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1.
Brain ; 143(1): 359-373, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31782760

RESUMO

Failure of Alzheimer's disease clinical trials to improve or stabilize cognition has led to the need for a better understanding of the driving forces behind cognitive decline in the presence of active disease processes. To dissect contributions of individual pathologies to cognitive function, we used the TgF344-AD rat model, which recapitulates the salient hallmarks of Alzheimer's disease pathology observed in patient populations (amyloid, tau inclusions, frank neuronal loss, and cognitive deficits). scyllo-Inositol treatment attenuated amyloid-ß peptide in disease-bearing TgF344-AD rats, which rescued pattern separation in the novel object recognition task and executive function in the reversal learning phase of the Barnes maze. Interestingly, neither activities of daily living in the burrowing task nor spatial memory in the Barnes maze were rescued by attenuating amyloid-ß peptide. To understand the pathological correlates leading to behavioural rescue, we examined the neuropathology and in vivo electrophysiological signature of the hippocampus. Amyloid-ß peptide attenuation reduced hippocampal tau pathology and rescued adult hippocampal neurogenesis and neuronal function, via improvements in cross-frequency coupling between theta and gamma bands. To investigate mechanisms underlying the persistence of spatial memory deficits, we next examined neuropathology in the entorhinal cortex, a region whose input to the hippocampus is required for spatial memory. Reduction of amyloid-ß peptide in the entorhinal cortex had no effect on entorhinal tau pathology or entorhinal-hippocampal neuronal network dysfunction, as measured by an impairment in hippocampal response to entorhinal stimulation. Thus, rescue or not of cognitive function is dependent on regional differences of amyloid-ß, tau and neuronal network dysfunction, demonstrating the importance of staging disease in patients prior to enrolment in clinical trials. These results further emphasize the need for combination therapeutic approaches across disease progression.

2.
Hypertension ; 74(4): 1041-1051, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31476904

RESUMO

Transient hypertension is a risk factor for Alzheimer disease (AD), but the effects of this interaction on brain vasculature are understudied. Addressing vascular pathology is a promising avenue to potentiate the efficacy of treatments for AD. We used arterial spin labeling magnetic resonance imaging to longitudinally assess brain vascular function and immunohistopathology to examine cerebrovascular remodeling and amyloid load. Hypertension was induced for 1 month by administration of l-NG-nitroarginine-methyl-ester in TgF344-AD rats at the prodromal stage. Following hypertension, nontransgenic rats showed transient cerebrovascular changes, whereas TgF344-AD animals exhibited sustained alterations in cerebrovascular function. Human umbilical cord perivascular cells in combination with scyllo-inositol, an inhibitor of Aß oligomerization, resulted in normalization of hippocampal vascular function and remodeling, in contrast to either treatment alone. Prodromal stage hypertension exacerbates latter AD pathology, and the combination of human umbilical cord perivascular cells with amyloid clearance promotes cerebrovascular functional recovery.


Assuntos
Doença de Alzheimer/fisiopatologia , Hipertensão/fisiopatologia , Doença de Alzheimer/etiologia , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Animais , Encéfalo/fisiopatologia , Modelos Animais de Doenças , Hemodinâmica/fisiologia , Hipertensão/complicações , Hipertensão/terapia , Imagem por Ressonância Magnética , Ratos , Marcadores de Spin
3.
Neuroimage ; 192: 135-144, 2019 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-30669007

RESUMO

The rapid growth in the use of optogenetics for neuroscience applications is largely driven by two important advantages: highly specific cellular targeting through genetic manipulations; and precise temporal control of neuronal activation via temporal modulation of the optical stimulation. The difference between the most commonly used stimulation modalities, namely diffused (i.e. synchronous) and focused (i.e. asynchronous) stimulation has not been described. Furthermore, full realization of optogenetics' potential is hindered by our incomplete understanding of the cellular and network level response to photoactivation. Here we address these gaps by examining the neuronal and cerebrovascular responses to focused and diffuse photostimulation of channelrhodopsin in the Thy1-ChR2 mouse. We presented the responses of photoactivation via 470-nm fiber optic illumination (diffuse) alongside 458-nm raster-scan (focused) stimulation of the barrel field. Local field potentials (LFP) assessment of intracerebral electrophysiology and two-photon fluorescence microscopy measurements of red blood cell (RBC) speed (vRBC) in cortical penetrating vessels revealed ∼40% larger LFP responses (p = 0.05) and twice as large cerebrovascular responses (p = 0.002) under focused vs. diffuse photostimulation (focused: 1.64 ±â€¯0.84 mV LFP amplitude and 75 ±â€¯48% increase in vRBC; diffuse: 1.14 ±â€¯0.75 mV LFP amplitude and 35 ±â€¯23% increase in vRBC). Compared to diffuse photostimulation, focused photostimulation resulted in a ∼65% increase in the yield of cerebrovascular responses (73 ±â€¯10% for focused and 42 ±â€¯29% for diffuse photostimulation) and a doubling of the signal-to-noise ratio of the cerebrovascular response (20.9 ±â€¯14.7 for focused and 10.4 ±â€¯1.4 for diffuse photostimulation). These data reveal important advantages of focused optogenetic photoactivation, which can be easily integrated into single- or two-photon fluorescence microscopy platforms, as a means of assessing neuronal excitability and cerebrovascular reactivity, thus paving the way for broader application of optogenetics in preclinical models of CNS diseases.


