Frontiers in neuroenergetics最新文献

{"title":"A comprehensive metabolic profile of cultured astrocytes using isotopic transient metabolic flux analysis and C-labeled glucose.","authors":"Ana I Amaral,&nbsp;Ana P Teixeira,&nbsp;Bjørn I Håkonsen,&nbsp;Ursula Sonnewald,&nbsp;Paula M Alves","doi":"10.3389/fnene.2011.00005","DOIUrl":"https://doi.org/10.3389/fnene.2011.00005","url":null,"abstract":"<p><p>Metabolic models have been used to elucidate important aspects of brain metabolism in recent years. This work applies for the first time the concept of isotopic transient 13C metabolic flux analysis (MFA) to estimate intracellular fluxes in primary cultures of astrocytes. This methodology comprehensively explores the information provided by 13C labeling time-courses of intracellular metabolites after administration of a 13C-labeled substrate. Cells were incubated with medium containing [1-13C]glucose for 24 h and samples of cell supernatant and extracts collected at different time points were then analyzed by mass spectrometry and/or high performance liquid chromatography. Metabolic fluxes were estimated by fitting a carbon labeling network model to isotopomer profiles experimentally determined. Both the fast isotopic equilibrium of glycolytic metabolite pools and the slow labeling dynamics of TCA cycle intermediates are described well by the model. The large pools of glutamate and aspartate which are linked to the TCA cycle via reversible aminotransferase reactions are likely to be responsible for the observed delay in equilibration of TCA cycle intermediates. Furthermore, it was estimated that 11% of the glucose taken up by astrocytes was diverted to the pentose phosphate pathway. In addition, considerable fluxes through pyruvate carboxylase [PC; PC/pyruvate dehydrogenase (PDH) ratio = 0.5], malic enzyme (5% of the total pyruvate production), and catabolism of branched-chained amino acids (contributing with ∼40% to total acetyl-CoA produced) confirmed the significance of these pathways to astrocytic metabolism. Consistent with the need of maintaining cytosolic redox potential, the fluxes through the malate-aspartate shuttle and the PDH pathway were comparable. Finally, the estimated glutamate/α-ketoglutarate exchange rate (∼0.7 μmol mg prot-1 h-1) was similar to the TCA cycle flux. In conclusion, this work demonstrates the potential of isotopic transient MFA for a comprehensive analysis of energy metabolism.</p>","PeriodicalId":88242,"journal":{"name":"Frontiers in neuroenergetics","volume":"3 ","pages":"5"},"PeriodicalIF":0.0,"publicationDate":"2011-09-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3389/fnene.2011.00005","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"30161704","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Compartmentalized Cerebral Metabolism of [1,6-(13)C]Glucose Determined by in vivo (13)C NMR Spectroscopy at 14.1 T.","authors":"João M N Duarte,&nbsp;Bernard Lanz,&nbsp;Rolf Gruetter","doi":"10.3389/fnene.2011.00003","DOIUrl":"https://doi.org/10.3389/fnene.2011.00003","url":null,"abstract":"<p><p>Cerebral metabolism is compartmentalized between neurons and glia. Although glial glycolysis is thought to largely sustain the energetic requirements of neurotransmission while oxidative metabolism takes place mainly in neurons, this hypothesis is matter of debate. The compartmentalization of cerebral metabolic fluxes can be determined by (13)C nuclear magnetic resonance (NMR) spectroscopy upon infusion of (13)C-enriched compounds, especially glucose. Rats under light α-chloralose anesthesia were infused with [1,6-(13)C]glucose and (13)C enrichment in the brain metabolites was measured by (13)C NMR spectroscopy with high sensitivity and spectral resolution at 14.1 T. This allowed determining (13)C enrichment curves of amino acid carbons with high reproducibility and to reliably estimate cerebral metabolic fluxes (mean error of 8%). We further found that TCA cycle intermediates are not required for flux determination in mathematical models of brain metabolism. Neuronal tricarboxylic acid cycle rate (V(TCA)) and neurotransmission rate (V(NT)) were 0.45 ± 0.01 and 0.11 ± 0.01 μmol/g/min, respectively. Glial V(TCA) was found to be 38 ± 3% of total cerebral oxidative metabolism, accounting for more than half of neuronal oxidative metabolism. Furthermore, glial anaplerotic pyruvate carboxylation rate (V(PC)) was 0.069 ± 0.004 μmol/g/min, i.e., 25 ± 1% of the glial TCA cycle rate. These results support a role of glial cells as active partners of neurons during synaptic transmission beyond glycolytic metabolism.