RESUMO
Traumatic brain injury (TBI) can lead to chronic diseases, including neurodegenerative disorders and epilepsy. The hippocampus, one of the most affected brain region after TBI, plays a critical role in learning and memory and is one of the only two regions in the brain in which new neurons are generated throughout life from neural stem cells (NSC) in the dentate gyrus (DG). These cells migrate into the granular layer where they integrate into the hippocampus circuitry. While increased proliferation of NSC in the hippocampus is known to occur shortly after injury, reduced neuronal maturation and aberrant migration of progenitor cells in the hilus contribute to cognitive and neurological dysfunctions, including epilepsy. Here, we tested the ability of a novel, proprietary non-invasive nano-pulsed laser therapy (NPLT), that combines near-infrared laser light (808 nm) and laser-generated, low-energy optoacoustic waves, to mitigate TBI-driven impairments in neurogenesis and cognitive function in the rat fluid percussion injury model. We show that injured rats treated with NPLT performed significantly better in a hippocampus-dependent cognitive test than did sham rats. In the DG, NPLT significantly decreased TBI-dependent impaired maturation and aberrant migration of neural progenitors, while preventing TBI-induced upregulation of specific microRNAs (miRNAs) in NSC. NPLT did not significantly reduce TBI-induced microglia activation in the hippocampus. Our data strongly suggest that NPLT has the potential to be an effective therapeutic tool for the treatment of TBI-induced cognitive dysfunction and dysregulation of neurogenesis, and point to modulation of miRNAs as a possible mechanism mediating its neuroprotective effects.
Assuntos
Lesões Encefálicas Traumáticas/fisiopatologia , Movimento Celular/fisiologia , Cognição/fisiologia , Hipocampo/fisiopatologia , Terapia a Laser , Células-Tronco Neurais/fisiologia , Animais , Masculino , Memória de Curto Prazo/fisiologia , Atividade Motora/fisiologia , Neurogênese/fisiologia , Ratos , Ratos Sprague-Dawley , Recuperação de Função Fisiológica/fisiologiaRESUMO
With the increasing incidence of traumatic brain injury (TBI) in both civilian and military populations, TBI is now considered a chronic disease; however, few studies have investigated the long-term effects of injury in rodent models of TBI. Shown here are behavioral measures that are well-established in TBI research for times early after injury, such as two weeks, until two months. Some of these methods have previously been used at later times after injury, up to one year, but by very few laboratories. The methods demonstrated here are a short neurological assessment to test reflexes, a Beam-Balance to test balance, a Beam-Walk to test balance and motor coordination, and a working memory version of the Morris water maze that can be sensitive to deficits in reference memory. Male rats were handled and pre-trained to neurological, balance, and motor coordination tests prior to receiving parasagittal fluid percussion injury (FPI) or sham injury. Rats can be tested on the short neurological assessment (neuroscore), the beam-balance, and the Beam-Walk multiple times, while testing on the water maze can only be done once. This difference is because rats can remember the task, thus confounding the results if repeated testing is attempted in the same animal. When testing from one to three days after injury, significant differences are detected in all three non-cognitive tasks. However, differences in the Beam-Walk task were not detectable at later time points (after 3 months). Deficits were detected at 3 months in the Beam-Balance and at 6 months in the neuroscore. Deficits in working memory were detected out to 12 months after injury, and a deficit in a reference memory first appeared at 12 months. Thus, standard behavioral tests can be useful measures of persistent behavioral deficits after FPI.
Assuntos
Comportamento Animal/fisiologia , Lesões Encefálicas Traumáticas/fisiopatologia , Animais , Modelos Animais de Doenças , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Although traumatic brain injury (TBI) is now considered a chronic disease, few studies have investigated the long-term behavioral deficits elicited by a well-established rodent model of injury. Here we evaluate behavioral measures, commonly used in TBI research, to determine which tests are useful for studying long-term effects of brain injury in rats. Male Sprague-Dawley rats were handled and pre-trained to neurological, balance, and motor coordination tests prior to receiving parasagittal fluid-percussion injury (FPI), sham injury, or maintenance as naïve cohorts. Rats underwent neuroscore, beam-balance, and beam-walk tests for 3 days after injury. Subsequently, in separate groups at 3, 6, or 12 months, they were re-tested on the same tasks followed by a working memory version of the Morris water maze. On post-injury days (PIDs) 1-3, significant effects of injury on neuroscore, beam-balance, and beam-walk were observed. Differences in the beam-walk task were not detectable at any of the later time-points. However, deficits persisted in beam-balance out to 3 months and neuroscore out to 6 months. Working memory deficits persisted out to 12 months, at which time a reference memory deficit was also evident. These data suggest that balance and motor coordination recovered more quickly than neurological deficits. Furthermore, while deficits in working memory remained stable over the 12-month period, the late onset of the reference memory deficit points to the progressive nature of the injury, or an age/TBI interaction. In conclusion, standard behavioral tests are useful measures of persistent behavioral deficits after parasagittal FPI and provide evidence that TBI is a chronic condition that can change over time and worsen with age.