Assuntos
Encéfalo/irrigação sanguínea , Circulação Cerebrovascular/fisiologia , Channelrhodopsins/metabolismo , Optogenética/métodos , Animais , Encéfalo/metabolismo , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos
4.
Front Mol Neurosci ; 11: 338, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30271324

RESUMO

Although epidemiological evidence suggests significant sex and gender-based differences in stroke risk and recovery, females have been widely under-represented in preclinical stroke research. The neurovascular sequelae of brain ischemia in females, in particular, are largely uncertain. We set out to address this gap by a multimodal in vivo study of neurovascular recovery from endothelin-1 model of cortical focal-stroke in sham vs. ovariectomized female rats. Three weeks post ischemic insult, sham operated females recapitulated the phenotype previously reported in male rats in this model, of normalized resting perfusion but sustained peri-lesional cerebrovascular hyperreactivity. In contrast, ovariectomized (Ovx) females showed reduced peri-lesional resting blood flow, and elevated cerebrovascular responsivity to hypercapnia in the peri-lesional and contra-lateral cortices. Electrophysiological recordings showed an attenuation of theta to low-gamma phase-amplitude coupling in the peri-lesional tissue of Ovx animals, despite relative preservation of neuronal power. Further, this chronic stage neuronal network dysfunction was inversely correlated with serum estradiol concentration. Our pioneering data demonstrate dramatic differences in spontaneous recovery in the neurovascular unit between Ovx and Sham females in the chronic stage of stroke, underscoring the importance of considering hormonal-dependent aspects of the ischemic sequelae in the development of novel therapeutic approaches and patient recruitment in clinical trials.

5.
Stroke ; 49(9): 2173-2181, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-30354983

RESUMO

Background and Purpose- Recent evidence suggests great potential of metabolically targeted interventions for treating neurological disorders. We investigated the use of the endogenous ketone body ß-hydroxybutyrate (BHB) as an alternate metabolic substrate for the brain in the acute phase of ischemia because postischemic hyperglycemia and brain glucose metabolism elevation compromise functional recovery. Methods- We delivered BHB (or vehicle) 1 hour after ischemic insult induced by cortical microinjection of endothelin-1 in sensorimotor cortex of rats. Two days after ischemic insult, the rats underwent multimodal characterization of the BHB effects. We examined glucose uptake on 2-Deoxy-d-glucose chemical exchange saturation transfer magnetic resonance imaging, cerebral hemodynamics on continuous arterial spin labeling magnetic resonance imaging, resting-state field potentials by intracerebral multielectrode arrays, Neurological Deficit Score, reactive oxygen species production, and astrogliosis and neuronal death. Results- When compared with vehicle-administered animals, BHB-treated cohort showed decreased peri-infarct neuronal glucose uptake which was associated with reduced oxidative stress, diminished astrogliosis and neuronal death. Functional examination revealed ameliorated neuronal functioning, normalized perilesional resting perfusion, and ameliorated cerebrovascular reactivity to hypercapnia, suggesting improved functioning. Cellular and functional recovery of the neurogliovascular unit in the BHB-treated animals was associated with improved performance on the withdrawal test. Conclusions- We characterize the effects of the ketone body BHB administration at cellular and system levels after focal cortical stroke. The results demonstrate that BHB curbs the peri-infarct glucose-metabolism driven production of reactive oxygen species and astrogliosis, culminating in improved neurogliovascular and functional recovery.


