on muscle-fibre typology, neural drive, and mechanical state. While the influence of fibre-type and mechanical state on energy use has been extensively characterised in isolated muscle preparations, these experiments fail to capture the influence of realistic in vivo motor unit recruitment strategies. Hence, this study aims to capture neural drive in the tibialis anterior, in particular motor unit recruitment thresholds and rate coding, and the corresponding changes in energy use across a range of mechanical demands. Fixed ankle angle dorsiflexion contractions were performed at varying rates of torque development, while high density electromyography characterized motor unit spiking activity, B-mode ultrasound captured muscle fascicle dynamics, and indirect calorimetry measured energetic rates. Faster rates of torque development required earlier recruitment of motor units with increased motor unit firing rates which coincided with increased fascicle strain rates. At higher rates of torque development, these altered motor unit discharge patterns and muscle mechanics coincided with increased muscle energy use. Together, these findings highlight that in vivo muscle energetics emerge from the dynamic interplay between neural control and mechanical demands.

The size principle is a theory of motor unit (MU) recruitment that suggests MUs are recruited in an orderly manner from the smallest (lower threshold) to the largest (higher threshold) MUs. A consequence of this biophysical theory is that, for isometric contractions, recruitment is dependent on the intensity of actual effort required to meet task demands. This concept has been supported by modelling work demonstrating that, in tasks performed to momentary failure, full MU recruitment will have occurred upon reaching failure irrespective of the force requirements of the task. However, in vivo studies examining this are limited. Therefore, the aim of the current study was to examine MU recruitment of the quadriceps under both higher- and lower-torque (70% and 30% of MVC, respectively) isometric knee extension, performed to momentary failure. Specifically, we compared surface electromyography (sEMG) frequency characteristics, determined by wavelet analysis, across the two continuous isometric knee extension tasks to identify potential differences in recruitment patterns. A convenience sample of 10 recreationally active adult males (height: 179.6{+/-}6.0 cm; mass: 76.8{+/-}7.3 kg; age: 26{+/-}7 years) with previous resistance training experience (6{+/-}3 years) were recruited. Using a within-session, repeated-measures, randomised crossover design participants performed the knee extension tasks whilst sEMG was collected from the vastus medialis (VM), rectus femoris (RF) and vastus lateralis (VL). Myoelectric signals were decomposed into intensities as a function of time and frequency using an EMG-specific wavelet transformation. Our first analysis compared the mean frequency at momentary failure; second, we investigated the effects of load on relative changes in wavelet intensities; finally, we quantified the degree of wavelet similarity over time. Wavelet-based calculation of the mean signal frequency appeared to show similar mean frequency characteristics occurring when reaching momentary failure. However, individual wavelets revealed that different changes in frequency components occurred between the two tasks, suggesting that patterns of recruitment differed. Low-torque conditions resulted in an increase in intensity of all frequency components across the trials for each muscle whereas high-torque conditions resulted in a wider range of frequency components contained within the myoelectric signals at the beginning of the trials. However, as the low-torque trial neared momentary failure there was an increased agreement between conditions across wavelets. Our results corroborate modelling studies as well as recent biopsy evidence, suggesting overall MU recruitment may largely be similar for isometric tasks performed to momentary failure with the highest threshold MUs likely recruited, despite being achieved with differences in the pattern of recruitment over time utilised.

