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1.
Life Sci ; 231: 116581, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31220524

RESUMO

AIMS: The aims of this study were to investigate the effect of colonic electrical stimulation (CES) on delayed colonic transit in Parkinson's disease (PD) model induced by rotenone and its possible mechanisms. MAIN METHODS: Sprague-Dawley male rats were implanted with a pair of electrodes on the serosa at the proximal colon and rotenone was subcutaneously injected for 6 weeks to induce the PD model. Behavior activity, stool volume and open-field test were recorded during the injection. Colonic propulsion rate was measured 6 weeks after rotenone injection. Colon samples of all rats were collected for the measurement of phosphorylated alpha-synuclein, choline acetyltransferase (CHAT), neuronal nitric oxide synthase (nNOS), and tyrosine hydroxylase (TH). The protocols of control rats were the same as the PD rats except that no electrodes were implanted and no rotenone was injected. KEY FINDINGS: (1) Rotenone-induced PD rats demonstrated weight loss, significant decrease of the dopaminergic neurons in substantia nigra, and impairment of colon movement. (2) CES significantly accelerated the delayed colonic transmit (91.67 ±â€¯5.58% vs 51.33 ±â€¯4.18%), superior to Macrogol-4000. (3) CES significantly upregulated the expression of CHAT, nNOS and TH protein in colon of PD rats. (4) In colon of PD rats, the phosphorylated alpha-synuclein was significantly upregulated, but CES had no significant effect on phosphorylated alpha-synuclein. SIGNIFICANCE: Our data show that CES can normalize the delayed colonic transit and this normalization may attribute to affecting enteric excitatory and inhibitory neurons.


Assuntos
Colo/metabolismo , Sistema Nervoso Entérico/efeitos dos fármacos , Doença de Parkinson/fisiopatologia , Animais , Colina O-Acetiltransferase/metabolismo , Colo/fisiopatologia , Constipação Intestinal/fisiopatologia , Modelos Animais de Doenças , Estimulação Elétrica , Motilidade Gastrointestinal/efeitos dos fármacos , Intestino Delgado/metabolismo , Masculino , Neurônios/efeitos dos fármacos , Óxido Nítrico Sintase/metabolismo , Óxido Nítrico Sintase Tipo I/metabolismo , Ratos , Ratos Sprague-Dawley , Rotenona/metabolismo , Rotenona/farmacologia , Substância Negra/efeitos dos fármacos , Tirosina 3-Mono-Oxigenase/metabolismo , alfa-Sinucleína/metabolismo
2.
Life Sci ; 232: 116600, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31251998

RESUMO

Neuroinflammation is one of the significant neuropathological conditions in Parkinson's disease (PD) which is due to microglial and astrocytes activation leads to progressive dopaminergic neuronal loss. To date, Current PD drugs offers only symptomatic relief with adverse effects and lack of ability to prevent the progression of neurodegeneration. Therefore, a better approach to develop a multi potent drug of natural origin would be beneficial in managing the disease. Therefore, the present study aimed to investigate the neuroprotective and anti-inflammatory effects of PHL by exploring its neuroprotective mechanism in 1-methyl-4-phenyl-1,2,3,6-tetrahydro pyridine (MPTP) induced PD in mice. MPTP intoxication in mice cause motor abnormalities, decreased dopamine (DA) levels, reduced tyrosine hydroxylase (TH) enzyme protein expression and inflammation which were effectively restored by PHL. Moreover gliotic specific inflammatory markers like glial fibrillary acidic protein (GFAP), ionized calcium-binding adaptor protein-1 (Iba-1), iNOS and COX-2 were found to be expressed more in MPTP intoxicated mice, Further the levels of proinflammatory cytokines like IL-ß, IL-6, and TNF-α were significantly upregulated in MPTP intoxicated mice, these deleterious responses were diminished to extend neuroprotection by PHL treatment. Our findings strongly suggest PHL as a potent therapeutic agent in treating PD.


Assuntos
Transtornos Parkinsonianos/tratamento farmacológico , Floretina/farmacologia , Animais , Comportamento Animal/efeitos dos fármacos , Proteínas de Ligação ao Cálcio/metabolismo , Ciclo-Oxigenase 2/metabolismo , Citocinas/metabolismo , Modelos Animais de Doenças , Dopamina/metabolismo , Neurônios Dopaminérgicos/efeitos dos fármacos , Proteína Glial Fibrilar Ácida/metabolismo , Inflamação/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas dos Microfilamentos/metabolismo , Microglia/efeitos dos fármacos , Neuroimunomodulação/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Óxido Nítrico Sintase Tipo II/metabolismo , Transtornos Parkinsonianos/metabolismo , Transtornos Parkinsonianos/patologia , Tirosina 3-Mono-Oxigenase/metabolismo
3.
Nat Commun ; 10(1): 2046, 2019 05 03.
Artigo em Inglês | MEDLINE | ID: mdl-31053723

RESUMO

Impaired neuronal processes, including dopamine imbalance, are central to the pathogenesis of major psychosis, but the molecular origins are unclear. Here we perform a multi-omics study of neurons isolated from the prefrontal cortex in schizophrenia and bipolar disorder (n = 55 cases and 27 controls). DNA methylation, transcriptomic, and genetic-epigenetic interactions in major psychosis converged on pathways of neurodevelopment, synaptic activity, and immune functions. We observe prominent hypomethylation of an enhancer within the insulin-like growth factor 2 (IGF2) gene in major psychosis neurons. Chromatin conformation analysis revealed that this enhancer targets the nearby tyrosine hydroxylase (TH) gene responsible for dopamine synthesis. In patients, we find hypomethylation of the IGF2 enhancer is associated with increased TH protein levels. In mice, Igf2 enhancer deletion disrupts the levels of TH protein and striatal dopamine, and induces transcriptional and proteomic abnormalities affecting neuronal structure and signaling. Our data suggests that epigenetic activation of the enhancer at IGF2 may enhance dopamine synthesis associated with major psychosis.


