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1.
bioRxiv ; 2024 Oct 12.
Artigo em Inglês | MEDLINE | ID: mdl-39416088

RESUMO

Cocaine use disorder is an intersecting issue in populations with HIV-1, further exacerbating the clinical course of the disease, contributing to neurotoxicity and neuroinflammation. Cocaine and HIV neurotoxins play roles in neuronal damage during neuroHIV progression by disrupting glutamate homeostasis in the brain. Even with cART, HIV-1 Nef, an early viral protein expressed in approximately 1% of infected astrocytes, remains a key neurotoxin. This study investigates the relationship that exists between Nef, glutamate homeostasis, and cocaine in the NAc, a critical brain region associated with drug motivation and reward. Using a rat model, we compared the effects of astrocytic Nef and cocaine by molecular analysis of glutamate transporters in the NAc. We further conducted behavioral assessments for cocaine self-administration to evaluate cocaine-seeking behavior. Our findings indicate that both cocaine and Nef independently decrease the expression of the glutamate transporter GLT-1 in the NAc. Additionally, rats with astrocytic Nef expression exhibited increased cocaine-seeking behavior but demonstrated sex dependent molecular differences after behavioral paradigm. In conclusion, our results suggest the expression of Nef intensifies cocaine-induced alterations in glutamate homeostasis in the NAc, potentially underlying increased cocaine-seeking. Understanding these interactions better may inform therapeutic strategies for managing cocaine use disorder in HIV-infected individuals.

2.
Acta Neuropathol Commun ; 12(1): 166, 2024 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-39434170

RESUMO

The recently discovered interaction between presenilin 1 (PS1), a subunit of γ-secretase involved in amyloid-ß (Aß) peptide production, and GLT-1, the major brain glutamate transporter (EAAT2 in the human), may link two pathological aspects of Alzheimer's disease: abnormal Aß occurrence and neuronal network hyperactivity. In the current study, we employed a FRET-based fluorescence lifetime imaging microscopy (FLIM) to characterize the PS1/GLT-1 interaction in brain tissue from sporadic AD (sAD) patients. sAD brains showed significantly less PS1/GLT-1 interaction than those with frontotemporal lobar degeneration or non-demented controls. Familial AD (fAD) PS1 mutations, inducing a "closed" PS1 conformation similar to that in sAD brain, and gamma-secretase modulators (GSMs), inducing a "relaxed" conformation, respectively reduced and increased the interaction. Furthermore, PS1 influences GLT-1 cell surface expression and homomultimer formation, acting as a chaperone but not affecting GLT-1 stability. The diminished PS1/GLT-1 interaction suggests that these functions may not work properly in AD.


Assuntos
Doença de Alzheimer , Secretases da Proteína Precursora do Amiloide , Encéfalo , Transportador 2 de Aminoácido Excitatório , Presenilina-1 , Humanos , Doença de Alzheimer/metabolismo , Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Transportador 2 de Aminoácido Excitatório/metabolismo , Transportador 2 de Aminoácido Excitatório/genética , Presenilina-1/genética , Presenilina-1/metabolismo , Secretases da Proteína Precursora do Amiloide/metabolismo , Secretases da Proteína Precursora do Amiloide/genética , Masculino , Idoso , Feminino , Encéfalo/metabolismo , Encéfalo/patologia , Idoso de 80 Anos ou mais , Chaperonas Moleculares/metabolismo , Chaperonas Moleculares/genética , Transferência Ressonante de Energia de Fluorescência , Pessoa de Meia-Idade , Mutação , Células HEK293
3.
Cells ; 13(19)2024 Sep 24.
Artigo em Inglês | MEDLINE | ID: mdl-39404364

RESUMO

Glutamate transporter-1 (GLT-1) dynamics are implicated in excitotoxicity and Alzheimer's disease (AD) progression. Early stages of AD are often marked by hyperactivity and increased epileptiform activity preceding cognitive decline. Previously, we identified a direct interaction between GLT-1 and Presenilin 1 (PS1) in the brain, highlighting GLT-1 as a promising target in AD research. This study reports the significance of this interaction and uncovers a novel role of GLT-1 in modulating amyloid-beta (Aß) production. Overexpression of GLT-1 in cells reduces the levels of Aß40 and Aß42 by decreasing γ-secretase activity pertinent to APP processing and induces a more "open" PS1 conformation, resulting in decreased Aß42/40 ratio. Inhibition of the GLT-1/PS1 interaction using cell-permeable peptides produced an opposing effect on Aß, highlighting the pivotal role of this interaction in regulating Aß levels. These findings emphasize the potential of targeting the GLT-1/PS1 interaction as a novel therapeutic strategy for AD.


