Multiple sclerosis is a chronic, autoimmune and neurodegenerative disease affecting multiple functional systems and resulting in motor impairments associated with muscle weakness and lack of movement ...coordination. We quantified upper limb motor deficits with a robot-based assessment including behavioral and muscle synergy analysis in 11 multiple sclerosis subjects with mild to moderate upper limb impairment (9 female; 50 ± 10 years) compared to 11 age- and gender- matched controls (9 female; 50 ± 9 years). All subjects performed planar reaching tasks by moving their upper limb or applying force while grasping the handle of a robotic manipulandum that generated four different environments: free space, assistive or resistive forces, and rigid constraint. We recorded the activity of 15 upper body muscles. Multiple sclerosis subjects generated irregular trajectories. While activities in isolated arm muscles appeared generally normal, shoulder muscle coordination with arm motions was impaired and there was a marked co-activation of the biceps and triceps in extension movements. Systematic differences in timing and organization of muscle synergies have also been observed. This study supports the definition of new biomarkers and rehabilitative treatments for improving upper limb motor coordination in multiple sclerosis.
Astrocytic glycogen, the only storage form of glucose in the brain, has been shown to play a fundamental role in supporting learning and memory, an effect achieved by providing metabolic support for ...neurons. We have examined the interplay between glycogenolysis and the bioenergetics of astrocytic Ca2+ homeostasis, by analyzing interdependency of glycogen and store‐operated Ca2+ entry (SOCE), a mechanism in cellular signaling that maintains high endoplasmatic reticulum (ER) Ca2+ concentration and thus provides the basis for store‐dependent Ca2+ signaling. We stimulated SOCE in primary cultures of murine cerebellar and cortical astrocytes, and determined glycogen content to investigate the effects of SOCE on glycogen metabolism. By blocking glycogenolysis, we tested energetic dependency of SOCE‐related Ca2+ dynamics on glycogenolytic ATP. Our results show that SOCE triggers astrocytic glycogenolysis. Upon inhibition of adenylate cyclase with 2',5'‐dideoxyadenosine, glycogen content was no longer significantly different from that in unstimulated control cells, indicating that SOCE triggers astrocytic glycogenolysis in a cAMP‐dependent manner. When glycogenolysis was inhibited in cortical astrocytes by 1,4‐dideoxy‐1,4‐imino‐D‐arabinitol, the amount of Ca2+ loaded into ER via sarco/endoplasmic reticulum Ca2‐ATPase (SERCA) was reduced, which suggests that SERCA pumps preferentially metabolize glycogenolytic ATP. Our study demonstrates SOCE as a novel pathway in stimulating astrocytic glycogenolysis. We also provide first evidence for a new functional role of brain glycogen, in providing local ATP to SERCA, thus establishing the bioenergetic basis for astrocytic Ca2+ signaling. This mechanism could offer a novel explanation for the impact of glycogen on learning and memory. GLIA 2014;62:526–534
Main Points
Store‐operated ca2+ entry triggers glycogenolysis
The signaling pathway is cAMP‐dependent
Glycogenolytic ATP supports store‐dependent Ca2+ signaling
Glycogen phosphorylase (GP) is activated to degrade glycogen in response to different stimuli, to support both the astrocyte's own metabolic demand and the metabolic needs of neurons. The regulatory ...mechanism allowing such a glycogenolytic response to distinct triggers remains incompletely understood. In the present study, we used siRNA‐mediated differential knockdown of the two isoforms of GP expressed in astrocytes, muscle isoform (GPMM), and brain isoform (GPBB), to analyze isoform‐specific regulatory characteristics in a cellular setting. Subsequently, we tested the response of each isoform to phosphorylation, triggered by incubation with norepinephrine (NE), and to AMP, increased by glucose deprivation in cells in which expression of one GP isoform had been silenced. Successful knockdown was demonstrated on the protein level by Western blot, and on a functional level by determination of glycogen content showing an increase in glycogen levels following knockdown of either GPMM or GPBB. NE triggered glycogenolysis within 15 min in control cells and after GPBB knockdown. However, astrocytes in which expression of GPMM had been silenced showed a delay in response to NE, with glycogen levels significantly reduced only after 60 min. In contrast, allosteric activation of GP by AMP, induced by glucose deprivation, seemed to mainly affect GPBB, as only knockdown of GPBB, but not of GPMM, delayed the glycogenolytic response to glucose deprivation. Our results indicate that the two GP isoforms expressed in astrocytes respond to different physiological triggers, therefore conferring distinct metabolic functions of brain glycogen. GLIA 2015;63:154–162
Main Points
This is the first evidence in a cellular system of distinct regulation of the two isoforms of glycogen phosphorylase expressed in astrocytes.
