Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 20
Filtrar
1.
Pharmacol Res ; 165: 105434, 2021 03.
Artículo en Inglés | MEDLINE | ID: mdl-33484816

RESUMEN

The dopamine transporter (DAT) is a membrane glycoprotein in dopaminergic neurons, which modulates extracellular and intracellular dopamine levels. DAT is regulated by different presynaptic proteins, including dopamine D2 (D2R) and D3 (D3R) receptors. While D2R signalling enhances DAT activity, some data suggest that D3R has a biphasic effect. However, despite the extensive therapeutic use of D2R/D3R agonists in neuropsychiatric disorders, this phenomenon has been little studied. In order to shed light on this issue, DAT activity, expression and posttranslational modifications were studied in mice and DAT-D3R-transfected HEK cells. Consistent with previous reports, acute treatment with D2R/D3R agonists promoted DAT recruitment to the plasma membrane and an increase in DA uptake. However, when the treatment was prolonged, DA uptake and total striatal DAT protein declined below basal levels. These effects were inhibited in mice by genetic and pharmacological inactivation of D3R, but not D2R, indicating that they are D3R-dependent. No changes were detected in mesostriatal tyrosine hydroxylase (TH) protein expression and midbrain TH and DAT mRNAs, suggesting that the dopaminergic system is intact and DAT is posttranslationally regulated. The use of immunoprecipitation and cell surface biotinylation revealed that DAT is phosphorylated at serine residues, ubiquitinated and released into late endosomes through a PKCß-dependent mechanism. In sum, the results indicate that long-term D3R activation promotes DAT down-regulation, an effect that may underlie neuroprotective and antidepressant actions described for some D2R/D3R agonists.


Asunto(s)
Agonistas de Dopamina/farmacología , Proteínas de Transporte de Dopamina a través de la Membrana Plasmática/metabolismo , Proteína Quinasa C/metabolismo , Proteolisis/efectos de los fármacos , Receptores de Dopamina D3/metabolismo , Ubiquitinación/fisiología , Animales , Relación Dosis-Respuesta a Droga , Esquema de Medicación , Células HEK293 , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Pramipexol/farmacología , Receptores de Dopamina D3/agonistas , Ubiquitinación/efectos de los fármacos
2.
Neurobiol Dis ; 88: 44-54, 2016 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-26777664

RESUMEN

The dopamine (DA) transporter (DAT) is a plasma membrane glycoprotein expressed in dopaminergic (DA-) cells that takes back DA into presynaptic neurons after its release. DAT dysfunction has been involved in different neuro-psychiatric disorders including Parkinson's disease (PD). On the other hand, numerous studies support that the glial cell line-derived neurotrophic factor (GDNF) has a protective effect on DA-cells. However, studies in rodents show that prolonged GDNF over-expression may cause a tyrosine hydroxylase (TH, the limiting enzyme in DA synthesis) decline. The evidence of TH down-regulation suggests that another player in DA handling, DAT, may also be regulated by prolonged GDNF over-expression, and the possibility that this effect is induced at GDNF expression levels lower than those inducing TH down-regulation. This issue was investigated here using intrastriatal injections of a tetracycline-inducible adeno-associated viral vector expressing human GDNF cDNA (AAV-tetON-GDNF) in rats, and doxycycline (DOX; 0.01, 0.03, 0.5 and 3mg/ml) in the drinking water during 5weeks. We found that 3mg/ml DOX promotes an increase in striatal GDNF expression of 12× basal GDNF levels and both DA uptake decrease and TH down-regulation in its native and Ser40 phosphorylated forms. However, 0.5mg/ml DOX promotes a GDNF expression increase of 3× basal GDNF levels with DA uptake decrease but not TH down-regulation. The use of western-blot under non-reducing conditions, co-immunoprecipitation and in situ proximity ligation assay revealed that the DA uptake decrease is associated with the formation of DAT dimers and an increase in DAT-α-synuclein interactions, without changes in total DAT levels or its compartmental distribution. In conclusion, at appropriate GDNF transduction levels, DA uptake is regulated through DAT protein-protein interactions without interfering with DA synthesis.


Asunto(s)
Cuerpo Estriado/metabolismo , Proteínas de Transporte de Dopamina a través de la Membrana Plasmática/metabolismo , Factor Neurotrófico Derivado de la Línea Celular Glial/metabolismo , Tirosina 3-Monooxigenasa/metabolismo , Análisis de Varianza , Animales , Membrana Celular/metabolismo , Cuerpo Estriado/citología , Dopamina/metabolismo , Ensayo de Inmunoadsorción Enzimática , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Inmunoprecipitación , Ligadura , Masculino , Ratas , Ratas Sprague-Dawley , Transducción Genética , Tritio/metabolismo , alfa-Sinucleína/metabolismo
3.
Neurobiol Dis ; 74: 325-35, 2015 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-25511804