Assuntos
Transtorno Bipolar/genética , Dopamina/biossíntese , Elementos Facilitadores Genéticos/genética , Epigênese Genética , Fator de Crescimento Insulin-Like II/genética , Esquizofrenia/genética , Tirosina 3-Mono-Oxigenase/genética , Adulto , Idoso , Animais , Transtorno Bipolar/patologia , Metilação de DNA , Feminino , Perfilação da Expressão Gênica , Humanos , Masculino , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Neurônios/patologia , Córtex Pré-Frontal/citologia , Córtex Pré-Frontal/patologia , Proteômica , Esquizofrenia/patologia , Transcriptoma/genética , Tirosina 3-Mono-Oxigenase/metabolismo , Adulto Jovem
4.
Int J Mol Sci ; 20(7)2019 Mar 27.
Artigo em Inglês | MEDLINE | ID: mdl-30934738

RESUMO

Parkinson's disease (PD), a multifactorial movement disorder that involves progressive degeneration of the nigrostriatal system affecting the movement ability of the patient. Oxidative stress and neuroinflammation both are shown to be involved in the etiopathogenesis of PD. The aim of this study was to evaluate the therapeutic potential of thymol, a dietary monoterpene phenol in rotenone (ROT)-induced neurodegeneration in rats that precisely mimics PD in humans. Male Wistar rats were injected ROT at a dose of 2.5 mg/kg body weight for 4 weeks, to induce PD. Thymol was co-administered for 4 weeks at a dose of 50 mg/kg body weight, 30 min prior to ROT injection. The markers of dopaminergic neurodegeneration, oxidative stress and inflammation were estimated using biochemical assays, enzyme-linked immunosorbent assay, western blotting and immunocytochemistry. ROT challenge increased the oxidative stress markers, inflammatory enzymes and cytokines as well as caused significant damage to nigrostriatal dopaminergic system of the brain. Thymol treatment in ROT challenged rats appears to significantly attenuate dopaminergic neuronal loss, oxidative stress and inflammation. The present study showed protective effects of thymol in ROT-induced neurotoxicity and neurodegeneration mediated by preservation of endogenous antioxidant defense networks and attenuation of inflammatory mediators including cytokines and enzymes.


Assuntos
Dieta , Neurônios Dopaminérgicos/patologia , Degeneração Neural/prevenção & controle , Fármacos Neuroprotetores/uso terapêutico , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/patologia , Timol/uso terapêutico , Animais , Catalase/metabolismo , Ciclo-Oxigenase 2/metabolismo , Citocinas/metabolismo , Modelos Animais de Doenças , Neurônios Dopaminérgicos/efeitos dos fármacos , Neurônios Dopaminérgicos/metabolismo , Glutationa/metabolismo , Mediadores da Inflamação/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Malondialdeído/metabolismo , Neostriado/efeitos dos fármacos , Neostriado/patologia , Degeneração Neural/patologia , Neuroglia/efeitos dos fármacos , Neuroglia/metabolismo , Fármacos Neuroprotetores/farmacologia , Óxido Nítrico Sintase Tipo II/metabolismo , Ratos Wistar , Rotenona , Substância Negra/efeitos dos fármacos , Substância Negra/patologia , Superóxido Dismutase/metabolismo , Timol/química , Timol/farmacologia , Tirosina 3-Mono-Oxigenase/metabolismo
5.
J Therm Biol ; 81: 137-145, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30975411

RESUMO

Brown adipose tissue (BAT) plays an important role in thermoregulation and many metabolic processes in small mammals, especially in cold adaptation. However, in warm adaptation, ambient temperature cannot directly activate BAT by sympathetic nervous system. Mongolian gerbils exhibit a wider thermoneutral zone (26.5-38.9 °C). We hypothesized that BAT atrophied near the lower critical temperature and further atrophied near the upper critical temperature. Male gerbils were acclimated to 23 °C, 27 °C or 37 °C, respectively, for 3 weeks. Results showed that regulatory non-shivering thermogenesis did not change in gerbils acclimated to 27 °C compared with 23 °C group, whereas it was reduced by 43.5% in gerbils acclimated to 37 °C. Bigger lipid droplet in BAT was observed in gerbils acclimated to 27 °C and 37 °C compared with 23 °C group, while the expression of uncoupling protein 1 and tyrosine hydroxylase was only reduced in gerbils acclimated to 37 °C. Further, thermoneutral acclimation did not change BAT thermogenesis by down-regulation of peroxisome proliferator-activated receptor gamma coactivator-1α, PR domain containing 16, peroxisome proliferator-activated receptor-α or peroxisome proliferator activated receptor-γ gene expression in BAT. In addition, body temperature was reduced in gerbils acclimated to 37 °C compared with 23 °C group, which was associated with a decreased resting metabolic rate and regulatory non-shivering thermogenesis. In conclusion, BAT does not atrophy near the lower critical temperature, whereas it atrophies near the upper critical temperature, suggesting that BAT may play thermoregulatory role within the TNZ in Mongolian gerbils.


