Journal of Physiology-London最新文献

Intrinsic biological rhythms regulate key physiological and behavioural processes, yet the influence of sex and age on these rhythms is not fully understood. We comprehensively examined 24 h (circadian) and >24 h (infradian; 5 and 10 day) rhythms in wheel-running and ingestive behaviours in single-housed young and middle-aged male and female mice. Circadian analysis revealed that middle-aged mice, particularly females, exhibited more precise daily rhythms and shifted a greater proportion of activity and feeding to the lights-on phase than young female mice. Middle-aged animals also ran for longer durations per day, suggesting age-related changes in activity regulation. Analysis of infradian rhythms further highlighted sex- and age-specific differences. Young female mice displayed robust 5 day rhythms in wheel-running activity, which were absent in middle-aged females. In contrast, few males (young or middle-aged) showed significant 5 day rhythms. Ten-day rhythms were most prominent in male mice, while females rarely expressed this periodicity. Physiologically, middle-aged mice lost more body weight in response to single housing, with middle-aged females being most affected. Interactions among behavioural rhythms in females also showed greater complexity, which increased with age. These findings reveal distinct sex- and age-dependent patterns in circadian and infradian rhythms as well as in physiological responses to isolation. Our work highlights the need to account for sex and age in chronobiological research, with broader implications for understanding vulnerability to age-related metabolic and behavioural disorders. KEY POINTS: Physiological findings: -Middle-aged mice lost more body weight after single housing, with females most affected. Circadian findings: -Older mice show more daytime activity. -Precision in daily rhythm differs by sex and age. -Middle-aged females showed prolonged daily wheel running. Infradian findings: -Young females had robust 5 day rhythms, absent in middle-aged females. -Some males showed 5 day rhythms, but 10 day rhythms were most prominent in males. Complexity of rhythms: -Complexity of interactions among behavioural rhythms increases with age, particularly in females.

Leucine-rich repeat-containing protein 8A (LRRC8A) is an essential subunit of the ubiquitously expressed volume-regulated anion channels (VRACs). Previous work has shown that LRRC8A is overexpressed in several cancers and is associated with poor survival outcomes. However, the underlying mechanisms remain obscure. In the present study, we investigated the role of LRRC8A and VRACs in the progression of glioblastoma (GBM), the most common and aggressive primary brain tumour. We found that, compared with healthy brain tissue, LRRC8A mRNA is significantly upregulated in surgical GBM specimens, patient-derived GBM cell lines and GBM datasets from The Cancer Genome Atlas. GBM patients in the lowest quartile of LRRC8A expression exhibited a trend toward longer survival. In patient-derived GBM cultures, RNA interference-mediated knockdown of LRRC8A or pharmacological blockade of VRAC with 4,4'-diisothiocyano-2,2'-stilbenedisulfonic acid (DIDS) reduced cell proliferation, lowered intracellular chloride levels and inhibited activity of mammalian target of rapamycin (mTOR) complex 2 (mTORC2). The anti-proliferative effects of LRRC8A knockdown and DIDS were non-additive, suggesting a shared mechanism. Biochemical and molecular analyses revealed that LRRC8A-containing VRACs promote GBM cell proliferation through a new non-enzymatic function of the chloride-sensitive protein kinase WNK1. Specifically, VRAC activity facilitates WNK1-dependent activation of mTORC2 and its downstream kinases AKT and SGK. In support of this model, either downregulation of WNK1 or pharmacological inhibition of mTOR or SGK/AKT suppressed GBM cell proliferation and mimicked the effect of LRRC8A knockdown. Together, these findings establish a new mTORC2-centric signalling axis for VRAC-dependent control of cellular functions and highlight several potential molecular targets for limiting GBM proliferation. KEY POINTS: Volume-regulated anion channels (VRACs) are considered to contribute to the progression of several human cancers. The essential VRAC subunit LRRC8A is significantly overexpressed in clinical specimens of glioblastoma, the most common and aggressive primary brain malignancy. RNA interference-mediated downregulation of LRRC8A reduces proliferation in patient-derived GBM cell cultures, suggesting that VRACs promote cancer cell growth. LRRC8A/VRAC-mediated effects on cell proliferation are driven by a mechanism involving the chloride-sensitive protein kinase WNK1, mTOR complex 2 and activation of downstream kinases AKT and SGK.

