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1.
Proc Natl Acad Sci U S A ; 121(27): e2403777121, 2024 Jul 02.
Artículo en Inglés | MEDLINE | ID: mdl-38916998

RESUMEN

Spinal cord dorsal horn inhibition is critical to the processing of sensory inputs, and its impairment leads to mechanical allodynia. How this decreased inhibition occurs and whether its restoration alleviates allodynic pain are poorly understood. Here, we show that a critical step in the loss of inhibitory tone is the change in the firing pattern of inhibitory parvalbumin (PV)-expressing neurons (PVNs). Our results show that PV, a calcium-binding protein, controls the firing activity of PVNs by enabling them to sustain high-frequency tonic firing patterns. Upon nerve injury, PVNs transition to adaptive firing and decrease their PV expression. Interestingly, decreased PV is necessary and sufficient for the development of mechanical allodynia and the transition of PVNs to adaptive firing. This transition of the firing pattern is due to the recruitment of calcium-activated potassium (SK) channels, and blocking them during chronic pain restores normal tonic firing and alleviates chronic pain. Our findings indicate that PV is essential for controlling the firing pattern of PVNs and for preventing allodynia. Developing approaches to manipulate these mechanisms may lead to different strategies for chronic pain relief.


Asunto(s)
Dolor Crónico , Parvalbúminas , Parvalbúminas/metabolismo , Animales , Dolor Crónico/metabolismo , Dolor Crónico/fisiopatología , Ratones , Neuronas/metabolismo , Neuronas/fisiología , Hiperalgesia/metabolismo , Hiperalgesia/fisiopatología , Masculino , Potenciales de Acción/fisiología , Canales de Potasio de Pequeña Conductancia Activados por el Calcio/metabolismo
2.
J Neurosci ; 43(31): 5608-5622, 2023 08 02.
Artículo en Inglés | MEDLINE | ID: mdl-37451982

RESUMEN

Parvalbumin-expressing interneurons (PVINs) play a crucial role within the dorsal horn of the spinal cord by preventing touch inputs from activating pain circuits. In both male and female mice, nerve injury decreases PVINs' output via mechanisms that are not fully understood. In this study, we show that PVINs from nerve-injured male mice change their firing pattern from tonic to adaptive. To examine the ionic mechanisms responsible for this decreased output, we used a reparametrized Hodgkin-Huxley type model of PVINs, which predicted (1) the firing pattern transition is because of an increased contribution of small conductance calcium-activated potassium (SK) channels, enabled by (2) impairment in intracellular calcium buffering systems. Analyzing the dynamics of the Hodgkin-Huxley type model further demonstrated that a generalized Hopf bifurcation differentiates the two types of state transitions observed in the transient firing of PVINs. Importantly, this predicted mechanism holds true when we embed the PVIN model within the neuronal circuit model of the spinal dorsal horn. To experimentally validate this hypothesized mechanism, we used pharmacological modulators of SK channels and demonstrated that (1) tonic firing PVINs from naive male mice become adaptive when exposed to an SK channel activator, and (2) adapting PVINs from nerve-injured male mice return to tonic firing on SK channel blockade. Our work provides important insights into the cellular mechanism underlying the decreased output of PVINs in the spinal dorsal horn after nerve injury and highlights potential pharmacological targets for new and effective treatment approaches to neuropathic pain.SIGNIFICANCE STATEMENT Parvalbumin-expressing interneurons (PVINs) exert crucial inhibitory control over Aß fiber-mediated nociceptive pathways at the spinal dorsal horn. The loss of their inhibitory tone leads to neuropathic symptoms, such as mechanical allodynia, via mechanisms that are not fully understood. This study identifies the reduced intrinsic excitability of PVINs as a potential cause for their decreased inhibitory output in nerve-injured condition. Combining computational and experimental approaches, we predict a calcium-dependent mechanism that modulates PVINs' electrical activity following nerve injury: a depletion of cytosolic calcium buffer allows for the rapid accumulation of intracellular calcium through the active membranes, which in turn potentiates SK channels and impedes spike generation. Our results therefore pinpoint SK channels as potential therapeutic targets for treating neuropathic symptoms.