Assuntos
Tecido Adiposo Marrom/fisiologia , Regulação da Temperatura Corporal , Gerbillinae/fisiologia , Termotolerância , Tecido Adiposo Marrom/citologia , Animais , Metabolismo Basal , Ingestão de Alimentos , Gerbillinae/metabolismo , Masculino , Tamanho do Órgão , Temperatura Ambiente , Termogênese , Tirosina 3-Mono-Oxigenase/metabolismo , Proteína Desacopladora 1/metabolismo
6.
Bull Exp Biol Med ; 166(6): 811-815, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31020581

RESUMO

We assessed changes of olfactory bulbs in rata with 6-hydroxydopamine destruction of the substantia nigra. The expression of marker proteins of immature and differentiated neurons and glia (vimentin, PSA-NCAM, tyrosine hydroxylase, and S100) was analyzed by immunohistochemical and morphometric methods. The number of periglomerular dopamine neurons and astroglia in the olfactory bulbs increased on the side of toxin injection and expression of PSA-NCAM and vimentin increased in the rostral migratory stream. Destruction of the substantia nigra shifted differentiation of neuronal progenitors towards the dopaminergic phenotype and increased their survival in the olfactory bulbs, which can be explained by increased expression of PSA-NCAM.


Assuntos
Neuroglia/patologia , Neurônios/patologia , Bulbo Olfatório/patologia , Doença de Parkinson Secundária/patologia , Substância Negra/patologia , Adaptação Fisiológica , Animais , Biomarcadores/metabolismo , Modelos Animais de Doenças , Expressão Gênica , Humanos , Imuno-Histoquímica , Injeções Intraventriculares , Masculino , Atividade Motora/fisiologia , Molécula L1 de Adesão de Célula Nervosa/genética , Molécula L1 de Adesão de Célula Nervosa/metabolismo , Neuroglia/metabolismo , Neurônios/metabolismo , Bulbo Olfatório/metabolismo , Oxidopamina/administração & dosagem , Doença de Parkinson Secundária/induzido quimicamente , Doença de Parkinson Secundária/genética , Doença de Parkinson Secundária/metabolismo , Ratos , Ratos Wistar , Proteínas S100/genética , Proteínas S100/metabolismo , Ácidos Siálicos/genética , Ácidos Siálicos/metabolismo , Técnicas Estereotáxicas , Substância Negra/metabolismo , Tirosina 3-Mono-Oxigenase/genética , Tirosina 3-Mono-Oxigenase/metabolismo , Vimentina/genética , Vimentina/metabolismo
7.
Chem Biol Interact ; 305: 134-147, 2019 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-30922767

RESUMO

Methiopropamine (MPA) is structurally categorized as a thiophene ring-based methamphetamine (MA) derivative. Although abusive potential of MPA was recognized, little is known about the neurotoxic potential of MPA up to now. We investigated whether MPA induces dopaminergic neurotoxicity, and whether MPA activates a specific dopamine receptor. Here, we observed that treatment with MPA resulted in dopaminergic neurotoxicity in a dose-dependent manner. MPA treatment potentiated oxidative parameters (i.e., increases in the level of reactive oxygen species, 4-hydroxynonenal, and protein carbonyl), M1 phenotype-related microglial activity, and pro-apoptotic property (i.e., increases in Bax- and cleaved caspase-3-expressions, while a decrease in Bcl-2-expression). Moreover, treatment with MPA resulted in significant impairments in dopaminergic parameters [i.e., changes in dopamine level, dopamine turnover rate, tyrosine hydroxylase (TH) levels, dopamine transporter (DAT) expression, and vesicular monoamine transporter-2 (VMAT-2) expression], and in behavioral deficits. Both dopamine D1 receptor antagonist SCH23390 and D2 receptor antagonist sulpiride protected from these neurotoxic consequences. Therefore, our results suggest that dopamine D1 and D2 receptors simultaneously mediate MPA-induced dopaminergic neurodegeneration in mice via oxidative burdens, microgliosis, and pro-apoptosis.


Assuntos
Metanfetamina/toxicidade , Estresse Oxidativo/efeitos dos fármacos , Receptores de Dopamina D1/metabolismo , Receptores de Dopamina D2/metabolismo , Animais , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismo , Benzazepinas/farmacologia , Benzazepinas/uso terapêutico , Diferenciação Celular/efeitos dos fármacos , Corpo Estriado/efeitos dos fármacos , Corpo Estriado/metabolismo , Dopamina/metabolismo , Antagonistas dos Receptores de Dopamina D2/farmacologia , Antagonistas dos Receptores de Dopamina D2/uso terapêutico , Febre/prevenção & controle , Locomoção/efeitos dos fármacos , Masculino , Metanfetamina/síntese química , Metanfetamina/química , Camundongos , Camundongos Endogâmicos ICR , Microglia/citologia , Microglia/efeitos dos fármacos , Microglia/metabolismo , Espécies Reativas de Oxigênio/análise , Espécies Reativas de Oxigênio/metabolismo , Receptores de Dopamina D1/antagonistas & inibidores , Receptores de Dopamina D2/química , Sulpirida/farmacologia , Sulpirida/uso terapêutico , Tirosina 3-Mono-Oxigenase/metabolismo
8.
Oxid Med Cell Longev ; 2019: 8745376, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30911352

