Neuronal activity, astrocytic responses to this activity, and energy homeostasis are linked together during baseline, conscious conditions, and short-term rapid activation (as occurs with sensory or motor function). Nervous system energy homeostasis also varies during long-term physiological conditions (i.e., development and aging) and with adaptation to pathological conditions, such as ischemia or low glucose. Neuronal activation requires increased metabolism (i.e., ATP generation) which leads initially to substrate depletion, induction of a variety of signals for enhanced astrocytic function, and increased local blood flow and substrate delivery. Energy generation (particularly in mitochondria) and use during ATP hydrolysis also lead to considerable heat generation. The local increases in blood flow noted following neuronal activation can both enhance local substrate delivery but also provides a heat sink to help cool the brain and removal of waste by-products. In this review we highlight the interactions between short-term neuronal activity and energy metabolism with an emphasis on signals and factors regulating astrocyte function and substrate supply.

The use of energy substrates, such as lactate and pyruvate, has been shown to improve synaptic function when administered during glucose deprivation. In the present study, we investigatedwhether prolonged incubation with monocarboxylate (pyruvate or lactate) prior rather than duringglucose deprivation can also sustainsynaptic and metabolic function.Pyruvate pre-incubation(3-4 hr) significantly prolonged (>25 min) the tolerance of rat hippocampal slices to delayed glucose deprivation compared to control and lactate pre-incubated slices, as revealed by field excitatory post synaptic potentials (fEPSPs); pre-incubation with pyruvate alsoreduced the marked decrease in NAD(P)H fluorescence resulting from glucose deprivation. Moreover, pyruvate exposure led to enhancement of glycogen stores with time, compared to glucose alone(12 μmol/g tissue at 4 hr vs. 3.5 μmol/g tissue). Prolonged resistance to glucose deprivation following exogenous pyruvate incubation was prevented by glycogenolysis inhibitors, suggesting that enhanced glycogen mediates the delay in synaptic activity failure.The application of an adenosine A1 receptor antagonist enhanced glycogen utilization and prolonged the time to synaptic failure, further confirming this hypothesis of the importance of glycogen.Moreover, tissue levels of ATP werealso significantly maintained duringglucose deprivation in pyruvate pre-treated slices compared to control and lactate. In summary, these experiments indicate that pyruvate exposure prior to glucose deprivation significantly increased the energy buffering capacity of hippocampal slices, particularly by enhancing internal glycogen stores, delaying synaptic failure during glucose deprivation by maintaining ATP levels, and minimizing the decreasein the levels of NAD(P)H.

Prolonged hypoxia leads to irreversible loss of neuronal function and metabolic impairment of nicotinamide adenine dinucleotide recycling (between NAD+ and NADH) immediately after reoxygenation, resulting in NADH hyperoxidation. We test whether addition of nicotinamide (to enhance NAD+ levels) or PARP-1 inhibition (to prevent consumption of NAD+) can be effective in improving either loss of neuronal function or hyperoxidation following severe hypoxic injury in hippocampal slices. After severe, prolonged hypoxia (maintained for 3 min after spreading depression) there was hyperoxidation of NADH following reoxygenation, an increased soluble NAD+/NADH ratio, loss of neuronal field excitatory post-synaptic potential (fEPSP) and decreased ATP content. Nicotinamide incubation (5 mM) 2 hr prior to hypoxia significantly increased total NAD(H) content, improved neuronal recovery, enhanced ATP content, and prevented NADH hyperoxidation. The nicotinamide-induced increase in total soluble NAD(H) was more significant in the cytosolic compartment than within mitochondria. Prolonged incubation with PJ-34 (>1hr) led to enhanced baseline NADH fluorescence prior to hypoxia, as well as improved neuronal recovery, NADH hyperoxidation and ATP content on recovery from severe hypoxia and reoxygenation. In this acute model of severe neuronal dysfunction prolonged incubation with either nicotinamide or PJ-34 prior to hypoxia improved recovery of neuronal function, enhanced NADH reduction and ATP content, but neither treatment restored function when administered during or after prolonged hypoxia and reoxygenation.