</p>","PeriodicalId":88242,"journal":{"name":"Frontiers in neuroenergetics","volume":"3 ","pages":"3"},"PeriodicalIF":0.0,"publicationDate":"2011-06-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3389/fnene.2011.00003","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29970764","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Lactate fuels the neonatal brain.","authors":"Karl Kasischke","doi":"10.3389/fnene.2011.00004","DOIUrl":"https://doi.org/10.3389/fnene.2011.00004","url":null,"abstract":"The brain is a highly oxidative organ and the prevailing dogma is that under normal conditions glucose is its principal metabolic substrate. However, it is well established that the brain is capable of oxidizing alternative substrates such as acetate, glutamate, lactate, and ketone bodies. The brain's capability to use alternative substrates may enable adaptation to extraordinary metabolic conditions such as fasting with enhanced ketogenesis or special dietary conditions during the suckling period with enhanced availability of fatty acids and ketone bodies. During early postnatal development the plasma levels of lactate, acetoacetate, and hydroxybutyrate are elevated and they replace glucose as the primary metabolic fuel until the weaning period is reached. This period is further characterized by hyperexcitability with network driven bursts of neuronal activity and calcium oscillations (Erecinska et al., 2004). Thus, glucose alone may not be sufficient to sustain the energy demands of the postnatal brain. A related question is whether alternative oxidative substrates such as lactate or hydroxybutyrate per se can sustain synaptic function in this developmental period. \u0000 \u0000To address this question, Ivanov et al. (2011) simultaneously recorded oxygen tension, NAD(P)H fluorescence transients and local field potentials during electrical stimulation of the hippocampal Schaffer collateral pathway in neonatal brain tissue slices from mice. From the very beginning, the authors took great care to ensure both viability and functionality of their preparations. They convincingly demonstrated that surprisingly high superfusion rates with standard artificial cerebrospinal fluid (ACSF) in the slice chamber are required to ensure adequate oxygenation and complete electrical function in blood-free tissue slices. An important implication of this methodological tour de force is that under many previously reported experiments the requirements for viability may been met while the functionality may have been compromised. \u0000 \u0000After ideal experimental conditions were established, hippocampal slices were first superfused with ACSF containing 10 mM glucose and then with modified ACSF containing 5 mM glucose and 5 mM l-lactate. Their reasoning was if glucose alone is sufficient to sustain enhanced network activity, the addition of lactate should not change tissue metabolic states and synaptic function. To the contrary, the addition of lactate significantly increased oxygen consumption (+31%) and enhanced local field potentials (+41%) during the train stimulation (10 s, 10 Hz) and radically modified the biphasic NAD(P)H signaling: the early oxidation phase was strongly augmented while the overshoot was attenuated. \u0000 \u0000The authors went on to investigate whether lactate (10 mM) can sustain synaptic function in the absence of glucose as has been reported for mature brain tissue (Schurr et al., 1988). When lactate was the sole oxidative substrate, the changes were even more pronoun","PeriodicalId":88242,"journal":{"name":"Frontiers in neuroenergetics","volume":"3 ","pages":"4"},"PeriodicalIF":0.0,"publicationDate":"2011-06-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3389/fnene.2011.00004","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29947906","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Lactate Effectively Covers Energy Demands during Neuronal Network Activity in Neonatal Hippocampal Slices.","authors":"Anton Ivanov,&nbsp;Marat Mukhtarov,&nbsp;Piotr Bregestovski,&nbsp;Yuri Zilberter","doi":"10.3389/fnene.2011.00002","DOIUrl":"https://doi.org/10.3389/fnene.2011.00002","url":null,"abstract":"<p><p>Although numerous experimental data indicate that lactate is efficiently used for energy by the mature brain, the direct measurements of energy metabolism parameters during neuronal network