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2.
PLoS Biol ; 22(7): e3002687, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38991663

RESUMO

Reactive astrocytes are associated with neuroinflammation and cognitive decline in diverse neuropathologies; however, the underlying mechanisms are unclear. We used optogenetic and chemogenetic tools to identify the crucial roles of the hippocampal CA1 astrocytes in cognitive decline. Our results showed that repeated optogenetic stimulation of the hippocampal CA1 astrocytes induced cognitive impairment in mice and decreased synaptic long-term potentiation (LTP), which was accompanied by the appearance of inflammatory astrocytes. Mechanistic studies conducted using knockout animal models and hippocampal neuronal cultures showed that lipocalin-2 (LCN2), derived from reactive astrocytes, mediated neuroinflammation and induced cognitive impairment by decreasing the LTP through the reduction of neuronal NMDA receptors. Sustained chemogenetic stimulation of hippocampal astrocytes provided similar results. Conversely, these phenomena were attenuated by a metabolic inhibitor of astrocytes. Fiber photometry using GCaMP revealed a high level of hippocampal astrocyte activation in the neuroinflammation model. Our findings suggest that reactive astrocytes in the hippocampus are sufficient and required to induce cognitive decline through LCN2 release and synaptic modulation. This abnormal glial-neuron interaction may contribute to the pathogenesis of cognitive disturbances in neuroinflammation-associated brain conditions.


Assuntos
Astrócitos , Disfunção Cognitiva , Hipocampo , Lipocalina-2 , Potenciação de Longa Duração , Doenças Neuroinflamatórias , Neurônios , Animais , Astrócitos/metabolismo , Astrócitos/patologia , Disfunção Cognitiva/metabolismo , Disfunção Cognitiva/etiologia , Disfunção Cognitiva/patologia , Lipocalina-2/metabolismo , Lipocalina-2/genética , Camundongos , Hipocampo/metabolismo , Hipocampo/patologia , Doenças Neuroinflamatórias/patologia , Doenças Neuroinflamatórias/metabolismo , Neurônios/metabolismo , Neurônios/patologia , Camundongos Knockout , Masculino , Camundongos Endogâmicos C57BL , Receptores de N-Metil-D-Aspartato/metabolismo , Optogenética , Região CA1 Hipocampal/patologia , Região CA1 Hipocampal/metabolismo , Modelos Animais de Doenças
3.
Glia ; 70(10): 1864-1885, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-35638268

RESUMO

Although ATP and/or adenosine derived from astrocytes are known to regulate sleep, the precise mechanisms underlying the somnogenic effects of ATP and adenosine remain unclear. We selectively expressed channelrhodopsin-2 (ChR2), a light-sensitive ion channel, in astrocytes within the ventrolateral preoptic nucleus (VLPO), which is an essential brain nucleus involved in sleep promotion. We then examined the effects of photostimulation of astrocytic ChR2 on neuronal excitability using whole-cell patch-clamp recordings in two functionally distinct types of VLPO neurons: sleep-promoting GABAergic projection neurons and non-sleep-promoting local GABAergic neurons. Optogenetic stimulation of VLPO astrocytes demonstrated opposite outcomes in the two types of VLPO neurons. It led to the inhibition of non-sleep-promoting neurons and excitation of sleep-promoting neurons. These responses were attenuated by blocking of either adenosine A1 receptors or tissue-nonspecific alkaline phosphatase (TNAP). In contrast, exogenous adenosine decreased the excitability of both VLPO neuron populations. Moreover, TNAP was expressed in galanin-negative VLPO neurons, but not in galanin-positive sleep-promoting projection neurons. Taken together, these results suggest that astrocyte-derived ATP is converted into adenosine by TNAP in non-sleep-promoting neurons. In turn, adenosine decreases the excitability of local GABAergic neurons, thereby increasing the excitability of sleep-promoting GABAergic projection neurons. We propose a novel mechanism involving astrocyte-neuron interactions in sleep regulation, wherein endogenous adenosine derived from astrocytes excites sleep-promoting VLPO neurons, and thus decreases neuronal excitability in arousal-related areas of the brain.