Asunto(s)
Calcio , Neuralgia , Ratones , Masculino , Femenino , Animales , Parvalbúminas/metabolismo , Neuralgia/metabolismo , Interneuronas/fisiología , Asta Dorsal de la Médula Espinal/metabolismo
3.
Int J Toxicol ; 39(6): 530-541, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33063577

RESUMEN

INTRODUCTION: Corrected QT (QTc) interval is an essential proarrhythmic risk biomarker, but recent data have identified limitations to its use. The J to T-peak (JTp) interval is an alternative biomarker for evaluating drug-induced proarrhythmic risk. The aim of this study was to evaluate pharmacological effects using spatial magnitude leads and DII electrocardiogram (ECG) leads and common ECG confounders (ie, stress and body temperature changes) on covariate adjusted QT (QTca), covariate adjusted JTp (JTpca), and covariate adjusted T-peak to T-end (Tpeca) intervals. METHODS: Beagle dogs were exposed to body hyper- (42 °C) or hypothermic (33 °C) conditions or were administered epinephrine to assess confounding effects on heart rate corrected QTca, JTpca, and Tpeca intervals. Dofetilide (0.1, 0.3, 1.0 mg/kg), ranolazine (100, 140, 200 mg/kg), and verapamil (7, 15, 30, 43, 62.5 mg/kg) were administered to evaluate pharmacological effects. RESULTS: Covariate adjusted QT (slope -12.57 ms/°C) and JTpca (-14.79 ms/°C) were negatively correlated with body temperature but Tpeca was minimally affected. Epinephrine was associated with QTca and JTpca shortening, which could be related to undercorrection in the presence of tachycardia, while minimal effects were observed for Tpeca. There were no significant ECG change following ranolazine administration. Verapamil decreased QTca and JTpca intervals and increased Tpeca, whereas dofetilide increased QTca and JTpca intervals but had inconsistent effects on Tpeca. CONCLUSION: Results highlight potential confounders on QTc interval, but also on JTpca and Tpeca intervals in nonclinical studies. These potential confounding effects may be relevant to the interpretation of ECG data obtained from nonclinical drug safety studies with Beagle dogs.


Asunto(s)
Arritmias Cardíacas/etiología , Epinefrina/farmacología , Fenetilaminas/farmacología , Ranolazina/farmacología , Sulfonamidas/farmacología , Verapamilo/farmacología , Animales , Antiarrítmicos/administración & dosificación , Antiarrítmicos/farmacología , Arritmias Cardíacas/prevención & control , Biomarcadores , Temperatura Corporal , Perros , Relación Dosis-Respuesta a Droga , Electrocardiografía , Femenino , Frecuencia Cardíaca , Masculino , Fenetilaminas/administración & dosificación , Ranolazina/administración & dosificación , Estrés Fisiológico/efectos de los fármacos , Sulfonamidas/administración & dosificación , Verapamilo/administración & dosificación
4.
J Neurosci ; 37(26): 6277-6288, 2017 06 28.
Artículo en Inglés | MEDLINE | ID: mdl-28550169

RESUMEN

The NMDAR is thought to play a key role in the refinement of connectivity in developing neural circuits. Pharmacological blockade or genetic loss-of-function manipulations that prevent NMDAR function during development result in the disorganization of topographic axonal projections. However, because NMDARs contribute to overall glutamatergic neurotransmission, such loss-of-function experiments fail to adequately distinguish between the roles played by NMDARs and neural activity in general. The gliotransmitter d-serine is a coagonist of the NMDAR that is required for NMDAR channel opening, but which cannot mediate neurotransmission on its own. Here we demonstrate that acute administration of d-serine has no immediate effect on glutamate release or AMPA-mediated neurotransmission. We show that endogenous d-serine is normally present below saturating levels in the developing visual system of the Xenopus tadpole. Using an amperometric enzymatic biosensor, we demonstrate that glutamatergic activation elevates ambient endogenous d-serine levels in the optic tectum. Chronically elevating levels of d-serine promoted synaptic maturation and resulted in the hyperstabilization of developing axon branches in the tadpole visual system. Conversely, treatment with an enzyme that degrades endogenous d-serine resulted in impaired synaptic maturation. Despite the reduction in axon arbor complexity seen in d-serine-treated animals, tectal neuron visual receptive fields were expanded, suggesting a failure to prune divergent retinal inputs. Together, these findings positively implicate NMDAR-mediated neurotransmission in developmental synapse maturation and the stabilization of axonal inputs and reveal a potential role for d-serine as an endogenous modulator of circuit refinement.SIGNIFICANCE STATEMENT Activation of NMDARs is critical for the activity-dependent development and maintenance of highly organized topographic maps. d-Serine, a coagonist of the NMDAR, plays a significant role in modulating NMDAR-mediated synaptic transmission and plasticity in many brain areas. However, it remains unknown whether d-serine participates in the establishment of precise neuronal connections during development. Using an in vivo model, we show that glutamate receptor activation can evoke endogenous d-serine release, which promotes glutamatergic synapse maturation and stabilizes axonal structural and functional inputs. These results reveal a pivotal modulatory role for d-serine in neurodevelopment.