Incomplete spinal cord injury disrupts voluntary movement, in part through motoneuron dysfunction, yet the mechanisms underlying this dysfunction remain poorly understood. Using a non-invasive approach to decode the spiking activity of large populations of spinal motoneurons, we quantified the relative contributions of excitatory, inhibitory, and neuromodulatory inputs to motoneuron rate coding after chronic incomplete spinal cord injury. Eighteen participants with incomplete spinal cord injury and 18 age- and sex-matched control participants performed submaximal isometric plantar flexion tasks while high-density surface electromyography was recorded from the soleus and gastrocnemius medialis muscles. Motoneuron firing behaviour was analysed to estimate neuromodulatory drive and the balance between inhibitory and excitatory inputs. Participants with incomplete spinal cord injury exhibited lower rate coding, characterised by lower firing rates at recruitment, lower firing rate modulation, and lower peak firing rates compared with healthy controls. Although estimates of neuromodulatory drive did not differ between groups, individuals with spinal cord injury showed a shift in the inhibition-excitation balance toward greater inhibition compared with controls. Furthermore, increasing inhibitory input through muscle length changes and antagonist tendon vibration modulated motoneuron firing in controls, but not in individuals with incomplete spinal cord injury. Together, these findings suggest that impaired rate coding after incomplete spinal cord injury arises from an altered inhibitory-excitatory balance rather than reduced neuromodulatory drive. Taking advantage of methodological advances to decode spinal motor neuron activity during voluntary contraction, this study identified excessive inhibitory input to spinal motoneurons as a key neural mechanism contributing to muscle weakness and impaired motor function in individuals with incomplete spinal cord injury.

We were interested in whether stretching the muscle via elastic tissue recoil during partial muscle deactivation could trigger stretch-induced mechanisms that subsequently enhance the muscles steady-state neuromechanical output. Two torque-controlled experiments were conducted to test this aim. In Experiment 1, fifteen participants performed fixed-end dorsiflexion contractions to a moderate or high-then-moderate level with torque-drop rates of 0, 10, 20, 40, or 80% MVC{middle dot}s-1 from 60-40% MVC, while net ankle joint torque, tibialis anterior (TA) muscle activity level, and TA ultrasound images were recorded. The same measurements were performed in Experiment 2, which tested twelve different participants who performed fixed-end contractions to three reference levels or to a higher-then-lower level (with torque-drop amplitudes of 85-45, 85-30, and 85-15% MVC at 20% MVC{middle dot}s-1). Increased fascicle shortening amplitudes (Experiment 1: 1-2 mm, p[&le;].049; Experiment 2: 5-10 mm, p<.001) to initially higher joint torques and different fascicle stretch rates (0.6 to 4.3 mm{middle dot}s-1, p[&le;].041) and amplitudes (4-9 mm; p[&le;].001) did not significantly affect TAs subsequent muscle activity level relative to the reference contractions at similar joint torques (0-1% MVC, p[&ge;].659 and p[&ge;].626). However, the torque steadiness relative to the reference conditions was significantly reduced after the 85-15% MVC (p=.036) and 85-30% MVC (p=.036) torque drops. Consequently, the history of force production affected the control of muscle force more than TAs steady-state neuromuscular output. These findings indicate that assessing fascicle kinematics concurrently with motor unit behavior during fixed-end contractions with large torque drops might provide unique insights into the neuromechanical contributors to impaired torque control. NEW & NOTEWORTHYThe history of force production affected the subsequent control of dorsiflexion torque more than tibialis anteriors (TAs) neuromuscular output following larger amounts of fascicle stretch during fixed-end contractions. Larger fascicle stretches also reduced the median frequency of TAs EMG signal, which tentatively indicates that the voluntary control of muscle force was altered in the steady state. Together, these findings indicate that voluntary force control might be impaired by stretch-induced and/or activity-level-dependent mechanisms during fixed-end contractions.