Assuntos
Ácido 3-Hidroxibutírico/farmacologia , Astrócitos/efeitos dos fármacos , Isquemia Encefálica/metabolismo , Encéfalo/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Ácido 3-Hidroxibutírico/metabolismo , Acetoacetatos/metabolismo , Animais , Astrócitos/patologia , Glicemia/metabolismo , Encéfalo/diagnóstico por imagem , Encéfalo/metabolismo , Encéfalo/patologia , Isquemia Encefálica/diagnóstico por imagem , Isquemia Encefálica/patologia , Morte Celular/efeitos dos fármacos , Circulação Cerebrovascular , Modelos Animais de Doenças , Fenômenos Eletrofisiológicos , Endotelina-1 , Hemodinâmica , Imagem por Ressonância Magnética , Espectroscopia de Ressonância Magnética , Microinjeções , Neurônios/patologia , Ratos , Espécies Reativas de Oxigênio/metabolismo , Córtex Sensório-Motor
6.
Theranostics ; 8(17): 4824-4836, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30279740

RESUMO

Traumatic brain injury (TBI) research has focused on moderate to severe injuries as their outcomes are significantly worse than those of a mild TBI (mTBI). However, recent epidemiological evidence has indicated that a series of even mild TBIs greatly increases the risk of neurodegenerative and psychiatric disorders. Neuropathological studies of repeated TBI have identified changes in neuronal ionic concentrations, axonal injury, and cytoskeletal damage as important determinants of later life neurological and mood compromise; yet, there is a paucity of data on the contribution of neurogliovascular dysfunction to the progression of repeated TBI and alterations of brain function in the intervening period. Methods: Here, we established a mouse model of repeated TBI induced via three electromagnetically actuated impacts delivered to the intact skull at three-day intervals and determined the long-term deficits in neurogliovascular functioning in Thy1-ChR2 mice. Two weeks post the third impact, cerebral blood flow and cerebrovascular reactivity were measured with arterial spin labelling magnetic resonance imaging. Neuronal function was investigated through bilateral intracranial electrophysiological responses to optogenetic photostimulation. Vascular density of the site of impacts was measured with in vivo two photon fluorescence microscopy. Pathological analysis of neuronal survival and astrogliosis was performed via NeuN and GFAP immunofluorescence. Results: Cerebral blood flow and cerebrovascular reactivity were decreased by 50±16% and 70±20%, respectively, in the TBI cohort relative to sham-treated animals. Concomitantly, electrophysiological recordings revealed a 97±1% attenuation in peri-contusional neuronal reactivity relative to sham. Peri-contusional vascular volume was increased by 33±2% relative to sham-treated mice. Pathological analysis of the peri-contusional cortex demonstrated astrogliosis, but no changes in neuronal survival. Conclusion: This work provides the first in-situ characterization of the long-term deficits of the neurogliovascular unit following repeated TBI. The findings will help guide the development of diagnostic markers as well as therapeutics targeting neurogliovascular dysfunction.


Assuntos
Lesões Encefálicas Traumáticas/patologia , Transtornos Cerebrovasculares/patologia , Modelos Animais de Doenças , Neuroglia/patologia , Neurônios/patologia , Animais , Lesões Encefálicas Traumáticas/diagnóstico por imagem , Transtornos Cerebrovasculares/diagnóstico por imagem , Imagem por Ressonância Magnética , Camundongos , Microscopia de Fluorescência , Optogenética , Recidiva
7.
J Neurochem ; 144(5): 669-679, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-28777881

RESUMO

Alzheimer's disease (AD) is pathologically characterized by amyloid-ß peptide (Aß) accumulation, neurofibrillary tangle formation, and neurodegeneration. Preclinical studies on neuronal impairments associated with progressive amyloidosis have demonstrated some Aß-dependent neuronal dysfunction including modulation of gamma-aminobutyric acid-ergic signaling. The present work focuses on the early stage of disease progression and uses TgF344-AD rats that recapitulate a broad repertoire of AD-like pathologies to investigate the neuronal network functioning using simultaneous intracranial recordings from the hippocampus (HPC) and the medial prefrontal cortex (mPFC), followed by pathological analyses of gamma-aminobutyric acid (GABAA ) receptor subunits α1, α5, and δ, and glutamic acid decarboxylases (GAD65 and GAD67). Concomitant to amyloid deposition and tau hyperphosphorylation, low-gamma band power was strongly attenuated in the HPC and mPFC of TgF344-AD rats in comparison to those in non-transgenic littermates. In addition, the phase-amplitude coupling of the neuronal networks in both areas was impaired, evidenced by decreased modulation of theta band phase on gamma band amplitude in TgF344-AD animals. Finally, the gamma coherence between HPC and mPFC was attenuated as well. These results demonstrate significant neuronal network dysfunction at an early stage of AD-like pathology. This network dysfunction precedes the onset of cognitive deficits and is likely driven by Aß and tau pathologies. This article is part of the Special Issue "Vascular Dementia".


