Glutamate is involved in cerebral ischemic injury, but its role has not been completely clarified and studies are required to understand how to minimize its detrimental effects, contemporarily boosting the positive ones. In fact, glutamate is not only a neurotransmitter, but primarily a key metabolite for brain bioenergetics. Thus, we investigated the relationships between glutamate and brain energy metabolism in an in vivo model of complete cerebral ischemia of 15 min and during post‐ischemic recovery after 1, 24, 48, 72, and 96 h in 1‐year‐old adult and 2‐year‐old aged rats. The maximum rates (V max) of glutamate dehydrogenase (GlDH ), glutamate‐oxaloacetate transaminase, and glutamate‐pyruvate transaminase were assayed in somatic mitochondria (FM ) and in intra‐synaptic ‘Light’ mitochondria and intra‐synaptic ‘Heavy’ mitochondria ones purified from cerebral cortex, distinguishing post‐ and pre‐synaptic compartments. During ischemia, none of the enzymes were modified in adult animals. In aged ones, glutamate‐oxaloacetate transaminase was increased in FM and GlDH in intra‐synaptic ‘Heavy’ mitochondria, stimulating glutamate catabolism. During post‐ischemic recovery, FM did not show modifications at both ages while, in intra‐synaptic mitochondria of adult animals, glutamate catabolism was increased after 1 h of recirculation and decreased after 48 and 72 h, whereas it remained decreased up to 96 h in aged rats. These results, with those previously published about Krebs’ cycle and Electron Transport Chain (Villa et al ., [2013] Neurochem. Int . 63, 765–781), demonstrate that: (i) V max of energy‐linked enzymes are different in the various cerebral mitochondria, which (ii) respond differently to ischemia and post‐ischemic recovery, also (iii) with respect to aging.
Cocaine is a recreational drug of abuse that binds to the dopamine transporter, preventing reuptake of dopamine into pre‐synaptic terminals. The increased presence of synaptic dopamine results in stimulation of both pre‐ and post‐synaptic dopamine receptors, considered an important mechanism by which cocaine elicits its reinforcing properties. However, the effects of acute cocaine administration on pre‐synaptic dopamine function remain unclear. Non‐invasive imaging techniques such as positron emission tomography have revealed impaired pre‐synaptic dopamine function in chronic cocaine users. Similar impairments have been seen in animal studies, with microdialysis experiments indicating decreased basal dopamine release. Here we use micro positron emission tomography imaging techniques in mice to measure dopamine synthesis capacity and determine the effect of acute cocaine administration of pre‐synaptic dopamine function. We show that a dose of 20 mg/kg cocaine is sufficient to elicit hyperlocomotor activity, peaking 15–20 min post treatment (p < 0.001). However, dopamine synthesis capacity in the striatum was not significantly altered by acute cocaine treatment (: 0.0097 per min vs. 0.0112 per min in vehicle controls, p > 0.05). Furthermore, expression levels of two key enzymes related to dopamine synthesis, tyrosine hydroxylase and aromatic l ‐amino acid decarboxylase, within the striatum of scanned mice were not significantly affected by acute cocaine pre‐treatment (p > 0.05). Our findings suggest that while the regulation of dopamine synthesis and release in the striatum have been shown to change with chronic cocaine use, leading to a reduced basal tone, these adaptations to pre‐synaptic dopaminergic neurons are not initiated following a single exposure to the drug.
Retinal ischemia and reperfusion injuries (R‐IRI) damage neuronal tissue permanently. Recently, we demonstrated that Argon exerts anti‐apoptotic and protective properties. The molecular mechanism remains unclear. We hypothesized that Argon inhalation exert neuroprotective effects in rats retinal ganglion cells (RGC) via an ERK‐1/2 dependent regulation of heat‐shock proteins. Inhalation of Argon (75 Vol%) was performed after R‐IRI on the rats′ left eyes for 1 h immediately or with delay. Retinal tissue was harvested after 24 h to analyze mRNA and protein expression of heat‐shock proteins ?70, ?90 and heme‐oxygenase‐1, mitogen‐activated protein kinases (p38, JNK, ERK‐1/2) and histological changes. To analyze ERK dependent effects, the ERK inhibitor PD98059 was applicated prior to Argon inhalation. RGC count was analyzed 7 days after injury. Statistics were performed using anova . Argon significantly reduced the R‐IRI‐affected heat‐shock protein expression (p < 0.05). While Argon significantly induced ERK‐1/2 expression (p < 0.001), inhibition of ERK‐1/2 before Argon inhalation resulted in significantly lower vital RGCs (p < 0.01) and increase in heme‐oxygenase‐1 (p < 0.05). R‐IRI‐induced RGC loss was reduced by Argon inhalation (p < 0.001). Immunohistochemistry suggested ERK‐1/2 activation in Müller cells. We conclude, that Argon treatment protects R‐IRI‐induced apoptotic loss of RGC via an ERK‐1/2 dependent regulation of heme‐oxygenase‐1.