Asunto(s)
Axones/fisiología , Neurogénesis/fisiología , Neuroglía/fisiología , Receptores de N-Metil-D-Aspartato/metabolismo , Serina/metabolismo , Sinapsis/fisiología , Animales , Neurotransmisores/metabolismo , Xenopus laevis
5.
J Neurosci ; 35(26): 9676-88, 2015 Jul 01.
Artículo en Inglés | MEDLINE | ID: mdl-26134650

RESUMEN

Activity-dependent strengthening of central synapses is a key factor driving neuronal circuit behavior in the vertebrate CNS. At fast inhibitory synapses, strengthening is thought to occur by increasing the number of GABAA receptors (GABARs) of the same subunit composition to preexisting synapses. Here, we show that strengthening of mouse cerebellar granule cell GABAergic synapses occurs by a different mechanism. Specifically, we show that the neuropeptide hormone, insulin, strengthens inhibitory synapses by recruiting α6-containing GABARs rather than accumulating more α1-containing receptors that are resident to the synapse. Because α6-receptors are targeted to functionally distinct postsynaptic sites from α1-receptors, we conclude that only a subset of all inhibitory synapses are strengthened. Together with our recent findings on stellate cells, we propose a general mechanism by which mature inhibitory synapses are strengthened. In this scenario, α1-GABARs resident to inhibitory synapses form the hardwiring of neuronal circuits with receptors of a different composition fulfilling a fundamental, but unappreciated, role in synapse strengthening.


Asunto(s)
Cerebelo/citología , Hipoglucemiantes/farmacología , Potenciales Postsinápticos Inhibidores/efectos de los fármacos , Insulina/farmacología , Neuronas/efectos de los fármacos , Receptores de GABA-A/metabolismo , 2-Amino-5-fosfonovalerato/farmacología , 6-Ciano 7-nitroquinoxalina 2,3-diona/farmacología , Animales , Animales Recién Nacidos , Antagonistas de Aminoácidos Excitadores/farmacología , Furosemida/farmacología , Técnicas In Vitro , Potenciales Postsinápticos Inhibidores/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Subunidades de Proteína/genética , Subunidades de Proteína/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Receptores de GABA-A/genética , Inhibidores del Simportador de Cloruro Sódico y Cloruro Potásico/farmacología , Sinapsis/efectos de los fármacos , Sinapsis/genética , Transmisión Sináptica/efectos de los fármacos , Transmisión Sináptica/genética , Factores de Tiempo , Ácido gamma-Aminobutírico/farmacología
6.
Cell Rep Methods ; 4(6): 100791, 2024 Jun 17.
Artículo en Inglés | MEDLINE | ID: mdl-38848714

RESUMEN

Characterizing neurons by their electrophysiological phenotypes is essential for understanding the neural basis of behavioral and cognitive functions. Technological developments have enabled the collection of hundreds of neural recordings; this calls for new tools capable of performing feature extraction efficiently. To address the urgent need for a powerful and accessible tool, we developed ElecFeX, an open-source MATLAB-based toolbox that (1) has an intuitive graphical user interface, (2) provides customizable measurements for a wide range of electrophysiological features, (3) processes large-size datasets effortlessly via batch analysis, and (4) yields formatted output for further analysis. We implemented ElecFeX on a diverse set of neural recordings; demonstrated its functionality, versatility, and efficiency in capturing electrical features; and established its significance in distinguishing neuronal subgroups across brain regions and species. ElecFeX is thus presented as a user-friendly toolbox to benefit the neuroscience community by minimizing the time required for extracting features from their electrophysiological datasets.


Asunto(s)
Fenómenos Electrofisiológicos , Análisis de la Célula Individual , Programas Informáticos , Fenómenos Electrofisiológicos/fisiología , Animales , Análisis de la Célula Individual/métodos , Neuronas/fisiología , Humanos , Encéfalo/fisiología , Ratones , Ratas
7.
Neuron ; 111(1): 8-9, 2023 01 04.
Artículo en Inglés | MEDLINE | ID: mdl-36603551

RESUMEN

Spinal cord circuits that process cold inputs from the periphery are poorly understood. In this issue of Neuron, Albisetti et al.1 identify a subset of inhibitory interneurons essential to this function.