Assuntos
Peptídeos beta-Amiloides , Transportador 2 de Aminoácido Excitatório , Peptídeos beta-Amiloides/metabolismo , Humanos , Transportador 2 de Aminoácido Excitatório/metabolismo , Presenilina-1/metabolismo , Presenilina-1/genética , Secretases da Proteína Precursora do Amiloide/metabolismo , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Animais , Fragmentos de Peptídeos/metabolismo , Células HEK293
4.
CNS Neurosci Ther ; 30(9): e70024, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-39218798

RESUMO

AIMS: Type 2 diabetes mellitus (T2DM) is related to an increased risk of postoperative cognitive dysfunction (POCD), which may be caused by neuronal hyperexcitability. Astrocyte glutamate transporter 1 (GLT-1) plays a crucial role in regulating neuron excitability. We investigated if T2DM would magnify the increased neuronal excitability induced by anesthesia/surgery (A/S) and lead to POCD in young adult mice, and if so, determined whether these effects were associated with GLT-1 expression. METHODS: T2DM model was induced by high fat diet (HFD) and injecting STZ. Then, we evaluated the spatial learning and memory of T2DM mice after A/S with the novel object recognition test (NORT) and object location test (OLT). Western blotting and immunofluorescence were used to analyze the expression levels of GLT-1 and neuronal excitability. Oxidative stress reaction and neuronal apoptosis were detected with SOD2 expression, MMP level, and Tunel staining. Hippocampal functional synaptic plasticity was assessed with long-term potentiation (LTP). In the intervention study, we overexpressed hippocampal astrocyte GLT-1 in GFAP-Cre mice. Besides, AAV-Camkllα-hM4Di-mCherry was injected to inhibit neuronal hyperexcitability in CA1 region. RESULTS: Our study found T2DM but not A/S reduced GLT-1 expression in hippocampal astrocytes. Interestingly, GLT-1 deficiency alone couldn't lead to cognitive decline, but the downregulation of GLT-1 in T2DM mice obviously enhanced increased hippocampal glutamatergic neuron excitability induced by A/S. The hyperexcitability caused neuronal apoptosis and cognitive impairment. Overexpression of GLT-1 rescued postoperative cognitive dysfunction, glutamatergic neuron hyperexcitability, oxidative stress reaction, and apoptosis in hippocampus. Moreover, chemogenetic inhibition of hippocampal glutamatergic neurons reduced oxidative stress and apoptosis and alleviated postoperative cognitive dysfunction. CONCLUSIONS: These findings suggest that the adult mice with type 2 diabetes are at an increased risk of developing POCD, perhaps due to the downregulation of GLT-1 in hippocampal astrocytes, which enhances increased glutamatergic neuron excitability induced by A/S and leads to oxidative stress reaction, and neuronal apoptosis.


Assuntos
Astrócitos , Diabetes Mellitus Tipo 2 , Regulação para Baixo , Transportador 2 de Aminoácido Excitatório , Hipocampo , Camundongos Endogâmicos C57BL , Complicações Cognitivas Pós-Operatórias , Animais , Transportador 2 de Aminoácido Excitatório/metabolismo , Transportador 2 de Aminoácido Excitatório/biossíntese , Transportador 2 de Aminoácido Excitatório/genética , Astrócitos/metabolismo , Complicações Cognitivas Pós-Operatórias/etiologia , Complicações Cognitivas Pós-Operatórias/metabolismo , Hipocampo/metabolismo , Camundongos , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/metabolismo , Masculino , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/metabolismo , Dieta Hiperlipídica/efeitos adversos , Camundongos Transgênicos
5.
Eur J Pharmacol ; 984: 177017, 2024 Sep 28.
Artigo em Inglês | MEDLINE | ID: mdl-39349114

RESUMO

The glutamatergic system, located throughout the brain including the prefrontal cortex and nucleus accumbens, plays a critical role in reward and reinforcement processing, and mediates the psychotropic effects of addictive drugs such as cocaine. Glutamate transporters, including EAAT2/GLT-1, are responsible for removing glutamate from the synaptic cleft. Reduced expression of GLT-1 following chronic cocaine use and abstinence has been reported. Here, we demonstrate that targeting GLT-1 with a novel positive allosteric modulator (PAM), NA-014, results in reduction of cocaine-associated behaviors in rats. Pharmacokinetic analysis demonstrated that NA-014 is brain-penetrant and suitable for in vivo studies.We found that 15 and 30 mg/kg NA-014 significantly reduced cocaine-induced locomotion in males. Only the 15 mg/kg dose was effective in females and 60 mg/kg was ineffective in both sexes. Furthermore, 30 and 60 mg/kg NA-014 reduced expression of cocaine conditioned place preference (CPP) in males. 30 mg/kg NA-014 reduced expression of cocaine CPP in females and 15 mg/kg did not affect cocaine CPP in either sex, suggesting GLT-1 influences cocaine-associated behaviors in a sex-dependent manner. NA-014 did not elicit rewarding behavior, nor alter baseline locomotion. Twice daily/7-day administration of 100 mg/kg of NA-014 did not alter GLT-1 or GLAST expression in either sex in the prefrontal cortex (PFC). Collectively, these studies demonstrated that NA-014 reduced the locomotor stimulant and rewarding effects of cocaine in male and female rats. In the context of psychostimulant use disorders, our study suggests studying GLT-1 PAMs as alternatives to ß-lactam compounds that increase GLT-1 protein levels.