RESUMEN

The dopamine (DA) transporter (DAT), a membrane glycoprotein expressed in dopaminergic neurons, clears DA from extracellular space and is regulated by diverse presynaptic proteins like protein kinases, α-synuclein, D2 and D3 autoreceptors. DAT dysfunction is implicated in Parkinson's disease and depression, which are therapeutically treated by dopaminergic D2/D3 receptor (D2/D3R) agonists. It is, then, important to improve our understanding of interactions between D3R and DAT. We show that prolonged administration of pramipexole (0.1mg/kg/day, 6 to 21 days), a preferential D3R agonist, leads to a decrease in DA uptake in mouse striatum that reflects a reduction in DAT affinity for DA in the absence of any change in DAT density or subcellular distribution. The effect of pramipexole was absent in mice with genetically-deleted D3R (D3R(-/-)), yet unaffected in mice genetically deprived of D2R (D2R(-/-)). Pramipexole treatment induced a physical interaction between D3R and DAT, as assessed by co-immunoprecipitation and in situ proximity ligation assay. Furthermore, it promoted the formation of DAT dimers and DAT association with both D2R and α-synuclein, effects that were abolished in D3R(-/-) mice, yet unaffected in D2R(-/-) mice, indicating dependence upon D3R. Collectively, these data suggest that prolonged treatment with dopaminergic D3 agonists provokes a reduction in DA reuptake by dopaminergic neurons related to a hitherto-unsuspected modification of the DAT interactome. These observations provide novel insights into the long-term antiparkinson, antidepressant and additional clinical actions of pramipexole and other D3R agonists.


Asunto(s)
Autorreceptores/metabolismo , Benzotiazoles/farmacología , Cuerpo Estriado/efectos de los fármacos , Agonistas de Dopamina/farmacología , Dopamina/metabolismo , Receptores de Dopamina D3/metabolismo , Animales , Antidepresivos/farmacología , Antiparkinsonianos/farmacología , Cuerpo Estriado/metabolismo , Dimerización , Proteínas de Transporte de Dopamina a través de la Membrana Plasmática/metabolismo , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Pramipexol , Receptores de Dopamina D3/agonistas , Receptores de Dopamina D3/genética , alfa-Sinucleína/metabolismo
4.
Ann Anat ; 250: 152127, 2023 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-37355144

RESUMEN

BACKGROUND: The anterior lobe of the insular cortex (aINS) is a cortical region that has reciprocal connections with limbic centers such as the anterior cingulate cortex, prefrontal cortex, amygdala and nucleus accumbens (NAc). In fact, the aINS has been involved in the integration of autonomic information for emotional and motivational functions. The compulsive consumption of drugs or high-fat foods induces alterations at both behavioural and brain levels. Brain reward circuits are altered in response to continued intake, in particular the dopaminergic projections from the ventral tegmental area (VTA) to the NAc. The aINS has multiple connections with the components of this system. In recent years, efforts have been made to better understand the fundamental role of the aINS in addiction, making it one of the key centres of interest for research into new treatments for addiction. OBJECTIVES: The present work focuses on studying 1.- whether the human aINS expresses orexigenic peptides such as neuropeptide Y (NPY), a peptide known to induce hyperphagia, and which has been implicated in the onset and development of obesity, 2.- the long-term effect of an obesogenic diet on NPY expression in the aINS and NAc of C57BL/6 mice. METHODS: A total of 17 female C57BL/6 J mice were used in this study. Female mice were fed ad libitum with water and, either a standard diet (SD) or a high-fat diet (HFD) to induce obesity. There were seven female mice on the SD and ten on the HFD. The duration of the experiment was 180 days. We also studied 3 human adult brains (1 male and 2 females, mean age 55.7 ± 5.2 years). The morphological study was performed using immunohistochemistry and double immunofluorescence techniques to study the neurochemical profile of NPY neurons of the aINS and NAc of humans and mice. RESULTS: Our morphological analysis demonstrates for the first time the basal expression of NPY in different layers of the human cortex (II, III, IV, V/VI), in a pattern similar to previous studies in other species. Furthermore, we observed an increase in the number of NPY-positive cells and their intracytoplasmic signal in the aINS and NAc of the obese mice subjected to a long-term obesogenic diet. CONCLUSIONS: To our knowledge, this is the first study to show the distribution and expression of NPY in the human INS and how its expression is altered after prolonged treatment with an obesogenic diet in obese mice. Our findings may contribute to the understanding of the pathophysiological mechanisms underlying obesity in regions related to the reward system and associated with uncontrolled intake of high-fat foods, thus facilitating the identification of novel therapeutic targets.


Asunto(s)
Neuropéptido Y , Núcleo Accumbens , Humanos , Ratones , Masculino , Femenino , Animales , Persona de Mediana Edad , Núcleo Accumbens/metabolismo , Ratones Obesos , Corteza Insular , Ratones Endogámicos C57BL , Obesidad/metabolismo , Dieta Alta en Grasa/efectos adversos
5.
Ann Anat ; 250: 152135, 2023 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-37460044