Standing posture control critically depends on the activation of the soleus (SOL) and medial gastrocnemius (MG), which serve distinct functional roles. Identifying underlying neural mechanisms has been challenging, as conventional invasive techniques sample only limited motor units (MUs). Recent advances in high-density surface electromyography (HDsEMG) have enabled analysis of MU activity and estimation of common synaptic inputs to spinal motoneurons. Therefore, we aimed to elucidate the common synaptic inputs underlying the distinct MU behaviors of SOL and MG during standing. We recorded HDsEMG from the SOL and MG, alongside electroencephalography, from 20 male participants during standing and isometric voluntary contractions. EMG signals were decomposed into individual MU activity, with common synaptic inputs estimated through intramuscular and corticomuscular coherence analyses (IMC and CMC). Compared to SOL, the MG exhibited significantly higher delta-, alpha-, and beta-band IMC during standing. In task comparisons, alpha-band IMC increased during standing specifically in the MG. Furthermore, although beta-band CMC decreased in both muscles while standing, IMC was preserved in the MG but markedly reduced in SOL. This dissociation suggests that the common neural drive to the MG during standing is likely derived from subcortical rather than cortical pathways. These results demonstrate that the SOL and MG are governed by distinct neural control strategies, which likely underlie their functional roles. Given the low CMC, the MG relies on strong common synaptic input from subcortical pathways (e.g., vestibulospinal and reticulospinal) to produce rapid corrective torque, whereas the SOL functions with lower neural synchrony to ensure steady ankle stiffness. Key pointsO_LIThe soleus and medial gastrocnemius play distinct roles in standing control, however, due to technical limitations, it has been difficult to identify the underlying neural mechanisms responsible for these differences. C_LIO_LIUsing high-density surface electromyography, we examined motor unit activity and neural inputs to these muscles during standing. C_LIO_LIThe medial gastrocnemius shows greater common synaptic input, potentially facilitating rapid ankle plantarflexion torque generation to correct postural sway. C_LIO_LIThe soleus exhibits lower motor unit synchrony, enabling stable and continuous ankle plantarflexion torque generation for body weight support. C_LIO_LIThis study demonstrates that the soleus and medial gastrocnemius are governed by distinct neural control strategies, which likely underlie their distinct functional roles. C_LI Abstract figure legend O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=131 SRC="FIGDIR/small/698550v1_ufig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@be53aorg.highwire.dtl.DTLVardef@f6793aorg.highwire.dtl.DTLVardef@190e8f9org.highwire.dtl.DTLVardef@af5e4b_HPS_FORMAT_FIGEXP M_FIG C_FIG Motor unit spike trains were decomposed from high-density surface electromyograms recorded from the medial gastrocnemius (MG; left; red) muscle and soleus (SOL; right; blue). To compare the neural input to the spinal motor neurons between them, we quantified the intramuscular coherence (IMC) of motor unit spike trains within the delta, alpha, and beta bands. MG exhibited greater IMC than SOL during standing, indicating stronger common synaptic input, likely mediated by vestibulospinal and reticulospinal pathways. In contrast, SOL showed lower IMC, suggesting a greater contribution of independent synaptic input. As a consequence, high motor-unit synchrony in the MG supports rapid, phasic torque generation for postural sway attenuation, whereas low synchrony in the SOL enables smooth, steady torque production for weight bearing during standing.

Active control of frontal plane mechanics regulates balance in destabilizing environments, such as during asymmetric split-belt walking. Compared to sagittal plane mechanics, mediolateral (ML) kinematic and kinetic adaptations to split-belt perturbations are not as extensively reported. Moreover, the associated metabolic cost of these adaptations as well as the retention of previously learned ML adaptations upon re-exposure to the same perturbation have not been concurrently examined. We investigated adaptations in step width and peak ML ground reaction forces (GRF) during an initial and subsequent perturbation in order to characterize motor learning and motor savings, respectively. Additionally, we examined the extent to which a neuroplasticity inducing stimulus, acute intermittent hypoxia (AIH), affected the magnitude of each adaptation. Although we observed bilateral increases in step width during the initial adaptation, only the slow leg significantly reduced step width during the subsequent perturbation. Distinct interlimb differences emerged as only the slow leg modulated ML GRF during the braking phase whereas the fast leg increased ML GRF during the propulsive phase. The AIH group uniquely demonstrated greater motor savings of reduced step width and peak ML GRF strategies during the propulsive phase, suggesting greater retention of prior strategies. Furthermore, we find significant associations between ML kinetic adaptations and reductions in metabolic cost. Together, our findings suggest that unlike the sagittal plane, asymmetrical frontal plane adaptations contribute to ML stability as well as reductions in metabolic cost during split-belt walking. These insights could inform clinical training approaches to improve balance and prevent falls in clinical populations. NEW & NOTEWORTHYWe investigated adaptations in step width and mediolateral ground reaction forces during the braking and propulsive phases of split-belt walking across an initial and subsequent perturbation. We observe that the initial learning and savings of unique interlimb frontal plane coordination strategies contribute to stability and are associated with a reduction in metabolic cost.