Assuntos
Galanina , Área Pré-Óptica , Adenosina/farmacologia , Trifosfato de Adenosina/farmacologia , Astrócitos , Neurônios GABAérgicos , Galanina/farmacologia , Área Pré-Óptica/fisiologia
4.
J Neural Eng ; 18(6)2021 11 19.
Artigo em Inglês | MEDLINE | ID: mdl-34795067

RESUMO

Objective. Neural interfaces are an essential tool to enable the human body to directly communicate with machines such as computers or prosthetic robotic arms. Since invasive electrodes can be located closer to target neurons, they have advantages such as precision in stimulation and high signal-to-noise ratio (SNR) in recording, while they often exhibit unstable performance in long-termin-vivoimplantation because of the tissue damage caused by the electrodes insertion. In the present study, we investigated the electrical functionality of flexible penetrating microelectrode arrays (FPMAs) up to 3 months inin-vivoconditions.Approach. Thein-vivoexperiment was performed by implanting FPMAs in five rats. Thein-vivoimpedance as well as the action potential (AP) amplitude and SNR were analyzed over weeks. Additionally, APs were tracked over time to investigate the possibility of single neuron recording.Main results. It was observed that the FPMAs exhibited dramatic increases in impedance for the first 4 weeks after implantation, accompanied by decreases in AP amplitude. However, the increase/decrease in AP amplitude was always accompanied by the increase/decrease in background noise, resulting in quite consistently maintained SNRs. After 4 weeks of implantation, we observed two distinctive issues regarding long-term implantation, each caused by chronic tissue responses or by the delamination of insulation layer. The results demonstrate that the FPMAs successfully recorded neuronal signals up to 12 weeks, with very stably maintained SNRs, reduced by only 16.1% on average compared to the first recordings, although biological tissue reactions or physical degradation of the FPMA were present.Significance. The fabricated FPMAs successfully recorded intracortical signals for 3 months. The SNR was maintained up to 3 months and the chronic function of FPMA was comparable with other silicon based implantable electrodes.


Assuntos
Neurônios , Silício , Potenciais de Ação/fisiologia , Animais , Eletrodos Implantados , Microeletrodos , Neurônios/fisiologia , Ratos
5.
J Neurosci ; 40(47): 8994-9011, 2020 11 18.
Artigo em Inglês | MEDLINE | ID: mdl-33067363

RESUMO

Although ventrolateral preoptic (VLPO) nucleus is regarded as a center for sleep promotion, the exact mechanisms underlying the sleep regulation are unknown. Here, we used optogenetic tools to identify the key roles of VLPO astrocytes in sleep promotion. Optogenetic stimulation of VLPO astrocytes increased sleep duration in the active phase in naturally sleep-waking adult male rats (n = 6); it also increased the extracellular ATP concentration (n = 3) and c-Fos expression (n = 3-4) in neurons within the VLPO. In vivo microdialysis analyses revealed an increase in the activity of VLPO astrocytes and ATP levels during sleep states (n = 4). Moreover, metabolic inhibition of VLPO astrocytes reduced ATP levels (n = 4) and diminished sleep duration (n = 4). We further show that tissue-nonspecific alkaline phosphatase (TNAP), an ATP-degrading enzyme, plays a key role in mediating the somnogenic effects of ATP released from astrocytes (n = 5). An appropriate sample size for all experiments was based on statistical power calculations. Our results, taken together, indicate that astrocyte-derived ATP may be hydrolyzed into adenosine by TNAP, which may in turn act on VLPO neurons to promote sleep.SIGNIFICANCE STATEMENT Glia have recently been at the forefront of neuroscience research. Emerging evidence illustrates that astrocytes, the most abundant glial cell type, are the functional determinants for fates of neurons and other glial cells in the central nervous system. In this study, we newly identified the pivotal role of hypothalamic ventrolateral preoptic (VLPO) astrocytes in the sleep regulation, and provide novel insights into the mechanisms underlying the astrocyte-mediated sleep regulation.


Assuntos
Astrócitos/fisiologia , Área Pré-Óptica/fisiologia , Sono/fisiologia , Adenosina/metabolismo , Trifosfato de Adenosina/metabolismo , Fosfatase Alcalina/biossíntese , Fosfatase Alcalina/genética , Animais , Citocinas/metabolismo , Masculino , Proteínas de Membrana/biossíntese , Proteínas de Membrana/genética , Neurotransmissores/metabolismo , Optogenética , Técnicas de Patch-Clamp , Estimulação Luminosa , Área Pré-Óptica/citologia , Proteínas Proto-Oncogênicas c-fos/biossíntese , Proteínas Proto-Oncogênicas c-fos/genética , Ratos , Ratos Sprague-Dawley
6.
Glia ; 68(9): 1794-1809, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32077526