Assuntos
Comportamento Animal/fisiologia , Lesões Encefálicas Traumáticas/fisiopatologia , Memória de Curto Prazo/fisiologia , Atividade Motora/fisiologia , Memória Espacial/fisiologia , Animais , Doença Crônica , Modelos Animais de Doenças , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
BACKGROUND: The present study was designed to examine the functional recovery following spinal cord injury (SCI) by adjusting the parameters of impact force and dwell-time using the Infinite Horizon (IH) impactor device. METHODS: Sprague-Dawley rats (225-240 g) were divided into eight injury groups based on force of injury (Kdyn) and dwell time (seconds), indicated as Force-Dwell time: 150-4, 150-3, 150-2, 150-1, 150-0, 200-0, 90-2 and sham controls, respectively. RESULTS: After T10 SCI, higher injury force produced greater spinal cord displacement (P < 0.05) and showed a significant correlation (r = 0.813) between the displacement and the force (P < 0.05). In neuropathic pain-like behavior, the percent of paw withdrawals scores in the hindpaw for the 150-4, 150-3, 150-2, 150-1 and the 200-0 injury groups were significantly lowered compared with sham controls (P < 0.05). The recovery of locomotion had a significant within-subjects effect of time (P < 0.05) and the 150-0 group had increased recovery compared to other groups (P < 0.05). In addition, the 200-0 and the 90-2 recovered significantly better than all the 150 kdyn impact groups that included a dwell-time (P < 0.05). In recovery of spontaneous bladder function, the 150-4 injury group took significantly longer recovery time whereas the 150-0 and the 90-2 groups had the shortest recovery times. CONCLUSIONS: The present study demonstrates SCI parameters optimize development of mechanical allodynia and other pathological outcomes.
RESUMO
Chronic neuropathic pain is a common consequence of spinal cord injury (SCI), develops over time and negatively impacts quality of life, often leading to substance abuse and suicide. Recent evidence has demonstrated that reactive oxygen species (ROS) play a role in contributing to neuropathic pain in SCI animal models. This investigation examines four compounds that reduce ROS and the downstream lipid peroxidation products, apocynin, 4-oxo-tempo, U-83836E, and tirilazad, and tests if these compounds can reduce nocioceptive behaviors in chronic SCI animals. Apocynin and 4-oxo-tempo significantly reduced abnormal mechanical hypersensitivity measured in forelimbs and hindlimbs in a model of chronic SCI-induced neuropathic pain. Thus, compounds that inhibit ROS or lipid peroxidation products can be used to ameliorate chronic neuropathic pain. We propose that the application of compounds that inhibit reactive oxygen species (ROS) and related downstream molecules will also reduce the behavioral measures of chronic neuropathic pain. Injury or trauma to nervous tissue leads to increased concentrations of ROS in the surviving tissue. Further damage from ROS molecules to dorsal lamina neurons leads to membrane excitability, the physiological correlate of chronic pain. Chronic pain is difficult to treat with current analgesics and this research will provide a novel therapy for this disease.
Assuntos
Acetofenonas/uso terapêutico , Ácidos Graxos/uso terapêutico , Hiperalgesia , Neuralgia/complicações , Espécies Reativas de Oxigênio/antagonistas & inibidores , Animais , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Hiperalgesia/tratamento farmacológico , Hiperalgesia/etiologia , Hiperalgesia/metabolismo , Injeções Espinhais , Masculino , Neuralgia/etiologia , Medição da Dor/efeitos dos fármacos , Limiar da Dor/efeitos dos fármacos , Estimulação Física/efeitos adversos , Ratos , Ratos Sprague-Dawley , Traumatismos da Medula Espinal/complicaçõesRESUMO
Mild traumatic brain injury (mTBI), particularly mild "blast type" injuries resulting from improvised exploding devices and many sport-caused injuries to the brain, result in long-term impairment of cognition and behavior. Our central hypothesis is that there are inflammatory consequences to mTBI that persist over time and, in part, are responsible for resultant pathogenesis and clinical outcomes. We used an adaptation (1 atmosphere pressure) of a well-characterized moderate-to-severe brain lateral fluid percussion (LFP) brain injury rat model. Our mild LFP injury resulted in acute increases in interleukin-1α/ß and tumor necrosis factor alpha levels, macrophage/microglial and astrocytic activation, evidence of heightened cellular stress, and blood-brain barrier (BBB) dysfunction that were evident as early as 3-6 h postinjury. Both glial activation and BBB dysfunction persisted for 18 days postinjury.