RESUMO

This study examined the effects of lithium on gene expression and activity of the antioxidant enzymes copper zinc superoxide dismutase (SOD1), manganese superoxide dismutase (SOD2), catalase (CAT), glutathione peroxidase (GPx), and glutathione reductase (GR) in the hippocampus of chronically stressed rats. In addition, we examined the effects of lithium on anxiety behaviors, hippocampal concentrations of dopamine (DA) and malondialdehyde (MDA), protein levels of brain-derived neurotrophic factor (BDNF), tyrosine hydroxylase (TH), dopamine transporter (DAT), and catechol-O-methyltransferase (COMT), as well as activity of monoamine oxidase (MAO) in chronically stressed rats. The investigated parameters were quantified by real-time RT-PCR, Western blot analyses, and assays of enzyme activities. We found that lithium did not change gene expression of SOD1, CAT, GPx, and GR but decreased gene expression of SOD2 in chronically stressed rats. A very important result in this study was that lithium treatment decreased the enzyme activities of SOD1 and SOD2 but increased the enzyme activities of GPx and GR in stress condition, which indicates the control of redox balance. The reduced concentration of MDA confirms this. In addition, we found that lithium treatment decreased high protein levels of BDNF and DAT in chronically stressed rats to the level found in unstressed animals. Also, lithium treatment increased the expression of TH but decreased the enzyme activity of MAO B, which contributed to the increase of hippocampal concentration of DA in chronically stressed rats to the level of unstressed animals. Finally, lithium treatment in animals exposed to chronic stress increased the time spent in open arms. Lithium-induced modulation of hippocampal antioxidant status and attenuation of oxidative stress stabilized behavior in animals with high anxiety index. In addition, reduced oxidative stress was followed by the changes of both turnover of DA and levels of BDNF protein in chronically stressed rats treated with lithium. These findings may be important in preclinical research of the effects of lithium on oxidative stress level in pathological conditions.


Assuntos
Antioxidantes/metabolismo , Hipocampo/metabolismo , Lítio/uso terapêutico , Estresse Psicológico/tratamento farmacológico , Animais , Ansiedade/tratamento farmacológico , Comportamento Animal , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Catalase/genética , Catalase/metabolismo , Catecol O-Metiltransferase/metabolismo , Doença Crônica , Dopamina/metabolismo , Proteínas da Membrana Plasmática de Transporte de Dopamina/metabolismo , Glutationa Peroxidase/genética , Glutationa Peroxidase/metabolismo , Lítio/farmacologia , Masculino , Malondialdeído/metabolismo , Monoaminoxidase/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos Wistar , Estresse Psicológico/enzimologia , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo
9.
Int J Mol Sci ; 20(3)2019 Jan 29.
Artigo em Inglês | MEDLINE | ID: mdl-30699944

RESUMO

Parkinson's disease (PD) is a neurodegenerative disorder defined by progressive deterioration of dopaminergic neurons in the substantia nigra pars compacta (SNpc). Dental pulp stem cells (DPSCs) have been proposed to replace the degenerated dopaminergic neurons due to its inherent neurogenic and regenerative potential. However, the effective delivery and homing of DPSCs within the lesioned brain has been one of the many obstacles faced in cell-based therapy of neurodegenerative disorders. We hypothesized that DPSCs, delivered intranasally, could circumvent these challenges. In the present study, we investigated the therapeutic efficacy of intranasally administered DPSCs in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced mouse model of PD. Human deciduous DPSCs were cultured, pre-labelled with PKH 26, and intranasally delivered into PD mice following MPTP treatment. Behavioural analyses were performed to measure olfactory function and sensorimotor coordination, while tyrosine hydroxylase (TH) immunofluorescence was used to evaluate MPTP neurotoxicity in SNpc neurons. Upon intranasal delivery, degenerated TH-positive neurons were ameliorated, while deterioration in behavioural performances was significantly enhanced. Thus, the intranasal approach enriched cell delivery to the brain, optimizing its therapeutic potential through its efficacious delivery and protection against dopaminergic neuron degeneration.


Assuntos
Polpa Dentária/citologia , Intoxicação por MPTP/terapia , Doença de Parkinson/terapia , Parte Compacta da Substância Negra/citologia , Células-Tronco/fisiologia , Animais , Comportamento Animal , Diferenciação Celular/fisiologia , Células Cultivadas , Neurônios Dopaminérgicos/metabolismo , Humanos , Intoxicação por MPTP/metabolismo , Masculino , Camundongos , Degeneração Neural/metabolismo , Degeneração Neural/terapia , Doença de Parkinson/metabolismo , Parte Compacta da Substância Negra/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo
10.
Artigo em Inglês | MEDLINE | ID: mdl-30735775

RESUMO

Tyrosine hydroxylase (TH) is the initial enzyme responsible for cuticle sclerotization and pigmentation in many insect species, but to date, no direct functional studies have focused on TH in Zeugodacus tau. Here, the 3336-bp full-length cDNA of TH was isolated from Z. tau, a notorious horticultural pest infesting fruits and vegetables. qRT-polymerase chain reaction revealed that ZtTH transcripts were highly abundant at the time of pupal tanning and during adult emergence and were expressed in the midgut, integument and head of molting larvae. The pupation and eclosion rates gradually decreased when the 1st-instar larvae were fed diets containing higher concentrations of the TH inhibitor 3-iodo-tyrosine (3-IT). Moreover, pupal weights were significantly decreased, and abnormal uncolored phenotypes were observed after 20 mg/g 3-IT was incorporated into the diet. In addition, the suppression of TH function (mediated by RNA interference) led to a decrease in TH mRNAs and eclosion rates, accompanied by less-pigmented phenotypes. There was a severe impairment of larval-pupal cuticle tanning, leading to pupae with less yellowish pigment or that were completely white and transparent, when we injected 2 µL of 24.4 mM or 73.27 mM 3-IT into 3rd-instar larvae or prepupae. These results suggest that TH is an important enzyme for the normal growth and pupal pigmentation of Z. tau and that TH is a potential gene target for use in the control of Z. tau.