Force production relies on the interaction between neural control of spinal motoneurons and the mechanical properties of the muscles. Changes in muscle length provide a useful model for exploring this interaction; however, studies typically assess these properties in the same muscle undergoing length change. This study investigated how altering the length of one muscle influences motor unit discharge behavior of its synergists. Eighteen healthy participants performed submaximal isometric knee extensions with the hip joint positioned at 90{degrees} (shortened rectus femoris, RF) and 180{degrees} (lengthened RF). At each hip position, participants followed trapezoidal force profiles at 10% and 30% of maximal voluntary contraction (MVC), while high-density surface electromyography (HDsEMG) was recorded from the synergistic vastus medialis and vastus lateralis (VL). Motor unit spike trains were decomposed from HDsEMG, tracked across hip positions, and analyzed for mean discharge rate and coefficient of variation of interspike interval (CoV-ISI). Lengthening the RF led to increased discharge rates of vasti motor units at 10% MVC, but not 30% MVC, with no changes in CoV-ISI. To further explore these force-dependent changes in discharge rate, two sets of experiments were conducted. The first showed that the discharge rate at recruitment during ramp-up contractions increased with RF lengthening, but only for vasti units recruited below 20% MVC. In the second, electrically evoked twitch contractions in the vasti revealed reduced twitches at 180{degrees} during low-frequency, but not high-frequency stimulation. These findings collectively suggest that the force-dependent changes in the vasti motor unit discharge rates are likely driven by RF-length dependent changes in the vasti muscles contractile properties.

In order to investigate the mechanisms governing energy and redox balance in skeletal muscle, we developed a computational model describing the coupled biochemical reaction network of glycolysis and mitochondrial oxidative phosphorylation (OxPhos) in fast-twitch oxidative glycolytic (FOG) muscle fibers. The model was identified against dynamic in vivo recordings of Phosphocreatine (PCr), inorganic Phosphate (Pi), and pH in rodent hindlimb muscle and verified against independent data from in vivo experiments and muscle biopsies. Step response testing revealed that mass action kinetics in combination with feedback control were sufficient to accomplish myoplasmic ATP homeostasis over a 100-fold range of ATP turnover rates. This vital emergent property of the metabolic model was associated with dynamic behaviour of intermediary metabolite concentrations similar to a second-order underdamped system that remains to be verified. The simulations additionally predicted that the lactate dehydrogenase (LDH) reaction makes substantial contributions to redox balance across the physiological range of ATP demands in this myofiber phenotype, while its role in slowing cellular acidification is minimal. Yet, LDH knock-out simulations revealed that oxidative recycling of myoplasmic NADH in and by itself sufficed to maintain redox balance over ATP turnover rates in the range of mitochondrial ATP synthesis. We conclude that aerobic lactate production in working muscles is a byproduct of the metabolic flexibility of FOG myofibers afforded by expression of high levels of LDH and OxPhos enzymes to support continual myoplasmic redox balance and ATP synthesis under conditions of high-intensity mechanical work. In the future, the presented simulation framework may be used to further enhance the understanding of how experimental observations in muscle emerge from the integrative behaviour of the metabolic network for carbohydrate metabolism in FOG myofibers.