Assuntos
Concussão Encefálica/patologia , Inflamação/patologia , Animais , Barreira Hematoencefálica/patologia , Encéfalo/patologia , Concussão Encefálica/complicações , Citocinas/análise , Citocinas/biossíntese , Modelos Animais de Doenças , Imunoensaio , Inflamação/etiologia , Masculino , Microscopia Confocal , Atividade Motora/fisiologia , Ratos , Ratos Sprague-DawleyRESUMO
Chronic central neuropathic pain after central nervous system injuries remains refractory to therapeutic interventions. A novel approach would be to target key intracellular signaling proteins that are known to contribute to continued activation by phosphorylation of kinases, transcription factors, and/or receptors that contribute to changes in membrane excitability. We demonstrate that one signaling kinase, calcium/calmodulin-dependent kinase II (CaMKII), is critical in maintaining aberrant dorsal horn neuron hyperexcitability in the neuropathic pain condition after spinal cord injury (SCI). After contusion SCI at spinal level T10, activated CaMKII (phosphorylated, pCaMKII) expression is significantly upregulated in the T7/8 spinal dorsal horn in neurons, but not glial cells, and in oligodendrocytes in the dorsal column in the same rats that displayed at-level mechanical allodynia. Furthermore, identified spinothalamic neurons demonstrated significant increases of pCaMKII after SCI compared to sham-treated control animals. However, neither astrocytes nor microglia showed pCaMKII expression in either sham-treated or SCI rats. To demonstrate causality, treatment of SCI rats with KN-93, which prevents CaMKII activation, significantly attenuated at-level mechanical allodynia and aberrant wide dynamic range neuronal activity evoked by brush, pressure, and pinch stimuli and a graded series of von Frey stimuli, respectively. Persistent CaMKII activation contributes to chronic central neuropathic pain by mechanisms that involve maintained hyperexcitability of wide dynamic range dorsal horn neurons. Furthermore, targeting key signaling proteins is a novel, useful therapeutic strategy for treating chronic central neuropathic pain.
Assuntos
Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Neuralgia/enzimologia , Neuralgia/etiologia , Traumatismos da Medula Espinal/complicações , Potenciais de Ação/efeitos dos fármacos , Análise de Variância , Animais , Benzilaminas/farmacologia , Benzilaminas/uso terapêutico , Antígeno CD11b/metabolismo , Modelos Animais de Doenças , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/uso terapêutico , Regulação da Expressão Gênica/efeitos dos fármacos , Proteína Glial Fibrilar Ácida/metabolismo , Hiperalgesia/tratamento farmacológico , Hiperalgesia/etiologia , Hiperalgesia/metabolismo , Masculino , Medição da Dor , Células do Corno Posterior/efeitos dos fármacos , Células do Corno Posterior/fisiologia , Ratos , Ratos Sprague-Dawley , Medula Espinal/patologia , Estilbamidinas , Sulfonamidas/farmacologia , Sulfonamidas/uso terapêutico , Fatores de TempoRESUMO
Amiloride is a drug approved by the United States Food and Drug Administration, which has shown neuroprotective effects in different neuropathological conditions, including brain injury or brain ischemia, but has not been tested in spinal cord injury (SCI). We tested amiloride's therapeutic potential in a clinically relevant rat model of contusion SCI inflicted at the thoracic segment T10. Rats receiving daily administration of amiloride from 24 h to 35 days after SCI exhibited a significant improvement in hindlimb locomotor ability at 21, 28, and 35 days after injury, when compared to vehicle-treated SCI rats. Rats receiving amiloride treatment also exhibited a significant increase in myelin oligodendrocyte glycoprotein (MOG) levels 35 days after SCI at the site of injury (T10) when compared to vehicle-treated controls, which indicated a partial reverse in the decrease of MOG observed with injury. Our data indicate that higher levels of MOG correlate with improved locomotor recovery after SCI, and that this may explain the beneficial effects of amiloride after SCI. Given that amiloride treatment after SCI caused a significant preservation of myelin levels, and improved locomotor recovery, it should be considered as a possible therapeutic intervention after SCI.