activity in early postnatal development have not been performed. Therefore, the role of lactate in the energy metabolism of neurons at this age remains unclear. In this study, we monitored field potentials and contents of oxygen and NAD(P)H in correlation with oxidative metabolism during intense network activity in the CA1 hippocampal region of neonatal brain slices. We show that in the presence of glucose, lactate is effectively utilized as an energy substrate, causing an augmentation of oxidative metabolism. Moreover, in the absence of glucose lactate is fully capable of maintaining synaptic function. Therefore, during network activity in neonatal slices, lactate can be an efficient energy substrate capable of sustaining and enhancing aerobic energy metabolism.</p>","PeriodicalId":88242,"journal":{"name":"Frontiers in neuroenergetics","volume":" ","pages":"2"},"PeriodicalIF":0.0,"publicationDate":"2011-05-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3389/fnene.2011.00002","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40101082","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A guide to delineate the logic of neurovascular signaling in the brain.","authors":"David Kleinfeld,&nbsp;Pablo Blinder,&nbsp;Patrick J Drew,&nbsp;Jonathan D Driscoll,&nbsp;Arnaud Muller,&nbsp;Philbert S Tsai,&nbsp;Andy Y Shih","doi":"10.3389/fnene.2011.00001","DOIUrl":"https://doi.org/10.3389/fnene.2011.00001","url":null,"abstract":"<p><p>The neurovascular system may be viewed as a distributed nervous system within the brain. It transforms local neuronal activity into a change in the tone of smooth muscle that lines the walls of arterioles and microvessels. We review the current state of neurovascular coupling, with an emphasis on signaling molecules that convey information from neurons to neighboring vessels. At the level of neocortex, this coupling is mediated by: (i) a likely direct interaction with inhibitory neurons, (ii) indirect interaction, via astrocytes, with excitatory neurons, and (iii) fiber tracts from subcortical layers. Substantial evidence shows that control involves competition between signals that promote vasoconstriction versus vasodilation. Consistent with this picture is evidence that, under certain circumstances, increased neuronal activity can lead to vasoconstriction rather than vasodilation. This confounds naïve interpretations of functional brain images. We discuss experimental approaches to detect signaling molecules in vivo with the goal of formulating an empirical basis for the observed logic of neurovascular control.</p>","PeriodicalId":88242,"journal":{"name":"Frontiers in neuroenergetics","volume":"3 ","pages":"1"},"PeriodicalIF":0.0,"publicationDate":"2011-04-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3389/fnene.2011.00001","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29871909","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Microglia in close vicinity of glioma cells: correlation between phenotype and metabolic alterations.","authors":"Pierre Voisin,&nbsp;Véronique Bouchaud,&nbsp;Michel Merle,&nbsp;Philippe Diolez,&nbsp;Laura Duffy,&nbsp;Kristian Flint,&nbsp;Jean-Michel Franconi,&nbsp;Anne-Karine Bouzier-Sore","doi":"10.3389/fnene.2010.00131","DOIUrl":"https://doi.org/10.3389/fnene.2010.00131","url":null,"abstract":"<p><p>Microglia are immune cells within the central nervous system. In brain-developing tumors, gliomas are able to silence the defense and immune functions of microglia, a phenomenon which strongly contributes to tumor progression and treatment resistance. Being activated and highly motile, microglia infiltrate tumors and secrete macrophagic chemoattractant factors. Thereafter, the tumor cells shut down their immune properties and stimulate the microglia to release tumor growth-promoting factors. The result of such modulation is that a kind of symbiosis occurs between microglia and tumor cells, in favor of tumor growth. However, little is known about microglial phenotype and metabolic modifications in a tumoral environment. Co-cultures were performed using CHME5 microglia cells grown on collagen beads or on coverslips and placed on monolayer of C6 cells, limiting cell/cell contacts. Phagocytic behavior and expression of macrophagic and cytoskeleton markers were monitored. Respiratory properties and energetic metabolism were also studied with regard to the activated phenotype of microglia. In co-cultures, transitory modifications of microglial morphology