Asunto(s)
Interneuronas , Médula Espinal , Médula Espinal/fisiología , Interneuronas/fisiología
8.
J Pharmacol Toxicol Methods ; 121: 107266, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-36963703

RESUMEN

INTRODUCTION: Characterization of the incidence of spontaneous arrhythmias to identify possible drug-related effects is often an important part of the analysis in safety pharmacology studies using telemetry. METHODS: A retrospective analysis in non-clinical species with and without telemetry transmitters was conducted. Electrocardiograms (24 h) from male and female beagle dogs (n = 131), Göttingen minipigs (n = 108) and cynomolgus non-human primates (NHP; n = 78) were analyzed. RESULTS: Ventricular tachycardia (VT) was observed in 3% of the dogs but was absent in minipigs and NHPs. Ventricular fibrillation (VF) was not observed in the 3 species. Ventricular premature beats (VPBs) were more frequent during daytime and atrioventricular blocks (AVBs) were more frequent at night in all species. A limited number of animals exhibited a high arrhythmia frequency and there was no correlation between animals with higher frequency of an arrhythmia type and the frequency of other arrythmias in the same animals. Clinical chemistry or hematology parameters were not different with or without telemetry devices. NHP with a transmural left ventricular pressure (LVP) catheter exhibited a greater incidence of VPBs and PJCs compared to telemetry animals without LVP. DISCUSSION: All species were similar with regards to the frequency of ventricular ectopic beats (26-46%) while the dog seemed to have more frequent junctional complexes and AVB compared to NHP and minipigs. Arrhythmia screening may be considered during pre-study evaluations, to exclude animals with abnormally high arrhythmia incidence.


Asunto(s)
Arritmias Cardíacas , Telemetría , Animales , Perros , Porcinos , Masculino , Femenino , Porcinos Enanos , Incidencia , Estudios Retrospectivos , Electrocardiografía
9.
J Neurosci ; 29(8): 2519-27, 2009 Feb 25.
Artículo en Inglés | MEDLINE | ID: mdl-19244526

RESUMEN

Dynamic mechanical allodynia is a widespread and intractable symptom of neuropathic pain for which there is a lack of effective therapy. We recently provided a novel perspective on the mechanisms of this symptom by showing that a simple switch in trigeminal glycine synaptic inhibition can turn touch into pain by unmasking innocuous input to superficial dorsal horn nociceptive specific neurons through a local excitatory, NMDA-dependent neural circuit involving neurons expressing the gamma isoform of protein kinase C. Here, we further investigated the clinical relevance and processing of glycine disinhibition. First, we showed that glycine disinhibition with strychnine selectively induced dynamic but not static mechanical allodynia. The induced allodynia was resistant to morphine. Second, morphine did not prevent the activation of the neural circuit underlying allodynia as shown by study of Fos expression and extracellular-signal regulated kinase phosphorylation in dorsal horn neurons. Third, in contrast to intradermal capsaicin injections, light, dynamic mechanical stimuli applied under disinhibition did not produce neurokinin 1 (NK1) receptor internalization in dorsal horn neurons. Finally, light, dynamic mechanical stimuli applied under disinhibition induced Fos expression only in neurons that did not express NK1 receptor. To summarize, the selectivity and morphine resistance of the glycine-disinhibition paradigm adequately reflect the clinical characteristics of dynamic mechanical allodynia. The present findings thus reveal the involvement of a selective dorsal horn circuit in dynamic mechanical allodynia, which operates through superficial lamina nociceptive-specific neurons that do not bear NK1 receptor and provide an explanation for the differences in the pharmacological sensitivity of neuropathic pain symptoms.


Asunto(s)
Analgésicos Opioides/administración & dosificación , Glicina/metabolismo , Hiperalgesia/metabolismo , Hiperalgesia/fisiopatología , Morfina/administración & dosificación , Umbral del Dolor/fisiología , Receptores de Neuroquinina-1/fisiología , Animales , Bicuculina/farmacología , Presión Sanguínea/efectos de los fármacos , Capsaicina/farmacología , Modelos Animales de Enfermedad , Resistencia a Medicamentos , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Antagonistas del GABA/farmacología , Hiperalgesia/inducido químicamente , Hiperalgesia/tratamiento farmacológico , Masculino , Dinámicas no Lineales , Proteínas Oncogénicas v-fos/metabolismo , Dimensión del Dolor , Umbral del Dolor/efectos de los fármacos , Estimulación Física/métodos , Células del Asta Posterior/metabolismo , Ratas , Ratas Sprague-Dawley , Estricnina
10.
J Pharmacol Toxicol Methods ; 105: 106883, 2020 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-32461084