RESUMEN

BACKGROUND: Parkinson's disease (PD) is a neurodegenerative disorder that affects primarily the dopaminergic (DAergic) neurons of the mesostriatal system, among other nuclei of the brain. Although it is considered an idiopathic disease, oxidative stress is believed to be involved in DAergic neuron death and therefore plays an important role in the onset and development of the disease. RAD9B is a paralog of the RAD9 checkpoint, sharing some similar functions related to DNA damage resistance and apoptosis, as well as the ability to form 9-1-1 heterotrimers with RAD1 and HUS1. METHODS: In addition to immunohistochemistry, immunofluorescence and Western-blot analysis, we implemented Quantitative RT-PCR and in situ hybridization techniques. RESULTS: We demonstrated RAD9B expression in rat and human mesencephalic DAergic cells using specific markers. Additionally, we observed significant overexpression of RAD9B mRNA (p<0.01) and protein (p<0.01) in the midbrain 48 h after inducing damage with 150 µg of 6-hydroxydopamine (6-OHDA) injected in a rat model of PD. Regarding protein expression, the increased levels were observed in neurons of the mesostriatal system and returned to normal 5 days post-injury. CONCLUSIONS: This response to a neurotoxin, known to produce oxidative stress specifically on DAergic neurons indicates the potential importance of RAD9B in this highly vulnerable population to cell death. In this model, RAD9B function appears to provide neuroprotection, as the induced lesion resulted in only mild degeneration. This observation highlights the potential of RAD9B checkpoint protein as a valuable target for future therapeutic interventions aimed at promoting neuroprotection.


Asunto(s)
Enfermedad de Parkinson , Animales , Humanos , Ratas , Modelos Animales de Enfermedad , Dopamina/metabolismo , Dopamina/uso terapéutico , Neuronas Dopaminérgicas/metabolismo , Neuronas Dopaminérgicas/patología , Estrés Oxidativo , Oxidopamina/toxicidad , Oxidopamina/metabolismo , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/tratamiento farmacológico , Enfermedad de Parkinson/metabolismo
6.
Brain Res Bull ; 202: 110756, 2023 10 01.
Artículo en Inglés | MEDLINE | ID: mdl-37678442

RESUMEN

The suprachiasmatic nucleus (SCN) is the main region for the regulation of circadian rhythms. Although the SCN contains a heterogeneous neurochemical phenotype with a wide variety of neuropeptides, a key role has been suggested for the vasoactive intestinal neuropeptide (VIP) as a modulator circadian, reproductive, and seasonal rhythms. VIP is a 28-amino acid polypeptide hormone that belongs to the secretin-glucagon peptide superfamily and shares 68 % homology with the pituitary adenylate cyclase-activating polypeptide (PACAP). VIP acts as an endogenous appetite inhibitor in the central nervous system, where it participates in the control of appetite and energy homeostasis. In recent years, significant efforts have been made to better understand the role of VIP in the regulation of appetite/satiety and energy balance. This study aimed to elucidate the long-term effect of an obesogenic diet on the distribution and expression pattern of VIP in the SCN and nucleus accumbens (NAc) of C57BL/6 mice. A total of 15 female C57BL/6J mice were used in this study. Female mice were fed ad libitum with water and, either a standard diet (SD) or a high-fat diet (HFD) to induce obesity. There were 7 female mice on the SD and 8 on the HFD. The duration of the experiment was 365 days. The morphological study was performed using immunohistochemistry and double immunofluorescence techniques to study the neurochemical profile of VIP neurons of the SCN of C57BL/6 mice. Our data show that HFD-fed mice gained weight and showed reduced VIP expression in neurons of the SCN and also in fibres located in the NAc. Moreover, we observed a loss of neuropeptide Y (NPY) expression in fibres surrounding the SCN. Our findings on VIP may contribute to the understanding of the pathophysiological mechanisms underlying obesity in regions associated with uncontrolled intake of high-fat foods and the reward system, thus facilitating the identification of novel therapeutic targets.


Asunto(s)
Dieta Alta en Grasa , Péptido Intestinal Vasoactivo , Femenino , Animales , Ratones , Ratones Endogámicos C57BL , Dieta Alta en Grasa/efectos adversos , Ratones Obesos , Núcleo Supraquiasmático
7.
Am J Physiol Cell Physiol ; 299(4): C779-90, 2010 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-20631247

RESUMEN

The δ-subunit of the epithelial Na(+) channel (ENaC) is expressed in neurons of the human and monkey central nervous system and forms voltage-independent, amiloride-sensitive Na(+) channels when expressed in heterologous systems. It has been proposed that δ-ENaC could affect neuronal excitability and participate in the transduction of ischemic signals during hypoxia or inflammation. The regulation of δ-ENaC activity is poorly understood. ENaC channels in kidney epithelial cells are regulated by the serum- and glucocorticoid-induced kinase 1 (SGK1). Recently, a new isoform of this kinase (SGK1.1) has been described in the central nervous system. Here we show that δ-ENaC isoforms and SGK1.1 are coexpressed in pyramidal neurons of the human and monkey (Macaca fascicularis) cerebral cortex. Coexpression of δßγ-ENaC and SGK1.1 in Xenopus oocytes increases amiloride-sensitive current and channel plasma membrane abundance. The kinase also exerts its effect when δ-subunits are expressed alone, indicating that the process is not dependent on accessory subunits or the presence of PY motifs in the channel. Furthermore, SGK1.1 action depends on its enzymatic activity and binding to phosphatidylinositol(4,5)-bisphosphate. Physiological or pharmacological activation of phospholipase C abrogates SGK1.1 interaction with the plasma membrane and modulation of δ-ENaC. Our data support a physiological role for SGK1.1 in the regulation of δ-ENaC through a pathway that differs from the classical one and suggest that the kinase could serve as an integrator of different signaling pathways converging on the channel.


