Function (Oxford, England)最新文献

Pattern separation, the ability of a network to distinguish similar inputs by transforming them into distinct outputs, was postulated by the Marr-Albus theory to be realized by divergent feedforward excitatory connectivity. Yet, there is evidence for strong but differential regulation of pattern separation by local circuit connectivity. How do we reconcile the conflicting views on local-circuit regulation of pattern separation in circuits receiving divergent feedforward connectivity? Here, we quantitatively examined a population of heterogeneous dentate gyrus (DG) spiking networks where identically divergent feedforward connectivity was enforced. We generated 20 000 random DG networks constructed with thousands of functionally validated, heterogeneous single-neuron models of 4 different DG neuronal subtypes. We recorded network outputs to morphed sets of input patterns and applied quantitative metrics that we developed to assess pattern separation performance of each network. Surprisingly, only 47 of these 20 000 networks (0.23%) manifested effective pattern separation showing that divergent feedforward connectivity alone does not guarantee pattern separation. Instead, our analyses unveiled strong contributions from the 3 interneuron subtypes toward granule cell sparsity and pattern separation, with pronounced network-to-network variability in such contributions. We traced this variability to differences in local synaptic weights across pattern-separating networks, highlighting synaptic degeneracy as a key mechanism that explains diversity in interneuronal regulation of pattern separation. Finally, we found heterogeneous DG networks to be more resilient to synaptic jitter compared to their homogeneous counterparts. Together, our findings reconcile conflicting evidence by revealing degeneracy in DG circuits, whereby similar pattern separation efficacy can arise through diverse interactions among granule cells and interneurons.

The proper functioning of lymphatic valves is critical for unidirectional lymph transport. Valve development and maintenance depends on multiple genes in lymphatic endothelium, including those controlling the expression of 4 connexin (Cx) isoforms-Cx37, Cx47, Cx43, and Cx45. The relative importance of these isoforms for valve function is undefined, but primary human lymphedema is linked to loss-of-function mutations in Cx47 or Cx43, while deficiencies in Cx43 or Cx45 produce functional valve defects in mice. Tests of back leak and closure for single lymphatic valves from mice with selective deficiency of each Cx isoform revealed defects associated with the loss of Cx37 or Cx43, but not Cx47. Combined deletion of multiple isoforms, including Cx45 but not Cx47, produced even more severe valve defects in certain genotypes, sometimes with nearly complete regression of valves within 6 d. Back leak across connexin-deficient LVs correlated highly with gaps between the commissures formed by leaflet insertion into the vessel wall, indicating that connexin function may be critical for the formation and/or maintenance of leaflet commissures. Our results reveal the following hierarchy of Cx importance in valve function: Cx37 = Cx43 > Cx45 > Cx47 and predict that patients with loss of function mutations in Cx37 (GJA4) should develop lymphedema. We propose a general classification scheme describing 4 stages of progressive valve dysfunction.

Sodium-glucose cotransporter 2 inhibitors (SGLT2i) exhibit cardiorenal protective effects that likely involve mechanisms aside from SGLT2 inhibition. Still, many details surrounding these clinically important pleiotropic effects remain unclear. We previously showed that several SGLT2-independent proximal tubular transport functions are inhibited by canagliflozin, but not empagliflozin. Here, we demonstrate a canagliflozin-specific reduction in Sgk1 abundance in both opossum kidney proximal tubule and mouse cortical collecting duct (mCCDcl1) cells, pointing to a possible underlying mechanism. Given the role of Sgk1 in the distal nephron, we hypothesized that canagliflozin would also inhibit epithelial Na+ channel (ENaC)-dependent Na+ transport. Canagliflozin inhibited ENaC-dependent Na+ transport (amiloride-sensitive short circuit current; ISC) in mCCDcl1 cells while empagliflozin had no effect. Selective membrane permeabilization revealed canagliflozin-induced inhibition of both apical conductance through ENaC and basolateral transport via the Na+/K+ ATPase. These effects were mimicked by the selective Sgk1 inhibitor, GSK650394. Surface labeling studies demonstrated reduced membrane localization of ENaC, but not Na+/K+ ATPase subunits, consistent with a mechanism involving Sgk1. Canagliflozin reduced ISC in the presence and absence of rotenone, suggesting inhibition occurs independently of effects on mitochondrial complex I, another known target of canagliflozin. ENaC activity in mouse distal colon was also inhibited by canagliflozin, confirming these effects in native tissue. We identify Na+ transport through ENaC and the Na+/K+ ATPase as novel targets of inhibition by canagliflozin, with Sgk1 as a likely upstream mechanistic component. Canagliflozin-specific effects on transport mediated via this mechanism may contribute to non-class effects of this drug observed clinically.

Abnormalities of Ca2+ signaling in the heart lead to common cardiac remodeling in the pathogenesis of cardiovascular disorders. The activation of calmodulin-dependent protein kinase II (CaMKII) is regulated by elevated intracellular Ca2+ level in cardiomyocytes, driving the progression of myocardial dysfunction. In this study, using models of β2 adrenergic receptor (β2AR) deficiency in cardiomyocytes (β2AR-CKO), we observed an increased phosphorylation of CaMKII and upregulation of gene expression and protein level of the fibrotic marker connective tissue growth factor (CTGF) in the myocytes. In vivo treatment with the CaMKII inhibitor KN93 attenuated the upregulation of CTGF protein expression in β2AR-CKO hearts. Enhanced L-type calcium channel (LTCC) current was observed in β2AR-CKO cardiomyocytes following adrenergic stimulation, indicating a disruption of Ca2+ signaling. Treatment with the LTCC blocker nifedipine attenuated CaMKII activity and the expression of CTGF in β2AR-CKO hearts, confirming the upstream role of abnormal LTCC-Ca2+ signaling. Additionally, 8-month-old β2AR-CKO mice exhibited cardiac fibrosis and diastolic dysfunction. One month of in vivo nifedipine treatment improved both cardiac dysfunction and fibrosis in β2AR-CKO mice. These findings highlight the critical role of cardiomyocyte β2AR in maintaining LTCC-Ca2+ homeostasis. Loss of β2AR amplifies the Ca2+-CaMKII axis, promoting fibrosis and cardiomyopathy in aging hearts.