and metabolism were observed linked to a concomitant transitory increase of phagocytic properties. Therefore, after 1 h of co-culture, microglia were activated but when longer in contact with tumor cells, phagocytic properties appear silenced. Like the behavior of the phenotype, microglial respiration showed a transitory readjustment although the mitochondria maintained their perinuclear relocation. Nevertheless, the energetic metabolism of the microglia was altered, suggesting a new energetic steady state. The results clearly indicate that like the depressed immune properties, the macrophagic and metabolic status of the microglia is quickly driven by the glioma environment, despite short initial phagocytic activation. Such findings question the possible contribution of diffusible tumor factors to the microglial metabolism.</p>","PeriodicalId":88242,"journal":{"name":"Frontiers in neuroenergetics","volume":"2 ","pages":"131"},"PeriodicalIF":0.0,"publicationDate":"2010-10-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3389/fnene.2010.00131","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29429548","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Pathophysiological interference with neurovascular coupling - when imaging based on hemoglobin might go blind.","authors":"Ute Lindauer,&nbsp;Ulrich Dirnagl,&nbsp;Martina Füchtemeier,&nbsp;Caroline Böttiger,&nbsp;Nikolas Offenhauser,&nbsp;Christoph Leithner,&nbsp;Georg Royl","doi":"10.3389/fnene.2010.00025","DOIUrl":"https://doi.org/10.3389/fnene.2010.00025","url":null,"abstract":"<p><p>Assessing neuronal activity by non-invasive functional brain imaging techniques which are based on the hemodynamic response depends totally on the physiological cascade of metabolism and blood flow. At present, functional brain imaging with near infrared spectroscopy (NIRS) or BOLD-fMRI is widely used in cognitive neuroscience in healthy subjects where neurovascular coupling and cerebrovascular reactivity can be assumed to be intact. Local activation studies as well as studies investigating functional connectivity between brain regions of the resting brain provide a rapidly increasing body of knowledge on brain function in humans and animals. Furthermore, functional NIRS and MRI techniques are increasingly being used in patients with severe brain diseases and this use might gain more and more importance for establishing their use in the clinical routine. However, more and more experimental evidence shows that changes in baseline physiological parameters, pharmacological interventions, or disease-related vascular changes may significantly alter the normal response of blood flow and blood oxygenation and thus may lead to misinterpretation of neuronal activity. In this article we present examples of recent experimental findings on pathophysiological changes of neurovascular coupling parameters in animals and discuss their potential implications for functional imaging based on hemodynamic signals such as fNIRS or BOLD-fMRI. To enable correct interpretation of neuronal activity by vascular signals, future research needs to deepen our understanding of the basic mechanisms of neurovascular coupling and the specific characteristics of disturbed neurovascular coupling in the diseased brain.</p>","PeriodicalId":88242,"journal":{"name":"Frontiers in neuroenergetics","volume":"2 ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2010-10-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3389/fnene.2010.00025","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29360989","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Recent Advances in High-Resolution MR Application and Its Implications for Neurovascular Coupling Research.","authors":"Noam Harel,&nbsp;Patrick J Bolan,&nbsp;Robert Turner,&nbsp;Kamil Ugurbil,&nbsp;Essa Yacoub","doi":"10.3389/fnene.2010.00130","DOIUrl":"https://doi.org/10.3389/fnene.2010.00130","url":null,"abstract":"<p><p>The current understanding of fMRI, regarding its vascular origins, is based on numerous assumptions and theoretical modeling, but little experimental validation exists to support or challenge these models. The known functional properties of cerebral vasculature are limited mainly to the large pial surface and the small capillary level vessels. However, a significant lack of knowledge exists regarding the cluster of intermediate-sized vessels, mainly the intracortical, connecting these two groups of vessels and where, arguably, key blood flow