Assuntos
Amilorida/farmacologia , Transtornos Neurológicos da Marcha/tratamento farmacológico , Transtornos Neurológicos da Marcha/fisiopatologia , Fármacos Neuroprotetores/farmacologia , Recuperação de Função Fisiológica/fisiologia , Traumatismos da Medula Espinal/tratamento farmacológico , Traumatismos da Medula Espinal/fisiopatologia , Amilorida/uso terapêutico , Animais , Modelos Animais de Doenças , Transtornos Neurológicos da Marcha/etiologia , Masculino , Fármacos Neuroprotetores/uso terapêutico , Ratos , Ratos Sprague-Dawley , Recuperação de Função Fisiológica/efeitos dos fármacos , Traumatismos da Medula Espinal/etiologiaRESUMO
Elevation of extracellular glutamate contributes to cell death and functional impairments generated by spinal cord injury (SCI), in part through the activation of the neurotoxic cytokine interleukin-1beta (IL-1beta). This study examines the participation of IL-1beta and its regulation by the endogenous interleukin-1 receptor antagonist (IL-1ra) in glutamate toxicity following SCI. Glutamate, glutamatergic agonists and SCI had similar effects on levels of IL-1beta and IL-1ra. Following spinal cord contusion or exposure to elevated glutamate, concentrations of IL-1beta first increased as IL-1ra decreased, and both then changed in the opposite directions. Applying the glutamate agonists NMDA and S-AMPA to the spinal cord caused changes in IL-1beta and IL-1ra levels very similar to those produced by contusion and glutamate. The glutamate antagonists MK801 and NBQX blocked the glutamate-induced changes in IL-1beta and IL-1ra levels. Administering IL-1beta elevated IL-1ra, and administering IL-1ra depressed IL-1beta levels. Infusing IL-beta into the spinal cord impaired locomotion, and infusing IL-1ra improved recovery from glutamate-induced motor impairments. We hypothesize that elevating IL-1ra opposes the damage caused by IL-1beta in SCI by reducing IL-1beta levels as well as by blocking binding of IL-1beta to its receptor. Our results demonstrate that IL-1beta contributes to glutamate damage following SCI; blocking IL-1beta may usefully counteract glutamate toxicity.
Assuntos
Citoproteção/efeitos dos fármacos , Proteína Antagonista do Receptor de Interleucina 1/metabolismo , Interleucina-1beta/metabolismo , Traumatismos da Medula Espinal/fisiopatologia , Animais , Modelos Animais de Doenças , Regulação para Baixo/efeitos dos fármacos , Agonistas de Aminoácidos Excitatórios/toxicidade , Antagonistas de Aminoácidos Excitatórios/farmacologia , Transtornos Neurológicos da Marcha/induzido quimicamente , Transtornos Neurológicos da Marcha/tratamento farmacológico , Ácido Glutâmico/toxicidade , Proteína Antagonista do Receptor de Interleucina 1/efeitos dos fármacos , Proteína Antagonista do Receptor de Interleucina 1/farmacologia , Interleucina-1beta/efeitos dos fármacos , Interleucina-1beta/farmacologia , Masculino , Fármacos Neuroprotetores/farmacologia , Neurotoxinas/toxicidade , Ratos , Ratos Sprague-Dawley , Receptores de Interleucina-1/antagonistas & inibidores , Recuperação de Função Fisiológica/efeitos dos fármacos , Traumatismos da Medula Espinal/induzido quimicamente , Traumatismos da Medula Espinal/tratamento farmacológico , Regulação para Cima/efeitos dos fármacosRESUMO
Recent work regarding chronic central neuropathic pain (CNP) following spinal cord injury (SCI) suggests that activation of key signaling molecules such as members of the mitogen activated protein kinase (MAPK) family play a role in the expression of at-level mechanical allodynia. Previously, we have shown that the development of at-level CNP following moderate spinal cord injury is correlated with increased expression of the activated (and thus phosphorylated) forms of the MAPKs extracellular signal related kinase and p38 MAPK. The current study extends this work by directly examining the role of p38 MAPK in the maintenance of at-level CNP following spinal cord injury. Using a combination of behavioral, immunocytochemical, and electrophysiological measures we demonstrate that increased activation of p38 MAPK occurs in the spinal cord just rostral to the site of injury in rats that develop at-level mechanical allodynia after moderate SCI. Immunocytochemical analyses indicate that the increases in p38 MAPK activation occurred in astrocytes, microglia, and dorsal horn neurons in the spinal cord rostral to the site of injury. Inhibiting the enzymatic activity of p38 MAPK dose dependently reverses the behavioral expression of at-level mechanical allodynia and also decreases the hyperexcitability seen in thoracic dorsal horn neurons after moderate SCI. Taken together, these novel data are the first to demonstrate causality that increased activation of p38 MAPK in multiple cell types play an important role in the maintenance of at-level CNP following spinal cord injury.
