Senescence atlas reveals an aged-like inflamed niche that blunts muscle regeneration.

Tissue regeneration requires coordination between resident stem cells and local niche cells1,2. Here we identify that senescent cells are integral components of the skeletal muscle regenerative niche that repress regeneration at all stages of life. The technical limitation of senescent-cell scarcity3 was overcome by combining single-cell transcriptomics and a senescent-cell enrichment sorting protocol. We identified and isolated different senescent cell types from damaged muscles of young and old mice. Deeper transcriptome, chromatin and pathway analyses revealed conservation of cell identity traits as well as two universal senescence hallmarks (inflammation and fibrosis) across cell type, regeneration time and ageing. Senescent cells create an aged-like inflamed niche that mirrors inflammation associated with ageing (inflammageing4) and arrests stem cell proliferation and regeneration. Reducing the burden of senescent cells, or reducing their inflammatory secretome through CD36 neutralization, accelerates regeneration in young and old mice. By contrast, transplantation of senescent cells delays regeneration. Our results provide a technique for isolating in vivo senescent cells, define a senescence blueprint for muscle, and uncover unproductive functional interactions between senescent cells and stem cells in regenerative niches that can be overcome. As senescent cells also accumulate in human muscles, our findings open potential paths for improving muscle repair throughout life.

Temporal and Spatial Information Elicit Different Power and Connectivity Profiles during Working Memory Maintenance.

Working memory (WM) is the cognitive ability to store and manipulate information necessary for ongoing tasks. Although frontoparietal areas are involved in the retention of visually presented information, oscillatory neural activity differs for temporal and spatial WM processing. In this study, we corroborated previous findings describing the modulation of neural oscillations and expanded our investigation to the network organization underlying the cognitive processing of temporal and spatial information. We utilized MEG recordings during a Sternberg visual WM task. The spectral oscillatory activity in the maintenance phase revealed increased frontal theta (4-8 Hz) and parietal beta (13-30 Hz) in the temporal condition. Source level coherence analysis delineated the prominent role of parietal areas in all frequency bands during the maintenance of temporal information, whereas frontal and central areas showed major contributions in theta and beta ranges during the maintenance of spatial information. Our study revealed distinct spectral profiles of neural oscillations for separate cognitive subdomains of WM processing. The delineation of specific functional networks might have important implications for clinical applications, enabling the development of stimulation protocols targeting cognitive disabilities associated with WM impairments.

Regionally specific cortical lateralization of abstract and concrete verb processing: Magnetic mismatch negativity study.

The neural underpinnings of processing concrete and abstract semantics remain poorly understood. Previous fMRI studies have shown that multimodal and amodal neural networks respond differentially to different semantic types; importantly, abstract semantics activates more left-lateralized networks, as opposed to more bilateral activity for concrete words. Due to the lack of temporal resolution, these fMRI results do not allow to easily separate language- and task-specific brain responses and to disentangle early processing stages from later post-comprehension phenomena. To tackle this, we used magnetoencephalography (MEG), a time-resolved neuroimaging technique, in combination with a task-free oddball mismatch negativity (MMN) paradigm, an established approach to tracking early automatic activation of word-specific memory traces in the brain. We recorded the magnetic MMN responses in 30 healthy adults to auditorily presented abstract and concrete action verbs to assess lateralization of word-specific lexico-semantic processing in a set of neocortical areas. We found that MMN responses to these stimuli showed different lateralization patterns of activity in the upper limb motor area (BA4) and parts of Broca's area (BA45/BA47) within ∼100-350 ms after the word disambiguation point. Importantly, the greater leftward response lateralization for abstract semantics was due to the lesser involvement of the right-hemispheric homologues, not increased left-hemispheric activity. These findings suggest differential region-specific involvement of bilateral sensorimotor systems already in the early automatic stages of processing abstract and concrete action semantics.

Context-dependent roles of cellular senescence in normal, aged, and disease states.

Cellular senescence is a state of irreversible cell cycle arrest that often emerges after tissue damage and in age-related diseases. Through the production of a multicomponent secretory phenotype (SASP), senescent cells can impact the regeneration and function of tissues. However, the effects of senescent cells and their SASP are very heterogeneous and depend on the tissue environment and type as well as the duration of injury, the degree of persistence of senescent cells and the organism's age. While the transient presence of senescent cells is widely believed to be beneficial, recent data suggest that it is detrimental for tissue regeneration after acute damage. Furthermore, although senescent cell persistence is typically associated with the progression of age-related chronic degenerative diseases, it now appears to be also necessary for correct tissue function in the elderly. Here, we discuss what is currently known about the roles of senescent cells and their SASP in tissue regeneration in ageing and age-related diseases, highlighting their (negative and/or positive) contributions. We provide insight for future research, including the possibility of senolytic-based therapies and cellular reprogramming, with aims ranging from enhancing tissue repair to extending a healthy lifespan.