RESUMEN

INTRODUCTION: Pre-clinically, safety risk assessment of a drug is primarily tested in vivo using functional evaluation of adult animals while the mechanistic etiology of drug-induced CNS adverse effects is often uncharacterized. In vitro electrophysiology may provide a better understanding of drug effects without additional animal use. However, in vitro protocols are typically designed for using embryonic or juvenile animals. METHODS: We examined whether brain tissue isolated from adult rats (3-5 months old) and adult non-human primates (NHPs) (2-8 years old) can generate qualitatively equivalent readouts for electrophysiology to characterize AMPAR synaptic and single channel currents. We used a known positive AMPAR allosteric modulator (LY451395) to template a response profile and provide proof-of-concept data to assess responses of these native AMPARs in a drug context. RESULTS: Brain slices from adult animals provided a support to measure AMPAR-driven excitatory post-synaptic currents (EPSCs), and can be dissociated into primary neuronal cultures for AMPAR single channel characterization. Additionally, similarities and differences in AMPAR basal kinetics and responses to LY451395 were seen between the two animal species. DISCUSSION: Glutamatergic synaptic activity and AMPAR biophysical properties in adult animals may be used to characterize test-article-mediated alterations in CNS responses. The use of older animals opens the possibility for in vivo test-article administration, either acutely or repeatedly, before in vitro electrophysiological assessment in order to reveal cumulative or delayed-onset effects, adding versatility to safety pharmacology assessment of the CNS.


Asunto(s)
Sistema Nervioso Central/metabolismo , Sistema Nervioso Central/fisiología , Neuronas/metabolismo , Neuronas/fisiología , Primates/metabolismo , Primates/fisiología , Animales , Células Cultivadas , Electrofisiología/métodos , Femenino , Masculino , Técnicas de Placa-Clamp/métodos , Ratas , Receptores AMPA
11.
J Pharmacol Toxicol Methods ; 99: 106589, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-31154034

RESUMEN

Spontaneous arrhythmia characterization in healthy rats can support interpretation when studying novel therapies. Male (n = 55) and female (n = 40) Sprague-Dawley rats with telemetry transmitters for a derivation II ECG. Arrhythmias were assessed from continuous ECG monitoring over a period of 24-48 h, and data analyzed using an automated detection algorithm with 100% manual over-read. While a total of 1825 spontaneous ventricular premature beats (VPB) were identified, only 7 rats (or 7.4%) did not present with any over the recording period. Spontaneous episode(s) of ventricular tachycardia (VT) were noted in males (27%) and females (3%). The incidence of VPB was significantly higher (p < 0.01) during the night time (7 pm-7 am) compared to daytime, while males presented with significantly (p < 0.001) more VPB than females. Most VPB were observed as single ectopic beats, followed by salvos (2 or 3 consecutive VPBs), and VT (i.e. 4 consecutive VPBs). Most VPBs were single premature ventricular contractions (PVCs) (57%), while the remaining were escape complexes (43%). Spontaneous premature junctional complexes (PJC) were also observed and were significantly more frequent during the night, and in males. Lastly, 596 episodes of spontaneous 2nd-degree atrioventricular (AV) block were identified and were significantly more frequent during the day time in males. Most 2nd-degree AV block episodes were Mobitz type I (57%), with a significantly (p < 0.05) higher incidence in males. This work emphasizes the importance of obtaining sufficient baseline data when undertaking arrhythmia analysis in safety study and provides a better understanding of both sex- and time- dependent effects of spontaneous arrhythmias in rats.

12.
Elife ; 52016 08 08.
Artículo en Inglés | MEDLINE | ID: mdl-27501334

RESUMEN

Type 1 cannabinoid receptors (CB1Rs) are widely expressed in the vertebrate retina, but the role of endocannabinoids in vision is not fully understood. Here, we identified a novel mechanism underlying a CB1R-mediated increase in retinal ganglion cell (RGC) intrinsic excitability acting through AMPK-dependent inhibition of NKCC1 activity. Clomeleon imaging and patch clamp recordings revealed that inhibition of NKCC1 downstream of CB1R activation reduces intracellular Cl(-) levels in RGCs, hyperpolarizing the resting membrane potential. We confirmed that such hyperpolarization enhances RGC action potential firing in response to subsequent depolarization, consistent with the increased intrinsic excitability of RGCs observed with CB1R activation. Using a dot avoidance assay in freely swimming Xenopus tadpoles, we demonstrate that CB1R activation markedly improves visual contrast sensitivity under low-light conditions. These results highlight a role for endocannabinoids in vision and present a novel mechanism for cannabinoid modulation of neuronal activity through Cl(-) regulation.