Assuntos
Neuralgia/enzimologia , Traumatismos da Medula Espinal/enzimologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Doenças do Sistema Nervoso Central/enzimologia , Doenças do Sistema Nervoso Central/etiologia , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/fisiologia , Imidazóis/farmacologia , Masculino , Neuralgia/etiologia , Dor/enzimologia , Dor/etiologia , Medição da Dor/métodos , Piridinas/farmacologia , Ratos , Ratos Sprague-Dawley , Traumatismos da Medula Espinal/complicações , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidoresRESUMO
Long-term functional impairments due to spinal cord injury (SCI) in the rat result from secondary apoptotic death regulated, in part, by SCI-induced decreases in protein levels of the antiapoptotic protein Bcl-xL. We have shown that exogenous administration of Bcl-xL spares neurons 24 h after SCI. However, long-term effects of chronic application of Bcl-xL have not been characterized. To counteract SCI-induced decreases in Bcl-xL and resulting apoptosis, we used the TAT protein transduction domain fused to the Bcl-xL protein (Tat-Bcl-xL), or its antiapoptotic domain BH4 (Tat-BH4). We used intrathecal delivery of Tat-Bcl-xL, or Tat-BH4, into injured spinal cords for 24 h or 7 days, and apoptosis, neuronal death and locomotor recovery were assessed up to 2 months after injury. Both, Tat-Bcl-xL and Tat-BH4, significantly decreased SCI-induced apoptosis in thoracic segments containing the site of injury (T10) at 24 h or 7 days after SCI. However, the 7-day delivery of Tat-Bcl-xL, or Tat-BH4, also induced a significant impairment of locomotor recovery that lasted beyond the drug delivery time. We found that the 7-day administration of Tat-Bcl-xL, or Tat-BH4, significantly increased non-apoptotic neuronal loss and robustly augmented microglia/macrophage activation. These results indicate that the antiapoptotic treatment targeting Bcl-xL shifts neuronal apoptosis to necrosis, increases the inflammatory response and impairs locomotor recovery. Our results suggest that a combinatorial treatment consisting of antiapoptotic and anti-inflammatory agents may be necessary to achieve tissue preservation and significant improvement in functional recovery after SCI.
Assuntos
Apoptose/fisiologia , Traumatismos da Medula Espinal/fisiopatologia , Traumatismos da Medula Espinal/terapia , Proteína bcl-X/administração & dosagem , Análise de Variância , Animais , Apoptose/efeitos dos fármacos , Comportamento Animal , Antígeno CD11b/metabolismo , Modelos Animais de Doenças , Esquema de Medicação , Ensaio de Imunoadsorção Enzimática/métodos , Regulação da Expressão Gênica/efeitos dos fármacos , Produtos do Gene tat/administração & dosagem , Masculino , Atividade Motora/efeitos dos fármacos , Atividade Motora/fisiologia , Estrutura Terciária de Proteína/fisiologia , Ratos , Ratos Sprague-Dawley , Recuperação de Função Fisiológica/efeitos dos fármacos , Recuperação de Função Fisiológica/fisiologia , Traumatismos da Medula Espinal/patologia , Fatores de TempoRESUMO
Spinal cord injury (SCI) results in a pathophysiology characterized by multiple locomotor and sensory deficits, resulting in altered nociception and hyperalgesia. SCI triggers an early and prolonged inflammatory response, with increased interleukin-1beta levels. Transient changes are observed in subunit populations of the transcription factor nuclear factor-kappaB (NF-kappaB). There were decreases in neuronal c-Rel levels and inverse increases in p65 and p50 levels. There were no changes in neuronal p52 or RelB subunits after SCI at any time point tested. Similarly, SCI had no effect on oligodendroglial levels of any NF-kappaB subunit. There were significant early increases in COX-2 and inducible nitric oxide synthase mRNA and protein levels after SCI. We used synthetic double-stranded "decoy" deoxyoligonucleotides containing selective NF-kappaB protein dimer binding consensus sequences. Decoys targeting the p65/p50 binding site on the COX-2 promoter decreased SCI-induced cell losses, NF-kappaB p65/p50 DNA-binding activity, and COX-2 and iNOS protein levels. NF-kappaB p65/p50 targeted decoys improved early locomotor recovery after moderate but not severe SCI, yet ameliorated SCI-induced hypersensitization after both moderate and severe SCI. To determine whether changes in GABA activity played a role in decreased hypersensitivity after SCI and p65/p50 targeted decoy, we counted gamma-aminobutyric acid (GABA)-containing neurons in laminae 1-3. There were significantly more GABAergic neurons in the p65/p50 targeted decoy-treated group at the level of injury.
Assuntos
Comportamento Animal/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Inflamação/etiologia , Inflamação/metabolismo , Oligodesoxirribonucleotídeos/farmacologia , Traumatismos da Medula Espinal/complicações , Animais , Sítios de Ligação/efeitos dos fármacos , Ciclo-Oxigenase 2/efeitos dos fármacos , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , DNA/metabolismo , Expressão Gênica/efeitos dos fármacos , Masculino , Atividade Motora/efeitos dos fármacos , Subunidade p50 de NF-kappa B/metabolismo , Neurônios/metabolismo , Óxido Nítrico Sintase Tipo II/antagonistas & inibidores , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas c-rel/metabolismo , Ratos , Ratos Sprague-Dawley , Recuperação de Função Fisiológica , Traumatismos da Medula Espinal/genética , Traumatismos da Medula Espinal/fisiopatologia , Fator de Transcrição RelA/metabolismo , Ácido gama-Aminobutírico/metabolismoRESUMO
A neuroprotective factor is shown to be present in mammalian serum. This factor is identified by Western blotting to be serum albumin. The serum factor and albumin both protected cultured spinal cord neurons against the toxicity of glutamate. The inability of K252a, a blocker of the high affinity tyrosine kinase receptor for members of the nerve growth factor family, to block the neuroprotective effect of the serum factor established that the serum factor is not a member of the nerve growth factor family. Post-injury injection of albumin intravenously or into the site of injury immediately after injury both improved significantly locomotor function according to Basso-Beattie-Bresnahan assessment and spontaneous locomotor activity recorded with a photobeam activity system. Albumin has multiple mechanisms whereby it may be neuroprotective, and it is a potentially useful agent for treating neurotraumas.