Despite the critical role of persistent inward currents (PICs) in modulating motor neuron output, and thus neuromuscular performance, it remains unknown whether their contribution to motor neuron discharge behaviour varies throughout the day. This study aimed to determine whether PIC-related effects on motor neuron activity during submaximal dorsiflexion tasks differ between the early morning and late afternoon. Eighteen healthy adults (4 females; 27.4{+/-}5.6 years) performed triangular isometric contractions at two time-points: early morning (7:00-8:30 a.m.) and late afternoon (5:00-7:30 p.m.). Two conditions were tested: (1) a relative condition, where the target force corresponded to 40% of the maximal voluntary force (MVF) measured during that session, and (2) an absolute condition, where the target force was 40% of the MVF recorded during the first session. High-density surface electromyography signals were recorded from the tibialis anterior and decomposed into motor unit spike trains. The prolongation effect of PICs, estimated via {Delta}F, was significantly greater in the late afternoon in both the relative and absolute force conditions. The amplification effect of PICs, estimated by the acceleration phase of the discharge trajectory, was also higher in the late afternoon, but only in the relative force condition. No time-of-day differences were found for brace height, while attenuation was reduced in the late afternoon in the relative force condition. Collectively, these findings provide evidence for a diurnal modulation of the influence of PICs on motor neuron discharge behaviour, likely mediated by reduced inhibitory input in the late afternoon rather than by changes in neuromodulatory drive. Key pointsO_LI- Human neuromuscular performance fluctuates throughout the day, with higher force output typically observed in the late afternoon. C_LIO_LI- While circadian effects on peripheral determinants of muscle function (e.g., muscle- tendon properties, metabolic efficiency) are well documented, the neural mechanisms underlying these diurnal variations remain poorly understood. C_LIO_LI- Persistent inward currents (PICs) play a critical role in regulating motor neuron behaviour, yet their potential diurnal modulation has not been previously investigated. C_LIO_LI- Our results provide novel evidence for a diurnal modulation of the influence of PICs on motor neuron discharge behaviour, likely mediated by reduced inhibitory input in the late afternoon rather than by changes in neuromodulatory drive. C_LIO_LI- These findings suggest that intrinsic motor neuron excitability (and therefore spinal gain modulation) fluctuates across the day. C_LI

Neuromuscular reflexes elicited by sensory nerve stimulation provide valuable insights into neural motor control pathways. Analysis at the level of individual motor units (MUs) is feasible via electromyographic decomposition, but the factors shaping MU-specific reflex responses remain poorly understood. We investigated long-latency responses to cutaneous electrical stimulation in a large population of tibialis anterior MUs from nine healthy subjects during isometric ankle dorsiflexion at 10-30% of maximum voluntary contraction. Individual MU reflex responses differed markedly. Using 1000 stimulation pulses per trial, substantially more than the 150-300 typically reported in previous studies, provided more reliable estimates of cutaneous reflex characteristics. Across the motor pool, reflex magnitude increased with force level (p < 0.001) while excitation probability correlated significantly with MU recruitment threshold in 78% of subjects (p = 0.012). Furthermore, excitation probability increased systematically with contraction intensity (p < 0.001) for individually tracked MUs. Post-excitatory depression (PED) magnitude correlated significantly with excitation probability (r = 0.50, p < 0.001) of individual MUs. A targeted reflex-removal analysis, validated by MU simulations incorporating realistic excitation probabilities into ordinary firing patterns, reduced the PED by 84.2% in simulated data but only by 34.7% in recorded units. These findings suggest that the PED is a complex, hybrid phenomenon, resulting from synchronization-induced discharge resetting and additional independent inhibitory components. These findings demonstrate that MU-level reflex excitability to somatosensory input is influenced by state- and identity-dependent motor neuron characteristics, underscoring the importance of using sufficient stimulation pulses for reliable reflex measures and MU population analysis.