Assuntos
Pigmentação , Pupa/metabolismo , Tephritidae/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo , Animais , Fenótipo , Interferência de RNA , Análise de Sequência , Tephritidae/enzimologia , Tephritidae/genética , Tirosina 3-Mono-Oxigenase/deficiência , Tirosina 3-Mono-Oxigenase/genética
11.
Neurosci Bull ; 35(2): 205-215, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30706412

RESUMO

The locus coeruleus (LC) has been studied in major depressive disorder (MDD) and bipolar disorder (BD). A major problem of immunocytochemical studies in the human LC is interference with the staining of the immunocytochemical end-product by the omnipresent natural brown pigment neuromelanin. Here, we used a multispectral method to untangle the two colors: blue immunocytochemical staining and brown neuromelanin. We found significantly increased tyrosine hydroxylase (TH) in the LC of MDD patients-thus validating the method-but not in BD patients, and we did not find significant changes in the receptor tyrosine-protein kinase ErbB4 in the LC in MDD or BD patients. We observed clear co-localization of ErbB4, TH, and neuromelanin in the LC neurons. The different stress-related molecular changes in the LC may contribute to the different clinical symptoms in MDD and BD.


Assuntos
Transtorno Bipolar/metabolismo , Transtorno Depressivo Maior/metabolismo , Locus Cerúleo/metabolismo , Melaninas/metabolismo , Receptor ErbB-4/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo , Idoso , Idoso de 80 Anos ou mais , Transtorno Bipolar/patologia , Transtorno Depressivo Maior/patologia , Feminino , Humanos , Processamento de Imagem Assistida por Computador , Imuno-Histoquímica/métodos , Locus Cerúleo/patologia , Masculino , Microscopia/métodos , Pessoa de Meia-Idade , Neurônios/metabolismo , Neurônios/patologia , Sensibilidade e Especificidade , Análise Espectral/métodos
12.
Artigo em Inglês | MEDLINE | ID: mdl-30774584

RESUMO

Dopaminergic and serotonergic neurons modulate and control processes ranging from reward signaling to regulation of motor outputs. Further, dysfunction of these neurons is involved in both degenerative and psychiatric disorders. Elucidating the roles of these neurons has been greatly facilitated by bacterial artificial chromosome (BAC) transgenic mouse lines expressing channelrhodopsin to readily enable cell-type specific activation. However, corresponding lines to silence these monoaminergic neurons have been lacking. We have generated two BAC transgenic mouse lines expressing the outward proton pump, enhanced ArchT3.0 (eArchT3.0), and GFP under control of the regulatory elements of either the dopamine transporter (DAT; Jax# 031663) or the tryptophan hydroxylase 2 (TPH2; Jax# 031662) gene locus. We demonstrate highly faithful and specific expression of these lines in dopaminergic and serotonergic neurons respectively. Additionally we validate effective and sensitive eArchT3.0-mediated silencing of these neurons using slice electrophysiology as well as with a well-established behavioral assay. These new transgenic tools will help expedite the study of dopaminergic and serotonergic system function in normal behavior and disease.


Assuntos
Neurônios Dopaminérgicos/fisiologia , Optogenética , Neurônios Serotoninérgicos/fisiologia , Potenciais de Ação/genética , Animais , Encéfalo/citologia , Channelrhodopsins/genética , Channelrhodopsins/metabolismo , Proteínas da Membrana Plasmática de Transporte de Dopamina/genética , Proteínas da Membrana Plasmática de Transporte de Dopamina/metabolismo , Estimulação Elétrica , Vetores Genéticos/genética , Vetores Genéticos/metabolismo , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Células HEK293 , Humanos , Técnicas In Vitro , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Técnicas de Patch-Clamp , RNA Mensageiro/metabolismo , Transfecção , Triptofano Hidroxilase/genética , Triptofano Hidroxilase/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo
13.
Kidney Blood Press Res ; 44(1): 1-11, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30808844