Asunto(s)
Canales Epiteliales de Sodio/metabolismo , Proteínas Inmediatas-Precoces/metabolismo , Neuronas/enzimología , Isoformas de Proteínas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Transducción de Señal/fisiología , Fosfolipasas de Tipo C/metabolismo , Secuencias de Aminoácidos , Secuencia de Aminoácidos , Animales , Línea Celular , Corteza Cerebral/citología , Canales Epiteliales de Sodio/química , Canales Epiteliales de Sodio/genética , Humanos , Proteínas Inmediatas-Precoces/genética , Macaca fascicularis , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Neuronas/citología , Oocitos/citología , Oocitos/fisiología , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Proteínas Serina-Treonina Quinasas/genética , Alineación de Secuencia , Fosfolipasas de Tipo C/genética , Xenopus laevis
8.
Neurobiol Dis ; 40(3): 518-30, 2010 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-20688165

RESUMEN

The dopamine transporter (DAT) is a transmembrane glycoprotein responsible for dopamine (DA) uptake, which has been shown to be involved in DA-cell degeneration in Parkinson's disease (PD). At the same time, some studies suggest that DAT may be regulated in response to dopaminergic injury. We have investigated the mechanisms underlying DAT regulation after different degrees of dopaminergic lesion. DAT is persistently down-regulated in surviving midbrain DA-neurons after substantial (62%) loss of striatal DA-terminals, and transiently after slight (11%) loss of DA-terminals in rats. Transient DAT down-regulation consisted of a decrease of glycosylated (mature) DAT in the plasma membrane with accumulation of non-glycosylated (immature) DAT in the endoplasmic reticulum-Golgi (ERG) compartment, and recovery of the normal expression pattern 5 days after lesion. DAT redistribution to the ERG was also observed in HEK cells expressing rat DAT exposed to MPP(+), but not after exposure to DAT-unrelated neurotoxins. In contrast to other midbrain DA-cells, those in the ventrolateral region of the substantia nigra do not regulate DAT and degenerate shortly after slight DA-lesion. These data suggest that DAT down-regulation is a post-translational event induced by DA-analogue toxins, consisting of a stop in its glycosylation and trafficking to the plasma membrane. Its persistence after substantial DA-lesion may act as a compensatory mechanism helping maintain striatal DA levels. The fact that neurons which do not regulate DAT die shortly after lesion suggests a relationship between DAT down-regulation and neuroprotection.


Asunto(s)
Adrenérgicos/toxicidad , Proteínas de Transporte de Dopamina a través de la Membrana Plasmática/metabolismo , Regulación de la Expresión Génica , Oxidopamina/toxicidad , Adrenérgicos/administración & dosificación , Animales , Western Blotting , Regulación hacia Abajo , Expresión Génica , Regulación de la Expresión Génica/efectos de los fármacos , Células HEK293 , Humanos , Inmunohistoquímica , Hibridación in Situ , Inyecciones Intraventriculares , Oxidopamina/administración & dosificación , Transporte de Proteínas/efectos de los fármacos , Transporte de Proteínas/fisiología , Ratas , Ratas Sprague-Dawley , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transfección
9.
Autophagy ; 16(7): 1279-1295, 2020 07.
Artículo en Inglés | MEDLINE | ID: mdl-31538542

RESUMEN

Growing evidence shows that autophagy is deficient in neurodegenerative and psychiatric diseases, and that its induction may have beneficial effects in these conditions. However, as autophagy shares signaling pathways with cell death and interferes with protein synthesis, prolonged use of autophagy inducers available nowadays is considered unwise. The search for novel autophagy inducers indicates that DRD2 (dopamine receptor 2)-DRD3 ligands may also activate autophagy, though critical aspects of the action mechanisms and effects of dopamine ligands on autophagy are still unknown. In order to shed light on this issue, DRD2- and DRD3-overexpressing cells and drd2 KO, drd3 KO and wild-type mice were treated with the DRD2-DRD3 agonist pramipexole. The results revealed that pramipexole induces autophagy through MTOR inhibition and a DRD3-dependent but DRD2-independent mechanism. DRD3 activated AMPK followed by inhibitory phosphorylation of RPTOR, MTORC1 and RPS6KB1 inhibition and ULK1 activation. Interestingly, despite RPS6KB1 inhibition, the activity of RPS6 was maintained through activation of the MAPK1/3-RPS6KA pathway, and the activity of MTORC1 kinase target EIF4EBP1 along with protein synthesis and cell viability, were also preserved. This pattern of autophagy through MTORC1 inhibition without suppression of protein synthesis, contrasts with that of direct allosteric and catalytic MTOR inhibitors and opens up new opportunities for G protein-coupled receptor ligands as autophagy inducers in the treatment of neurodegenerative and psychiatric diseases. ABBREVIATIONS: AKT/Protein kinase B: thymoma viral proto-oncogene 1; AMPK: AMP-activated protein kinase; BECN1: beclin 1; EGFP: enhanced green fluorescent protein; EIF4EBP1/4E-BP1: eukaryotic translation initiation factor 4E binding protein 1; GPCR; G protein-coupled receptor; GFP: green fluorescent protein; HEK: human embryonic kidney; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MAP2K/MEK: mitogen-activated protein kinase kinase; MAPK1/ERK2: mitogen-activated protein kinase 1; MAPK3/ERK1: mitogen-activated protein kinase 3; MDA: malonildialdehyde; MTOR: mechanistic target of rapamycin kinase; MTT: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; PPX: pramipexole; RPTOR/raptor: regulatory associated protein of MTOR, complex 1; RPS6: ribosomal protein S6; RPS6KA/p90S6K: ribosomal protein S6 kinase A; RPS6KB1/p70S6K: ribosomal protein S6 kinase B1; SQSTM1/p62: sequestosome 1; ULK1: unc-51 like autophagy activating kinase 1; WT: wild type.