Asunto(s)
Proteínas Quinasas Activadas por AMP/metabolismo , Sensibilidad de Contraste/fisiología , Endocannabinoides/metabolismo , Receptor Cannabinoide CB1/metabolismo , Células Ganglionares de la Retina/metabolismo , Miembro 2 de la Familia de Transportadores de Soluto 12/metabolismo , Proteínas Quinasas Activadas por AMP/genética , Animales , Cloruros/metabolismo , Regulación de la Expresión Génica , Transporte Iónico , Larva/fisiología , Potenciales de la Membrana/fisiología , Imagen Molecular , Técnicas de Placa-Clamp , Receptor Cannabinoide CB1/genética , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Células Ganglionares de la Retina/citología , Transducción de Señal , Miembro 2 de la Familia de Transportadores de Soluto 12/genética , Visión Ocular/fisiología , Xenopus laevis/fisiología
13.
Nat Commun ; 5: 5262, 2014 Oct 31.
Artículo en Inglés | MEDLINE | ID: mdl-25358432

RESUMEN

The introduction of calcium ion (Ca(2+)) indicators based on red fluorescent proteins (RFPs) has created new opportunities for multicolour visualization of intracellular Ca(2+) dynamics. However, one drawback of these indicators is that they have optimal two-photon excitation outside the near-infrared window (650-1,000 nm) where tissue is most transparent to light. To address this shortcoming, we developed a long Stokes shift RFP-based Ca(2+) indicator, REX-GECO1, with optimal two-photon excitation at <1,000 nm. REX-GECO1 fluoresces at 585 nm when excited at 480 nm or 910 nm by a one- or two-photon process, respectively. We demonstrate that REX-GECO1 can be used as either a ratiometric or intensiometric Ca(2+) indicator in organotypic hippocampal slice cultures (one- and two-photon) and the visual system of albino tadpoles (two-photon). Furthermore, we demonstrate single excitation wavelength two-colour Ca(2+) and glutamate imaging in organotypic cultures.


Asunto(s)
Calcio/análisis , Proteínas Luminiscentes/química , Imagen Óptica , Ingeniería de Proteínas , Protones , Animales , Células HeLa , Humanos , Indicadores y Reactivos , Microscopía Confocal , Estructura Molecular , Ratas , Ratas Sprague-Dawley , Proteína Fluorescente Roja
14.
Pain ; 155(2): 275-291, 2014 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-24120461

RESUMEN

Bone cancer pain is a common and disruptive symptom in cancer patients. In cancer pain animal models, massive reactive astrogliosis in the dorsal horn of the spinal cord has been reported. Because astrocytes may behave as driving partners for pathological pain, we investigated the temporal development of pain behavior and reactive astrogliosis in a rat bone cancer pain model induced by injecting MRMT-1 rat mammary gland carcinoma cells into the tibia. Along with the development of bone lesions, a gradual mechanical and thermal allodynia and hyperalgesia as well as a reduced use of the affected limb developed in bone cancer-bearing animals, but not in sham-treated animals. Dorsal horn Fos expression after nonpainful palpation of the injected limb was also increased in bone cancer-bearing animals. However, at any time during the evolution of tumor, there was no increase in glial fibrillary acidic protein (GFAP) immunoreactivity in the dorsal horn. Further analysis at 21days after injection of the tumor showed no increase in GFAP and interleukin (IL) 1ß transcripts, number of superficial dorsal horn S100ß protein immunoreactive astrocytes, or immunoreactivity for microglial markers (OX-42 and Iba-1). In contrast, all these parameters were increased in the dorsal horn of rats 2weeks after sciatic nerve ligation. This suggests that in some cases, bone cancer pain may not be correlated with spinal overexpression of reactive glia markers, whereas neuropathic pain is. Glia may thus play different roles in the development and maintenance of chronic pain in these 2 situations.