The muscle isoform of glycogen phosphorylase (GPMM) responds primarily to phosphorylation, whereas the brain isoform (GPBB) primarily responds to allosteric activation by AMP.
Thus, in astrocytes GPBB responds mainly to the energy status of the cell, whereas GPMM responds mostly to receptor‐mediated signaling cascades.
As top predators, falcons possess unique morphological, physiological and behavioral adaptations that allow them to be successful hunters: for example, the peregrine is renowned as the world's ...fastest animal. To examine the evolutionary basis of predatory adaptations, we sequenced the genomes of both the peregrine (Falco peregrinus) and saker falcon (Falco cherrug), and we present parallel, genome-wide evidence for evolutionary innovation and selection for a predatory lifestyle. The genomes, assembled using Illumina deep sequencing with greater than 100-fold coverage, are both approximately 1.2 Gb in length, with transcriptome-assisted prediction of approximately 16,200 genes for both species. Analysis of 8,424 orthologs in both falcons, chicken, zebra finch and turkey identified consistent evidence for genome-wide rapid evolution in these raptors. SNP-based inference showed contrasting recent demographic trajectories for the two falcons, and gene-based analysis highlighted falcon-specific evolutionary novelties for beak development and olfaction and specifically for homeostasis-related genes in the arid environment-adapted saker.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Extracellular ATP plays important roles in coordinating the activities of astrocytes and neurons, and aberrant signalling is associated with neurodegenerative diseases. In rodents, ATP stimulates ...opening of Ca2+‐permeable channels formed by P2X receptor subunits in the plasma membrane. It is widely assumed, but not verified, that P2X receptors also evoke Ca2+ signals in human astrocytes. Here, we directly assess this hypothesis. We showed that cultured foetal cortical human astrocytes express mRNA for several P2X receptor subunits (P2X4, P2X5, P2X6) and G protein‐coupled P2Y receptors (P2Y1, P2Y2, P2Y6, P2Y11). In these astrocytes, ATP stimulated Ca2+ release from intracellular stores through IP3 receptors and store‐operated Ca2+ entry. These responses were entirely mediated by P2Y1 and P2Y2 receptors. Agonists of P2X receptors did not evoke Ca2+ signals, and nor did ATP when Ca2+ release from intracellular stores and store‐operated Ca2+ entry were inhibited. We conclude that ATP‐evoked Ca2+ signals in cultured human foetal astrocytes are entirely mediated by P2Y1 and P2Y2 receptors, with no contribution from P2X receptors.
Extracellular ATP evokes Ca2+ signals in astrocytes, thereby coordinating their activity with neurons. It is assumed that ATP opens plasma membrane Ca2+‐permeable channels formed by P2X receptors. We demonstrate that in cultured foetal cortical human astrocytes, all ATP‐evoked Ca2+ signals are mediated by G protein‐coupled P2Y1 and P2Y2 receptors, which stimulate formation of IP3 and release of Ca2+ from the ER.
Extracellular ATP plays important roles in coordinating the activities of astrocytes and neurons, and aberrant signalling is associated with neurodegenerative diseases. In rodents, ATP stimulates ...opening of Ca
-permeable channels formed by P2X receptor subunits in the plasma membrane. It is widely assumed, but not verified, that P2X receptors also evoke Ca
signals in human astrocytes. Here, we directly assess this hypothesis. We showed that cultured foetal cortical human astrocytes express mRNA for several P2X receptor subunits (P2X
, P2X
, P2X
) and G protein-coupled P2Y receptors (P2Y
, P2Y
, P2Y
, P2Y
). In these astrocytes, ATP stimulated Ca
release from intracellular stores through IP
receptors and store-operated Ca
entry. These responses were entirely mediated by P2Y
and P2Y
receptors. Agonists of P2X receptors did not evoke Ca
signals, and nor did ATP when Ca
release from intracellular stores and store-operated Ca
entry were inhibited. We conclude that ATP-evoked Ca
signals in cultured human foetal astrocytes are entirely mediated by P2Y
and P2Y
receptors, with no contribution from P2X receptors.
Highlights • Vesicular release of glutamate is reduced by NAX-5055 in vitro. • The extracellular glutamate concentration is reduced by NAX-5055 in cultured neurons. • The extracellular GABA ...concentration is elevated by NAX-5055 in cultured neurons. • The in vitro effects of NAX-5055 are in line with an anticonvulsant action in vivo. • NAX-5055 does not induce toxicity in cultured neurons.