Asunto(s)
Autofagia , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Biosíntesis de Proteínas , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D3/metabolismo , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Autofagia/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Células HEK293 , Humanos , Masculino , Ratones Noqueados , Proteínas Asociadas a Microtúbulos/metabolismo , Modelos Biológicos , Fosforilación/efectos de los fármacos , Pramipexol/farmacología , Biosíntesis de Proteínas/efectos de los fármacos , Proto-Oncogenes Mas , Proteína S6 Ribosómica/metabolismo , Proteína Sequestosoma-1/metabolismo , Transducción de Señal/efectos de los fármacos , Sirolimus/farmacología
10.
Neurobiol Dis ; 36(3): 494-508, 2009 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-19766189

RESUMEN

The dopamine transporter (DAT) is a membrane glycoprotein responsible for dopamine (DA) uptake, which has been involved in the degeneration of DA cells in Parkinson's disease (PD). Given that DAT activity depends on its glycosylation status and membrane expression, and that not all midbrain DA cells show the same susceptibility to degeneration in PD, we have investigated a possible relationship between DAT glycosylation and function and the differential vulnerability of DA cells. Glycosylated DAT expression, DA uptake, and DAT V(max) were significantly higher in terminals of nigrostriatal neurons than in those of mesolimbic neurons. No differences were found in non-glycosylated DAT expression and DAT K(m), and DA uptake differences disappeared after deglycosylation of nigrostriatal synaptosomes. The expression pattern of glycosylated DAT in the human midbrain and striatum showed a close anatomical relationship with DA degeneration in parkinsonian patients. This relationship was confirmed in rodent and monkey models of PD, and in HEK cells expressing the wild-type and a partially deglycosylated DAT form. These results strongly suggest that DAT glycosylation is involved in the differential vulnerability of midbrain DA cells in PD.


Asunto(s)
Proteínas de Transporte de Dopamina a través de la Membrana Plasmática/metabolismo , Dopamina/metabolismo , Mesencéfalo/metabolismo , Neuronas/metabolismo , Enfermedad de Parkinson/metabolismo , Anciano , Animales , Cuerpo Estriado/metabolismo , Modelos Animales de Enfermedad , Femenino , Glicosilación , Humanos , Macaca fascicularis , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Ratas , Ratas Sprague-Dawley , Especificidad de la Especie
11.
J Neural Transm Suppl ; (73): 21-37, 2009.
Artículo en Inglés | MEDLINE | ID: mdl-20411765

RESUMEN

The degeneration of nigral dopaminergic (DA-) neurons is the histopathologic hallmark of Parkinson's disease (PD), but not all nigral DA-cells show the same susceptibility to degeneration. This starts in DA-cells in the ventrolateral and caudal regions of the susbtantia nigra (SN) and progresses to DA-cells in the dorsomedial and rostral regions of the SN and the ventral tegmental area, where many neurons remain intact until the final stages of the disease. This fact indicates a relationship between the topographic distribution of midbrain DA-cells and their differential vulnerability, and the possibility that this differential vulnerability is associated with phenotypic differences between different subpopulations of nigral DA-cells. Studies carried out during the last two decades have contributed to establishing the existence of different compartments of nigral DA-cells according to their neurochemical profile, and a possible relationship between the expression of some factors and the relative vulnerability or resistance of DA-cell subpopulations to degeneration. These aspects are reviewed and discussed here.


Asunto(s)
Dopamina/metabolismo , Regulación de la Expresión Génica/fisiología , Neuronas/fisiología , Fenotipo , Sustancia Negra/citología , Sustancia Negra/fisiología , Animales , Humanos
12.
Exp Neurol ; 299(Pt A): 137-147, 2018 01.
Artículo en Inglés | MEDLINE | ID: mdl-29056363

RESUMEN

Huntington's disease (HD) is a neurodegenerative disorder caused by abnormal expansion of the polyglutamine tract in the huntingtin protein (HTT). The toxicity of mutant HTT (mHTT) is associated with intermediate mHTT soluble oligomers that subsequently form intranuclear inclusions. Thus, interventions promoting the clearance of soluble mHTT are regarded as neuroprotective. Striatal neurons are particularly vulnerable in HD. Their degeneration underlies motor symptoms and striatal atrophy, the anatomical hallmark of HD. Recent studies indicate that autophagy may be activated by dopamine D2 and D3 receptor (D2R/D3R) agonists. Since autophagy plays a central role in the degradation of misfolded proteins, and striatal neurons express D2R and D3R, D2R/D3R agonists may promote the clearance of mHTT in striatal neurons. Here, this hypothesis was tested by treating 8-week old R6/1 mice with the D2R/D3R agonist pramipexole for 4weeks. Pramipexole reduced striatal levels of soluble mHTT and increased the size of intranuclear inclusions in R6/1 mice. Furthermore, striatal DARPP-32 levels and motor functions were recovered. These effects were accompanied by an increase in LC3-II and a decrease in p62 in the striatum. Tollip, a selective adaptor of ubiquitinated polyQ proteins to LC3, was also reduced in the striata of R6/1mice but not in their wild-type littermates. No changes were detected in the cerebral cortex where D3R expression is very low, and behavioral and biochemical effects in the striatum were prevented by a D3R antagonist. The findings indicate that PPX protects striatal neurons by promoting the clearance of soluble mHTT through a D3R-mediated mechanism. The evidence of autophagy markers suggests that autophagy is activated, although it is not efficient at removing all mHTT recruited by the autophagic machinery as indicated by the increase in the size of intranuclear inclusions.