6.
Horm Behav ; 165: 105618, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-39180889

RESUMO

Infusion of 17ß-estradiol (E2) into the dorsal hippocampus (DH) of ovariectomized (OVX) mice enhances memory consolidation, an effect that depends on rapid phosphorylation of extracellular signal-regulated kinase (ERK) and Akt. Astrocytic glutamate transporter 1 (GLT-1) modulates neurotransmission via glutamate uptake from the synaptic cleft. However, little is known about the contribution of DH astrocytes, and astrocytic glutamate transport, to the memory-enhancing effects of E2. This study was designed to test whether DH astrocytes contribute to estrogenic modulation of memory consolidation by determining the extent to which DH GLT-1 is necessary for E2 to enhance memory in object recognition and object placement tasks and trigger rapid phosphorylation events in DH astrocytes. OVX female mice were bilaterally cannulated into the DH or the DH and dorsal third ventricle (ICV). Post-training DH infusion of the GLT-1 inhibitor dihydrokainic acid (DHK) dose-dependently impaired memory consolidation in both tasks. Moreover, the memory-enhancing effects of ICV-infused E2 in each task were blocked by DH DHK infusion. E2 increased p42 ERK and Akt phosphorylation in DH astrocytes, and these effects were blocked by DHK. Results suggest the necessity of DH GLT-1 activity for object and spatial memory consolidation, and for E2 to enhance consolidation of these memories and to rapidly activate cell signaling in DH astrocytes. Findings indicate that astrocytic function in the DH of OVX females is necessary for memory formation and is regulated by E2, and suggest an essential role for DH astrocytic GLT-1 activity in the memory-enhancing effects of E2.


Assuntos
Astrócitos , Estradiol , Transportador 2 de Aminoácido Excitatório , Ácido Glutâmico , Hipocampo , Ovariectomia , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Feminino , Estradiol/farmacologia , Camundongos , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Ácido Glutâmico/metabolismo , Transportador 2 de Aminoácido Excitatório/metabolismo , Consolidação da Memória/efeitos dos fármacos , Consolidação da Memória/fisiologia , Fosforilação/efeitos dos fármacos , Memória/efeitos dos fármacos , Memória/fisiologia , Camundongos Endogâmicos C57BL , Reconhecimento Psicológico/efeitos dos fármacos , Reconhecimento Psicológico/fisiologia , Ácido Caínico/análogos & derivados
7.
Biomedicines ; 12(8)2024 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-39200176

RESUMO

Background: Ceftriaxone upregulates GLT1 glutamate transporter in the brain and may have anti-CFC and anti-OCD effects. Methods: Twenty WZ-5HT rats were used to investigate the effects of ceftriaxone on obsessive-compulsive (OCD)-like behaviour in the marble-burying (MB) test, freezing behaviour in contextual fear conditioning (CFC) and expression of GLT1 protein in the hippocampus or amygdala using immunoblots. Fifteen DBA/2J mice were used in the MB test. We also compared diazepam with ceftriaxone in open-field, beam-walking, and wire-hanging tests on 47 DBA/2J mice. Ceftriaxone (200 mg/kg) and saline were applied intraperitoneally, once daily for 7 (rats) or 5 (mice) consecutive days. A single dose of diazepam (1.5-3.0 mg/kg) or saline was injected 30 min before the behavioural tests. Results: Ceftriaxone significantly diminished OCD-like behaviour (↓ number of marbles buried) and freezing behaviour in CFC context session (↑ latencies, ↓ total duration, ↓ duration over four 2 min periods of the session) but increased GLT1 protein expression in the amygdala and hippocampus of rats. Diazepam induced sedation, ataxia and myorelaxation in mice. Ceftriaxone did not have these side effects. Conclusions: The results of this study confirm the anti-CFC and anti-OCD effects of ceftriaxone, which did not produce the unwanted effects typical of diazepam.

8.
Toxics ; 12(8)2024 Aug 19.
Artigo em Inglês | MEDLINE | ID: mdl-39195706

RESUMO

Opioid-related deaths are attributed to overdoses, and fentanyl overdose has been on the rise in many parts of the world, including the USA. Glutamate transporter 1 (GLT-1) has been identified as a therapeutic target in several preclinical models of substance use disorders, and ß-lactams effectively enhance its expression and function. In the current study, we characterized the metabolomic profile of the nucleus accumbens (NAc) in fentanyl-overdose mouse models, and we evaluated the protective effects of the functional enhancement of GLT-1 using ß-lactams, ceftriaxone, and MC-100093. BALB/c mice were divided into four groups: control, fentanyl, fentanyl/ceftriaxone, and fentanyl/MC-100093. While the control group was intraperitoneally (i.p.) injected with normal saline simultaneously with other groups, all fentanyl groups were i.p. injected with 1 mg/kg of fentanyl as an overdose after habituation with four repetitive non-consecutive moderate doses (0.05 mg/kg) of fentanyl for a period of seven days. MC-100093 (50 mg/kg) and ceftriaxone (200 mg/kg) were i.p. injected from days 5 to 9. Gas chromatography-mass spectrometry (GC-MS) was used for metabolomics, and Western blotting was performed to determine the expression of target proteins. Y-maze spontaneous alternation performance and the open field activity monitoring system were used to measure behavioral manifestations. Fentanyl overdose altered the abundance of about 30 metabolites, reduced the expression of GLT-1, and induced the expression of inflammatory mediators IL-6 and TLR-4 in the NAc. MC-100093 and ceftriaxone attenuated the effects of fentanyl-induced downregulation of GLT-1 and upregulation of IL-6; however, only ceftriaxone attenuated fentanyl-induced upregulation of TRL4 expression. Both of the ß-lactams attenuated the effects of fentanyl overdose on locomotor activities but did not induce significant changes in the overall metabolomic profile. Our findings revealed that the exposure to a high dose of fentanyl causes alterations in key metabolic pathways in the NAc. Pretreatment with ceftriaxone and MC-100093 normalized fentanyl-induced downregulation of GLT-1 expression with subsequent attenuation of neuroinflammation as well as the hyperactivity, indicating that ß-lactams may be promising drugs for treating fentanyl use disorder.