This editorial highlights a study by Rodriguez, Sanchez‐Moran et al. (2019) in the current issue of the Journal of Neurochemistry, in which the authors describe a microcephalic boy carrying the novel heterozygous de novo missense mutation c.560A> G; p.Asp187Gly in Cdh1/Fzr1 encoding the APC/C E3‐ubiquitin ligase cofactor CDH1. A functional characterization of mutant APC/CCDH1 confirms an aberrant division of neural progenitor cells, a condition known to determine the mouse brain cortex size. These data suggest that APC/CCDH1 may contribute to the regulation of the human brain size.
CDP‐choline has shown neuroprotective effects in cerebral ischemia. In humans, although a recent trial International Citicoline Trial on Acute Stroke (ICTUS) has shown that global recovery is similar in CDP‐choline and placebo groups, CDP‐choline was shown to be more beneficial in some patients, such as those with moderate stroke severity and not treated with t‐PA. Several mechanisms have been proposed to explain the beneficial actions of CDP‐choline. We have now studied the participation of Sirtuin1 (SIRT1) in the neuroprotective actions of CDP‐choline. Fischer rats and Sirt1?/? mice were subjected to permanent focal ischemia. CDP‐choline (0.2 or 2 g/kg), sirtinol (a SIRT1 inhibitor; 10 mg/kg), and resveratrol (a SIRT1 activator; 2.5 mg/kg) were administered intraperitoneally. Brains were removed 24 and 48 h after ischemia for western blot analysis and infarct volume determination. Treatment with CDP‐choline increased SIRT1 protein levels in brain concomitantly to neuroprotection. Treatment with sirtinol blocked the reduction in infarct volume caused by CDP‐choline, whereas resveratrol elicited a strong synergistic neuroprotective effect with CDP‐choline. CDP‐choline failed to reduce infarct volume in Sirt1?/? mice. Our present results demonstrate a robust effect of CDP‐choline like SIRT1 activator by up‐regulating its expression. Our findings suggest that therapeutic strategies to activate SIRT1 may be useful in the treatment of stroke.
Parkinson's disease (PD) is a chronic neurodegenerative disorder characterized by the loss of nigrostriatal dopaminergic neurons and consequent motor dysfunction. Zonisamide (1,2‐benzisoxazole‐3‐methanesulfonamide), which was originally developed as an antiepileptic drug, has been found to have therapeutic benefits for PD. However, the pharmacological mechanisms behind the beneficial actions of zonisamide in PD are not fully understood. Here, we investigated the neuroprotective effects of zonisamide on nigrostriatal dopaminergic neurons of the Engrailed mutant mouse, a genetic model of PD. Chronic administration of zonisamide in Engrailed mutant mice was shown to improve the survival of nigrostriatal dopaminergic neurons compared with that under saline treatment. In addition, dopaminergic terminals in the striatum and the motor function were improved in zonisamide‐treated Engrailed mutant mice to the levels of those in control mice. To clarify the mechanism behind the neuroprotective effects of zonisamide, the contents of neurotrophic factors were determined after chronic administration of zonisamide. Brain‐derived neurotrophic factor content was increased in the striatum and ventral midbrain of the zonisamide‐treated mice compared to saline‐treated mice. These findings imply that zonisamide reduces nigrostriatal dopaminergic cell death through brain‐derived neurotrophic factor signaling and may have similar beneficial effects in human parkinsonian patients as well.