Brown adipose tissue (BAT) plays a central role in mammalian non-shivering thermogenesis, dissipating mitochondrial membrane potentials through the activity of uncoupling protein UCP1 to release heat. Inner membranes of mitochondria are known to be permeable to potassium ions (K+), which enter the matrix either through ATP-sensitive channels (MitoKATP) or leakage across the bilayer driven by inner membrane potentials. Mitochondrial K+ influx is associated with increased osmotic pressure, promoting water influx and increasing matrix volume. Since BAT mitochondria have lower inner membrane potentials due to uncoupling protein 1 (UCP1) activity, we hypothesized this could involve compensatory changes in MitoKATP activity, and thus tested MitoKATP involvement in brown adipocyte activities under basal and stimulated conditions. We find that cold exposure and adrenergic stimulation in mice modulate BAT MitoK levels, the channel portion of MitoKATP. Genetic ablation of the gene that codes for the pore-forming subunit of MitoKATP in human pre-adipocytes decreased cellular respiration and proliferation, compromising differentiation into mature adipocytes. In mouse cell lines, the absence of the protein limited cellular oxygen consumption in the precursor stage, but not in mature adipocytes. Interestingly, inhibition of MitoKATP in mature adipocytes increased adrenergic-stimulated oxygen consumption, indicating that shutdown of this pathway is important for full BAT thermogenesis. Similarly, MitoKATP inhibition increased oxygen consumption in BAT mitochondria isolated from mice treated with beta 3 adrenergic receptor agonist CL316,243. Overall, our results suggest that the activity of MitoKATP regulates differentiation and metabolism of brown adipocytes, impacting on thermogenesis. New and NoteworthyBrown fat cells are important to maintain healthy body weight by promoting mitochondrial uncoupling. Here, we demonstrate that mitochondrial ATP-sensitive potassium channels (MitoKATP) have important roles both in the differentiation of brown fat cells and in the activation of energy-dissipating uncoupling in this tissue.

The contributions of intrinsic muscle fiber resistance during mechanical perturbations to standing and other postural behaviors are unclear. Muscle stiffness, a traditional metric for estimating muscles intrinsic resistance to stretch, is known to vary depending on the current level and history of the muscles activation, as well as the muscles recent movement history; this property has been referred to as history dependence or muscle thixotropy. However, we currently lack sufficient data about the degree to which muscle stiffness is modulated across posturally-relevant characteristics of muscle stretch and activation. Here, we characterized the history dependence of muscles resistance to stretch in single, permeabilized, activated, muscle fibers in posturally-relevant stretch conditions and activation levels. We used a classic paired muscle stretch paradigm, varying the amplitude of a "conditioning" triangular stretch-shorten cycle followed by a "test" ramp-and-hold imposed after a variable inter-stretch interval. We tested low (<15%), intermediate (15-50%) and high (>50%) muscle fiber activation levels, evaluating short-range stiffness and total impulse in the test stretch. Muscle fiber resistance to stretch remained high at conditioning amplitudes of <1% L0 and inter-stretch intervals of >1 s, characteristic of healthy standing postural sway. A ~70% attenuation of muscle resistance to stretch was reached at conditioning amplitudes of >3% L0 and inter-stretch intervals of <0.1s, characteristic of larger, faster postural sway in balance-impaired individuals. Overall, amplitude and inter-stretch interval interact to disrupt myofilaments such that intrinsic resistance to stretch is attenuated if the stretch is large enough and/or frequent enough. Summary StatementIntrinsic muscle fiber resistance to stretch is preserved after small, slow pre-movements based on healthy postural sway, but markedly reduced as pre-movements increase to emulate abnormal postural sway.

While maximal force increases following short-term isometric strength training, the rate of force development (RFD) may remain relatively unaffected. The underlying neural and muscular mechanisms during rapid contractions after strength training are largely unknown. Since strength training increases the neural drive to muscles, it may be hypothesized that there are distinct neural or muscular adaptations determining the change in RFD independently of an increase in maximal force. Therefore, we examined motor unit population data during the rapid generation of force before and after four weeks of strength training. We observed that strength training did not change the RFD because it did not influence the number of motor units recruited per second or their initial discharge rate during rapid contractions. While strength training did not change motoneuron behaviour in the force increase phase of rapid contractions, it increased the discharge rate of motoneurons (by [~]4 spikes/s) when reaching the plateau phase ([~]150 ms) of the rapid contractions, determining an increase in maximal force production. Computer simulations with a motor unit model that included neural and muscular properties, closely matched the experimental observations and demonstrated that the lack of change in RFD following training is primarily mediated by an unchanged maximal recruitment speed of motoneurons. These results demonstrate that maximal force and contraction speed are determined by different adaptations in motoneuron behaviour following strength training and indicate that increases in the recruitment speed of motoneurons are required to evoke training-induced increases in RFD.