9.
Neuroscience ; 552: 89-99, 2024 Aug 06.
Artigo em Inglês | MEDLINE | ID: mdl-38909675

RESUMO

Chronic ethanol consumption increased extracellular glutamate concentrations in several reward brain regions. Glutamate homeostasis is regulated in majority by astrocytic glutamate transporter 1 (GLT-1) as well as the interactive role of cystine/glutamate antiporter (xCT). In this study, we aimed to determine the attenuating effects of a novel beta-lactam MC-100093, lacking the antibacterial properties, on ethanol consumption and GLT-1 and xCT expression in the subregions of nucleus accumbens (NAc core and NAc shell) and medial prefrontal cortex (Infralimbic, mPFC-IL and Prelimbic, mPFC-PL) in male and female alcohol-preferring (P) rats. Female and male rats were exposed to free access to ethanol (15% v/v) and (30% v/v) and water for five weeks, and on Week 6, rats were administered 100 mg/kg (i.p) of MC-100093 or saline for five days. MC-100093 reduced ethanol consumption in both male and female P rats from Day 1-5. Additionally, MC-100093 upregulated GLT-1 and xCT expression in the mPFC and NAc subregions as compared to ethanol-saline groups in female and male rats. Chronic ethanol intake reduced GLT-1 and xCT expression in the IL and PL in female and male rats, except there was no reduction in GLT-1 expression in the mPFC-PL in female rats. Although, MC-100093 upregulated GLT-1 and xCT expression in the subregions of NAc, we did not observe any reduction in GLT-1 and xCT expression with chronic ethanol intake in female rats. These findings strongly suggest that MC-100093 treatment effectively reduced ethanol intake and upregulated GLT-1 and xCT expression in the mPFC and NAc subregions in male and female P rats.


Assuntos
Consumo de Bebidas Alcoólicas , Astrócitos , Etanol , Transportador 2 de Aminoácido Excitatório , Núcleo Accumbens , Animais , Feminino , Masculino , Consumo de Bebidas Alcoólicas/metabolismo , Ratos , Astrócitos/metabolismo , Astrócitos/efeitos dos fármacos , Transportador 2 de Aminoácido Excitatório/metabolismo , Transportador 2 de Aminoácido Excitatório/biossíntese , Etanol/administração & dosagem , Etanol/farmacologia , Núcleo Accumbens/metabolismo , Núcleo Accumbens/efeitos dos fármacos , Córtex Pré-Frontal/metabolismo , Córtex Pré-Frontal/efeitos dos fármacos , Depressores do Sistema Nervoso Central/farmacologia , Depressores do Sistema Nervoso Central/administração & dosagem , Caracteres Sexuais
10.
Saudi Pharm J ; 32(7): 102108, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38868175

RESUMO

Chronic exposure to opioids can lead to downregulation of astrocytic glutamate transporter 1 (GLT-1), which regulates the majority of glutamate uptake. Studies from our lab revealed that beta-lactam antibiotic, ceftriaxone, attenuated hydrocodone-induced downregulation of GLT-1 as well as cystine/glutamate antiporter (xCT) expression in central reward brain regions. In this study, we investigated the effects of escalating doses of morphine and tested the efficacy of novel synthetic non-antibiotic drug, MC-100093, and ceftriaxone in attenuating the effects of morphine exposure in the expression of GLT-1, xCT, and neuroinflammatory factors (IL-6 and TGF-ß) in the nucleus accumbens (NAc). This study also investigated the effects of morphine and beta-lactams in locomotor activity, spontaneous alternation percentage (SAP) and number of entries in Y maze since opioids have effects in locomotor sensitization. Mice were exposed to moderate dose of morphine (20 mg/kg, i.p.) on days 1, 3, 5, 7, and a higher dose of morphine (150 mg/kg, i.p.) on day 9, and these mice were then behaviorally tested and euthanized on Day 10. Western blot analysis showed that exposure to morphine downregulated GLT-1 and xCT expression in the NAc, and both MC-100093 and ceftriaxone attenuated these effects. In addition, morphine exposure increased IL-6 mRNA and TGF-ß mRNA expression, and MC-100093 and ceftriaxone attenuated only the effect on IL-6 mRNA expression in the NAc. Furthermore, morphine exposure induced an increase in distance travelled, and MC-100093 and ceftriaxone attenuated this effect. In addition, morphine exposure decreased the SAP and increased the number of arm entries in Y maze, however, neither MC-100093 nor ceftriaxone showed any attenuating effect. Our findings demonstrated for the first time that MC-100093 and ceftriaxone attenuated morphine-induced downregulation of GLT-1 and xCT expression, and morphine-induced increase in neuroinflammatory factor, IL-6, as well as hyperactivity. These findings revealed the beneficial therapeutic effects of MC-100093 and ceftriaxone against the effects of exposure to escalated doses of morphine.