RESUMO

Finding causative genetic mutations is important in the diagnosis and treatment of hereditary peripheral neuropathies. This study was conducted to find new genes involved in the pathophysiology of hereditary peripheral neuropathy. We identified a new mutation in the EBP50 gene, which is co-segregated with neuropathic phenotypes, including motor and sensory deficit in a family with Charcot-Marie-Tooth disease. EBP50 is known to be important for the formation of microvilli in epithelial cells, and the discovery of this gene mutation allowed us to study the function of EBP50 in the nervous system. EBP50 was strongly expressed in the nodal and paranodal regions of sciatic nerve fibers, where Schwann cell microvilli contact the axolemma, and at the growth tips of primary Schwann cells. In addition, EBP50 expression was decreased in mouse models of peripheral neuropathy. Knockout mice were used to study EBP50 function in the peripheral nervous system. Interestingly motor function deficit and abnormal histology of nerve fibers were observed in EBP50+/- heterozygous mice at 12 months of age, but not 3 months. in vitro studies using Schwann cells showed that NRG1-induced AKT activation and migration were significantly reduced in cells overexpressing the I325V mutant of EBP50 or cells with knocked-down EBP50 expression. In conclusion, we show for the first time that loss of function due to EBP50 gene deficiency or mutation can cause peripheral neuropathy.


Assuntos
Doença de Charcot-Marie-Tooth , Animais , Doença de Charcot-Marie-Tooth/genética , Camundongos , Camundongos Knockout , Mutação , Nervos Periféricos , Sistema Nervoso Periférico
7.
Materials (Basel) ; 12(3)2019 Feb 12.
Artigo em Inglês | MEDLINE | ID: mdl-30759744

RESUMO

Intracerebral local field potential (LFP) measurements are commonly used to monitor brain activity, providing insight into the flow of information across neural networks. Herein we describe synthesis and application of a neural electrode possessing a nano/micro-scale porous surface topology for improved LFP measurement. Compared with conventional brain electrodes, the porous electrodes demonstrate higher measured amplitudes with lower noise levels.

8.
Clin Psychopharmacol Neurosci ; 17(1): 93-104, 2019 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-30690944

RESUMO

OBJECTIVE: Brain activity is known to be voluntarily controllable by neurofeedback, a kind of electroencephalographic (EEG) operant conditioning. Although its efficacy in clinical effects has been reported, it is yet to be uncovered whether or how a specific band activity is controllable. Here, we examined EEG spectral profiles along with conditioning training of a specific brain activity, theta band (4-8 Hz) amplitude, in rats. METHODS: During training, the experimental group received electrical stimulation to the medial forebrain bundle contingent to suppression of theta activity, while the control group received stimulation non-contingent to its own band activity. RESULTS: In the experimental group, theta activity gradually decreased within the training session, while there was an increase of theta activity in the control group. There was a significant difference in theta activity during the sessions between the two groups. The spectral theta peak, originally located at 7 Hz, shifted further towards higher frequencies in the experimental group. CONCLUSION: Our results showed that an operant conditioning technique could train rats to control their specific EEG activity indirectly, and it may be used as an animal model for studying how neuronal systems work in human neurofeedback.

9.
Korean J Physiol Pharmacol ; 22(6): 649-660, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30402025

RESUMO

Migraine is a neurological disorder characterized by recurrent and disabling severe headaches. Although several anticonvulsant drugs that block voltage-dependent Na+ channels are widely used for migraine, far less is known about the therapeutic actions of carbamazepine on migraine. In the present study, therefore, we characterized the effects of carbamazepine on tetrodotoxin-resistant (TTX-R) Na+ channels in acutely isolated rat dural afferent neurons, which were identified by the fluorescent dye DiI. The TTX-R Na+ currents were measured in medium-sized DiIpositive neurons using the whole-cell patch clamp technique in the voltage-clamp mode. While carbamazepine had little effect on the peak amplitude of transient Na+ currents, it strongly inhibited steady-state currents of transient as well as persistent Na+ currents in a concentration-dependent manner. Carbamazepine had only minor effects on the voltage-activation relationship, the voltage-inactivation relationship, and the use-dependent inhibition of TTX-R Na+ channels. However, carbamazepine changed the inactivation kinetics of TTX-R Na+ channels, significantly accelerating the development of inactivation and delaying the recovery from inactivation. In the current-clamp mode, carbamazepine decreased the number of action potentials without changing the action potential threshold. Given that the sensitization of dural afferent neurons by inflammatory mediators triggers acute migraine headaches and that inflammatory mediators potentiate TTX-R Na+ currents, the present results suggest that carbamazepine may be useful for the treatment of migraine headaches.