Asunto(s)
Biomarcadores de Tumor/metabolismo , Neoplasias Óseas/metabolismo , Neuroglía/metabolismo , Dimensión del Dolor/métodos , Dolor/metabolismo , Médula Espinal/metabolismo , Animales , Neoplasias Óseas/patología , Femenino , Regulación Neoplásica de la Expresión Génica , Masculino , Neuroglía/patología , Dolor/patología , Ratas , Ratas Sprague-Dawley , Médula Espinal/patología , Células Tumorales Cultivadas
15.
PLoS One ; 7(1): e29086, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-22242157

RESUMEN

The developing retinotectal system of the Xenopus laevis tadpole is a model of choice for studying visual experience-dependent circuit maturation in the intact animal. The neurotransmitter gamma-aminobutyric acid (GABA) has been shown to play a critical role in the formation of sensory circuits in this preparation, however a comprehensive neuroanatomical study of GABAergic cell distribution in the developing tadpole has not been conducted. We report a detailed description of the spatial expression of GABA immunoreactivity in the Xenopus laevis tadpole brain at two key developmental stages: stage 40/42 around the onset of retinotectal innervation and stage 47 when the retinotectal circuit supports visually-guided behavior. During this period, GABAergic neurons within specific brain structures appeared to redistribute from clusters of neuronal somata to a sparser, more uniform distribution. Furthermore, we found that GABA levels were regulated by recent sensory experience. Both ELISA measurements of GABA concentration and quantitative analysis of GABA immunoreactivity in tissue sections from the optic tectum show that GABA increased in response to a 4 hr period of enhanced visual stimulation in stage 47 tadpoles. These observations reveal a remarkable degree of adaptability of GABAergic neurons in the developing brain, consistent with their key contributions to circuit development and function.


Asunto(s)
Sensación/fisiología , Vías Visuales/crecimiento & desarrollo , Vías Visuales/metabolismo , Ácido gamma-Aminobutírico/metabolismo , Animales , Encéfalo/crecimiento & desarrollo , Encéfalo/metabolismo , Ensayo de Inmunoadsorción Enzimática , Técnica del Anticuerpo Fluorescente , Neuronas GABAérgicas/metabolismo , Larva/citología , Larva/metabolismo , Estimulación Luminosa , Retina/citología , Retina/crecimiento & desarrollo , Retina/metabolismo , Colículos Superiores/crecimiento & desarrollo , Colículos Superiores/metabolismo , Vías Visuales/citología , Xenopus laevis/crecimiento & desarrollo , Xenopus laevis/metabolismo
16.
Pain ; 152(6): 1340-1348, 2011 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-21392888

RESUMEN

Glycine inhibitory dysfunction provides a useful experimental model for studying the mechanism of dynamic mechanical allodynia, a widespread and intractable symptom of neuropathic pain. In this model, allodynia expression relies on N-methyl-d-aspartate receptors (NMDARs), and it has been shown that astrocytes can regulate their activation through the release of the NMDAR coagonist d-serine. Recent studies also suggest that astrocytes potentially contribute to neuropathic pain. However, the involvement of astrocytes in dynamic mechanical allodynia remains unknown. Here, we show that after blockade of glycine inhibition, orofacial tactile stimuli activated medullary dorsal horn (MDH) astrocytes, but not microglia. Accordingly, the glia inhibitor fluorocitrate, but not the microglia inhibitor minocycline, prevented allodynia. Fluorocitrate also impeded activation of astrocytes and blocked activation of the superficial MDH neural circuit underlying allodynia, as revealed by study of Fos expression. MDH astrocytes are thus required for allodynia. They may also produce d-serine because astrocytic processes were selectively immunolabeled for serine racemase, the d-serine synthesizing enzyme. Accordingly, selective degradation of d-serine with d-amino acid oxidase applied in vivo prevented allodynia and activation of the underlying neural circuit. Conversely, allodynia blockade by fluorocitrate was reversed by exogenous d-serine. These results suggest the following scenario: removal of glycine inhibition makes tactile stimuli able to activate astrocytes; activated astrocytes may provide d-serine to enable NMDAR activation and thus allodynia. Such a contribution of astrocytes to pathological pain fuels the emerging concept that astrocytes are critical players in pain signaling. Glycine disinhibition makes tactile stimuli able to activate astrocytes, which may provide d-serine to enable NMDA receptor activation and thus allodynia.


Asunto(s)
Citratos/uso terapéutico , Glicina/metabolismo , Hiperalgesia/tratamiento farmacológico , Serina/metabolismo , Análisis de Varianza , Animales , Astrocitos/química , Astrocitos/efectos de los fármacos , Antígeno CD11b/metabolismo , Citratos/farmacología , Modelos Animales de Enfermedad , Inhibidores Enzimáticos/farmacología , Proteína Ácida Fibrilar de la Glía/metabolismo , Masculino , Proteínas Oncogénicas v-fos/metabolismo , Dimensión del Dolor , Ratas , Ratas Sprague-Dawley , Raíces Nerviosas Espinales/citología , Estricnina/uso terapéutico , Factores de Tiempo
17.
Pain ; 137(2): 340-351, 2008 Jul 15.
Artículo en Inglés | MEDLINE | ID: mdl-17977663