Assuntos
Doença de Alzheimer/fisiopatologia , Hipocampo/fisiopatologia , Neurônios/fisiologia , Córtex Pré-Frontal/fisiopatologia , Doença de Alzheimer/patologia , Animais , Ondas Encefálicas , Modelos Animais de Doenças , Feminino , Glutamato Descarboxilase/metabolismo , Hipocampo/metabolismo , Hipocampo/patologia , Masculino , Vias Neurais/fisiopatologia , Placa Amiloide/metabolismo , Córtex Pré-Frontal/patologia , Ratos Endogâmicos F344 , Ratos Transgênicos , Receptores de GABA-A/metabolismo
8.
Int J Mol Sci ; 18(11)2017 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-29143800

RESUMO

Epilepsy afflicts up to 1.6% of the population and the mechanisms underlying the appearance of seizures are still not understood. In past years, many efforts have been spent trying to understand the mechanisms underlying the excessive and synchronous firing of neurons. Traditionally, attention was pointed towards synaptic (dys)function and extracellular ionic species (dys)regulation. Recently, novel clinical and preclinical studies explored the role of brain metabolism (i.e., glucose utilization) of seizures pathophysiology revealing (in most cases) reduced metabolism in the inter-ictal period and increased metabolism in the seconds preceding and during the appearance of seizures. In the present review, we summarize the clinical and preclinical observations showing metabolic dysregulation during epileptogenesis, seizure initiation, and termination, and in the inter-ictal period. Recent preclinical studies have shown that 2-Deoxyglucose (2-DG, a glycolysis blocker) is a novel therapeutic approach to reduce seizures. Furthermore, we present initial evidence for the effectiveness of 2-DG in arresting 4-Aminopyridine induced neocortical seizures in vivo in the mouse.


Assuntos
Metabolismo Energético , Neurônios/metabolismo , Animais , Modelos Animais de Doenças , Humanos , Convulsões/etiologia , Convulsões/metabolismo , Convulsões/fisiopatologia
9.
J Magn Reson Imaging ; 46(2): 505-517, 2017 08.
Artigo em Inglês | MEDLINE | ID: mdl-28703413

RESUMO

PURPOSE: Stroke is the leading cause of adult disability worldwide. The absence of more effective interventions in the chronic stage-that most patients stand to benefit from-reflects uncertainty surrounding mechanisms that govern recovery. The present work investigated the effects of a novel treatment (selective cyclooxygenase-1, COX-1, inhibition) in a model of focal ischemia. MATERIALS AND METHODS: FR122047 (COX-1 inhibitor) was given beginning 7 days following stroke (cortical microinjection of endothelin-1) in 23 adult male rats. Longitudinal continuous-arterial-spin-labeling was performed prior to treatment (7 days), and repeated following treatment (21 days) on a 7T magnetic resonance imaging (MRI) system to estimate resting perfusion and reactivity to hypercapnia. These in vivo measurements were buttressed by immunohistochemistry. RESULTS: Stroke caused an increase in perilesional resting perfusion (peri-/contralesional perfusion ratio of 170 ± 10%) and perfusion responses to hypercapnia (180 ± 10%) at 7 days. At 21 days, placebo-administered rats showed normalized perilesional perfusion (100 ± 20%) but persistent hyperreactivity (190 ± 20%). Treated animals exhibited sustained perilesional hyperperfusion (180 ± 10%). Further, reactivity lateralization did not persist following treatment (peri- vs. contralesional reactivity: P = 0.002 at 7 vs. P = 0.2 at 21 days). Hemodynamic changes were accompanied by neuronal loss, increased endothelial density, and widespread microglial and astrocytic activation. Moreover, relative to controls, treated rats showed increased perilesional neuronal survival (22 ± 1% vs. 14.9 ± 0.8%, P = 0.02) and decreased microglia/macrophage recruitment (17 ± 1% vs. 20 ± 1%, P = 0.05). Finally, perilesional perfusion was correlated with neuronal survival (slope = 0.14 ± 0.05; R2 = 0.7, P = 0.03). CONCLUSION: These findings shed light on the role of COX-1 in chronic ischemic injury and suggest that delayed selective COX-1 inhibition exerts multiple beneficial effects on the neurogliovascular unit. LEVEL OF EVIDENCE: 1 Technical Efficacy: Stage 4 J. MAGN. RESON. IMAGING 2017;46:505-517.