10.
Eur J Pharmacol ; 838: 69-77, 2018 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-30194938

RESUMO

Migraine is characterized by recurrent and disabling headaches; therefore, several drugs have been widely prescribed to prevent acute migraine attacks. Amitriptyline, a tricyclic antidepressant, is among the most commonly administered. It is poorly known, however, whether amitriptyline modulates the excitability of dural afferent neurons that transmit pain signals from the dura mater. In this study, the effects of amitriptyline on tetrodotoxin-resistant (TTX-R) Na+ channels were examined in acutely isolated rat dural afferent neurons, which were identified by the fluorescent dye DiI. The TTX-R Na+ currents (INa) were recorded from medium-sized DiI-positive neurons using a whole-cell patch clamp technique. Amitriptyline (3 µM) slightly reduced the peak component of transient INa and induced a marked decrease in the steady-state component of transient TTX-R INa, as well as in the slow ramp-induced TTX-R INa. Our findings suggest that amitriptyline specifically inhibits persistent Na+ currents mediated by TTX-R Na+ channels. While amitriptyline had minor effects on voltage-activation/inactivation, it increased the extent of the use-dependent inhibition of TTX-R Na+ channels. Amitriptyline also affected the inactivation kinetics of TTX-R Na+ channels by significantly accelerating the inactivation of TTX-R Na+ channels and slowing the subsequent recovery. Amitriptyline decreased the number of action potentials by increasing the threshold for their generation. In conclusion, the amitriptyline-mediated diverse modulation of TTX-R Na+ channels would be, at least in part, responsible for its prophylactic efficacy for migraine attacks.


Assuntos
Amitriptilina/farmacologia , Antidepressivos Tricíclicos/farmacologia , Transtornos de Enxaqueca/tratamento farmacológico , Nociceptores/efeitos dos fármacos , Canais de Sódio/efeitos dos fármacos , Amitriptilina/uso terapêutico , Animais , Antidepressivos Tricíclicos/uso terapêutico , Dura-Máter/citologia , Dura-Máter/metabolismo , Masculino , Potenciais da Membrana/efeitos dos fármacos , Transtornos de Enxaqueca/patologia , Nociceptores/metabolismo , Técnicas de Patch-Clamp , Ratos , Ratos Sprague-Dawley , Bloqueadores dos Canais de Sódio/farmacologia , Canais de Sódio/metabolismo , Tetrodotoxina/farmacologia
11.
Glia ; 65(9): 1471-1490, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28581123

RESUMO

Lipocalin-2 (LCN2) has diverse functions in multiple pathophysiological conditions; however, its pathogenic role in vascular dementia (VaD) is unknown. Here, we investigated the role of LCN2 in VaD using rodent models of global cerebral ischemia and hypoperfusion with cognitive impairment and neuroinflammation. Mice subjected to transient bilateral common carotid artery occlusion (tBCCAo) for 50 min showed neuronal death and gliosis in the hippocampus at 7 days post-tBCCAo. LCN2 expression was observed predominantly in the hippocampal astrocytes, whereas its receptor was mainly detected in neurons, microglia, and astrocytes. Furthermore, Lcn2-deficient mice, compared with wild-type animals, showed significantly weaker CA1 neuronal loss, cognitive decline, white matter damage, blood-brain barrier permeability, glial activation, and proinflammatory cytokine production in the hippocampus after tBCCAo. Lcn2 deficiency also attenuated hippocampal neuronal death and cognitive decline at 30 days after unilateral common carotid artery occlusion (UCCAo). Furthermore, intracerebroventricular (i.c.v) injection of recombinant LCN2 protein elicited CA1-neuronal death and a cognitive deficit. Our studies using cultured glia and hippocampal neurons supported the decisive role of LCN2 in hippocampal neurotoxicity and microglial activation, and the role of the HIF-1α-LCN2-VEGFA axis of astrocytes in vascular injury. Additionally, plasma levels of LCN2 were significantly higher in patients with VaD than in the healthy control subjects. These results indicate that hippocampal damage and cognitive impairment are mediated by LCN2 secreted from reactive astrocytes in VaD.


Assuntos
Astrócitos/metabolismo , Disfunção Cognitiva/metabolismo , Demência Vascular/metabolismo , Hipocampo/metabolismo , Lipocalina-2/metabolismo , Animais , Astrócitos/patologia , Biomarcadores/sangue , Células Cultivadas , Cognição/fisiologia , Disfunção Cognitiva/patologia , Demência Vascular/patologia , Modelos Animais de Doenças , Hipocampo/irrigação sanguínea , Hipocampo/patologia , Humanos , Lipocalina-2/administração & dosagem , Lipocalina-2/genética , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microglia/metabolismo , Microglia/patologia , Microvasos/metabolismo , Microvasos/patologia , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo
12.
Cell Rep ; 17(11): 3049-3061, 2016 12 13.
Artigo em Inglês | MEDLINE | ID: mdl-27974216