Assuntos
Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Recuperação de Função Fisiológica/efeitos dos fármacos , Albumina Sérica/farmacologia , Traumatismos da Medula Espinal/tratamento farmacológico , Traumatismos da Medula Espinal/patologia , Animais , Células Cultivadas , Ácido Glutâmico/toxicidade , Masculino , Neurônios/patologia , Neurotoxinas/toxicidade , Ratos , Ratos Sprague-Dawley , Recuperação de Função Fisiológica/fisiologiaRESUMO
Grafted human neural stem cells (hNSCs) may help to alleviate functional deficits resulting from spinal cord injury by bridging gaps, replacing lost neurons or oligodendrocytes, and providing neurotrophic factors. Previously, we showed that primed hNSCs differentiated into cholinergic neurons in an intact spinal cord. In this study, we tested the fate of hNSCs transplanted into a spinal cord T10 contusion injury model. When grafted into injured spinal cords of adult male rats on either the same day or 3 or 9 days after a moderate contusion injury, both primed and unprimed hNSCs survived for 3 months postengraftment only in animals that received grafts at 9 days postinjury. Histological analyses revealed that primed hNSCs tended to survive better and differentiated at higher rates into neurons and oligodendrocytes than did unprimed counterparts. Furthermore, only primed cells gave rise to cholinergic neurons. Animals receiving primed hNSC grafts on the ninth day postcontusion improved trunk stability, as determined by rearing activity measurements 3 months after grafting. This study indicates that human neural stem cell fate determination in vivo is influenced by the predifferentiation stage of stem cells prior to grafting. Furthermore, stem cell-mediated facilitation of functional improvement depends on the timing of transplantation after injury, the grafting sites, and the survival of newly differentiated neurons and oligodendrocytes.
Assuntos
Comportamento Animal/fisiologia , Neurônios/transplante , Traumatismos da Medula Espinal/cirurgia , Transplante de Células-Tronco , Células-Tronco/fisiologia , Animais , Antígeno CD11b , Contagem de Células/métodos , Células Cultivadas , Colina O-Acetiltransferase/metabolismo , Claudinas , Comportamento Exploratório/fisiologia , Feto , Proteína Glial Fibrilar Ácida/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Humanos , Imuno-Histoquímica/métodos , Indóis , Proteína 2 de Membrana Associada ao Lisossomo/metabolismo , Masculino , Atividade Motora/fisiologia , Proteínas do Tecido Nervoso/metabolismo , Neurônios/fisiologia , Ratos , Traumatismos da Medula Espinal/fisiopatologia , Transfecção/métodos , Transplante HeterólogoRESUMO
Rats given moderate spinal cord injury (SCI) display increases in the expression of the activated form of the transcription factor cyclic AMP responsive element binding protein (CREB) in spinal segments of dermatomes corresponding to permanent mechanical allodynia, a model of chronic central neuropathic pain (CNP; (Crown, E.D., Ye, Z., Johnson, K.M., Xu, G.Y., McAdoo, D.J., Westlund, K.N., Hulsebosch, C.E., 2005. Upregulation of the phosphorylated form of CREB in spinothalamic tract cells following spinal cord injury: relation to central neuropathic pain. Neurosci. Lett. 384, 139-144)). Given that not all rats that receive moderate SCI develop CNP, the current study was designed to further analyze changes in persistent CREB activation and in the activation state of upstream intracellular signaling cascades (e.g., mitogen-activated protein kinases [MAPKs]) in populations of rats that receive SCI and weeks later develop CNP and rats that receive SCI but do not develop CNP. The results indicate that activated kinases such as pERK 1/2, p-p38 MAPK, but not pJNK, are upregulated in injured rats that develop CNP as compared to injured rats that fail to develop CNP. In addition, the current results replicated our previous finding that activated CREB is upregulated following SCI, however, only in SCI rats that developed CNP. Taken together, these results indicate that activation of intracellular signaling cascades traditionally associated with long-term potentiation and memory is associated with the expression of chronic CNP following SCI.