RESUMEN

Windup is a progressive, frequency-dependent increase in the excitability of trigeminal and spinal dorsal horn wide dynamic range (WDR) nociceptive neurons to repetitive stimulation of primary afferent nociceptive C-fibers. Superficial dorsal horn neurokinin 1 receptor (NK1R)-expressing neurons were recently shown to regulate sensitization of WDR nociceptive neurons through activation of a defined spino-bulbo-spinal loop. However, the windup of WDR nociceptive neurons was not regulated through this loop. In the present study, we sought to identify the alternative circuit activated by dorsal horn NK1Rs that mediates WDR neuron windup. As a model we used the rat spinal trigeminal nucleus, in which the subnucleus oralis (Sp5O) contains a pool of WDR neurons that receive their nociceptive C-input indirectly via interneurons located in the medullary dorsal horn (MDH). First, we found that intravenous injection of NK1R antagonists (SR140333 and RP67580) produced a reversible inhibition of Sp5O WDR neuron windup. Second, we anatomically identified in the MDH lamina III a subpopulation of NK1R-expressing local interneurons that relay nociceptive information from the MDH to downstream Sp5O neurons. Third, using microinjections of NK1R antagonists during in vivo electrophysiological recordings from Sp5O WDR neurons, we showed that WDR neuron windup depends on activation of NK1Rs located in the MDH laminae I-III. We conclude that, in contrast to central sensitization that is controlled by a spino-bulbo-spinal loop, Sp5O WDR neuron windup is regulated through a local circuit activated by MDH lamina III NK1Rs.


Asunto(s)
Vías Aferentes/metabolismo , Nociceptores/metabolismo , Dolor/metabolismo , Células del Asta Posterior/metabolismo , Receptores de Neuroquinina-1/metabolismo , Núcleo Caudal del Trigémino/metabolismo , Vías Aferentes/citología , Animales , Masculino , Fibras Nerviosas Amielínicas/metabolismo , Fibras Nerviosas Amielínicas/ultraestructura , Vías Nerviosas/citología , Vías Nerviosas/metabolismo , Antagonistas del Receptor de Neuroquinina-1 , Neuronas Aferentes/citología , Neuronas Aferentes/efectos de los fármacos , Neuronas Aferentes/metabolismo , Nociceptores/citología , Dolor/fisiopatología , Piperidinas/farmacología , Células del Asta Posterior/efectos de los fármacos , Terminales Presinápticos/metabolismo , Terminales Presinápticos/ultraestructura , Quinuclidinas/farmacología , Ratas , Ratas Sprague-Dawley , Sustancia P/metabolismo , Transmisión Sináptica/fisiología , Núcleo Caudal del Trigémino/citología , Nervio Trigémino/citología , Nervio Trigémino/metabolismo , Núcleos del Trigémino/citología , Núcleos del Trigémino/metabolismo
18.
PLoS One ; 2(11): e1116, 2007 Nov 07.
Artículo en Inglés | MEDLINE | ID: mdl-17987109

RESUMEN

Dynamic mechanical allodynia is a widespread and intractable symptom of neuropathic pain for which there is a lack of effective therapy. During tactile allodynia, activation of the sensory fibers which normally detect touch elicits pain. Here we provide a new behavioral investigation into the dynamic component of tactile allodynia that developed in rats after segmental removal of glycine inhibition. Using in vivo electrophysiological recordings, we show that in this condition innocuous mechanical stimuli could activate superficial dorsal horn nociceptive specific neurons. These neurons do not normally respond to touch. We anatomically show that the activation was mediated through a local circuit involving neurons expressing the gamma isoform of protein kinase C (PKCgamma). Selective inhibition of PKCgamma as well as selective blockade of glutamate NMDA receptors in the superficial dorsal horn prevented both activation of the circuit and allodynia. Thus, our data demonstrates that a normally inactive circuit in the dorsal horn can be recruited to convert touch into pain. It also provides evidence that glycine inhibitory dysfunction gates tactile input to nociceptive specific neurons through PKCgamma-dependent activation of a local, excitatory, NMDA receptor-dependent, circuit. As a consequence of these findings, we suggest that pharmacological inhibition of PKCgamma might provide a new tool for alleviating allodynia in the clinical setting.


Asunto(s)
Glicina/antagonistas & inhibidores , Interneuronas/enzimología , Dolor/metabolismo , Proteína Quinasa C/metabolismo , Animales , Masculino , Dolor/enzimología , Ratas , Ratas Sprague-Dawley
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