RESUMO

While glial activation is an integral part of pain pathogenesis, the existence of a causal relationship between glia and pain processing has yet to be demonstrated in vivo. Here, we have investigated whether the activation of spinal astrocytes could directly evoke pain hypersensitivity in vivo via the use of optogenetic techniques. Optogenetic stimulation of channelrhopdopsin-2 (ChR)-expressing spinal astrocytes induced pain hypersensitivity in a reversible and time-dependent manner, which was accompanied by glial activation, NR1 phosphorylation, ATP release, and the production of proalgesic mediators. Photostimulation of ChR2-expressing astrocytes in culture and spinal slices recapitulated in vivo findings, demonstrating the release of proalgesic mediators and electrophysiological disinhibition of spinal projection neurons. These findings deepen our understanding of the role of astrocytes in pain pathogenesis and provide the scientific basis for an astrocyte-oriented pain treatment.


Assuntos
Astrócitos/metabolismo , Hipersensibilidade/genética , Dor/genética , Rodopsina/genética , Trifosfato de Adenosina/metabolismo , Astrócitos/patologia , Regulação da Expressão Gênica , Humanos , Hipersensibilidade/fisiopatologia , Proteínas do Tecido Nervoso/genética , Neuroglia/metabolismo , Neuroglia/fisiologia , Neurônios/metabolismo , Neurônios/patologia , Optogenética , Dor/fisiopatologia , Medula Espinal/metabolismo , Medula Espinal/fisiopatologia
13.
J Neurosci ; 36(20): 5608-22, 2016 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-27194339

RESUMO

UNLABELLED: Lipocalin-2 (LCN2) is a member of the highly heterogeneous secretory protein family of lipocalins and increases in its levels can contribute to neurodegeneration in the adult brain. However, there are no reports on the role of LCN2 in Parkinson's disease (PD). Here, we report for the first time that LCN2 expression is increased in the substantia nigra (SN) of patients with PD. In mouse brains, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treatment for a neurotoxin model of PD significantly upregulated LCN2 expression, mainly in reactive astrocytes in both the SN and striatum. The increased LCN2 levels contributed to neurotoxicity and neuroinflammation, resulting in disruption of the nigrostriatal dopaminergic (DA) projection and abnormal locomotor behaviors, which were ameliorated in LCN2-deficient mice. Similar to the effects of MPTP treatment, LCN2-induced neurotoxicity was also observed in the 6-hydroxydopamine (6-OHDA)-treated animal model of PD. Moreover, treatment with the iron donor ferric citrate (FC) and the iron chelator deferoxamine mesylate (DFO) increased and decreased, respectively, the LCN2-induced neurotoxicity in vivo In addition to the in vivo results, 1-methyl-4-phenylpyridinium (MPP(+))-induced neurotoxicity in cocultures of mesencephalic neurons and astrocytes was reduced by LCN2 gene deficiency in the astrocytes and conditioned media derived from MPP(+)-treated SH-SY5Y neuronal enhanced glial expression of LCN2 in vitro Therefore, our results demonstrate that astrocytic LCN2 upregulation in the lesioned DA system may play a role as a potential pathogenic factor in PD and suggest that inhibition of LCN2 expression or activity may be useful in protecting the nigrostriatal DA system in the adult brain. SIGNIFICANCE STATEMENT: Lipocalin-2 (LCN2), a member of the highly heterogeneous secretory protein family of lipocalins, may contribute to neuroinflammation and neurotoxicity in the brain. However, LCN2 expression and its role in Parkinson's disease (PD) are largely unknown. Here, we report that LCN2 is upregulated in the substantia nigra of patients with PD and neurotoxin-treated animal models of PD. Our results suggest that LCN2 upregulation might be a potential pathogenic mechanism of PD, which would result in disruption of the nigrostriatal dopaminergic system through neurotoxic iron accumulation and neuroinflammation. Therefore, inhibition of LCN2 expression or activity may be useful in protecting the nigrostriatal dopaminergic projection in PD.


Assuntos
Lipocalina-2/metabolismo , Neuroglia/metabolismo , Doença de Parkinson/metabolismo , Regulação para Cima , Idoso , Idoso de 80 Anos ou mais , Animais , Estudos de Casos e Controles , Neurônios Dopaminérgicos/metabolismo , Feminino , Humanos , Lipocalina-2/genética , Intoxicação por MPTP/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Doença de Parkinson/patologia , Substância Negra/citologia , Substância Negra/metabolismo
14.
J Vis Exp ; (107): e53064, 2016 Jan 12.
Artigo em Inglês | MEDLINE | ID: mdl-26862716