11.
Am J Physiol Endocrinol Metab ; 327(1): E27-E41, 2024 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-38690938

RESUMO

Pancreatic endocrine cells employ a sophisticated system of paracrine and autocrine signals to synchronize their activities, including glutamate, which controls hormone release and ß-cell viability by acting on glutamate receptors expressed by endocrine cells. We here investigate whether alteration of the excitatory amino acid transporter 2 (EAAT2), the major glutamate clearance system in the islet, may occur in type 2 diabetes mellitus and contribute to ß-cell dysfunction. Increased EAAT2 intracellular localization was evident in islets of Langerhans from T2DM subjects as compared with healthy control subjects, despite similar expression levels. Chronic treatment of islets from healthy donors with high-glucose concentrations led to the transporter internalization in vesicular compartments and reduced [H3]-d-glutamate uptake (65 ± 5% inhibition), phenocopying the findings in T2DM pancreatic sections. The transporter relocalization was associated with decreased Akt phosphorylation protein levels, suggesting an involvement of the phosphoinositide 3-kinase (PI3K)/Akt pathway in the process. In line with this, PI3K inhibition by a 100-µM LY294002 treatment in human and clonal ß-cells caused the transporter relocalization in intracellular compartments and significantly reduced the glutamate uptake compared to control conditions, suggesting that hyperglycemia changes the trafficking of the transporter to the plasma membrane. Upregulation of the glutamate transporter upon treatment with the antibiotic ceftriaxone rescued hyperglycemia-induced ß-cells dysfunction and death. Our data underscore the significance of EAAT2 in regulating islet physiology and provide a rationale for potential therapeutic targeting of this transporter to preserve ß-cell survival and function in diabetes.NEW & NOTEWORTHY The glutamate transporter SLC1A2/excitatory amino acid transporter 2 (EAAT2) is expressed on the plasma membrane of pancreatic ß-cells and controls islet glutamate clearance and ß-cells survival. We found that the EAAT2 membrane expression is lost in the islets of Langerhans from type 2 diabetes mellitus (T2DM) patients due to hyperglycemia-induced downregulation of the phosphoinositide 3-kinase/Akt pathway and modification of its intracellular trafficking. Pharmacological rescue of EAAT2 expression prevents ß-cell dysfunction and death, suggesting EAAT2 as a new potential target of intervention in T2DM.


Assuntos
Diabetes Mellitus Tipo 2 , Transportador 2 de Aminoácido Excitatório , Ácido Glutâmico , Hiperglicemia , Ilhotas Pancreáticas , Transportador 2 de Aminoácido Excitatório/metabolismo , Humanos , Diabetes Mellitus Tipo 2/metabolismo , Ácido Glutâmico/metabolismo , Hiperglicemia/metabolismo , Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/efeitos dos fármacos , Masculino , Pessoa de Meia-Idade , Feminino , Transporte Proteico , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Idoso , Adulto , Animais , Fosfatidilinositol 3-Quinases/metabolismo
12.
Pharmacol Rep ; 76(4): 665-678, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38758470

RESUMO

Neurodegenerative diseases (NDDs) encompass a range of conditions characterized by the specific dysfunction and continual decline of neurons, glial cells, and neural networks within the brain and spinal cord. The majority of NDDs exhibit similar underlying causes, including oxidative stress, neuroinflammation, and malfunctioning of mitochondria. Elevated levels of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), alongside decreased expression of brain-derived neurotrophic factor (BDNF) and glutamate transporter subtype 1 (GLT-1), constitute significant factors contributing to the pathogenesis of NDDs. Additionally, the dual-specificity tyrosine phosphorylation-regulated kinase 1 A (DYRK1A) gene has emerged as a significant target for the treatment of NDDs at the preclinical level. It significantly contributes to developmental brain defects, early onset neurodegeneration, neuronal loss, and dementia in Down syndrome. Moreover, an impaired ubiquitin-proteosome system (UPS) also plays a pathological role in NDDs. Malfunctioning of UPS leads to abnormal protein buildup or aggregation of α-synuclein. α-Synuclein is a highly soluble unfolded protein that accumulates in Lewy bodies and Lewy neurites in Parkinson's disease and other synucleinopathies. Recent research highlights the promising potential of natural products in combating NDDs relative to conventional therapies. Alkaloids have emerged as promising candidates in the fight against NDDs. Harmine is a tricyclic ß-carboline alkaloid (harmala alkaloid) with one indole nucleus and a six-membered pyrrole ring. It is extracted from Banisteria caapi and Peganum harmala L. and exhibits diverse pharmacological properties, encompassing neuroprotective, antioxidant, anti-inflammatory, antidepressant, etc. Harmine has been reported to mediate its neuroprotective via reducing the level of inflammatory mediators, NADPH oxidase, AChE, BChE and reactive oxygen species (ROS). Whereas, it has been observed to increase the levels of BDNF, GLT-1 and anti-oxidant enzymes, along with protein kinase-A (PKA)-mediated UPS activation. This review aims to discuss the mechanistic interplay of various mediators involved in the neuroprotective effect of harmine.