Mitochondria are dynamic organelles that undergo fusion and fission. Key proteins are needed to create mitochondrial networks, as well as facilitate biogenesis, fragmentation or movement within the cell. Septins are considered as the fourth component of the cytoskeleton, providing attachment sites for proteins. Besides that, they have important roles in different cellular processes, including mitochondrial fission and fusion (remodelling). Septins form oligomeric complexes comprising various septin subgroups, which can create higher-order structures. Septin7 is the sole member of its subgroup. We aimed to examine how mitochondrial dynamics and oxidative phosphorylation (OXPHOS) are affected in Septin7 downregulated C2C12 (S7-KD) myoblasts and terminally differentiated myotubes compared to scrambled short hairpin RNA-transfected control cells. We detected altered expression of genes related to mitochondrial biogenesis (PGC1α), dynamics (DRP1, OPA1 and MFN2) and autophagy (PINK1 and BNIP3); furthermore, a significant decrease in differentiation-dependent mRNA expression of OXPHOS markers (ATP synthase, COX1 and SDH). Septin7 downregulation also affected the expression of post-translational modifications of MFN2 and DRP1. Functional measurements of OXPHOS revealed decreased O₂ consumption (flux) and higher O₂ concentration in Septin7 KD cultures following selective inhibition of electron transport complexes. We observed significant alterations in basal respiration and OXPHOS pathways in Septin7 KD cultures. Our results suggest that Septin7, as a cytoskeletal protein, could be a significant regulator of mitochondrial dynamics and oxidative metabolism. Therefore, these molecules, as mitochondrial dynamics modulators, can serve as potential therapeutic targets in diseases related to changes in mitochondrial function. KEY POINTS: Knockdown of Septin7 results in altered gene and protein expression of markers controlling mitochondrial dynamics. Diminished level of Septin7 causes decreased gene expression of members of oxidative phosphorylation. Knockdown of Septin7 has an impact on microRNAs involved in the regulation of mitochondrial markers. Septin7 has an impact on mitochondrial respiration.

PIEZO2 is a principal mechanotransduction channel for somatosensory and interoceptive signalling, yet coordinated regulation of this channel remains poorly understood. Emerging work reveals that multilayered regulation of PIEZO2 coordinates the expression, function, gating and membrane localization of the channel to maintain homeostatic mechanosensitivity. This review highlights evidence for several interdependent mechanisms by which PIEZO2 is regulated. Specifically, alternative splicing generates PIEZO2 splice isoforms with distinct functions, post-translational modifications modulate gating kinetics of the protein, and a growing network of interacting and non-interacting protein partners influence PIEZO2 mechanosensitivity. Evidence that highlights the clinical importance of PIEZO2 regulation is presented. PIEZO2 variants cause congenital mechanosensory/proprioceptive disorders, often with respiratory complications; and altered PIEZO2 activity or expression has been implicated in tactile allodynia/mechanical hypersensitivity and reported across multiple cancers. Overall, this review integrates current knowledge of PIEZO2 regulation and dysregulation, highlighting how convergent mechanisms shape tissue-specific and context-specific mechanosensitivity. A central conclusion is that PIEZO2 functions as the core of a larger mechanotransduction module and, therefore, the behaviour of the channel cannot be understood by examination of the pore alone. Discussion of these emerging studies highlights an opportunity to gain deeper insight into the regulatory mechanisms of PIEZO2 channels in vivo and may reveal strategies to selectively modulate PIEZO2-dependent mechanosensation in disease.

Astrocytes, the most abundant glial cells in the brain, are wired into neural circuits through close contact with neuronal pre- and post-synapses, called tripartite synapses. The mutual communication between neurons and astrocytes is crucial for neural circuit dynamics and animal behaviour. Recent advancements in imaging, manipulation and transcriptomics in astrocytes have revealed that astrocytes exhibit spatiotemporally complex computations and represent circuit-specialised functions and molecular makeups. However, understanding the neuron-astrocyte circuitry by means of conventional anatomical methods is hindered due to technical limitations. In this review, we highlight recently developed optical, genetic and viral techniques that enable high-throughput identification of connected neuron-astrocyte pairs with circuit and genetic specificity. These approaches will accelerate anatomical and functional dissections of the neuron-astrocyte circuits in health and disease in future studies.