Kiss1, a neuropeptide predominantly expressed in the habenula, modulates the serotonin (5‐HT) system to decrease odorant cue [alarm substance (AS)]‐evoked fear behaviour in the zebrafish. The purpose of this study was to assess the interaction of Kiss1 with the 5‐HT system as well as to determine the involvement of the 5‐HT receptor subtypes in AS‐evoked fear. We utilized 0. 28 mg/kg WAY 100635 (WAY), a selective 5‐HT1A receptor antagonist, to observe the effects of Kiss1 administration on AS‐evoked fear. We found WAY significantly inhibited the anxiolytic effects of Kiss1 (p <0.001) with an exception of freezing behaviour. Based on this, we utilized 92.79 mg/kg methysergide, a 5‐HT1 and 5‐HT2 receptor antagonist, and found that methysergide significantly blocked the anxiolytic effects of Kiss1 in the presence of the AS (p <0.001). From this, we conclude that Kiss1 modulates AS‐evoked fear responses mediated by the 5‐HT1A and 5‐HT2 receptors.
Expression of a familial Alzheimer's disease (AD)‐linked mutant of amyloid β precursor protein (APP) or the binding of transforming growth factor β2 to wild‐type (wt)‐APP causes neuronal death by activating an intracellular death signal (a APP‐mediated intracellular death signal) in the absence of the involvement of amyloid β (Aβ) toxicity in vitro. These neuronal death models may therefore be regarded as Aβ‐independent neuronal death models related to AD. A recent study has shown that the A673T mutation in the APP isoform APP770, corresponding to the A598T mutation in the most prevalent neuronal APP isoform APP695 (an AD‐protective mutant of APP), is linked to a reduction in the incidence rate of AD. Consistent with this, cells expressing the AD‐protective mutant of APP produce less Aβ than cells expressing wt‐APP. In this study, transforming growth factor β2 caused death in cultured neuronal cells expressing wt‐APP, but not in those expressing the AD‐protective mutant of APP. This result suggests that the AD‐protective mutation of APP reduces the incidence rate of AD by attenuating the APP‐mediated intracellular death signal. In addition, a mutation that causes hereditary cerebral hemorrhage with amyloidosis‐Dutch type also attenuated the APP‐mediated intracellular death signal.
Molecular imaging of vesicular acetylcholine transporter (VACh T) in the brain provides an important cholinergic biomarker for the pathophysiology and treatment of dementias including Alzheimer's disease. In this study, kinetics modeling methods were applied and compared for quantifying regional brain uptake of the VACh T‐specific positron emission tomography radiotracer, ((?)‐(1‐(‐8‐(2‐fluoroethoxy)‐3‐hydroxy‐1,2,3,4‐tetrahydronaphthalen‐2‐yl)piperidin‐4‐yl)(4‐fluorophenyl)‐methanone) ([18F]VAT ) in macaques. Total volume distribution (V T ) estimates were compared for one‐tissue compartment model (1TCM ), two‐tissue compartment model (2TCM ), Logan graphic analysis (LoganAIF ) and multiple linear analysis (MA 1) with arterial blood input function using data from three macaques. Using the cerebellum‐hemispheres as the reference region with data from seven macaques, three additional models were compared: reference tissue model (RTM ), simplified RTM (SRTM ), and Logan graphic analysis (LoganREF ). Model selection criterion indicated that a) 2TCM and SRTM were the most appropriate kinetics models for [18F]VAT ; and b) SRTM was strongly correlated with 2TCM (Pearson's coefficients r > 0.93, p < 0.05). Test–retest studies demonstrated that [18F]VAT has good reproducibility and reliability (TRV < 10%, ICC > 0.72). These studies demonstrate [18F]VAT is a promising VACh T positron emission tomography tracer for quantitative assessment of VACh T levels in the brain of living subjects.