Assuntos
Harmina , Doenças Neurodegenerativas , Fármacos Neuroprotetores , Humanos , Harmina/farmacologia , Fármacos Neuroprotetores/farmacologia , Fármacos Neuroprotetores/uso terapêutico , Animais , Doenças Neurodegenerativas/tratamento farmacológico , Doenças Neurodegenerativas/metabolismo , Estresse Oxidativo/efeitos dos fármacos
13.
Brain Sci ; 14(4)2024 Apr 05.
Artigo em Inglês | MEDLINE | ID: mdl-38672013

RESUMO

Chronic opioid treatments dysregulate the glutamatergic system, inducing a hyperglutamatergic state in mesocorticolimbic brain regions. This study investigated the effects of exposure to hydrocodone overdose on locomotor activity, expression of target proteins related to the glutamatergic system, signaling kinases, and neuroinflammatory factors in the nucleus accumbens. The locomotor activity of mice was measured using the Comprehensive Laboratory Animal Monitoring System (CLAMS). CLAMS data showed that exposure to hydrocodone overdose increased locomotion activity in mice. This study tested ceftriaxone, known to upregulate major glutamate transporter 1 (GLT-1), in mice exposed to an overdose of hydrocodone. Thus, ceftriaxone normalized hydrocodone-induced hyperlocomotion activity in mice. Furthermore, exposure to hydrocodone overdose downregulated GLT-1, cystine/glutamate antiporter (xCT), and extracellular signal-regulated kinase activity (p-ERK/ERK) expression in the nucleus accumbens. However, exposure to an overdose of hydrocodone increased metabotropic glutamate receptor 5 (mGluR5), neuronal nitric oxide synthase activity (p-nNOS/nNOS), and receptor for advanced glycation end products (RAGE) expression in the nucleus accumbens. Importantly, ceftriaxone treatment attenuated hydrocodone-induced upregulation of mGluR5, p-nNOS/nNOS, and RAGE, as well as hydrocodone-induced downregulation of GLT-1, xCT, and p-ERK/ERK expression. These data demonstrated that exposure to hydrocodone overdose can cause dysregulation of the glutamatergic system, neuroinflammation, hyperlocomotion activity, and the potential therapeutic role of ceftriaxone in attenuating these effects.

14.
J Neurochem ; 168(9): 2479-2494, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-38497582

RESUMO

Stressful life events contribute to the onset of major depressive disorder (MDD). We recently demonstrated abnormalities in ubiquitination in the pathophysiology of MDD. However, the underlying molecular mechanisms remain unclear. We investigated the involvement of the ubiquitination system-mediated glutamatergic dysfunction in social impairment induced by chronic social defeat stress (CSDS). Adult C57BL/6J mice were exposed to aggressor ICR male mice for 10 consecutive days. Social impairment was induced by CSDS in the social interaction test 1 days after the last stress exposure. In terms of brain microdialysis, CSDS reduced depolarization-evoked glutamate release in the prefrontal cortex (PFC), which was reversed by a glutamate transporter 1 (GLT-1) inhibitor. Interestingly, the expression of ubiquitinated, but not total GLT-1, was decreased in the PFC of mice exposed to CSDS. The expression of neural precursor cells expressing developmentally downregulated gene 4-like (Nedd4L: E3 ligase for GLT-1), and ubiquitin-conjugating enzyme E2D2 (Ube2d2: E2 ubiquitin-conjugating enzyme for Nedd4L) was also reduced in CSDS mice. Furthermore, the downregulation of the Nedd4L-GLT-1 ubiquitination pathway decreased SIT ratio, but up-regulation increased it even in non-CSDS mice. Taken together, the decrease in GLT-1 ubiquitination may reduce the release of extracellular glutamate induced by high-potassium stimulation, which may lead to social impairment, while we could not find differences in GLT-1 ubiquitination between susceptible and resistant CSDS mice. In conclusion, GLT-1 ubiquitination could play a crucial role in the pathophysiology of MDD and is an attractive target for the development of novel antidepressants.


Assuntos
Transportador 2 de Aminoácido Excitatório , Córtex Pré-Frontal , Derrota Social , Animais , Masculino , Camundongos , Regulação para Baixo , Transportador 2 de Aminoácido Excitatório/metabolismo , Transportador 2 de Aminoácido Excitatório/biossíntese , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos ICR , Ubiquitina-Proteína Ligases Nedd4/metabolismo , Córtex Pré-Frontal/metabolismo , Transdução de Sinais/fisiologia , Estresse Psicológico/metabolismo , Ubiquitinação
15.
J Biol Chem ; 300(4): 107172, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38499151

RESUMO

The recently discovered interaction between Presenilin 1 (PS1), a catalytic subunit of γ-secretase responsible for generating amyloid-ß peptides, and GLT-1, a major glutamate transporter in the brain (EAAT2), provides a mechanistic link between these two key factors involved in Alzheimer's disease (AD) pathology. Modulating this interaction can be crucial to understand the consequence of such crosstalk in AD context and beyond. However, the interaction sites between these two proteins are unknown. Herein, we utilized an alanine scanning approach coupled with FRET-based fluorescence lifetime imaging microscopy to identify the interaction sites between PS1 and GLT-1 in their native environment within intact cells. We found that GLT-1 residues at position 276 to 279 (TM5) and PS1 residues at position 249 to 252 (TM6) are crucial for GLT-1-PS1 interaction. These results have been cross validated using AlphaFold Multimer prediction. To further investigate whether this interaction of endogenously expressed GLT-1 and PS1 can be prevented in primary neurons, we designed PS1/GLT-1 cell-permeable peptides (CPPs) targeting the PS1 or GLT-1 binding site. We used HIV TAT domain to allow for cell penetration which was assayed in neurons. First, we assessed the toxicity and penetration of CPPs by confocal microscopy. Next, to ensure the efficiency of CPPs, we monitored the modulation of GLT-1-PS1 interaction in intact neurons by fluorescence lifetime imaging microscopy. We saw significantly less interaction between PS1 and GLT-1 with both CPPs. Our study establishes a new tool to study the functional aspect of GLT-1-PS1 interaction and its relevance in normal physiology and AD models.