RESUMO

BACKGROUND/AIMS: Dopamine (DA) is a natriuretic hormone that inhibits renal sodium reabsorption, being Angiotensin II (Ang II) its powerful counterpart. These two systems work together to maintain sodium homeostasis and consequently, the blood pressure (BP) within normal limits. We hypothesized that L-tyrosine (L-tyr) or L-dihydroxyphenylalanine (L-dopa) could inhibit the Na+/K+-ATPase activity. We also evaluated whether L-tyr treatment modulates Tyrosine Hydroxylase (TH). METHODS: Experiments involved cultured LLCPK1 cells treated with L-tyr or L-dopa for 30 minutes a 37°C. In experiments on the effect of Dopa Descarboxylase (DDC) inhibition, cells were pre incubated for 15 minutes with 3-Hydroxybenzylhydrazine dihydrochloride (HBH), and them L-dopa was added for 30 minutes. Na+/K+-ATPase activity was quantified colorimetrically. We used immunoblotting and immunocytochemistry to identify the enzymes TH, DDC and the dopamine receptor D1R in LLCPK1 cells. TH activity was accessed by immunoblotting (increase in the phosphorylation). TH and DDC activities were also evaluated by the modulation of the Na+/K+-ATPase activity, which can be ascribed to the synthesis of dopamine. RESULTS: LLCPK1 cells express the required machinery for DA synthesis: the enzymes TH, and (DDC) as well as its receptor D1R, were detected in control steady state cells. Cells treated with L-tyr or L-dopa showed an inhibition of the basolateral Na+/K+-ATPase activity. We can assume that DA formed in the cytoplasm from L-tyr or L-dopa led to inhibition of the Na+/K+-ATPase activity compared to control. L-tyr treatment increases TH phosphorylation at Ser40 by 100%. HBH, a specific DDC inhibitor; BCH, a LAT2 inhibitor; and Sch 23397, a specific D1R antagonist, totally suppressed the inhibition of Na+/K+-ATPase activity due to L-dopa or L-tyr administration, as indicated in the figures. CONCLUSION: The results indicate that DA formed mainly from luminal L-tyr or L-dopa uptake by LAT2, can inhibit the Na+/K+-ATPase. In addition, our results showed for the very first time that TH activity is also significantly increased when the cells were exposed to L-tyr.


Assuntos
Dopamina/biossíntese , Rim/citologia , Serina/metabolismo , ATPase Trocadora de Sódio-Potássio/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo , Tirosina/farmacologia , Animais , Linhagem Celular , Dopa Descarboxilase , Rim/metabolismo , Fosforilação/efeitos dos fármacos , Receptores de Dopamina D1 , ATPase Trocadora de Sódio-Potássio/antagonistas & inibidores , Suínos , Tirosina 3-Mono-Oxigenase/efeitos dos fármacos
14.
Neuroscience ; 400: 184-195, 2019 02 21.
Artigo em Inglês | MEDLINE | ID: mdl-30599270

RESUMO

Intrauterine growth restriction (IUGR) associates with increased preference for palatable foods and altered insulin sensitivity. Insulin modulates the central dopaminergic response and changes behavioral responses to reward. We measured the release of dopamine in the accumbens during palatable food intake in IUGR rats both at baseline and in response to insulin. From pregnancy day 10 until birth, gestating Sprague-Dawley rats received either an ad libitum (Control), or a 50% food restricted (FR) diet. In adulthood, palatable food consumption and feeding behavior entropy was assessed using an electronic food intake monitor (BioDAQ®), and dopamine response to palatable food was measured by chronoamperometry recordings in the nucleus accumbens (NAcc). FR rats eat more palatable foods during the dark phase, and their eating pattern has a higher entropy compared to control rats. There was a delayed dopamine release in the FR group in response to palatable food and insulin administration reverted this delayed effect. Western blot showed a decrease in suppressor of cytokine signaling 3 protein (SOCS3) in the ventral tegmental area (VTA) and an increase in the ratio of phospho-tyrosine hydroxylase to tyrosine hydroxylase (pTH/TH) in the NAcc of FR rats. Administration of insulin also abolished this latter effect in FR rats. FR rats showed metabolic alterations and a delay in the dopaminergic response to palatable foods. This could explain the increased palatable food intake and behavioral entropy found in FR rats. IUGR may lead to binge eating, obesity and its metabolic consequences by modifying the central dopaminergic response to sweet food.


Assuntos
Dopamina/metabolismo , Ingestão de Alimentos , Retardo do Crescimento Fetal/metabolismo , Insulina/metabolismo , Núcleo Accumbens/metabolismo , Animais , Comportamento Alimentar , Feminino , Privação de Alimentos/fisiologia , Insulina/administração & dosagem , Masculino , Ratos Sprague-Dawley , Proteína 3 Supressora da Sinalização de Citocinas/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo , Área Tegmentar Ventral/metabolismo
15.
Methods Mol Biol ; 1919: 119-128, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30656625

RESUMO

Genetic reporters offer attractive approaches to investigate gene expression, gene function, and spatiotemporal assessment in vitro and in vivo. Tyrosine hydroxylase (TH) is the rate-limiting enzyme for the biosynthesis of the dopamine neurotransmitter, and thus it has been used as a reliable marker for dopaminergic neurons in vitro and in vivo. Herein we describe a method for making iPSC lines with TH-green fluorescent protein reporter gene using CRISPR/Cas9 technique.


Assuntos
Diferenciação Celular , Neurônios Dopaminérgicos/citologia , Neurônios Dopaminérgicos/metabolismo , Expressão Gênica , Genes Reporter , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Sistemas CRISPR-Cas , Técnicas de Cultura de Células , Edição de Genes , Células HEK293 , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Doença de Parkinson/metabolismo , RNA Guia , Tirosina 3-Mono-Oxigenase/genética , Tirosina 3-Mono-Oxigenase/metabolismo
16.
Int J Mol Sci ; 20(2)2019 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-30634592