3-deazaadenosine (3DA) alleviates senescence to promote cellular fitness and cell therapy efficiency in mice.

Cellular senescence is a stable type of cell cycle arrest triggered by different stresses. As such, senescence drives age-related diseases and curbs cellular replicative potential. Here, we show that 3-deazaadenosine (3DA), an S-adenosyl homocysteinase (AHCY) inhibitor, alleviates replicative and oncogene-induced senescence. 3DA-treated senescent cells showed reduced global Histone H3 Lysine 36 trimethylation (H3K36me3), an epigenetic modification that marks the bodies of actively transcribed genes. By integrating transcriptome and epigenome data, we demonstrate that 3DA treatment affects key factors of the senescence transcriptional program. Remarkably, 3DA treatment alleviated senescence and increased the proliferative and regenerative potential of muscle stem cells from very old mice in vitro and in vivo. Moreover, ex vivo 3DA treatment was sufficient to enhance the engraftment of human umbilical cord blood (UCB) cells in immunocompromised mice. Together, our results identify 3DA as a promising drug enhancing the efficiency of cellular therapies by restraining senescence.

Activity Stabilization in a Population Model of Working Memory by Sinusoidal and Noisy Inputs.

According to mechanistic theories of working memory (WM), information is retained as stimulus-dependent persistent spiking activity of cortical neural networks. Yet, how this activity is related to changes in the oscillatory profile observed during WM tasks remains a largely open issue. We explore joint effects of input gamma-band oscillations and noise on the dynamics of several firing rate models of WM. The considered models have a metastable active regime, i.e., they demonstrate long-lasting transient post-stimulus firing rate elevation. We start from a single excitatory-inhibitory circuit and demonstrate that either gamma-band or noise input could stabilize the active regime, thus supporting WM retention. We then consider a system of two circuits with excitatory intercoupling. We find that fast coupling allows for better stabilization by common noise compared to independent noise and stronger amplification of this effect by in-phase gamma inputs compared to anti-phase inputs. Finally, we consider a multi-circuit system comprised of two clusters, each containing a group of circuits receiving a common noise input and a group of circuits receiving independent noise. Each cluster is associated with its own local gamma generator, so all its circuits receive gamma-band input in the same phase. We find that gamma-band input differentially stabilizes the activity of the "common-noise" groups compared to the "independent-noise" groups. If the inter-cluster connections are fast, this effect is more pronounced when the gamma-band input is delivered to the clusters in the same phase rather than in the anti-phase. Assuming that the common noise comes from a large-scale distributed WM representation, our results demonstrate that local gamma oscillations can stabilize the activity of the corresponding parts of this representation, with stronger effect for fast long-range connections and synchronized gamma oscillations.

Protocol for RNA-seq library preparation starting from a rare muscle stem cell population or a limited number of mouse embryonic stem cells.

It remains challenging to generate reproducible, high-quality cDNA libraries from RNA derived from rare cell populations. Here, we describe a protocol for high-throughput RNA-seq library preparation, including isolation of 200 skeletal muscle stem cells from mouse tibialis anterior muscle by fluorescence-activated cell sorting and cDNA preparation. We also describe RNA extraction and cDNA preparation from differentiating mouse embryonic stem cells. For complete details on the use and execution of this protocol, please refer to Juan et al. (2016) and Garcia-Prat et al. (2016).

Cortical plasticity elicited by acoustically cued monetary losses: an ERP study.

Both human and animal studies have demonstrated remarkable findings of experience-induced plasticity in the cortex. Here, we investigated whether the widely used monetary incentive delay (MID) task changes the neural processing of incentive cues that code expected monetary outcomes. We used a novel auditory version of the MID task, where participants responded to acoustic cues that coded expected monetary losses. To investigate task-induced brain plasticity, we presented incentive cues as deviants during passive oddball tasks before and after two sessions of the MID task. During the oddball task, we recorded the mismatch-related negativity (MMN) as an index of cortical plasticity. We found that two sessions of the MID task evoked a significant enhancement of MMN for incentive cues that predicted large monetary losses, specifically when monetary cue discrimination was essential for maximising monetary outcomes. The task-induced plasticity correlated with the learning-related neural activity recorded during the MID task. Thus, our results confirm that the processing of (loss)incentive auditory cues is dynamically modulated by previously learned monetary outcomes.

FoxO maintains a genuine muscle stem-cell quiescent state until geriatric age.