Resistance training (RT) promotes muscle protein accretion and myofiber hypertrophy, driven by dynamic processes of protein synthesis and degradation. While molecular studies have focused on acute signalling or long-term hypertrophy and strength gains, a critical gap remains in understanding the intermediate processes of muscle adaptation. Acute signalling does not always correlate directly with long-term outcomes, highlighting the need for a time-course analysis of protein abundance and turnover rates. To address this, we utilised deuterium oxide labelling and peptide mass spectrometry to quantify absolute protein content and synthesis rates in skeletal muscle. A daily programmed resistance training regimen was applied to the rat tibialis anterior (TA) via electrical stimulation of the left hind limb for 10, 20, and 30 days (5 sets of 10 repetitions daily). Muscle samples from stimulated (Stim) and contralateral control (Ctrl) limbs were analysed, quantifying 658 protein abundances and 215 protein synthesis rates. Unsupervised temporal clustering of protein responses revealed distinct phases of muscle adaptation, with early (0-10 days) and mid (10-20 days) responses driven by differential protein accretion rates in ribosomal and mitochondrial networks, respectively. These findings suggest that subsets of proteins exhibit distinct adaptation timelines due to variations in translation and/or degradation rates. A deeper understanding of these temporal shifts could improve strategies for optimising muscle growth and functional adaptation to resistance training.

Bistability in spinal motoneurons supports tonic spike activity in the absence of excitatory drive. Earlier work in adult preparations suggested that smaller motoneurons innervating slow antigravity muscle fibers are more likely to generate bistability for postural maintenance. However, whether large motoneurons innervating fast-fatigable muscle fibers display bistability related to postural tone is still controversial. To address this, we examined the relationship between soma size and bistability in lumbar ventrolateral -motoneurons of ChAT-GFP and Hb9-GFP mice across different developmental stages: neonatal (P2-P7), young (P7-P14) and mature (P21-P25). We found that as neuron size increases, the prevalence of bistability rises. Smaller -motoneurons lack bistability, while larger fast -motoneurons (MMP-9+/Hb9+) with a soma area [&ge;] 400{micro}m2 exhibit significantly higher bistability. Ionic currents associated with bistability, including the persistent Nav1.6 current, thermosensitive Trpm5 Ca2+-activated Na+ current and the slowly inactivating Kv1.2 current, also scale with cell size. Serotonin evokes full bistability in large motoneurons with partial bistable properties, but not in small motoneurons. Our study provides important insights into the neural mechanisms underlying bistability and how motoneuron size dictates this process. New and NoteworthyBistability is not a common feature of all mouse spinal motoneurons. It is absent in small, slow motoneurons but present in most large, fast motoneurons. This difference results from differential expression of ionic currents that enable bistability, which are highly expressed in large motoneurons but small or absent in small motoneurons. These results support a possible role for fast motoneurons in maintenance of tonic posture in addition to their known roles in fast movements.

EMG-based muscle force predictions are often inaccurate following active muscle stretch or shortening because of residual force enhancement (rFE) or depression (rFD), respectively, which can alter the neural drive to a muscle. However, the extent of neural drive modulation due to rFE or rFD remains unknown, making it difficult to correct EMG-based force predictions. Therefore, seventeen participants performed dorsiflexion contractions at 20 and 40% of maximum voluntary torque (MVT) in three conditions: stretch-hold, shortening-hold, and fixed-end reference (REF) conditions. The ankle dorsiflexion torques and angles were matched using dynamometry to the REF condition over a 10-s steady state following a 1-s 25{degrees} stretch or shortening, during which we recorded and decomposed tibialis anterior individual motor unit action potentials from high-density surface EMG recordings to gain insights into neural drive. Normalized EMG amplitudes were 2% lower following stretch and 1 or 3% higher following shortening relative to REF at 20 versus 40% MVT (p[&le;].008), respectively. Discharge rates (DRs) from 19 matched motor units per person on average obtained via DEMUSE and MUedit were similar (p=.871). Following stretch and shortening, DRs were [~]1 Hz lower (p[&le;].004) and 0 (p=.966) to 1 Hz higher relative to REF (p=.003), respectively. More unique motor units were also detected following shortening versus REF and in REF versus following stretch. These findings indicate that to account for rFE or rFD, neural drive is respectively decreased or increased via reduced or additional motor unit recruitment and DR modulation, with a contraction-intensity specific discharge rate modulation following active shortening.