Asunto(s)
Benzotiazoles/uso terapéutico , Agonistas de Dopamina/uso terapéutico , Proteína Huntingtina/genética , Proteína Huntingtina/metabolismo , Neostriado/citología , Neuronas/efectos de los fármacos , Fármacos Neuroprotectores/uso terapéutico , Receptores de Dopamina D3/efectos de los fármacos , Animales , Autofagia , Fosfoproteína 32 Regulada por Dopamina y AMPc/metabolismo , Humanos , Enfermedad de Huntington/tratamiento farmacológico , Enfermedad de Huntington/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Masculino , Ratones , Movimiento , Neostriado/efectos de los fármacos , Neostriado/metabolismo , Pramipexol , Complejo de la Endopetidasa Proteasomal
13.
J Histochem Cytochem ; 54(4): 427-41, 2006 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-16322601

RESUMEN

Nitric oxide synthase (NOS) and interleukin-6 (IL-6) are constitutively expressed in hypothalamic cells. However, phenotypic and functional aspects of these cells remain unknown. We have studied the expression pattern of these two molecules in hypothalamic cells expressing corticotropin-releasing factor (CRF) and arginin-vasopressin (AVP), two major regulatory peptides in the hypothalamus-pituitary system, using immunofluorescence, intracerebroventricular injection of colchicine, and the study in parallel of the labeling pattern of axons in the median eminence. Within AVP cells, we distinguished two different populations: large, intensely stained AVP cells coexpressing IL-6; and large, intensely stained AVP cells coexpressing IL-6 and NOS. Within the CRF cells, we distinguished three different populations: large, intensely stained CRF cells immunonegative for AVP, NOS, and IL-6; large cells weakly stained for CRF and AVP, immunopositive for NOS and immunonegative for IL-6; and small cells intensely stained for CRF and AVP and immunonegative for IL-6 and NOS. In addition, we also found AVP cells containing IL-6 in the suprachiasmatic nucleus. These results suggest that neuronal NOS and IL-6 may be involved in different modulatory processes in hypophysiotropic and non-hypophysiotropic cells.


Asunto(s)
Arginina Vasopresina/metabolismo , Hormona Liberadora de Corticotropina/metabolismo , Hipotálamo/metabolismo , Interleucina-6/biosíntesis , Óxido Nítrico Sintasa de Tipo I/biosíntesis , Animales , Colchicina/administración & dosificación , Colchicina/farmacología , Hipotálamo/anatomía & histología , Inmunohistoquímica , Inyecciones Intraventriculares , Masculino , Eminencia Media/metabolismo , Núcleo Hipotalámico Paraventricular/metabolismo , Ratas , Ratas Sprague-Dawley , Núcleo Supraquiasmático/metabolismo , Núcleo Supraóptico/metabolismo
14.
Front Neuroanat ; 8: 84, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-25206324

RESUMEN

Nowadays it is assumed that besides its roles in neuronal processing, dopamine (DA) is also involved in the regulation of cerebral blood flow. However, studies on the hemodynamic actions of DA have been mainly focused on the cerebral cortex, but the possibility that vessels in deeper brain structures receive dopaminergic axons and the origin of these axons have not been investigated. Bearing in mind the evidence of changes in the blood flow of basal ganglia in Parkinson's disease (PD), and the pivotal role of the dopaminergic mesostriatal pathway in the pathophysiology of this disease, here we studied whether striatal vessels receive inputs from midbrain dopaminergic neurons. The injection of an anterograde neuronal tracer in combination with immunohistochemistry for dopaminergic, vascular and astroglial markers, and dopaminergic lesions, revealed that midbrain dopaminergic axons are in close apposition to striatal vessels and perivascular astrocytes. These axons form dense perivascular plexuses restricted to striatal regions in rats and monkeys. Interestingly, they are intensely immunoreactive for tyrosine hydroxylase (TH) phosphorylated at Ser19 and Ser40 residues. The presence of phosphorylated TH in vessel terminals indicates they are probably the main source of basal TH activity in the striatum, and that after activation of midbrain dopaminergic neurons, DA release onto vessels precedes that onto neurons. Furthermore, the relative weight of this "vascular component" within the mesostriatal pathway suggests that it plays a relevant role in the pathophysiology of PD.