Tissue regeneration declines with ageing but little is known about whether this arises from changes in stem-cell heterogeneity. Here, in homeostatic skeletal muscle, we identify two quiescent stem-cell states distinguished by relative CD34 expression: CD34High, with stemness properties (genuine state), and CD34Low, committed to myogenic differentiation (primed state). The genuine-quiescent state is unexpectedly preserved into later life, succumbing only in extreme old age due to the acquisition of primed-state traits. Niche-derived IGF1-dependent Akt activation debilitates the genuine stem-cell state by imposing primed-state features via FoxO inhibition. Interventions to neutralize Akt and promote FoxO activity drive a primed-to-genuine state conversion, whereas FoxO inactivation deteriorates the genuine state at a young age, causing regenerative failure of muscle, as occurs in geriatric mice. These findings reveal transcriptional determinants of stem-cell heterogeneity that resist ageing more than previously anticipated and are only lost in extreme old age, with implications for the repair of geriatric muscle.

Simultaneous Isolation of Stem and Niche Cells of Skeletal Muscle: Applicability for Aging Studies.

The maintenance of adult stem cells in their normal quiescent state depends on intrinsic factors and extrinsic signals originating from their microenvironment (also known as the stem cell niche). In skeletal muscle, its stem cells (satellite cells) lose their regenerative potential with aging, and this has been attributed, at least in part, to both age-associated changes in the satellite cells as in the niche cells, which include resident fibro-adipogenic progenitors (FAPs), macrophages, and endothelial cells, among others. To understand the regenerative decline of skeletal muscle with aging, there is a need for methods to specifically isolate stem and niche cells from resting muscle. Here we describe a fluorescence-activated cell sorting (FACS) protocol to simultaneously isolate discrete populations of satellite cells and niche cells from skeletal muscle of aging mice.

The Monetary Incentive Delay (MID) Task Induces Changes in Sensory Processing: ERP Evidence.

Numerous cognitive studies have demonstrated experience-induced plasticity in the primary sensory cortex, indicating that repeated decisions could modulate sensory processing. In this context, we investigated whether an auditory version of the monetary incentive delay (MID) task could change the neural processing of the incentive cues that code expected monetary outcomes. To study sensory plasticity, we presented the incentive cues as deviants during oddball sessions recorded before and after training in the two MID task sessions. We found that after 2 days of training in the MID task, incentive cues evoked a larger P3a (compared with the baseline condition), indicating there was an enhancement of the involuntary attention to the stimuli that predict rewards. At the individual level, the training-induced change of mismatch-related negativity was correlated with the amplitude of the feedback-related negativity (FRN) recorded during the first MID task session. Our results show that the MID task evokes plasticity changes in the auditory system associated with better passive discrimination of incentive cues and with enhanced involuntary attention switching towards these cues. Thus, the sensory processing of incentive cues is dynamically modulated by previous outcomes.

Minimal Circuit Model of Reward Prediction Error Computations and Effects of Nicotinic Modulations.

Dopamine (DA) neurons in the ventral tegmental area (VTA) are thought to encode reward prediction errors (RPE) by comparing actual and expected rewards. In recent years, much work has been done to identify how the brain uses and computes this signal. While several lines of evidence suggest the interplay of the DA and the inhibitory interneurons in the VTA implements the RPE computation, it still remains unclear how the DA neurons learn key quantities, for example the amplitude and the timing of primary rewards during conditioning tasks. Furthermore, endogenous acetylcholine and exogenous nicotine, also likely affect these computations by acting on both VTA DA and GABA (γ -aminobutyric acid) neurons via nicotinic-acetylcholine receptors (nAChRs). To explore the potential circuit-level mechanisms for RPE computations during classical-conditioning tasks, we developed a minimal computational model of the VTA circuitry. The model was designed to account for several reward-related properties of VTA afferents and recent findings on VTA GABA neuron dynamics during conditioning. With our minimal model, we showed that the RPE can be learned by a two-speed process computing reward timing and magnitude. By including models of nAChR-mediated currents in the VTA DA-GABA circuit, we showed that nicotine should reduce the acetylcholine action on the VTA GABA neurons by receptor desensitization and potentially boost DA responses to reward-related signals in a non-trivial manner. Together, our results delineate the mechanisms by which RPE are computed in the brain, and suggest a hypothesis on nicotine-mediated effects on reward-related perception and decision-making.

MEG Signatures of a Perceived Match or Mismatch between Individual and Group Opinions.