Autonomic control of heart rate is mediated by cardioinhibitory parasympathetic cholinergic neurons located in the brainstem and stimulatory sympathetic noradrenergic neurons. During embryonic development the survival and cholinergic phenotype of brainstem autonomic neurons is promoted by brain‐derived neurotrophic factor (BDNF). We now provide evidence that BDNF regulates heart rate by a mechanism involving increased brainstem cardioinhibitory parasympathetic activity. Mice with a BDNF haploinsufficiency exhibit elevated resting heart rate, and infusion of BDNF intracerebroventricularly reduces heart rate in both wild‐type and BDNF+/? mice. The atropine‐induced elevation of heart rate is diminished in BDNF+/? mice and is restored by BDNF infusion, whereas the atenolol‐induced decrease in heart rate is unaffected by BDNF levels, suggesting that BDNF signaling enhances parasympathetic tone which is diminished with BDNF haploinsufficiency. Whole‐cell recordings from pre‐motor cholinergic cardioinhibitory vagal neurons in the nucleus ambiguus indicate that BDNF haploinsufficiency reduces cardioinhibitory vagal neuron activity by increased inhibitory GABAergic and diminished excitatory glutamatergic neurotransmission to these neurons. Our findings reveal a previously unknown role for BDNF in the control of heart rate by a mechanism involving increased activation of brainstem cholinergic parasympathetic neurons
Dopaminergic neurotransmission in the nucleus accumbens is important for various reward‐related cognitive processes including reinforcement learning. Repeated cocaine enhances hippocampal synaptic plasticity, and phasic elevations of accumbal dopamine evoked by unconditioned stimuli are dependent on impulse flow from the ventral hippocampus. Therefore, sensitized hippocampal activity may be one mechanism by which drugs of abuse enhance limbic dopaminergic activity. In this study, in vivo microdialysis in freely moving adult male Sprague–Dawley rats was used to investigate the effect of repeated cocaine on ventral hippocampus‐mediated dopaminergic transmission within the medial shell of the nucleus accumbens. Following seven daily injections of saline or cocaine (20 mg/kg, ip), unilateral infusion of N‐methyl‐d ‐aspartate (NMDA, 0.5 μg) into the ventral hippocampus transiently increased both motoric activity and ipsilateral dopamine efflux in the medial shell of the nucleus accumbens, and this effect was greater in rats that received repeated cocaine compared to controls that received repeated saline. In addition, repeated cocaine altered NMDA receptor subunit expression in the ventral hippocampus, reducing the NR2A : NR2B subunit ratio. Together, these results suggest that repeated exposure to cocaine produces maladaptive ventral hippocampal‐nucleus accumbens communication, in part through changes in glutamate receptor composition.
Parkinson's disease (PD) is a common neurodegenerative disease, but its pathogenesis remains elusive. A mutation in ubiquitin C‐terminal hydrolase L1 (UCH‐L1) is responsible for a form of genetic PD which strongly resembles the idiopathic PD. We previously showed that 1‐(3′,4′‐dihydroxybenzyl)‐1,2,3,4‐tetrahydroisoquinoline (3′,4′DHBnTIQ) is an endogenous parkinsonism‐inducing dopamine derivative. Here, we investigated the interaction between 3′,4′DHBnTIQ and UCH‐L1 and its possible role in the pathogenesis of idiopathic PD. Our results indicate that 3′,4′DHBnTIQ binds to UCH‐L1 specifically at Cys152 in vitro. In addition, 3′,4′DHBnTIQ treatment increased the amount of UCH‐L1 in the insoluble fraction of SH‐SY5Y cells and inhibited its hydrolase activity to 60%, reducing the level of ubiquitin in the soluble fraction of SH‐SY5Y cells. Catechol‐modified UCH‐L1 as well as insoluble UCH‐L1 were detected in the midbrain of 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine‐treated PD model mice. Structurally as well as functionally altered UCH‐L1 have been detected in the brains of patients with idiopathic PD. We suggest that conjugation of UCH‐L1 by neurotoxic endogenous compounds such as 3′,4′DHBnTIQ might play a key role in onset and progression of idiopathic PD.