Assuntos
Transportador 2 de Aminoácido Excitatório , Presenilina-1 , Animais , Humanos , Camundongos , Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Secretases da Proteína Precursora do Amiloide/metabolismo , Secretases da Proteína Precursora do Amiloide/genética , Sítios de Ligação , Transportador 2 de Aminoácido Excitatório/química , Transportador 2 de Aminoácido Excitatório/genética , Transportador 2 de Aminoácido Excitatório/metabolismo , Transferência Ressonante de Energia de Fluorescência , Células HEK293 , Neurônios/metabolismo , Presenilina-1/química , Presenilina-1/genética , Presenilina-1/metabolismo , Ligação Proteica , Peptídeos/metabolismo
16.
Biochem Biophys Res Commun ; 701: 149550, 2024 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-38310688

RESUMO

The beneficial effect of a beta-lactam antibiotic, Ceftriaxone (CEF), to improve depressive-like symptoms has been documented previously, attributed to its modulation of glutamate neurotransmission. Here, we aimed to determine whether CEF could improve LPS-altered glutamatergic signaling associated with neuroinflammation-allied depression. To assess our goals, we established a neuroinflammation-allied depression mice model by injecting lipopolysaccharides (LPS), followed by behavioral and biochemical analysis. LPS-treated mice displayed depressive symptoms, neuroinflammation, dysregulated glutamate and its transporter (GLT-1) expression, altered expression of astrocyte reactive markers (GFAP, cxcl10, steap4, GBP2, and SRGN), and dysregulated BDNF/TrkB signaling. However, these changes were rescued by CEF treatment, as we found decreased neuroinflammation, relief of depression symptoms, and improved GLT-1 and BDNF/TrkB signaling upon CEF treatment. Moreover, GLT-1 and BDNF/TrkB regulation role of CEF was validated by K252a and DHK treatment. In summary, the anti-depressive effects of glutamate modulators, like CEF, are closely related to their anti-inflammatory role.


Assuntos
Fator Neurotrófico Derivado do Encéfalo , Ceftriaxona , Camundongos , Animais , Ceftriaxona/farmacologia , Ceftriaxona/uso terapêutico , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Lipopolissacarídeos , Doenças Neuroinflamatórias , Ácido Glutâmico/metabolismo , Transportador 2 de Aminoácido Excitatório/metabolismo
17.
Neurochem Res ; 49(5): 1226-1238, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38393622

RESUMO

Both epidemiological investigation and animal experiments demonstrated that pre-/postnatal exposure to perfluorooctane sulfonic acid (PFOS) could induce neurodevelopmental disorders. Previous studies showed that astrocyte was involved in PFOS-induced neurotoxicity, while little information is available. In the present study, the role of astrocyte-derived calmodulin-dependent protein kinase II (CaMKII)-phosphorylated discs large homolog 1 (DLG1) signaling in PFOS eliciting cytotoxicity in neuron was explored with primary cultured hippocampal astrocyte and neuron. The application of PFOS showed a decreased cell viability, synapse length and glutamate transporter 1 (GLT-1) expression, but an increased CaMKII, DLG1 and cyclic AMP response element binding protein (CREB) expression in primary cultured astrocyte. With 2-(2-hydroxyethylamino)-6-aminohexylcarbamic acid tert-butyl ester-9-isopropylpurine (CK59), the CaMKII inhibitor, the disturbed cell viability and molecules induced by PFOS could be alleviated (CREB expression was excluded) in astrocytes. The cytotoxic effect of neuron exposed to astrocyte conditional medium collected from PFOS (PFOS-ACM) pretreated with CK59 was also decreased. These results indicated that PFOS mediated GLT-1 expression through astrocyte-derived CaMKII-DLG signaling, which might be associated with injuries on neurons. The present study gave an insight in further exploration of mechanism in PFOS-induced neurotoxicity.


Assuntos
Ácidos Alcanossulfônicos , Astrócitos , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina , Fluorocarbonos , Ratos , Animais , Astrócitos/metabolismo , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Hipocampo/metabolismo , Neurônios/metabolismo , Células Cultivadas
18.
Mol Neurobiol ; 61(9): 7181-7194, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-38374316