Pain significantly influences movement, yet the neural mechanisms underlying the range of observed motor adaptations remain unclear. This study combined experimental data and in silico models to investigate the contribution of inhibitory and neuromodulatory inputs to motor unit behaviour in response to nociceptive stimulation during contractions at 30% of maximal torque. Specifically, we aimed to unravel the distribution pattern of inhibitory inputs to the motor unit pool. Seventeen participants performed isometric knee extension tasks under three conditions: Control, Pain (induced by injecting hypertonic saline into the infra-patellar fat pad), and Washout. We identified large samples of motor units in the vastus lateralis (up to 53/participant) from high-density electromyographic signals, leading to three key observations. First, while motor unit discharge rates significantly decreased during Pain, a substantial proportion of motor units (14.8-24.8%) did not show this decrease and, in some cases, even exhibited an increase. Second, using complementary approaches, we found that pain did not significantly affect neuromodulation, making it unlikely to be a major contributor to the observed changes in motor unit behaviour. Third, we observed a significant reduction in the proportion of common inputs to motor units during Pain. To explore potential neurophysiological mechanisms underlying these results, we simulated the behaviour of motor unit pools with varying distribution patterns of inhibitory inputs. Our simulations support the hypothesis that a non-homogeneous distribution of inhibitory inputs, not strictly organised according to motor unit size, is a key mechanism underlying the motor response to nociceptive stimulation during moderate contraction intensity. Key pointsO_LIPain affects movement, but the neural mechanisms underlying these motor adaptations are not well defined. C_LIO_LIThe traditional view is that pain causes uniform (homogeneous) inhibition among motor units. C_LIO_LIRecent research has observed differential motor unit responses to experimental pain - some with decreased discharge rates and others with increased discharge rates. C_LIO_LICombining experimental data with modelling, we provide compelling evidence of increased inhibition that is non-uniformly distributed across motor units, regardless of their size. C_LI Legend of the abstract figureWe combined experimental data and in silico models to investigate the contribution of inhibitory and neuromodulatory inputs to motor unit behaviour in response to nociceptive stimulation during submaximal isometric contractions at 30% of maximal voluntary contraction. We identified large samples of motor units in the vastus lateralis, leading to three key observations. First, while motor unit discharge rates significantly decreased during Pain, a substantial proportion of motor units did not show this decrease and, in some cases, even exhibited an increase. Second, using complementary approaches, we found that pain did not significantly affect neuromodulation, making it unlikely to be a major contributor to the observed changes in motor unit behaviour. Third, we observed a significant reduction in the proportion of common inputs to motor units during Pain. Together with our simulations, these results provide evidence of increased inhibition that is non-uniformly distributed across motor units, regardless of their size. Ppp, pulses per second; MVC, maximal voluntary contraction.

The diversity of motor units arises from differences in the contractile properties of muscle fibers and the intrinsic electrical properties of their motoneurons. In mice, however, this relationship has not been quantitatively defined, and conventional classification often relies on subjective thresholds. Here, we combined in vivo intracellular recordings with supervised and unsupervised machine-learning methods to test whether motoneuron electrophysiology can predict the physiological identity of mouse motor units. Unbiased clustering identified four groups corresponding to slow (S), fast fatigue-resistant (FR), intermediate (FI), and fast fatigable (FF) types. A multinomial logistic regression model performed well, with most errors occurring between FI and FF types, which showed substantial overlap. Reducing the task to three classes improved accuracy. Feature selection revealed that four electrophysiological properties (input conductance, rheobase, AHP duration, maximal frequency) were sufficient for high predictive performance. Overall, this study provides a quantitative description of mouse motor-unit properties and a framework for incorporating motor-unit diversity into future investigations of neuromuscular physiology and disease.