After its uptake into the cytosol, intracellular glucose is phosphorylated to glucose-6-phosphate (G6P), trapping it within the cell and preparing it for metabolism. In glucose-exporting tissues, ...like liver, G6P is transported into the ER, where it is dephosphorylated by G6Pase-α. The glucose is then returned to the cytosol for export 1, 2. Defects in these pathways cause glycogen storage diseases 1. G6Pase-β, an isozyme of G6Pase-α, is widely expressed 3, 4. Its role in cells that do not export glucose is unclear, although mutations in G6Pase-β cause severe and widespread abnormalities 5–7. Astrocytes, the most abundant cells in the brain, provide metabolic support to neurons, facilitated by astrocytic endfeet that contact blood capillaries or neurons 8–12. Perivascular endfeet are the main site of glucose uptake by astrocytes 13, but in human brain they may be several millimeters away from the perineuronal processes 14. We show that cultured human fetal astrocytes express G6Pase-β, but not G6Pase-α. ER-targeted glucose sensors 15, 16 reveal that G6Pase-β allows the ER of human astrocytes to accumulate glucose by importing G6P from the cytosol. Glucose uptake by astrocytes, ATP production, and Ca2+ accumulation by the ER are attenuated after knockdown of G6Pase-β using lentivirus-delivered shRNA and substantially rescued by expression of G6Pase-α. We suggest that G6Pase-β activity allows effective uptake of glucose by astrocytes, and we speculate that it allows the ER to function as an intracellular “highway” delivering glucose from perivascular endfeet to the perisynaptic processes.
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•Glucose-6-phosphatase-β (G6Pase-β) is expressed in human astrocytes•G6P is sequestered by ER and dephosphorylated to glucose in the lumen by G6Pase-β•Loss of G6Pase-β reduces glucose uptake, intracellular ATP, and ER Ca2+ content•ER may provide a protected highway for long-range glucose transport in astrocytes
Müller et al. use targeted glucose sensors to show that glucose-6-phosphate uptake by the endoplasmic reticulum of human astrocytes and its dephosphorylation in the ER by glucose-6-phosphatase-β deliver glucose to the ER lumen and sustain cellular glucose uptake. The ER lumen may provide an intracellular protected highway for glucose transport.
Astrocytic glycogen degradation is an important factor in metabolic support of brain function, particularly during increased neuronal firing. In this context, glycogen is commonly thought of as a ...source for the provision of energy substrates, such as lactate, to neurons. However, the signalling pathways eliciting glycogen degradation inside astrocytes are themselves energy-demanding processes, a fact that has been emphasized in recent studies, demonstrating dependence of these signalling mechanisms on glycogenolytic ATP.
•Concentration levels of protoanemonin and anemonin in H.niger L. and P.vulgaris extracts were monitored by HPLC-DAD analysis.•Stability of protoanemonin and anemonin was studied under different ...storage conditions.•Protoanemonin levels decreased rapidly over time and in particular when initial concentration was high; low levels remained fairly constant over years.•Overview of protoanemonin and anemonin concentrations levels in a range of extract batches is reported.
The concentration levels and stability of protoanemonin, a characteristic constituent of Ranunculaceae species with antimicrobial and fungicidal properties, were studied for the first time in plant extracts prepared from Helleborus niger L. and Pulsatilla vulgaris Mill. using fermentative production processes. Protoanemonin levels quantified by HPLC-DAD analysis were 0.0345 and 0.0662 mg/g in two freshly prepared Helleborus (whole, flowering plant) extracts and 0.3875 mg/g and 0.4193 mg/g in Pulsatilla (flowers) extracts. Protoanemonin proved to be rather instable in aqueous-fermented extracts stored at 15 °C in the dark, and its concentration decreased rapidly over 12 months of storage independently of the plant species. The decrease was most pronounced when initial concentrations were high (decrease by about 70%). In contrast, low protoanemonin levels remained stable in solution for more than 12 months. Anemonin, the dimer of protoanemonin, was detected in increasing concentrations only in Pulsatilla samples, but its concentration only accounted for less than 50% of the theoretically expected amount. With respect to fermented extracts, both physical processes such as self-polymerization and adsorption/binding to other extract constituents as well as biodegradation were concluded to be responsible for protoanemonin decline. As opposed to plant extracts, both protoanemonin and anemonin levels decreased in 0.22 μm-filtered samples stored in vials. This may be explained by a reduced release from plant material in combination with physicochemically induced degradation. Reduction was most pronounced upon light exposure and elevated temperatures, clearly indicating that photochemical degradation is involved. Contents of protoanemonin in a set of extract batches were 0.0896 ± 0.0125 mg/g and 0.0618 ± 0.0180 mg/g in Helleborus and Pulsatilla extracts, and anemonin levels were 0.0230 ± 0.0076 mg/g and 0.0482 ± 0.0282 mg/g, respectively. Due to its antibiotic effects, but also its reactivity, protoanemonin is a therapeutically and toxicologically relevant constituent, and its concentration should therefore be carefully monitored.
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