RESUMO

Α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors (AMPARs) are crucial for properties of synaptic plasticity, such as long-term potentiation (LTP). LTP impairment can occur early in the onset of Alzheimer's disease (AD). The downregulation or decreased abundance of AMPAR expression in the postsynaptic membrane is closely associated with LTP impairment. Ceftriaxone (Cef) can improve LTP impairment in the early stages of AD in a mouse model. The purpose of this study was to explore the mechanism underlying this process from the aspects of AMPAR expression and ubiquitination degree. In this study, we found that ß-amyloid (Aß) treatment induced hippocampal LTP impairment and AMPAR downregulation and ubiquitination. Cef pretreatment ameliorated Aß-induced hippocampal LTP impairment, reduced AMPAR ubiquitination, and increased AMPAR expression, especially in the plasma membrane, in Aß-treated mice. Administration of USP46 siRNA and DHK (a specific blocker of glutamate transporter-1) significantly inhibited the above effects of Cef, suggesting a role for anti-AMPAR ubiquitination and upregulation of glutamate transporter-1 (GLT-1) in the Cef-induced improvements mentioned above. The above findings demonstrate that pretreatment with Cef effectively mitigated Aß-induced impairment of hippocampal LTP by suppressing the ubiquitination process of AMPARs in a GLT-1-dependent manner. These results provide novel insights into the underlying mechanisms elucidating the anti-AD by Cef.


Assuntos
Peptídeos beta-Amiloides , Ceftriaxona , Potenciação de Longa Duração , Camundongos Endogâmicos C57BL , Receptores de AMPA , Ubiquitinação , Animais , Ceftriaxona/farmacologia , Potenciação de Longa Duração/efeitos dos fármacos , Receptores de AMPA/metabolismo , Ubiquitinação/efeitos dos fármacos , Peptídeos beta-Amiloides/metabolismo , Masculino , Hipocampo/metabolismo , Hipocampo/efeitos dos fármacos , Transportador 2 de Aminoácido Excitatório/metabolismo , Camundongos
19.
Front Cell Dev Biol ; 12: 1334861, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38362041

RESUMO

Introduction: Astrocytic GLT-1 glutamate transporters ensure the fidelity of glutamic neurotransmission by spatially and temporally limiting glutamate signals. The ability to limit neuronal hyperactivity relies on the localization and diffusion of GLT-1 on the astrocytic surface, however, little is known about the underlying mechanisms. We show that two isoforms of GLT-1, GLT-1a and GLT-1b, form nanoclusters on the surface of transfected astrocytes and HEK-293 cells. Methods: We used both fixed and live cell super-resolution imaging of fluorescent protein and epitope tagged proteins in co-cultures of rat astrocytes and neurons. Immunofluorescence techniques were also used. GLT1 diffusion was assessed via single particle tracking and fluorescence recovery after photobleach (FRAP). Results: We found GLT-1a, but not GLT-1b, nanoclusters concentrated adjacent to actin filaments which was maintained after addition of glutamate. GLT-1a nanocluster concentration near actin filaments was prevented by expression of a cytosolic GLT-1a C-terminus, suggesting the C-terminus is involved in the localization adjacent to cortical actin. Using super-resolution imaging, we show that astrocytic GLT-1a and actin co-localize in net-like structures around neuronal Kv2.1 clusters at points of neuron/astrocyte contact. Conclusion: Overall, these data describe a novel relationship between GLT-1a and cortical actin filaments, which localizes GLT-1a near neuronal structures responsive to ischemic insult.

20.
Neurobiol Dis ; 193: 106452, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38401650

RESUMO

A common adverse effect of Parkinson's disease (PD) treatment is L-dopa-induced dyskinesia (LID). This condition results from both dopamine (DA)-dependent and DA-independent mechanisms, as glutamate inputs from corticostriatal projection neurons impact DA-responsive medium spiny neurons in the striatum to cause the dyskinetic behaviors. In this study, we explored whether suppression of presynaptic corticostriatal glutamate inputs might affect the behavioral and biochemical outcomes associated with LID. We first established an animal model in which 6-hydroxydopamine (6-OHDA)-lesioned mice were treated daily with L-dopa (10 mg/kg, i.p.) for 2 weeks; these mice developed stereotypical abnormal involuntary movements (AIMs). When the mice were pretreated with the NMDA antagonist, amantadine, we observed suppression of AIMs and reductions of phosphorylated ERK1/2 and NR2B in the striatum. We then took an optogenetic approach to manipulate glutamatergic activity. Slc17a6 (vGluT2)-Cre mice were injected with pAAV5-Ef1a-DIO-eNpHR3.0-mCherry and received optic fiber implants in either the M1 motor cortex or dorsolateral striatum. Optogenetic inactivation at either optic fiber implant location could successfully reduce the intensity of AIMs after 6-OHDA lesioning and L-dopa treatment. Both optical manipulation strategies also suppressed phospho-ERK1/2 and phospho-NR2B signals in the striatum. Finally, we performed intrastriatal injections of LDN 212320 in the dyskenesic mice to enhance expression of glutamate uptake transporter GLT-1. Sixteen hours after the LDN 212320 treatment, L-dopa-induced AIMs were reduced along with the levels of striatal phospho-ERK1/2 and phospho-NR2B. Together, our results affirm a critical role of corticostriatal glutamate neurons in LID and strongly suggest that diminishing synaptic glutamate, either by suppression of neuronal activity or by upregulation of GLT-1, could be an effective approach for managing LID.


Assuntos
Discinesias , Doença de Parkinson , Ratos , Camundongos , Animais , Levodopa/farmacologia , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/metabolismo , Oxidopamina/toxicidade , Ácido Glutâmico/metabolismo , Ratos Sprague-Dawley , Dopamina/metabolismo , Corpo Estriado/metabolismo , Modelos Animais de Doenças , Antiparkinsonianos/efeitos adversos
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