regulation takes place. In recent years, advances in MR technology and methodology have enabled the probing of the brain, both structurally and functionally, at resolutions and coverage not previously attainable. Functional MRI has been utilized to map functional units down to the levels of cortical columns and lamina. These capabilities open new possibilities for investigating neurovascular coupling and testing hypotheses regarding fundamental cerebral organization. Here, we summarize recent cutting-edge MR applications for studying neurovascular and functional imaging, both in humans as well as in animal models. In light of the described imaging capabilities, we put forward a theory in which a cortical column, an ensemble of neurons involved in a particular neuronal computation is spatially correlated with a specific vascular unit, i.e., a cluster of an emerging principle vein surrounded by a set of diving arteries. If indeed such a correlation between functional (neuronal) and structural (vascular) units exist as a fundamental intrinsic cortical feature, one could conceivably delineate functional domains in cortical areas that are not known or have not been identified.</p>","PeriodicalId":88242,"journal":{"name":"Frontiers in neuroenergetics","volume":"2 ","pages":"130"},"PeriodicalIF":0.0,"publicationDate":"2010-09-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3389/fnene.2010.00130","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29444708","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Astroglial Wiring is Adding Complexity to Neuroglial Networking.","authors":"Christian Giaume","doi":"10.3389/fnene.2010.00129","DOIUrl":"https://doi.org/10.3389/fnene.2010.00129","url":null,"abstract":"<p><p>Astrocytes are organized as networks of communicating cells due to their high expression level of connexins, the molecular constituents of gap junction channels. Based on their permeability properties for ions and small signaling molecules such astroglial wiring interferes with neuronal activity and survival. In this paper, I identify and discuss which future technical and conceptual progress or advances should be achieved in order to better understand how neuroglial networking contributes to brain functions and dysfunctions.</p>","PeriodicalId":88242,"journal":{"name":"Frontiers in neuroenergetics","volume":"2 ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2010-09-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3389/fnene.2010.00129","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29335688","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Imaging retinal blood flow with laser speckle flowmetry.","authors":"Anja I Srienc,&nbsp;Zeb L Kurth-Nelson,&nbsp;Eric A Newman","doi":"10.3389/fnene.2010.00128","DOIUrl":"https://doi.org/10.3389/fnene.2010.00128","url":null,"abstract":"<p><p>Laser speckle flowmetry (LSF) was initially developed to measure blood flow in the retina. More recently, its primary application has been to image baseline blood flow and activity-dependent changes in blood flow in the brain. We now describe experiments in the rat retina in which LSF was used in conjunction with confocal microscopy to monitor light-evoked changes in blood flow in retinal vessels. This dual imaging technique permitted us to stimulate retinal photoreceptors and measure vessel diameter with confocal microscopy while simultaneously monitoring blood flow with LSF. We found that a flickering light dilated retinal arterioles and evoked increases in retinal blood velocity with similar time courses. In addition, focal light stimulation evoked local increases in blood velocity. The spatial distribution of these increases depended on the location of the stimulus relative to retinal arterioles and venules. The results suggest that capillaries are largely unresponsive to local neuronal activity and that hemodynamic responses are mediated primarily by arterioles. The use of LSF to image retinal blood flow holds promise in elucidating the mechanisms mediating functional hyperemia in the retina and in characterizing changes in blood flow that occur during retinal pathology.</p>","PeriodicalId":88242,"journal":{"name":"Frontiers in neuroenergetics","volume":"2 ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2010-09-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.3389/fnene.2010.00128","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29349678","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}