15.
Front Neuroanat ; 4: 140, 2010.
Artículo en Inglés | MEDLINE | ID: mdl-21079748

RESUMEN

The term vulnerability was first associated with the midbrain dopaminergic neurons 85 years ago, before they were identified as monoaminergic neurons, when Foix and Nicolesco (1925) reported the loss of neuromelanin containing neurons in the midbrain of patients with post-encephalitic Parkinson's disease (PD). A few years later, Hassler (1938) showed that degeneration is more intense in the ventral tier of the substantia nigra compacta than in its dorsal tier and the ventral tegmental area (VTA), outlining the concept of differential vulnerability of midbrain dopaminergic (DA-) neurons. Nowadays, we know that other neuronal groups degenerate in PD, but the massive loss of nigral DA-cells is its pathological hallmark, having a pivotal position in the pathophysiology of the disease as it is responsible for the motor symptoms. Data from humans as well as cellular and animal models indicate that DA-cell degeneration is a complex process, probably precipitated by the convergence of different risk factors, mediated by oxidative stress, and involving pathogenic factors arising within the DA-neuron (intrinsic factors), and from its environment and distant interconnected brain regions (extrinsic factors). In light of current data, intrinsic factors seem to be preferentially involved in the first steps of the degenerative process, and extrinsic factors in its progression. A controversial issue is the relative weight of the impairment of common cell functions, such as energy metabolism and proteostasis, and specific dopaminergic functions, such as pacemaking activity and DA handling, in the pathogenesis of DA-cell degeneration. Here we will review the current knowledge about the relevance of these factors at the beginning and during the progression of PD, and in the differential vulnerability of midbrain DA-cells.

16.
Neurobiol Aging ; 30(6): 973-86, 2009 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-17976862

RESUMEN

Several studies report that the striatal dopamine (DA) uptake declines with age, but the underlying mechanisms are still unclear. The use of molecular, biochemical and morphological techniques, and antibodies which detect the glycosylated (80 kDa) and non-glycosylated (50 kDa) DA transporter (DAT) forms in the rat mesostriatal system, reveals that DAT is pre- and post-translationally damaged during aging. In middle age (18 months), the glycosylated DAT form decreases in the plasma membrane of striatal terminals, and the non-glycosylated form is accumulated in the endoplasmic reticulum-Golgi complex. Thereafter, in aged rats (24 months), DAT synthesis is also affected as the decrease in both DATmRNA and total DAT protein levels suggests. However, the evidence of a decrease in both DAT expression in the endosomal (vesicle-enriched) compartment and the phosphorylated DAT fraction from middle age, as well as its compartmental redistribution towards the terminal plasma membrane, with an increase in the membrane DAT/total DAT ratio in striatal synapotosomes, in aged rats, indicate that DA-cells activate compensatory mechanisms directed at maintaining DAT function during normal aging.


Asunto(s)
Envejecimiento/metabolismo , Cuerpo Estriado/metabolismo , Proteínas de Transporte de Dopamina a través de la Membrana Plasmática/metabolismo , Fracciones Subcelulares/metabolismo , Animales , Regulación de la Expresión Génica/fisiología , Masculino , Ratas , Ratas Sprague-Dawley , Distribución Tisular
17.
Neurobiol Aging ; 29(11): 1702-11, 2008 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-17582657

RESUMEN

The vesicular monoamine transporter type 2 (VMAT2) is a transmembrane glycoprotein responsible for the vesicular monoamine uptake in the brain. This function declines in the dopaminergic mesostriatal system during normal aging, but the mechanisms responsible for this deficit are unknown. We investigated possible age-related changes in the expression and subcellular distribution of VMAT2 in the rat mesostriatal system. VMAT2 is constitutively expressed as glycosylated (75 kDa), partially glycosylated (55 kDa) and native (45 kDa) forms, they are all present in both synaptosomal compartments (synaptosomal membrane and synaptic vesicle-enriched fractions) of the striatal terminals in young rats. In aged rats, no changes were found in midbrain VMAT2mRNA and VMAT2 total protein levels in whole striatal extracts. However, its subcellular distribution and glycosylation pattern were severely modified. The three VMAT2 forms virtually disappeared from the synaptic vesicle-enriched fraction, while the 55 kDa form was accumulated in the soluble compartment. These changes may be responsible for the loss of VMAT2 activity during aging and may contribute to the high susceptibility of aged midbrain dopaminergic cells to degeneration.


Asunto(s)
Envejecimiento/metabolismo , Cuerpo Estriado/metabolismo , Fracciones Subcelulares/metabolismo , Sinapsis/metabolismo , Proteínas de Transporte Vesicular de Monoaminas/metabolismo , Envejecimiento/patología , Animales , Glicosilación , Masculino , Ratas , Ratas Sprague-Dawley , Distribución Tisular
18.
Exp Neurol ; 204(1): 147-61, 2007 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-17112516

RESUMEN

The impairment of the mesostriatal dopaminergic system has been considered responsible for motor and affective disturbances associated with aging and a risk factor for Parkinson's disease. However, the basic mechanisms underlying this phenomenon are still unknown. Here we used biochemical, molecular and morphological techniques directed at detecting flaws in the dopamine synthesis route and signs of dopaminergic degeneration in the rat mesostriatal system during normal aging. We found two different age-related processes. One is characterized by a dopa decarboxylase decrease, and involves both the nigrostriatal and mesolimbic compartments, and is responsible for a moderate dopamine loss in the dorsal striatum, where other parameters of dopamine synthesis are not affected. The other is characterized by axonal degeneration with aggregation of phosphorylated forms of tyrosine hydroxylase (TH) and amyloid precursor protein in degenerate terminals, and alpha-synuclein in their original somata. This process is restricted to mesolimbic regions and is responsible for the decline of TH activity and l-dopa levels and the greater decrease in dopamine levels in this compartment. These findings suggest that both the nigrostriatal and the mesolimbic systems are vulnerable to aging, but in contrast to what occurs in Parkinson's disease, the mesolimbic system is more vulnerable to aging than the nigrostriatal one.