Assuntos
Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Expressão Gênica/fisiologia , Hiperestesia/fisiopatologia , Traumatismos da Medula Espinal , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Comportamento Animal , Western Blotting/métodos , Modelos Animais de Doenças , Masculino , Medição da Dor/métodos , Ratos , Ratos Sprague-Dawley , Traumatismos da Medula Espinal/metabolismo , Traumatismos da Medula Espinal/fisiopatologia , Estatística como Assunto , TatoRESUMO
Central neuropathic pain (CNP) is an important problem following spinal cord injury (SCI), because it severely affects the quality of life of SCI patients. As in the patient population, the majority of rats develop significant allodynia (CNP rats) after moderate SCI. However, about 10% of SCI rats do not develop allodynia, or develop significantly less allodynia than CNP rats (non-CNP rats). To identify transcriptional changes underlying CNP development after SCI, we used Affymetrix DNA microarrays and RNAs extracted from the spinal cords of CNP and non-CNP rats. DNA microarry analysis showed significantly increased expression of a number of genes associated with inflammation and astrocytic activation in the spinal cords of rats that developed CNP. For example, mRNA levels of glial fibrilary acidic protein (GFAP) and Aquaporin 4 (AQP4) significantly increased in CNP rats. We also found that GFAP, S100beta and AQP4 protein elevation persisted for at least 9 months throughout contused spinal cords, consistent with the chronic nature of CNP. Thus, we hypothesize that CNP development results, in part, from dysfunctional, chronically "over-activated" astrocytes. Although, it has been shown that activated astrocytes are associated with peripheral neuropathic pain, this has not previously been demonstrated in CNP after SCI.
Assuntos
Dor/metabolismo , Traumatismos da Medula Espinal/metabolismo , Ativação Transcricional/fisiologia , Animais , Western Blotting/métodos , Modelos Animais de Doenças , Imunofluorescência/métodos , Proteína Glial Fibrilar Ácida/genética , Proteína Glial Fibrilar Ácida/metabolismo , Masculino , Microscopia Confocal/métodos , Fatores de Crescimento Neural/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Dor/etiologia , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Subunidade beta da Proteína Ligante de Cálcio S100 , Proteínas S100/metabolismo , Traumatismos da Medula Espinal/complicações , Fatores de TempoRESUMO
Spinal cord injury (SCI) often leads to chronic central pain (CCP) syndromes such as allodynia and hyperalgesia. Although several experimental animal models for CCP studies exist, little is known about the effect of age on the development of CCP following SCI. In this study, we evaluated behavioral responses to mechanical and thermal stimuli following SCI using three different age groups of adult Sprague-Dawley rats: young (40 days), adult (60 days), and middle-age (12 months). SCI was produced by unilateral hemisection of the spinal cord at T13. Behavioral measures of locomotor function were assayed in open field tests and somatosensory function by paw withdrawal frequency (PWF) to innocuous mechanical stimuli and paw withdrawal latency (PWL) to radiant heat stimuli on both the forelimbs and hindlimbs. Prior to hemisection, the PWF was not different between the three groups; however, the PWL of the young group was significantly greater than the adult and middle-age group. After spinal hemisection, spontaneous locomotor recovery occurred more rapidly in young and adult than in middle-age rats. In both forelimbs and hindlimbs, the young group displayed a significant increase in PWF and a significant decrease in PWL compared to presurgical and sham values or values from the adult and middle-age groups. These results indicate that younger rats developed more robust neuropathic behaviors than middle-age rats, indicating that age selection is an important factor in animal models of CCP syndromes following SCI. Additionally, our data suggest that age at the time of injury may be one risk factor in predicting the development of CCP after SCI in people.
Assuntos
Envelhecimento/fisiologia , Hiperalgesia/etiologia , Hiperalgesia/fisiopatologia , Neuralgia/fisiopatologia , Traumatismos da Medula Espinal/complicações , Traumatismos da Medula Espinal/fisiopatologia , Fatores Etários , Animais , Comportamento Animal/fisiologia , Membro Anterior/inervação , Membro Anterior/fisiopatologia , Membro Posterior/inervação , Membro Posterior/fisiopatologia , Masculino , Atividade Motora/fisiologia , Plasticidade Neuronal/fisiologia , Medição da Dor , Estimulação Física , Valor Preditivo dos Testes , Prognóstico , Ratos , Ratos Sprague-Dawley , Recuperação de Função Fisiológica/fisiologia , Fatores de Risco , Medula Espinal/patologia , Medula Espinal/fisiopatologiaRESUMO
We tested the effect of age at the time of spinal cord injury (SCI) on locomotor recovery, in open field tests, and mechanical hyperalgesia, using paw withdrawal frequency (PWF) in response to noxious mechanical stimuli, in male Sprague-Dawley rats after spinal hemisection at T13 in young (40 days), adult (60 days) and middle-age (1 year) groups. Behavioral outcomes were measured weekly for 4 weeks in both SCI and sham groups. Following SCI, the young and adult groups recovered significantly more locomotor function, at a more rapid rate, than did the middle-age group. The PWF of the young group was significantly increased, the adult group was significantly decreased, and the middle-age group showed no significant change in fore- and hindlimbs when compared to other age groups, pre-injury and sham controls. These results support age-dependent behavioral outcomes after SCI.