Assuntos
Inibidores de Ciclo-Oxigenase/farmacologia , Isquemia/diagnóstico por imagem , Imagem por Ressonância Magnética , Proteínas de Membrana/antagonistas & inibidores , Acidente Vascular Cerebral/diagnóstico por imagem , Acidente Vascular Cerebral/fisiopatologia , Animais , Ciclo-Oxigenase 1 , Modelos Animais de Doenças , Endotelina-1/química , Macrófagos/patologia , Masculino , Microglia/patologia , Neuroglia/patologia , Neurônios/patologia , Perfusão , Piperazinas/química , Ratos , Ratos Sprague-Dawley , Marcadores de Spin , Tiazóis/química
10.
Sci Rep ; 7: 46427, 2017 04 12.
Artigo em Inglês | MEDLINE | ID: mdl-28401931

RESUMO

Alzheimer's disease (AD), pathologically characterized by amyloid-ß peptide (Aß) accumulation, neurofibrillary tangle formation, and neurodegeneration, is thought to involve early-onset neurovascular abnormalities. Hitherto studies on AD-associated neurovascular injury have used animal models that exhibit only a subset of AD-like pathologies and demonstrated some Aß-dependent vascular dysfunction and destabilization of neuronal network. The present work focuses on the early stage of disease progression and uses TgF344-AD rats that recapitulate a broader repertoire of AD-like pathologies to investigate the cerebrovascular and neuronal network functioning using in situ two-photon fluorescence microscopy and laminar array recordings of local field potentials, followed by pathological analyses of vascular wall morphology, tau hyperphosphorylation, and amyloid plaques. Concomitant to widespread amyloid deposition and tau hyperphosphorylation, cerebrovascular reactivity was strongly attenuated in cortical penetrating arterioles and venules of TgF344-AD rats in comparison to those in non-transgenic littermates. Blood flow elevation to hypercapnia was abolished in TgF344-AD rats. Concomitantly, the phase-amplitude coupling of the neuronal network was impaired, evidenced by decreased modulation of theta band phase on gamma band amplitude. These results demonstrate significant neurovascular network dysfunction at an early stage of AD-like pathology. Our study identifies early markers of pathology progression and call for development of combinatorial treatment plans.


Assuntos
Doença de Alzheimer/fisiopatologia , Encéfalo/fisiopatologia , Circulação Cerebrovascular/fisiologia , Rede Nervosa/fisiopatologia , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Animais , Encéfalo/metabolismo , Modelos Animais de Doenças , Feminino , Masculino , Rede Nervosa/metabolismo , Neurônios/metabolismo , Neurônios/fisiologia , Fosforilação , Ratos , Ratos Transgênicos , Proteínas tau/metabolismo
11.
Neuroimage ; 146: 869-882, 2017 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-27664828

RESUMO

Brain plasticity following focal cerebral ischaemia has been observed in both stroke survivors and in preclinical models of stroke. Endogenous neurovascular adaptation is at present incompletely understood yet its potentiation may improve long-term functional outcome. We employed longitudinal MRI, intracranial array electrophysiology, Montoya Staircase testing, and immunofluorescence to examine function of brain vessels, neurons, and glia in addition to forelimb skilled reaching during the subacute stage of ischemic injury progression. Focal ischemic stroke (~100mm3 or ~20% of the total brain volume) was induced in adult Sprague-Dawley rats via direct injection of endothelin-1 (ET-1) into the right sensori-motor cortex, producing sustained impairment in left forelimb reaching ability. Resting perfusion and vascular reactivity to hypercapnia in the peri-lesional cortex were elevated by approximately 60% and 80% respectively seven days following stroke. At the same time, the normal topological pattern of local field potential (LFP) responses to peripheral somatosensory stimulation was abolished and the average power of spontaneous LFP activity attenuated by approximately 50% relative to the contra-lesional cortex, suggesting initial response attenuation within the peri-infarct zone. By 21 days after stroke, perilesional blood flow resolved, but peri-lesional vascular reactivity remained elevated. Concomitantly, the LFP response amplitudes increased with distance from the site of ET-1 injection, suggesting functional remodelling from the core of the lesion to its periphery. This notion was further buttressed by the lateralization of spontaneous neuronal activity: by day 21, the average ipsi-lesional power of spontaneous LFP activity was almost twice that of the contra-lesional cortex. Over the observation period, the peri-lesional cortex exhibited increased vascular density, along with neuronal loss, astrocytic activation, and recruitment and activation of microglia and macrophages, with neuronal loss and inflammation extending beyond the peri-lesional cortex. These findings highlight the complex relationship between neurophysiological state and behaviour and provide evidence of highly dynamic functional changes in the peri-infarct zone weeks following the ischemic insult, suggesting an extended temporal window for therapeutic interventions.