The neurotransmitter serotonin underlies many of the brain's functions. Understanding serotonin neurochemistry is important for improving treatments for neuropsychiatric disorders such as depression. Antidepressants commonly target serotonin clearance via serotonin transporters and have variable clinical effects. Adjunctive therapies, targeting other systems including serotonin autoreceptors, also vary clinically and carry adverse consequences. Fast scan cyclic voltammetry is particularly well suited for studying antidepressant effects on serotonin clearance and autoreceptors by providing real‐time chemical information on serotonin kinetics in vivo. However, the complex nature of in vivo serotonin responses makes it difficult to interpret experimental data with established kinetic models. Here, we electrically stimulated the mouse medial forebrain bundle to provoke and detect terminal serotonin in the substantia nigra reticulata. In response to medial forebrain bundle stimulation we found three dynamically distinct serotonin signals. To interpret these signals we developed a computational model that supports two independent serotonin reuptake mechanisms (high affinity, low efficiency reuptake mechanism, and low affinity, high efficiency reuptake system) and bolsters an important inhibitory role for the serotonin autoreceptors. Our data and analysis, afforded by the powerful combination of voltammetric and theoretical methods, gives new understanding of the chemical heterogeneity of serotonin dynamics in the brain. This diverse serotonergic matrix likely contributes to clinical variability of antidepressants.
The E3 ubiquitin ligase Parkin plays a central role in the pathogenesis of many neurodegenerative diseases. Parkin promotes specific ubiquitination and affects the localization of transactivation response DNA‐binding protein 43 (TDP‐43), which controls the translation of thousands of mRNAs. Here we tested the effects of lentiviral Parkin and TDP‐43 expression on amino acid metabolism in the rat motor cortex using high frequency 13C NMR spectroscopy. TDP‐43 expression increased glutamate levels, decreased the levels of other amino acids, including glutamine, aspartate, leucine and isoleucine, and impaired mitochondrial tricarboxylic acid cycle. TDP‐43 induced lactate accumulation and altered the balance between excitatory (glutamate) and inhibitory (GABA) neurotransmitters. Parkin restored amino acid levels, neurotransmitter balance and tricarboxylic acid cycle metabolism, rescuing neurons from TDP‐43‐induced apoptotic death. Furthermore, TDP‐43 expression led to an increase in 4E‐BP levels, perhaps altering translational control and deregulating amino acid synthesis; while Parkin reversed the effects of TDP‐43 on the 4E‐BP signaling pathway. Taken together, these data suggest that Parkin may affect TDP‐43 localization and mitigate its effects on 4E‐BP signaling and loss of amino acid homeostasis.
The cadherin epidermal growth factor (EGF) laminin G (LAG) seven‐pass G‐type receptors (CELSRs) are a special subgroup of adhesion G protein‐coupled receptors, which are pivotal regulators of many biologic processes such as neuronal/endocrine cell differentiation, vessel valve formation, and the control of planar cell polarity during embryonic development. All three members of the CELSR family (CELSR1‐3) have large ecto‐domains that form homophilic interactions and encompass more than 2000 amino acids. Mutations in the ecto‐domain or other gene locations of CELSRs are associated with neural tube defects and other diseases in humans. Celsr knockout (KO) animals have many developmental defects. Therefore, specific agonists or antagonists of CELSR members may have therapeutic potential. Although significant progress has been made regarding the functions and biochemical properties of CELSRs, our knowledge of these receptors is still lacking, especially considering that they are broadly distributed but have few characterized functions in a limited number of tissues. The dynamic activation and inactivation of CELSRs and the presence of endogenous ligands beyond homophilic interactions remain elusive, as do the regulatory mechanisms and downstream signaling of these receptors. Given this motivation, future studies with more advanced cell biology or biochemical tools, such as conditional KO mice, may provide further insights into the mechanisms underlying CELSR function, laying the foundation for the design of new CELSR‐targeted therapeutic reagents.