Assuntos
Hiperalgesia/etiologia , Atividade Motora/fisiologia , Recuperação de Função Fisiológica/fisiologia , Traumatismos da Medula Espinal/fisiopatologia , Fatores Etários , Animais , Comportamento Animal , Membro Anterior/fisiopatologia , Membro Posterior/fisiopatologia , Masculino , Medição da Dor , Ratos , Ratos Sprague-Dawley , Fatores de Tempo , TatoRESUMO
Spinal cord injury (SCI) initiates a cascade of biochemical events that leads to an increase in extracellular excitatory amino acid (EAA) concentrations, which results in glutamate receptor-mediated excitotoxic events. An important division of these glutamate receptors is the metabotropic glutamate receptor (mGluR) class, which is divided into three groups. Of these three groups, group I (mGluR1 and mGluR5) activation can initiate a number of intracellular pathways that lead to increased extracellular EAA concentrations. To evaluate subtypes of group I mGluRs in SCI, we administered AIDA (group I antagonist), LY 367385 (mGluR1 specific antagonist), or MPEP (mGluR5 specific antagonist) by interspinal injection to adult male Sprague-Dawley rats (175-200 g) immediately following injury at T10 with an NYU impactor (12.5-mm drop, 10-g rod, 2 mm in diameter). AIDA- and LY 367385-treated subjects had improved locomotor scores and demonstrated an attenuation in the development of mechanical allodynia as measured by von Frey stimulation of the forelimbs; however, LY 367385 potentiated the development of thermal hyperalgesia. MPEP had no effect on locomotor recovery or mechanical allodynia, but attenuated the development of thermal hyperalgesia. AIDA and LY 367385 treatment resulted in a significant increase in tissue sparing compared to the vehicle-treated group at 4 weeks following SCI. These results suggest that mGluRs play an important role in EAA toxicity and have different acute pathophysiological roles following spinal cord injury.
Assuntos
Benzoatos , Glicina/análogos & derivados , Dor/fisiopatologia , Receptores de Glutamato Metabotrópico/fisiologia , Traumatismos da Medula Espinal/fisiopatologia , Animais , Doença Crônica , Antagonistas de Aminoácidos Excitatórios/farmacologia , Glicina/farmacologia , Hiperalgesia/tratamento farmacológico , Hiperalgesia/fisiopatologia , Locomoção/efeitos dos fármacos , Masculino , Fármacos Neuroprotetores/farmacologia , Neurotoxinas , Dor/tratamento farmacológico , Limiar da Dor/efeitos dos fármacos , Limiar da Dor/fisiologia , Estimulação Física , Piridinas/farmacologia , Ratos , Ratos Sprague-Dawley , Traumatismos da Medula Espinal/tratamento farmacológicoRESUMO
Spinal cord injury (SCI) produces an increase in extracellular excitatory amino acid (EAA) concentrations that results in glutamate receptor-mediated excitotoxic events. An important class of these receptors is the metabotropic glutamate receptors (mGluRs). mGluRs can activate a number of intracellular pathways that increase neuronal excitability and modulate neurotransmission. Group I mGluRs are known to modulate EAA release and the development of chronic central pain (CCP) following SCI; however, the role of group II and III mGluRs remains unclear. To begin evaluating group II and III mGluRs in SCI, we administered the specific agonists for group II, APDC, or group III, L-AP4, by interspinal injection immediately following SCI. Contusion injury was produced at spinal segment T10 with a New York University impactor (12.5-mm drop, 10-g rod 2 mm in diameter) in 30 adult male Sprague-Dawley rats (175-200 g). Evoked and spontaneous behavioral measures of CCP, locomotor recovery, changes in mGluR expression, and amount of spared tissue were examined. Neither APDC nor L-AP4 affected locomotor recovery or the development of thermal hyperalgesia; however, L-AP4 and APDC attenuated changes in mechanical thresholds and changes in exploratory behavior indicative of CCP. APDC- and L-AP4-treated groups had higher expression levels of mGluR2/3 at the epicenter of injury on post contusion day 28; however, there was no difference in the amount of spared tissue between treatment groups. These results demonstrate that treatment with agonists to group II and III mGluRs following SCI affects mechanical responses, exploratory behavior, and mGluR2/3 expression without affecting the amount of tissue spared, suggesting that the level of mGluR expression after SCI may modulate nociceptive responses.