Assuntos
Isquemia Encefálica/fisiopatologia , Encéfalo/irrigação sanguínea , Encéfalo/fisiopatologia , Córtex Somatossensorial/irrigação sanguínea , Córtex Somatossensorial/fisiopatologia , Acidente Vascular Cerebral/fisiopatologia , Remodelação Vascular , Animais , Encéfalo/metabolismo , Isquemia Encefálica/induzido quimicamente , Isquemia Encefálica/complicações , Ondas Encefálicas , Encefalite/complicações , Encefalite/metabolismo , Endotelina-1/administração & dosagem , Hipercapnia/fisiopatologia , Imagem por Ressonância Magnética , Masculino , Destreza Motora , Neuroglia/metabolismo , Neurônios/metabolismo , Estimulação Física , Ratos Sprague-Dawley , Recuperação de Função Fisiológica , Córtex Sensório-Motor/efeitos dos fármacos , Acidente Vascular Cerebral/induzido quimicamente , Acidente Vascular Cerebral/complicações , Percepção do Tato/fisiologia
12.
Front Cell Neurosci ; 11: 397, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29311830

RESUMO

The inferior olive (IO) is a nucleus located in the brainstem and it is part of the olivo-cerebellar loop. This circuit plays a fundamental role in generation and acquisition of coherent motor patterns and it relies on synchronous activation of groups of Purkinje cells (PC) in the cerebellar cortex. IO neurons integrate their intrinsic oscillatory activity with excitatory inputs coming from the somatosensory system and inhibitory feedback coming from the cerebellar nuclei. Alongside these chemical synaptic inputs, IO neurons are coupled to one another via connexin 36 (Cx36) containing gap junctions (GJs) that create a functional syncytium between neurons. Communication between olivary neurons is regulated by these GJs and their correct functioning contributes to coherent oscillations in the IO and proper motor learning. Here, we explore the cellular pathways that can regulate the coupling between olivary neurons. We combined in vitro electrophysiology and immunohistochemistry (IHC) on mouse acute brain slices to unravel the pathways that regulate olivary coupling. We found that enhancing the activity of the protein kinase A (PKA) pathway and blocking the Ca2+/calmodulin-dependent protein kinase II (CaMKII) pathway can both down-regulate the size of the coupled network. However, these two kinases follow different mechanisms of action. Our results suggest that activation of the PKA pathway reduces the opening probability of the Cx36 GJs, whereas inhibition of the CaMKII pathway reduces the number of Cx36 GJs. The low densities of Cx36 proteins and electrical synapses in ßCaMKII knock-out mice point towards an essential role for this protein kinase in regulating the density of GJs in the IO. Thus, the level of olivary coupling is a dynamic process and regulated by a variety of enzymes modulating GJs expression, docking and activity.

13.
Neurobiol Dis ; 101: 1-7, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28007587

RESUMO

Extracellular potassium concentration, [K+]o, is a major determinant of neuronal excitability. In the healthy brain, [K+]o levels are tightly controlled. During seizures, [K+]o increases up to 15mM and is thought to cause seizures due to its depolarizing effect. Although astrocytes have been suggested to play a key role in the redistribution (or spatial buffering) of excess K+ through Connexin-43 (Cx43)-based Gap Junctions (GJs), the relation between this dynamic regulatory process and seizure generation remains unknown. Here we contrasted the role of astrocytic GJs and hemichannels by studying the effect of GJ and hemichannel blockers on [K+]o regulation in vivo. [K+]o was measured by K+-sensitive microelectrodes. Neuronal excitability was estimated by local field potential (LFP) responses to forepaw stimulation and changes in the power of resting state activity. Starting at the baseline [K+]o level of 1.61±0.3mM, cortical microinjection of CBX, a broad spectrum connexin channel blocker, increased [K+]o to 11±3mM, Cx43 GJ/hemichannel blocker Gap27 increased it from 1.9±0.7 to 9±1mM. At these [K+]o levels, no seizures were observed. Cx43 hemichannel blockade with TAT-Gap19 increased [K+]o by only ~1mM. Microinjection of 4-aminopyridine, a known convulsant, increased [K+]o to ~10mM and induced spontaneously recurring seizures, whereas direct application of K+ did not trigger seizure activity. These findings are the first in vivo demonstration that astrocytic GJs are major determinants for the spatial buffering of [K+]o and that an increase in [K+]o alone does not trigger seizures in the neocortex.