Humans often adjust their opinions to the perceived opinions of others. Neural responses to a perceived match or mismatch between individual and group opinions have been investigated previously, but some findings are inconsistent. In this study, we used magnetoencephalographic source imaging to investigate further neural responses to the perceived opinions of others. We found that group opinions mismatching with individual opinions evoked responses in the anterior and posterior medial prefrontal cortices, as well as in the temporoparietal junction and ventromedial prefrontal cortex in the 220-320 and 380-530 ms time windows. Evoked responses were accompanied by an increase in the power of theta oscillations (4-8 Hz) over a number of frontal cortical sites. Group opinions matching with individual opinions evoked an increase in amplitude of beta oscillations (13-30 Hz) in the anterior cingulate and ventral medial prefrontal cortices. Based on these results, we argue that distinct valuation and performance-monitoring neural circuits in the medial cortices of the brain may monitor compliance of individual behavior to the perceived group norms.

Neurofeedback learning modifies the incidence rate of alpha spindles, but not their duration and amplitude.

Although the first experiments on alpha-neurofeedback date back nearly six decades ago, when Joseph Kamiya reported successful operant conditioning of alpha-rhythm in humans, the effectiveness of this paradigm in various experimental and clinical settings is still a matter of debate. Here, we investigated the changes in EEG patterns during a continuously administered neurofeedback of P4 alpha activity. Two days of neurofeedback training were sufficient for a significant increase in the alpha power to occur. A detailed analysis of these EEG changes showed that the alpha power rose because of an increase in the incidence rate of alpha episodes, whereas the amplitude and the duration of alpha oscillations remained unchanged. These findings suggest that neurofeedback facilitates volitional control of alpha activity onset, but alpha episodes themselves appear to be maintained automatically with no volitional control - a property overlooked by previous studies that employed continuous alpha-power neurofeedback. We propose that future research on alpha neurofeedback should explore reinforcement schedules based on detection of onsets and offsets of alpha waves, and employ these statistics for exploration and quantification of neurofeedback induced effects.

Long-Range Temporal Correlations in the amplitude of alpha oscillations predict and reflect strength of intracortical facilitation: Combined TMS and EEG study.

While variability of the motor responses to transcranial magnetic stimulation (TMS) is widely acknowledged, little is known about its central origin. One plausible explanation for such variability may relate to different neuronal states defining the reactivity of the cortex to TMS. In this study intrinsic spatio-temporal neuronal dynamics were estimated with Long-Range Temporal Correlations (LRTC) in order to predict the inter-individual differences in the strength of intra-cortical facilitation (ICF) and short-interval intracortical inhibition (SICI) produced by paired-pulse TMS (ppTMS) of the left primary motor cortex. LRTC in the alpha frequency range were assessed from multichannel electroencephalography (EEG) obtained at rest before and after the application of and single-pulse TMS (spTMS) and ppTMS protocols. For the EEG session, preceding TMS application, we showed a positive correlation across subjects between the strength of ICF and LRTC in the fronto-central and parietal areas. This in turn attests to the existence of subject-specific neuronal phenotypes defining the reactivity of the brain to ppTMS. In addition, we also showed that ICF was associated with the changes in neuronal dynamics in the EEG session after the application of the stimulation. This result provides a complementary evidence for the recent findings demonstrating that the cortical stimulation with sparse non-regular stimuli might have considerable long-lasting effects on the cortical activity.

Early and gender-specific differences in spinal cord mitochondrial function and oxidative stress markers in a mouse model of ALS.

INTRODUCTION: Amyotrophic lateral sclerosis (ALS) is a motor neuron disease with a gender bias towards major prevalence in male individuals. Several data suggest the involvement of oxidative stress and mitochondrial dysfunction in its pathogenesis, though differences between genders have not been evaluated. For this reason, we analysed features of mitochondrial oxidative metabolism, as well as mitochondrial chain complex enzyme activities and protein expression, lipid profile, and protein oxidative stress markers, in the Cu,Zn superoxide dismutase with the G93A mutation (hSOD1-G93A)- transgenic mice and Neuro2A(N2A) cells overexpressing hSOD1-G93A. RESULTS AND CONCLUSIONS: Our results show that overexpression of hSOD1-G93A in transgenic mice decreased efficiency of mitochondrial oxidative phosphorylation, located at complex I, revealing a temporal delay in females with respect to males associated with a parallel increase in selected markers of protein oxidative damage. Further, females exhibit a fatty acid profile with higher levels of docosahexaenoic acid at 30 days. Mechanistic studies showed that hSOD1-G93A overexpression in N2A cells reduced complex I function, a defect prevented by 17ß-estradiol pretreatment. In conclusion, ALS-associated SOD1 mutation leads to delayed mitochondrial dysfunction in female mice in comparison with males, in part attributable to the higher oestrogen levels of the former. This study is important in the effort to further understanding of whether different degrees of spinal cord mitochondrial dysfunction could be disease modifiers in ALS.