RESUMO

Head fixation is a technique of immobilizing animal's head by attaching a head-post on the skull for rigid clamping. Traditional head fixation requires surgical attachment of metallic frames on the skull. The attached frames are then clamped to a stationary platform resulting in immobilization of the head. However, metallic frames for head fixation have been technically difficult to design and implement in general laboratory environment. In this study, we provide a novel head fixation method. Using a custom-made head fixation bar, head mounter is constructed during implantation surgery. After the application of acrylic resin for affixing implants such as electrodes and cannula on the skull, additional resins applied on top of that to build a mold matching to the port of the fixation bar. The molded head mounter serves as a guide rails, investigators conveniently fixate the animal's head by inserting the head mounter into the port of the fixation bar. This method could be easily applicable if implantation surgery using dental acrylics is necessary and might be useful for laboratories that cannot easily fabricate CNC machined metal head-posts.


Assuntos
Resinas Acrílicas , Cabeça/fisiologia , Imobilização/instrumentação , Imobilização/métodos , Animais , Desenho de Equipamento , Próteses e Implantes , Ratos , Crânio
15.
J Biol Chem ; 291(11): 6011-6025, 2016 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-26769971

RESUMO

The dorsal root ganglion (DRG) is a highly vulnerable site in diabetic neuropathy. Under diabetic conditions, the DRG is subjected to tissue ischemia or lower ambient oxygen tension that leads to aberrant metabolic functions. Metabolic dysfunctions have been documented to play a crucial role in the pathogenesis of diverse pain hypersensitivities. However, the contribution of diabetes-induced metabolic dysfunctions in the DRG to the pathogenesis of painful diabetic neuropathy remains ill-explored. In this study, we report that pyruvate dehydrogenase kinases (PDK2 and PDK4), key regulatory enzymes in glucose metabolism, mediate glycolytic metabolic shift in the DRG leading to painful diabetic neuropathy. Streptozotocin-induced diabetes substantially enhanced the expression and activity of the PDKs in the DRG, and the genetic ablation of Pdk2 and Pdk4 attenuated the hyperglycemia-induced pain hypersensitivity. Mechanistically, Pdk2/4 deficiency inhibited the diabetes-induced lactate surge, expression of pain-related ion channels, activation of satellite glial cells, and infiltration of macrophages in the DRG, in addition to reducing central sensitization and neuroinflammation hallmarks in the spinal cord, which probably accounts for the attenuated pain hypersensitivity. Pdk2/4-deficient mice were partly resistant to the diabetes-induced loss of peripheral nerve structure and function. Furthermore, in the experiments using DRG neuron cultures, lactic acid treatment enhanced the expression of the ion channels and compromised cell viability. Finally, the pharmacological inhibition of DRG PDKs or lactic acid production substantially attenuated diabetes-induced pain hypersensitivity. Taken together, PDK2/4 induction and the subsequent lactate surge induce the metabolic shift in the diabetic DRG, thereby contributing to the pathogenesis of painful diabetic neuropathy.


Assuntos
Diabetes Mellitus Experimental/complicações , Neuropatias Diabéticas/metabolismo , Neuropatias Diabéticas/patologia , Gânglios Espinais/metabolismo , Gânglios Espinais/patologia , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Células Cultivadas , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/metabolismo , Neuropatias Diabéticas/genética , Glicólise , Hiperglicemia/complicações , Hiperglicemia/genética , Hiperglicemia/metabolismo , Ácido Láctico/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Serina-Treonina Quinases/genética , Piruvato Desidrogenase Quinase de Transferência de Acetil , Ratos Sprague-Dawley , Nervo Isquiático/metabolismo , Nervo Isquiático/patologia , Regulação para Cima
16.
J Neurosci ; 35(42): 14353-69, 2015 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-26490872