RESUMO

The ventral tegmental area (VTA), a critical portion of the mesencephalic dopamine system, is thought to be involved in the development and maintenance of addiction. It has been proposed that the dopaminergic regulatory factors TH, Nurr1, and Pitx3 are crucial for determining the survival and maintenance of dopaminergic neurons. Thus, the present study investigated whether abnormalities in these dopaminergic regulatory factors in the VTA were associated with neuronal injury induced by chronic morphine dependence. Rat models with different durations of morphine dependence were established. Thionine staining was used to observe morphological changes in the VTA neurons. Immunohistochemistry and western blot were used to observe changes in the expression of the dopaminergic regulatory proteins TH, Nurr1, and Pitx3. Thionine staining revealed that prolonged morphine dependence resulted in dopaminergic neurons with edema, a lack of Nissl bodies, and pyknosis. Immunohistochemistry showed that the number of TH⁺, Nurr1⁺, and Pitx3⁺ cells, and the number of TH⁺ cells expressing Nurr1 or Pitx3, significantly decreased in the VTA after a long period of morphine dependence. Western blot results were consistent with the immunohistochemistry findings. Chronic morphine exposure resulted in abnormalities in dopaminergic regulatory factors and pathological changes in dopaminergic neurons in the VTA. These results suggest that dysregulation of dopaminergic regulatory factors in the VTA are associated with neuronal injury induced by chronic morphine dependence.


Assuntos
Neurônios Dopaminérgicos/metabolismo , Proteínas de Homeodomínio/metabolismo , Dependência de Morfina/metabolismo , Dependência de Morfina/patologia , Membro 2 do Grupo A da Subfamília 4 de Receptores Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo , Área Tegmentar Ventral/metabolismo , Área Tegmentar Ventral/patologia , Animais , Expressão Gênica , Proteínas de Homeodomínio/genética , Imuno-Histoquímica , Masculino , Dependência de Morfina/genética , Membro 2 do Grupo A da Subfamília 4 de Receptores Nucleares/genética , Ratos , Fatores de Transcrição/genética , Tirosina 3-Mono-Oxigenase/genética
17.
Neuropharmacology ; 144: 219-232, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30366005

RESUMO

Methoxetamine (MXE) is a novel psychoactive substance that can induce several short-term effects on emotional states and behavior. However, little is known about the persistent emotional and behavioral effects of MXE. Moreover, neurotoxic effects of MXE have been hypothesized, but never demonstrated in vivo. To clarify these issues, rats received repeated treatment with MXE every other day (0.1-0.5 mg/kg, i.p., × 5), and 7 days later they were challenged with MXE (0.1-0.5 mg/kg, i.p.). Behavioral effects of MXE were first evaluated by measuring emission of ultrasonic vocalizations and locomotor activity after each administration. Thereafter, persistent behavioral effects of MXE were evaluated, starting 8 days after challenge, through elevated plus maze, spontaneous alternation, novel object recognition, and marble burying tests. After completion of behavioral analysis, neurotoxic effects of MXE were evaluated by measuring densities of dopamine transporter, tyrosine hydroxylase, and serotonin transporter in various brain regions. Repeated treatment and challenge with MXE affected neither calling behavior nor locomotor activity of rats. Conversely, rats previously treated with MXE exhibited behavioral alterations in the elevated plus maze, marble burying and novel object recognition tests, suggestive of increased anxiety and impaired non-spatial memory. Noteworthy, the same rats displayed dopaminergic damage in the medial prefrontal cortex, nucleus accumbens, caudate-putamen, substantia nigra pars compacta, and ventral tegmental area, along with accumbal serotonergic damage. Our findings show for the first time that repeated administration of MXE induces persistent behavioral abnormalities and neurotoxicity in rats, which can help elucidating the risks associated with human MXE consumption.


Assuntos
Comportamento Animal/efeitos dos fármacos , Cicloexanonas/efeitos adversos , Cicloexilaminas/efeitos adversos , Síndromes Neurotóxicas , Neurotoxinas/efeitos adversos , Psicotrópicos/efeitos adversos , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Encéfalo/patologia , Proteínas da Membrana Plasmática de Transporte de Dopamina/metabolismo , Relação Dose-Resposta a Droga , Emoções/efeitos dos fármacos , Masculino , Memória/efeitos dos fármacos , Atividade Motora/efeitos dos fármacos , Síndromes Neurotóxicas/metabolismo , Síndromes Neurotóxicas/patologia , Síndromes Neurotóxicas/psicologia , Proteínas de Ligação a RNA/metabolismo , Ratos Sprague-Dawley , Tirosina 3-Mono-Oxigenase/metabolismo
18.
Acta Histochem ; 121(2): 171-181, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30573341

RESUMO

Lead (Pb) is a metal element released into the atmosphere and a major source of environmental contamination. The accumulation and concentration of this metal in a food web may lead to the intoxication of the body, more precisely, the nervous system (NS). In addition, Pb-exposure can cause structural and functional disruption of the NS. Studies have shown that Pb-exposure could be a risk factor in the development of Parkinson's disease (PD). The latter is related to dopaminergic deficiency that may be triggered by genetic and environmental factors such as Pb intoxication. In this study, we have evaluated, in one hand, the neurotoxic effect of Pb (25 mg / kg B.W i.p) for three consecutive days on dopaminergic system and locomotor performance in Merione shawi. In the other hand, the possible restorative potential of C. sativus (CS) (50 mg / kg BW) by oral gavage. The immunohistochemical approach has revealed that Pb-intoxicated Meriones show a significant increase of Tyrosine Hydroxylase (TH) levels within the Substantia Nigra compacta (SNc), Ventral Tegmental Area (VTA), Locus Coeruleus (LC), Dorsal Striatum (DS) and Medial Forebrain Bundle (MFB), unlike the control meriones, a group intoxicated and treated with Crocus sativus hydroethanolic extract (CSHEE) and treated group by CSHEE. Treatment with CSHEE, has shown a real potential to prevent all Pb-induced damages. In fact, restores the TH levels by 92%, 90%, 88%, 90% and 93% in SNc, VTA, LC, DS and MFB respectively, similarly, locomotor activity dysfunction in Pb-intoxicaed meriones was reinstated by 90%. In this study, we have revealed a new pharmacological potential of Crocus sativus that can be used as a neuroprotective product for neurodegenerative disorders, especially, which implying dopaminergic and noradrenergic injuries, like PD, trigged by heavy metals.