Dietary load and composition are known contributors that accelerate cyst growth in polycystic kidney disease (PKD). High protein intake, which increases amino acid burden in the kidneys, is one such factor. Despite identical protein load, a plant-based wheat-gluten (WG) diet was recently reported to blunt the inflammatory response of animal-based casein diet in a hypertensive model. Considering the importance of pro-inflammatory signals on cystogenesis in PKD, we therefore sought to determine whether a WG compared to casein diet would decelerate cyst progression. Tamoxifen-inducible, global Pkd1 knockout mice were fed either a low casein (6%), high casein (60%), or high wheat-gluten (60%) protein diet for 6 wk. In a separate cohort, mice were gavaged daily with vehicle, lysine, or glutamine for 4 wk while maintained on a normal protein (18%) diet. Tissues were used for histology, flow cytometry, mitochondrial function, metabolomics, and various biochemical assays. WG-fed mice had better kidney function and reduced kidney macrophage percentages, proinflammatory cytokine expression, and cyst growth compared to casein-fed mice. Protein source did not alter kidney mitochondria function. Supplementation with lysine, the highest amino acid in casein versus WG diet, increased kidney cyst growth, acid production, and metabolic disarray. This did not occur with glutamine supplementation, the highest amino acid in WG versus casein diet, despite increased glomerular filtration rate with both amino acids. Neither supplementation mounted an inflammatory response. A plant-based, low-lysine diet slows disease burden in a murine model of PKD. This easily modifiable diet may be a beneficial intervention for PKD patients.

Low testosterone in males (hypogonadism) is associated with limb muscle mass loss, yet the underlying mechanisms of muscle mass loss remain largely unknown. We previously showed androgen deprivation disrupted limb muscle molecular clock function, and the disruption coincided with elevated levels of the primary molecular clock suppressor, Period 2 (Per2). The purposes herein were to determine if PER2 overexpression leads to muscle atrophy and if preventing PER2 accumulation blunts limb muscle mass loss in response to androgen deprivation. Here, we identify Per2 as a negative regulator of muscle size. Overexpression of Per2 in differentiated C2C12 myotubes reduced myotube diameter, while deletion of Per2 in male mice partially preserved tibialis anterior (TA) mass following castration. The muscle-sparing effect of Per2 deletion in vivo was specific to the TA despite evidence of molecular clock disruption and mass loss in other muscles. Subsequently, we show overexpression of the other primary clock suppressor, Period 1 (Per1) also reduced myotube diameter in differentiated C2C12 myotubes. Mechanistically, both Per1 and Per2 overexpression in vitro induced muscle atrophy in part by an autocrine-mediated mechanism likely involving inflammation as their overexpression induced an inflammatory gene expression signature and increased cytokine/chemokine secretion. Moreover, incubation of C2C12 myotubes in the media conditioned from Per1 or Per2 overexpressing myotubes reduced myotube diameter. Several inflammatory genes identified in vitro were also altered in the limb muscles in response to androgen deprivation. These findings identify a previously unrecognized role for Per1/2 in regulating skeletal muscle mass with implications for muscle loss during hypogonadism.

The fusion of skeletal muscle stem cell (MuSC) to myofibers during hypertrophy has exclusively focused on the transfer of the MuSC nucleus, leaving the fate of other MuSC organelles, such as mitochondria, largely unexplored. The objective of this study was to determine whether MuSCs transfer their mitochondria upon myofiber fusion in response to a hypertrophic stimulus. To achieve this goal, we specifically labeled MuSC mitochondria with Dendra2 fluorescence by crossing the MuSC-specific CreER (Pax7CreER/CreER) mouse with the Rosa26-Dendra2 mouse to generate the Pax7-Dendra2 mouse. To induce the fusion of MuSC to myofibers, Pax7-Dendra2 mice underwent synergist ablation surgery to induce mechanical overload (MOV) of plantaris muscle for 3, 7, and 14 d. To track MuSC proliferation, a mini-osmotic pump was implanted at the time of MOV to continuously deliver EdU. Our study revealed a progressive increase in Dendra2-positive fibers across the MOV time course. Three distinct patterns or domains of Dendra2 fluorescence within myofibers were identified and designated as newly fused, crescent, or diffuse. From these Dendra2+ domain types, we inferred MuSC fusion dynamics which indicated MuSC fusion occurred prior to mechanical overload day 3 (MOV-3) and preferentially with Type 2A fibers. Quantification of EdU+ myonuclei found the majority of early (MOV < 3 d) MuSC fusion was division-independent, while proliferating MuSCs contributed primarily to later fusion events. The results of this study provide the first evidence that MuSC mitochondria are transferred to myofibers upon fusion during hypertrophy while, unexpectedly, revealing a greater complexity in MuSC fusion than previously recognized.