RESUMO

Pyruvate dehydrogenase kinases (PDK1-4) are mitochondrial metabolic regulators that serve as decision makers via modulation of pyruvate dehydrogenase (PDH) activity to convert pyruvate either aerobically to acetyl-CoA or anaerobically to lactate. Metabolic dysregulation and inflammatory processes are two sides of the same coin in several pathophysiological conditions. The lactic acid surge associated with the metabolic shift has been implicated in diverse painful states. In this study, we investigated the role of PDK-PDH-lactic acid axis in the pathogenesis of chronic inflammatory pain. Deficiency of Pdk2 and/or Pdk4 in mice attenuated complete Freund's adjuvant (CFA)-induced pain hypersensitivities. Likewise, Pdk2/4 deficiency attenuated the localized lactic acid surge along with hallmarks of peripheral and central inflammation following intraplantar administration of CFA. In vitro studies supported the role of PDK2/4 as promoters of classical proinflammatory activation of macrophages. Moreover, the pharmacological inhibition of PDKs or lactic acid production diminished CFA-induced inflammation and pain hypersensitivities. Thus, a PDK-PDH-lactic acid axis seems to mediate inflammation-driven chronic pain, establishing a connection between metabolism and inflammatory pain. SIGNIFICANCE STATEMENT: The mitochondrial pyruvate dehydrogenase (PDH) kinases (PDKs) and their substrate PDH orchestrate the conversion of pyruvate either aerobically to acetyl-CoA or anaerobically to lactate. Lactate, the predominant end product of glycolysis, has recently been identified as a signaling molecule for neuron-glia interactions and neuronal plasticity. Pathological metabolic shift and subsequent lactic acid production are thought to play an important role in diverse painful states; however, their contribution to inflammation-driven pain is still to be comprehended. Here, we report that the PDK-PDH-lactic acid axis constitutes a key component of inflammatory pain pathogenesis. Our findings establish an unanticipated link between metabolism and inflammatory pain. This study unlocks a previously ill-explored research avenue for the metabolic control of inflammatory pain pathogenesis.


Assuntos
Inflamação/complicações , Ácido Láctico/metabolismo , Dor/etiologia , Dor/metabolismo , Proteínas Serina-Treonina Quinases/deficiência , Complexo Piruvato Desidrogenase/metabolismo , Animais , Células Cultivadas , Modelos Animais de Doenças , Edema/etiologia , Edema/patologia , Regulação da Expressão Gênica/fisiologia , Hiperalgesia/fisiopatologia , Inflamação/congênito , Macrófagos/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Condução Nervosa/genética , Medição da Dor , Limiar da Dor/fisiologia , Proteínas Serina-Treonina Quinases/genética , Piruvato Desidrogenase Quinase de Transferência de Acetil , Fatores de Tempo
17.
Eur J Pharmacol ; 762: 18-25, 2015 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-25962663

RESUMO

Acid-sensing ion channels (ASICs) are widely expressed in both the peripheral and central nervous system, and contribute to the modulation of central nociceptive transmission under both physiological and pathophysiological conditions. In this study, we characterized the proton-induced membrane currents in acutely isolated rat substantia gelatinosa (SG) neurons of the trigeminal subnucleus caudalis using the whole cell patch-clamp technique. Exposure to acidic conditions (pH<6.5) induced the inward currents in a pH-dependent manner. Amiloride, a general ASIC antagonist, significantly blocked the proton-induced currents in a non-competitive manner. The pH 6.0-induced membrane current (IpH6.0) was greatly attenuated in the Na(+)-free external solution, and the reversal potential of the proton-induced currents was similar to the theoretical Na(+) equilibrium potential. The IpH6.0 was reciprocally potentiated by a lower extracellular Ca(2+) concentration. The modulation of IpH6.0 by divalent cations and other modulators suggests that the proton-induced currents are mediated by multiple types of ASIC subunits, including ASIC1a and ASIC2a. Multi-cell RT-PCR analysis revealed that SG neurons express these subunits. Exposure to a pH 6.0 solution directly depolarized the membrane potential, and generated a burst of action potentials in a current-clamp mode. This acidic pH-induced depolarization was significantly blocked by amiloride. The present results suggest that ASICs expressed on SG neurons play important roles in the regulation of nociceptive transmission from the orofacial tissues.


Assuntos
Fenômenos Eletrofisiológicos/efeitos dos fármacos , Prótons , Substância Gelatinosa/citologia , Substância Gelatinosa/efeitos dos fármacos , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Ácido Araquidônico/farmacologia , Cátions Bivalentes/farmacologia , Relação Dose-Resposta a Droga , Feminino , Ácido Láctico/farmacologia , Masculino , Potenciais da Membrana/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley
18.
Neuropharmacology ; 93: 94-102, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25656480

RESUMO

Purine nucleotides, such as ATP and ADP, activate ionotropic P2X and metabotropic P2Y receptors to regulate neurotransmitter release in the peripheral as well as central nervous system. Here we report another type of ATP-induced presynaptic modulation of glutamate release in rat medullary dorsal horn neurons. Glutamatergic excitatory postsynaptic currents (EPSCs) induced by electrical stimulation of trigeminal tract were recorded from horizontal brain stem slices using a whole-cell patch clamp technique. ATP decreased the amplitude of glutamatergic EPSCs in a reversible and concentration dependent manner and increased the paired-pulse ratio. In addition, ATP reduced the frequency of miniature EPSCs without affecting the current amplitude, suggesting that ATP acts presynaptically to reduce the probability of glutamate release. The ATP-induced decrease in glutamatergic EPSCs was not affected by P2X and P2Y receptor antagonists, but was completely blocked by DPCPX, a selective adenosine A1 receptor antagonist. The ATP-induced decrease in glutamatergic EPSCs was also inhibited by an inhibitor of tissue nonspecific alkaline phosphatase but not by inhibitors of other enzymes such as ecto-nucleoside triphosphate diphosphohydrolases and ecto-5'-nucleotidases. The results suggest that exogenously applied purine nucleotides are rapidly converted to adenosine by specific enzymes, and subsequently act on presynaptic A1 receptors to inhibit glutamate release from primary afferent terminals. This type of modulation mediated by purine nucleotides may play an important role in regulating nociceptive transmission from orofacial tissues.