Assuntos
Astrócitos/metabolismo , Junções Comunicantes/metabolismo , Neocórtex/metabolismo , Potássio/metabolismo , Animais , Astrócitos/efeitos dos fármacos , Cátions Monovalentes/metabolismo , Conexina 43/antagonistas & inibidores , Conexina 43/metabolismo , Potenciais Somatossensoriais Evocados/efeitos dos fármacos , Potenciais Somatossensoriais Evocados/fisiologia , Junções Comunicantes/efeitos dos fármacos , Camundongos , Neocórtex/efeitos dos fármacos , Canais de Potássio/metabolismo , Convulsões/metabolismo
14.
Artigo em Inglês | MEDLINE | ID: mdl-27574307

RESUMO

Ischaemic stroke is the leading cause of adult disability worldwide. Effective rehabilitation is hindered by uncertainty surrounding the underlying mechanisms that govern long-term ischaemic injury progression. Despite its potential as a sensitive non-invasive in vivo marker of brain function that may aid in the development of new treatments, blood oxygenation level-dependent (BOLD) functional magnetic resonance imaging (fMRI) has found limited application in the clinical research on chronic stage stroke progression. Stroke affects each of the physiological parameters underlying the BOLD contrast, markedly complicating the interpretation of BOLD fMRI data. This review summarizes current progress on application of BOLD fMRI in the chronic stage of ischaemic injury progression and discusses means by which more information may be gained from such BOLD fMRI measurements. Concomitant measurements of vascular reactivity, neuronal activity and metabolism in preclinical models of stroke are reviewed along with illustrative examples of post-ischaemic evolution in neuronal, glial and vascular function. The realization of the BOLD fMRI potential to propel stroke research is predicated on the carefully designed preclinical research establishing an ischaemia-specific quantitative model of BOLD signal contrast to provide the framework for interpretation of fMRI findings in clinical populations.This article is part of the themed issue 'Interpreting BOLD: a dialogue between cognitive and cellular neuroscience'.


Assuntos
Mapeamento Encefálico/métodos , Imagem por Ressonância Magnética/métodos , Acidente Vascular Cerebral/diagnóstico por imagem , Progressão da Doença , Humanos , Oxigênio/sangue , Acidente Vascular Cerebral/fisiopatologia
15.
Front Cell Neurosci ; 10: 192, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27540355

RESUMO

The neurons in the inferior olive express subthreshold oscillations in their membrane potential. This oscillatory activity is known to drive synchronous activity in the cerebellar cortex and plays a role in motor learning and motor timing. In the past years, it was commonly thought that olivary neurons belonged to a unique population of oscillating units and that oscillation properties were exclusively dependent on network settings and/or synaptic inputs. The origin of olivary oscillations is now known to be a local phenomenon and is generated by a combination of conductances. In the present work, we show the existence of at least two neuronal populations that can be distinguished on the basis of the presence or absence of low-voltage activated Ca(2+) channels. The expression of this channel determines the oscillatory behavior of olivary neurons. Furthermore, the number of cells that express this channel is different between sub nuclei of the inferior olive. These findings clearly indicate the functional variability within and between olivary sub nuclei.

16.
Neuroimage ; 104: 110-6, 2015 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-25312775

RESUMO

Potassium homeostasis is fundamental for the physiological functioning of the brain. Increased [K(+)] in the extracellular fluid has a major impact on neuronal physiology and can lead to ictal events. Compromised regulation of extracellular [K(+)] is involved in generation of seizures in animal models and potentially also in humans. For this reason, the investigation of K(+) spatio-temporal dynamics is of fundamental importance for neuroscientists in the field of epilepsy and other related pathologies. To date, the majority of studies investigating changes in extracellular K(+) have been conducted using a micropipette filled with a K(+) sensitive solution. However, this approach presents a major limitation: the area of the measurement is circumscribed to the tip of the pipette and it is not possible to know the spatiotemporal distribution or origin of the focally measured K(+) signal. Here we propose a novel approach, based on wide field fluorescence, to measure extracellular K(+) dynamics in neural tissue. Recording the local field potential from the somatosensory cortex of the mouse, we compared responses obtained from a K(+)-sensitive microelectrode to the spatiotemporal increases in fluorescence of the fluorophore, Asante Potassium Green-2, in physiological conditions and during 4-AP induced ictal activity. We conclude that wide field imaging is a valuable and versatile tool to measure K(+) dynamics over a large area of the cerebral cortex and is capable of capturing fast dynamics such as during ictal events. Moreover, the present technique is potentially adaptable to address questions regarding spatiotemporal dynamics of other ionic species.


Assuntos
Química Encefálica/fisiologia , Neuroimagem/métodos , Potássio/metabolismo , 4-Aminopiridina , Animais , Córtex Cerebral/patologia , Cisterna Magna/patologia , Convulsivantes , Estimulação Elétrica , Eletrodos , Fenômenos Eletrofisiológicos , Líquido Extracelular/metabolismo , Fluorescência , Corantes Fluorescentes , Camundongos , Convulsões/induzido quimicamente , Convulsões/patologia
17.
PLoS Comput Biol ; 8(12): e1002814, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-23271962