Asunto(s)
Envejecimiento/metabolismo , Cuerpo Estriado/metabolismo , Sistema Límbico/metabolismo , Sustancia Negra/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Animales , Axones/patología , Dopa-Decarboxilasa/metabolismo , Dopamina/metabolismo , Levodopa/metabolismo , Sistema Límbico/patología , Masculino , Degeneración Nerviosa/metabolismo , Degeneración Nerviosa/patología , Fosforilación , Ratas , Ratas Sprague-Dawley , Tirosina 3-Monooxigenasa/metabolismo , alfa-Sinucleína/metabolismo
19.
J Neurochem ; 102(4): 1304-15, 2007 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-17472699

RESUMEN

Epithelial sodium channel (ENaC) is a member of the ENaC/degenerin family of amiloride-sensitive, non-voltage gated sodium ion channels. ENaC alpha, beta and gamma subunits are abundantly expressed in epithelial tissues, where they have been well characterized. An ENaC delta subunit has also been described in the human nervous system, although its histological distribution pattern remains unexplored. We have now isolated a novel ENaC delta isoform (delta2) from human brain and studied the expression pattern of both the known (delta1) and the new (delta2) isoforms in the human and monkey telencephalon. ENaC delta2 is produced by a combination of alternative transcription start sites, a frame shift in exon 3 and alternative splicing of exon 4. It forms functional amiloride-sensitive sodium channels when co-expressed with ENaC beta and gamma accessory subunits. Comparison with the classical ENaC channel (alphabetagamma) indicates that the interaction between delta2, beta and gamma is functionally inefficient. Both ENaC delta isoforms are widely expressed in pyramidal cells of the human and monkey cerebral cortex and in different neuronal populations of telencephalic subcortical nuclei, but double-labelling experiments demonstrated a low level of co-localization between isoforms (5-8%), suggesting specific functional roles for each of them.


Asunto(s)
Canales Epiteliales de Sodio/fisiología , Expresión Génica/fisiología , Neuronas/metabolismo , Telencéfalo/citología , Animales , Clonación Molecular/métodos , Femenino , Haplorrinos , Humanos , Hibridación in Situ/métodos , Masculino , Potenciales de la Membrana/fisiología , Microinyecciones/métodos , Persona de Mediana Edad , Datos de Secuencia Molecular , Oocitos , Técnicas de Placa-Clamp/métodos , Isoformas de Proteínas/fisiología , ARN Mensajero/biosíntesis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Xenopus laevis
20.
Eur J Neurosci ; 21(7): 1815-27, 2005 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-15869477

RESUMEN

Glial cell line-derived neurotrophic factor (GDNF) is a member of the transforming growth factor-beta superfamily that when exogenously administrated exerts a potent trophic action on dopaminergic (DA) cells. Although we know a lot about its signalling mechanisms and pharmacological effects, physiological actions of GDNF on the adult brain remain unclear. Here, we have used morphological and molecular techniques, and an experimental model of Parkinson's disease in rats, to investigate whether GDNF constitutively expressed in the adult mesostriatal system plays a neuroprotective role on midbrain DA cells. We found that although all midbrain DA cells express both receptor components of GDNF (GFRalpha1 and Ret), those in the ventral tegmental area (VTA) and rostromedial substantia nigra (SNrm) also contain GDNF but not GDNFmRNA. The levels of GDNFmRNA are significantly higher in the ventral striatum (vSt), the target region of VTA and SNrm cells, than in the dorsal striatum (dSt), the target region of DA cells in the caudoventral substantia nigra (SNcv). After fluoro-gold injection in striatum, VTA and SNrm DA cells show triple labelling for tyrosine hydroxylase, GDNF and fluoro-gold, and after colchicine injection in the lateral ventricle, they become GDNF-immunonegative, suggesting that GDNF in DA somata comes from their striatal target. As DA cells in VTA and SNrm are more resistant than those in SNcv to intracerebroventricular injection of 6-OHDA, as occurs in Parkinson's disease, we can suggest that the fact that they project to vSt, where GDNF expression is significantly higher than in the dSt, is a neuroprotective factor involved in the differential vulnerability of midbrain DA neurons.


Asunto(s)
Cuerpo Estriado/metabolismo , Dopamina/metabolismo , Regulación de la Expresión Génica/fisiología , Mesencéfalo/citología , Factores de Crecimiento Nervioso/metabolismo , Neuronas/metabolismo , Análisis de Varianza , Animales , Recuento de Células/métodos , Colchicina/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Factor Neurotrófico Derivado de la Línea Celular Glial , Receptores del Factor Neurotrófico Derivado de la Línea Celular Glial , Inmunohistoquímica/métodos , Hibridación in Situ/métodos , Masculino , Redes Neurales de la Computación , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-ret , ARN Mensajero/biosíntesis , Ratas , Ratas Sprague-Dawley , Proteínas Tirosina Quinasas Receptoras/genética , Proteínas Tirosina Quinasas Receptoras/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Estilbamidinas/metabolismo , Tirosina 3-Monooxigenasa/metabolismo
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA