Single nuclei profiling identifies cell specific markers of skeletal muscle aging, frailty, and senescence.

Aging is accompanied by a loss of muscle mass and function, termed sarcopenia, which causes numerous morbidities and economic burdens in human populations. Mechanisms implicated in age-related sarcopenia or frailty include inflammation, muscle stem cell depletion, mitochondrial dysfunction, and loss of motor neurons, but whether there are key drivers of sarcopenia are not yet known. To gain deeper insights into age-related muscle loss, we performed transcriptome profiling on lower limb muscle biopsies from 72 young, elderly, and frail human subjects using bulk RNA-seq (N = 72) and single-nuclei RNA-seq (N = 17). This combined approach revealed changes in gene expression that occur with age and frailty in multiple cell types comprising mature skeletal muscle. Notably, we found increased expression of the genes MYH8 and PDK4, and decreased expression of the gene IGFN1, in aged muscle. We validated several key genes changes in fixed human muscle tissue using digital spatial profiling. We also identified a small population of nuclei that express CDKN1A, present only in aged samples, consistent with p21cip1-driven senescence in this subpopulation. Overall, our findings identify unique cellular subpopulations in aged and sarcopenic skeletal muscle, which will facilitate the development of new therapeutic strategies to combat age-related frailty.

Senolysis induced by 25-hydroxycholesterol targets CRYAB in multiple cell types.

Cellular senescence is a driver of many age-related pathologies. There is an active search for pharmaceuticals termed senolytics that can mitigate or remove senescent cells in vivo by targeting genes that promote the survival of senescent cells. We utilized single-cell RNA sequencing to identify CRYAB as a robust senescence-induced gene and potential target for senolysis. Using chemical inhibitor screening for CRYAB disruption, we identified 25-hydroxycholesterol (25HC), an endogenous metabolite of cholesterol biosynthesis, as a potent senolytic. We then validated 25HC as a senolytic in mouse and human cells in culture and in vivo in mouse skeletal muscle. Thus, 25HC represents a potential class of senolytics, which may be useful in combating diseases or physiologies in which cellular senescence is a key driver.

Differential effects of chemogenetic inhibition of dopamine and norepinephrine neurons in the mouse 5-choice serial reaction time task.

Attention-deficit/hyperactivity disorder (ADHD) is a psychiatric disorder characterized by inattention, aberrant impulsivity, and hyperactivity. Although the underlying pathophysiology of ADHD remains unclear, dopamine and norepinephrine signaling originating from the ventral tegmental area (VTA) and locus coeruleus (LC) is thought to be critically involved. In this study, we employ Designer Receptor Exclusively Activated by Designer Drugs (DREADDs) together with the mouse 5-Choice Serial Reaction Time Task (5-CSRTT) to investigate the necessary roles of these catecholamines in ADHD-related behaviors, including attention, impulsivity, and motivation. By selective inhibition of tyrosine hydroxylase (TH)-positive VTA dopamine neurons expressing the Gi-coupled DREADD (hM4Di), we observed a marked impairment of effort-based motivation and subsequently speed and overall vigor of responding. At the highest clozapine N-oxide (CNO) dose tested (i.e. 2 mg/kg) to activate hM4Di, we detected a reduction in locomotor activity. DREADD-mediated inhibition of LC norepinephrine neurons reduced attentional performance in a variable stimulus duration test designed to increase task difficulty, specifically by increasing trials omissions, reducing mean score, and visual processing speed. These findings show that VTA dopamine and LC norepinephrine neurons differentially affect attention, impulsive and motivational control. In addition, this study highlights how molecular genetic probing of selective catecholamine circuits can provide valuable insights into the mechanisms underlying ADHD-relevant behaviors.