Hereditary neuropathies comprise a wide variety of chronic diseases associated to more than 80 genes identified to date. We herein examined 612 index patients with either a Charcot‐Marie‐Tooth phenotype, hereditary sensory neuropathy, familial amyloid neuropathy, or small fiber neuropathy using a customized multigene panel based on the next generation sequencing technique. In 121 cases (19.8%), we identified at least one putative pathogenic mutation. Of these, 54.4% showed an autosomal dominant, 33.9% an autosomal recessive, and 11.6% an X‐linked inheritance. The most frequently affected genes were PMP22 (16.4%), GJB1 (10.7%), MPZ, and SH3TC2 (both 9.9%), and MFN2 (8.3%). We further detected likely or known pathogenic variants in HINT1, HSPB1, NEFL, PRX, IGHMBP2, NDRG1, TTR, EGR2, FIG4, GDAP1, LMNA, LRSAM1, POLG, TRPV4, AARS, BIC2, DHTKD1, FGD4, HK1, INF2, KIF5A, PDK3, REEP1, SBF1, SBF2, SCN9A, and SPTLC2 with a declining frequency. Thirty‐four novel variants were considered likely pathogenic not having previously been described in association with any disorder in the literature. In one patient, two homozygous mutations in HK1 were detected in the multigene panel, but not by whole exome sequencing. A novel missense mutation in KIF5A was considered pathogenic because of the highly compatible phenotype. In one patient, the plasma sphingolipid profile could functionally prove the pathogenicity of a mutation in SPTLC2. One pathogenic mutation in MPZ was identified after being previously missed by Sanger sequencing. We conclude that panel based next generation sequencing is a useful, time‐ and cost‐effective approach to assist clinicians in identifying the correct diagnosis and enable causative treatment considerations.
Although bevacizumab initially shows high response rates in gliomas and other tumours, therapy resistance usually develops later. Because anti‐angiogenic agents are supposed to induce hypoxia, we asked whether rendering glioma cells independent of oxidative phosphorylation modulates their sensitivity against hypoxia and bevacizumab. LNT‐229 glioma cells without functional mitochondria (rho0) and control (rho+) cells were generated. LNT‐229 rho0‐cells displayed reduced expression of oxidative phosphorylation‐related genes and diminished oxygen consumption. Conversely, glycolysis was up‐regulated in these cells, as shown by increased lactate production and stronger expression of glucose transporter‐1 and lactate dehydrogenase‐A. However, hypoxia‐induced cell death in vitro was nearly completely abolished in the LNT‐229 rho0‐cells, these cells were more sensitive towards glucose restriction and the treatment with the glycolysis inhibitor 2‐deoxy‐D‐glucose. In an orthotopic mouse xenograft experiment, bevacizumab induced hypoxia as reflected by elevated Hypoxia‐inducible factor 1‐alpha staining in both, rho+‐ and rho0‐tumours. However, it prolonged survival only in the mice bearing rho+‐tumours (74 days vs. 105 days, p = 0.024 log‐rank test) and had no effect on survival in mice carrying LNT‐229 rho0‐tumours (75 days vs. 70 days, p = 0.52 log‐rank test). Interestingly, inhibition of glycolysis in vivo with 2‐deoxy‐D‐glucose re‐established sensitivity of rho0‐tumours against bevacizumab (98 days vs. 80 days, p = 0.0001). In summary, ablation of oxidative phosphorylation in glioma cells leads to a more glycolytic and hypoxia‐resistant phenotype and is sufficient to induce bevacizumab‐refractory tumours. These results add to increasing evidence that a switch towards glycolysis is one mechanism how tumour cells may evade anti‐angiogenic treatments and suggest anti‐glycolytic strategies as promising approaches to overcome bevacizumab resistance.
Fructose reacts spontaneously with proteins in the brain to form advanced glycation end products (AGE) that may elicit neuroinflammation and cause brain pathology, including Alzheimer's disease. We investigated whether fructose is eliminated by oxidative metabolism in neocortex. Injection of [14C]fructose or its AGE‐prone metabolite [14C]glyceraldehyde into rat neocortex in vivo led to formation of 14C‐labeled alanine, glutamate, aspartate, GABA, and glutamine. In isolated neocortical nerve terminals, [14C]fructose‐labeled glutamate, GABA, and aspartate, indicating uptake of fructose into nerve terminals and oxidative fructose metabolism in these structures. This was supported by high expression of hexokinase 1, which channels fructose into glycolysis, and whose activity was similar with fructose or glucose as substrates. By contrast, the fructose‐specific ketohexokinase was weakly expressed. The fructose transporter Glut5 was expressed at only 4% of the level of neuronal glucose transporter Glut3, suggesting transport across plasma membranes of brain cells as the limiting factor in removal of extracellular fructose. The genes encoding aldose reductase and sorbitol dehydrogenase, enzymes of the polyol pathway that forms glucose from fructose, were expressed in rat neocortex. These results point to fructose being transported into neocortical cells, including nerve terminals, and that it is metabolized and thereby detoxified primarily through hexokinase activity.
下载免费PDF全文