Assuntos
Neurônios Dopaminérgicos/metabolismo , Doença de Parkinson/metabolismo , Parte Compacta da Substância Negra/metabolismo , Área Tegmentar Ventral/metabolismo , Animais , Crocus/metabolismo , Dopamina/metabolismo , Masculino , Feixe Prosencefálico Mediano/metabolismo , Substância Negra/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo
19.
Artigo em Inglês | MEDLINE | ID: mdl-30296469

RESUMO

Methamphetamine, an amphetamine derivative, is a powerful psychomotor stimulant and commonly used drug of abuse. This study examined the effect of binge-like methamphetamine (MA) dosing (4 × 4 mg/kg, s.c., 2 h apart) on regional dopamine and dopaminergic metabolite levels in rat brain at a range of early time points after final dose (2-48 h). Body temperature was elevated when measured 2 h after the last dose. MA increased dopamine levels in the frontal cortex 2 and 24 h after the last dose. The dopamine level was also increased in the amygdala at 24 h. No change was observed in the striatum at any time point, but levels of the dopamine metabolite DOPAC were markedly reduced at 24 and 48 h. Tyrosine hydroxylase expression is induced downstream of dopamine activity, and it is the rate limiting enzyme in dopamine synthesis. The effect of MA binge-like dosing on the volume of tyrosine hydroxylase containing cell bodies and the area fraction of tyrosine hydroxylase containing fibres was also assessed. MA increased the area fraction of tyrosine hydroxylase fibres in the frontal cortex and reduced the volume of tyrosine hydroxylase containing cell bodies 2 h after last dose in the ventral tegmental area and the substantia nigra. An increase in cell body volume in the substantia nigra was observed 48 h after treatment. These findings collectively highlight the importance of the dopaminergic system in methamphetamine induced effects, identify the frontal cortex, amygdala and striatum as key regions that undergo early changes in response to binge-like methamphetamine dosing and provide evidence of time-dependent effects on the cell bodies and fibres of tyrosine hydroxylase expressing neurons.


Assuntos
Encéfalo/efeitos dos fármacos , Estimulantes do Sistema Nervoso Central/administração & dosagem , Dopamina/metabolismo , Metanfetamina/administração & dosagem , Neurônios/efeitos dos fármacos , Tirosina 3-Mono-Oxigenase/metabolismo , Transtornos Relacionados ao Uso de Anfetaminas/metabolismo , Transtornos Relacionados ao Uso de Anfetaminas/patologia , Animais , Temperatura Corporal/efeitos dos fármacos , Encéfalo/metabolismo , Encéfalo/patologia , Expressão Gênica/efeitos dos fármacos , Neurônios/metabolismo , Neurônios/patologia , Ratos Sprague-Dawley , Fatores de Tempo
20.
PLoS One ; 13(12): e0208891, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30586424

RESUMO

Histological evidence points to the presence of dopamine (DA) in the cephalic sensory organs of multiple gastropod molluscs, suggesting a possible sensory role for the neurotransmitter. We investigated the sensory function of DA in the nudipleuran Pleurobranchaea californica, in which the central neural correlates of sensation and foraging behavior have been well characterized. Tyrosine hydroxylase-like immunoreactivity (THli), a signature of the dopamine synthetic pathway, was similar to that found in two other opisthobranchs and two pulmonates previously studied: 1) relatively few (<100) THli neuronal somata were observed in the central ganglia, with those observed found in locations similar to those documented in the other snails but varying in number, and 2) the vast majority of THli somata were located in the peripheral nervous system, were associated with ciliated, putative primary sensory cells, and were highly concentrated in chemotactile sensory organs, giving rise to afferent axons projecting to the central nervous system. We extended these findings by observing that applying a selective D2/D3 receptor antagonist to the chemo- and mechanosensory oral veil-tentacle complex of behaving animals significantly delayed feeding behavior in response to an appetitive stimulus. A D1 blocker had no effect. Recordings of the two major cephalic sensory nerves, the tentacle and large oral veil nerves, in a deganglionated head preparation revealed a decrease of stimulus-evoked activity in the former nerve following application of the same D2/D3 antagonist. Broadly, our results implicate DA in sensation and engender speculation regarding the foraging-based decisions the neurotransmitter may serve in the nervous system of Pleurobranchaea and, by extension, other gastropods.


Assuntos
Dopamina/metabolismo , Sistema Nervoso Periférico/metabolismo , Pleurobranchaea/metabolismo , Células Receptoras Sensoriais/metabolismo , Animais , Agonistas de Dopamina/farmacologia , Antagonistas de Dopamina/farmacologia , Sistema Nervoso Periférico/efeitos dos fármacos , Pleurobranchaea/efeitos dos fármacos , Células Receptoras Sensoriais/efeitos dos fármacos , Tirosina 3-Mono-Oxigenase/metabolismo
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