Assuntos
Potenciais de Ação/efeitos dos fármacos , Trifosfato de Adenosina/farmacologia , Adenosina/metabolismo , Ácido Glutâmico/metabolismo , Células do Corno Posterior/efeitos dos fármacos , Células do Corno Posterior/metabolismo , Difosfato de Adenosina/farmacologia , Trifosfato de Adenosina/análogos & derivados , Animais , Animais Recém-Nascidos , Relação Dose-Resposta a Droga , Estimulação Elétrica , Inibidores Enzimáticos/farmacologia , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Feminino , Masculino , Técnicas de Patch-Clamp , Agonistas do Receptor Purinérgico P2X/farmacologia , Antagonistas do Receptor Purinérgico P2X/farmacologia , Ratos , Ratos Sprague-Dawley
19.
Neurosci Lett ; 591: 192-196, 2015 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-25703221

RESUMO

The roles of persistent Na(+) currents (INaP) in intrinsic membrane properties were examined in rat substantia gelatinosa (SG) neurons of the trigeminal subnucleus caudalis using a conventional whole-cell patch clamp technique. In a voltage-clamp mode, riluzole inhibited the slow voltage ramp-induced INaP but had little effect on the peak amplitude of transient Na(+) currents in SG neurons. In a current-clamp mode, most SG neurons exhibited spontaneous action potentials and tonic firing pattern. Riluzole reduced both spontaneous and elicited action potentials in a concentration-dependent manner. The present results suggest that the riluzole-sensitive INaP plays an important role in the excitability of SG neurons and are thus, likely to contribute to the modulation of nociceptive transmission from the orofacial tissues.


Assuntos
Potenciais de Ação , Neurônios/fisiologia , Substância Gelatinosa/fisiologia , Canais de Sódio Disparados por Voltagem/fisiologia , Animais , Neurônios/efeitos dos fármacos , Técnicas de Patch-Clamp , Ratos Sprague-Dawley , Riluzol/farmacologia
20.
J Clin Neurophysiol ; 31(6): 594-9, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25462148

RESUMO

PURPOSE: In this pilot study, we analyzed relationships between quantitative EEG measurements and clinical parameters in idiopathic normal pressure hydrocephalus patients, along with differences in these quantitative EEG markers between cerebrospinal fluid tap test responders and nonresponders. METHODS: Twenty-six idiopathic normal pressure hydrocephalus patients (9 cerebrospinal fluid tap test responders and 17 cerebrospinal fluid tap test nonresponders) constituted the final group for analysis. The resting EEG was recorded and relative powers were computed for seven frequency bands. RESULTS: Cerebrospinal fluid tap test nonresponders, when compared with responders, showed a statistically significant increase in alpha2 band power at the right frontal and centrotemporal regions. Higher delta2 band powers in the frontal, central, parietal, and occipital regions and lower alpha1 band powers in the right temporal region significantly correlated with poorer cognitive performance. Higher theta1 band powers in the left parietal and occipital regions significantly correlated with gait dysfunction. And higher delta1 band powers in the right frontal regions significantly correlated with urinary disturbance. CONCLUSIONS: Our findings may encourage further research using quantitative EEG in patients with ventriculomegaly as a potential electrophysiological marker for predicting cerebrospinal fluid tap test responders. This study additionally suggests that the delta, theta, and alpha bands are statistically correlated with the severity of symptoms in idiopathic normal pressure hydrocephalus patients.


Assuntos
Ondas Encefálicas , Encéfalo/fisiopatologia , Eletroencefalografia , Hidrocefalia de Pressão Normal/diagnóstico , Pressão Intracraniana , Punção Espinal , Idoso , Derivações do Líquido Cefalorraquidiano , Feminino , Humanos , Hidrocefalia de Pressão Normal/fisiopatologia , Hidrocefalia de Pressão Normal/cirurgia , Masculino , Seleção de Pacientes , Projetos Piloto , Valor Preditivo dos Testes
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