Ca 2؉ /calmodulin (CaM)-dependent protein kinase II (CaMKII) plays a critical role in neuronal signal transduction and synaptic plasticity. Here, we showed that this kinase was very susceptible to oxidative modulation. Treatment of mouse brain synaptosomes with H 2 O 2 , diamide, and sodium nitroprusside caused aggregation of CaMKII through formation of disulfide and non-disulfide linkages, and partial inhibition of the kinase activity. These CaMKII aggregates were found to associate with the post synaptic density. However, treatment of purified CaMKII with these oxidants did not replicate those effects observed in the synaptosomes. Using two previously identified potential mediators of oxidants in the brain, glutathione disulfide S-monoxide (GS-DSMO) and glutathione disulfide S-dioxide (GS-DSDO), we showed that they oxidized and inhibited CaMKII in a manner partly related to those of the oxidanttreated synaptosomes as well as the ischemia-elicited oxidative stress in the acutely prepared hippocampal slices. Interestingly, the autophosphorylated and activated CaMKII was relatively refractory to GS-DSMO-and GS-DSDO-mediated aggregation. Short term ischemia (10 min) caused a depression of basal synaptic response of the hippocampal slices, and re-oxygenation (after 10 min) reversed the depression. However, oxidation of CaMKII remained at above the pre-ischemic level throughout the treatment. Oxidation of CaMKII also prevented full recovery of CaMKII autophosphorylation after re-oxygenation. Subsequently, the high frequency stimulation-mediated synaptic potentiation in the hippocampal CA1 region was significantly reduced compared with the control without ischemia. Thus, ischemia-evoked oxidation of CaMKII, probably via the action of glutathione disulfide S-oxides or their analogues, may be involved in the suppression of synaptic plasticity.Ca 2ϩ /calmodulin (CaM) 2 -dependent protein kinase II (CaMKII) is one of the major Ca 2ϩ -sensing enzymes important in transducing neuronal, hormonal, and electrical signals in brain, heart, and other tissues. In the central nervous system, CaMKII plays a pivotal role in the facilitation of synaptic plasticity, learning, and memory and in activity-dependent developmental processes (1). CaMKII holoenzyme is a dodecamer composed of two stacked hexameric rings, in which each catalytic/regulatory domain from the upper ring interacts with the equivalent catalytic/regulatory domain in the lower ring by an antiparallel coiled-coil, which resides in regulatory domains (2). Binding of Ca 2ϩ /CaM to the regulatory domain separates the dimer pair and causes the exposure of Thr-286 or Thr-287 (within ␣ or ␤ subunit) for phosphorylation by another catalytic domain in the same ring. This inter-subunit phosphorylation of Thr-286/287 converts the kinase into a high affinity binding protein for Ca 2ϩ /CaM, and the kinase becomes an activatorindependent autonomous enzyme (3). The autophosphorylation also leads to increased affinity of the kinase for several proteins near the sites of elevated Ca 2...

Calmodulin (CaM) and neurogranin (Ng) are two abundant neuronal proteins in the forebrain whose interactions are implicated in the enhancement of synaptic plasticity. To gain further insight into the actions of these two proteins we investigated whether they co-localize in principle neurons and whether they respond to high frequency stimulation in a coordinated fashion. Immunohistochemical staining of CaM and Ng in mouse hippocampal slices revealed that CaM was highly concentrated in the nucleus of CA1 pyramidal neurons, whereas Ng was more broadly localized throughout the soma and dendrites. The asymmetrical localization of CaM in the nucleus of pyramidal neurons was in sharp contrast to the distribution observed in pyramidal cells of the neighboring subiculum, where CaM was uniformly localized throughout the soma and dendrites. The somatic concentrations of CaM and Ng in CA1 pyramidal neurons were approximately tenand two-fold greater than observed in the dendrites, respectively. High frequency stimulation (HFS) of hippocampal slices promoted mobilization of CaM and Ng from soma to dendrites. These responses were spatially restricted to the area close to the site of stimulation and were inhibited by the N-methyl-D-asparate receptor antagonist 2-amino-5-phosphonopentanoic acid. Furthermore, HFS failed to promote translocation of CaM from soma to dendrites of slices from Ng knockout mice, which also exhibited deficits in HFS-induced long-term potentiation. Translocated CaM and Ng exhibited distinct puncta decorating the apical dendrites of pyramidal neurons and appeared to be concentrated in dendritic spines. These findings suggest that mobilization of CaM and Ng to stimulated dendritic spines may enhance synaptic efficacy by increasing and prolonging the Ca 2+ transients and activation of Ca 2+ /CaM-dependent enzymes. Keywordscalmodulin; neurogranin; hippocampus; translocation; dendritic spines; LTP Calmodulin (CaM) is a highly conserved protein known to participate in a broad range of cellular functions. In contrast, neurogranin (Ng) is expressed predominantly in the forebrain of vertebrates and is believed to be involved primarily in the regulation of neural function.Correspondence: Kuo-Ping Huang (huangk@mail.nih.gov) or Freesia L. Huang (fhuang@mail.nih.gov), Building 49, Room 6A36, NIH, Bethesda, MD 20892-4510, Phone: 301-496-7827; Fax: 301-496-7434. 1 Present address: Division of Neurosurgery, Department of Surgery, Duke University Medical Center, Durham, North Carolina, 27710Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. NIH Public Access Author Manuscri...

Previous reports have indicated that with aging, intrinsic brain tissue changes in cellular bioenergetics may hamper the brain’s ability to cope with metabolic stress. Therefore, we analyzed the effects of age on neuronal sensitivity to glucose deprivation by monitoring changes in field excitatory postsynaptic potentials (fEPSPs), tissue Po2, and NADH fluorescence imaging in the CA1 region of hippocampal slices obtained from F344 rats (1–2, 3–6, 12–20, and >22 months). Forty minutes of moderate low glucose (2.5 mM) led to approximately 80% decrease of fEPSP amplitudes and NADH decline in all 4 ages that reversed after reintroduction of 10 mM glucose. However, tissue slices from 12 to 20 months and >22-month-old rats were more vulnerable to low glucose: fEPSPs decreased by 50% on average 8 minutes faster compared with younger slices. Tissue oxygen utilization increased after onset of 2.5 mM glucose in all ages of tissue slices, which persisted for 40 minutes in younger tissue slices. But, in older tissue slices the increased oxygen utilization slowly faded and tissue Po2 levels increased toward baseline values after approximately 25 minutes of glucose deprivation. In addition, with age the ability to regenerate NADH after oxidation was diminished. The NAD+/NADH ratio remained relatively oxidized after low glucose, even during recovery. In young slices, glycogen levels were stable throughout the exposure to low glucose. In contrast, with aging utilization of glycogen stores was increased during low glucose, particularly in hippocampal slices from >22 months old rats, indicating both inefficient metabolism and increased demand for glucose. Lactate addition (20 mM) improved oxidative metabolism by directly supplementing the mitochondrial NADH pool and maintained fEPSPs in young as well as aged tissue slices, indicating that inefficient metabolism in the aging tissue can be improved by directly enhancing NADH regeneration.

Disulfide S-monoxide (DSMO) and disulfide S-dioxide (DSDO) have been proposed as proximal mediators for the oxidant-mediated modification of proteins. These disulfide S-oxides (DSOs) derived from glutathione (GSH) and captopril (CPSH) were synthesized by iron- or methyltrioxorhenium (VII)-catalyzed oxidation of the thiols with H2O2. Treatment of mouse hippocampal extracts with [35S]GS-DSOs revealed that a large number of proteins were susceptible to thionylation; however, only a limited number of the them were detectable by the commonly used antibody against GS-associated proteins. Using protein kinase C (PKC) as a model, we found that DSOs derived from different thiols modified this kinase with different efficacy and specificity; for example, the inhibitory potency of the kinase was glutathione disulfide S-dioxide (GS-DSDO) (IC50, approximately 30 microM) > captopril disulfide S-dioxide (CPS-DSDO) (IC50, approximately 450 microM) > glutathione disulfide S-monoxide (GS-DSMO) and captopril disulfide S-monoxide (CPS-DSMO). The stoichiometries of thionylation of PKC beta mediated by [35S]GS-DSMO and [35S]GS-DSDO were approximately 1 and 5 mol/mol, respectively, and at least four glutathionylation sites were identified in the GS-DSDO-treated kinase. Modification of PKC by GS-DSDO and CPS-DSDO rendered the kinase very susceptible to limited proteolysis; the former preferentially caused the degradation of the catalytic and the latter the regulatory domain of the kinase. Furthermore, CPS-DSDO-mediated modification of PKC increased the autonomous kinase activity; this was not the case for GS-DSDO-mediated modification. Since DSOs of different oxidative states as well as those derived from different thiols exert different effects on a target protein, these molecules could cause distinct cellular responses if derived from endogenous cellular reactions or even if they arise from exogenous sources.

The aim of the project is to develop a grip measurement and data logging product which can be used as an exercise ball for patients who have undergone orthopedic surgery and as a recovery quantification tool for the doctor treating patients with the same condition. It provides the doctor with a measure of how effective the surgery has been and how well the patient has been recovering. and allows the doctor to quantify certain each force. The device logs the pressure applied on the ball for a period of time as a part of the recovering exercise and also has an attachment to measure the grip pressure between adjacent fingers and the thumb and other fingertips. The doctor has an interface on the other side on a computer that shows the logged data and allows the device to be put in various different modes to measure the different forces. It stores the parameters after every examination. Texas Instruments India Educators' Conference978-0-7695-5146-3/13 $26.00