RESUMO

The inferior olivary nucleus provides one of the two main inputs to the cerebellum: the so-called climbing fibers. Activation of climbing fibers is generally believed to be related to timing of motor commands and/or motor learning. Climbing fiber spikes lead to large all-or-none action potentials in cerebellar Purkinje cells, overriding any other ongoing activity and silencing these cells for a brief period of time afterwards. Empirical evidence shows that the climbing fiber can transmit a short burst of spikes as a result of an olivary cell somatic spike, potentially increasing the information being transferred to the cerebellum per climbing fiber activation. Previously reported results from in vitro studies suggested that the information encoded in the climbing fiber burst is related to the occurrence of the spike relative to the ongoing sub-threshold membrane potential oscillation of the olivary cell, i.e. that the phase of the oscillation is reflected in the size of the climbing fiber burst. We used a detailed three-compartmental model of an inferior olivary cell to further investigate the possible factors determining the size of the climbing fiber burst. Our findings suggest that the phase-dependency of the burst size is present but limited and that charge flow between soma and dendrite is a major determinant of the climbing fiber burst. From our findings it follows that phenomena such as cell ensemble synchrony can have a big effect on the climbing fiber burst size through dendrodendritic gap-junctional coupling between olivary cells.


Assuntos
Potenciais de Ação , Núcleo Olivar/fisiologia , Animais , Camundongos
18.
Artigo em Inglês | MEDLINE | ID: mdl-23189043

RESUMO

The inferior olive (IO) forms one of the major gateways for information that travels to the cerebellar cortex. Olivary neurons process sensory and motor signals that are subsequently relayed to Purkinje cells. The intrinsic subthreshold membrane potential oscillations of the olivary neurons are thought to be important for gating this flow of information. In vitro studies have revealed that the phase of the subthreshold oscillation determines the size of the olivary burst and may gate the information flow or encode the temporal state of the olivary network. Here, we investigated whether the same phenomenon occurred in murine olivary cells in an intact olivocerebellar system using the in vivo whole-cell recording technique. Our in vivo findings revealed that the number of wavelets within the olivary burst did not encode the timing of the spike relative to the phase of the oscillation but was related to the amplitude of the oscillation. Manipulating the oscillation amplitude by applying Harmaline confirmed the inverse relationship between the amplitude of oscillation and the number of wavelets within the olivary burst. Furthermore, we demonstrated that electrotonic coupling between olivary neurons affect this modulation of the olivary burst size. Based on these results, we suggest that the olivary burst size might reflect the "expectancy" of a spike to occur rather than the spike timing, and that this process requires the presence of gap junction coupling.

19.
PLoS One ; 7(9): e46360, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-23029495

RESUMO

The inferior olivary nucleus (IO) forms the gateway to the cerebellar cortex and receives feedback information from the cerebellar nuclei (CN), thereby occupying a central position in the olivo-cerebellar loop. Here, we investigated the feedback input from the CN to the IO in vivo in mice using the whole-cell patch-clamp technique. This approach allows us to study how the CN-feedback input is integrated with the activity of olivary neurons, while the olivo-cerebellar system and its connections are intact. Our results show how IO neurons respond to CN stimulation sequentially with: i) a short depolarization (EPSP), ii) a hyperpolarization (IPSP) and iii) a rebound depolarization. The latter two phenomena can also be evoked without the EPSPs. The IPSP is sensitive to a GABA(A) receptor blocker. The IPSP suppresses suprathreshold and subthreshold activity and is generated mainly by activation of the GABA(A) receptors. The rebound depolarization re-initiates and temporarily phase locks the subthreshold oscillations. Lack of electrotonical coupling does not affect the IPSP of individual olivary neurons, nor the sensitivity of its GABA(A) receptors to blockers. The GABAergic feedback input from the CN does not only temporarily block the transmission of signals through the IO, it also isolates neurons from the network by shunting the junction current and re-initiates the temporal pattern after a fixed time point. These data suggest that the IO not only functions as a cerebellar controlled gating device, but also operates as a pattern generator for controlling motor timing and/or learning.


Assuntos
Núcleos Cerebelares/fisiologia , Neurônios/fisiologia , Núcleo Olivar/fisiologia , Receptores de GABA-A/fisiologia , Potenciais de Ação/efeitos dos fármacos , Animais , Núcleos Cerebelares/efeitos dos fármacos , Retroalimentação Fisiológica/efeitos dos fármacos , Antagonistas de Receptores de GABA-A/farmacologia , Aprendizagem/efeitos dos fármacos , Aprendizagem/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Microeletrodos , Vias Neurais/efeitos dos fármacos , Vias Neurais/fisiologia , Neurônios/citologia , Neurônios/efeitos dos fármacos , Núcleo Olivar/efeitos dos fármacos , Técnicas de Patch-Clamp , Estilbenos/farmacologia , Transmissão Sináptica/efeitos dos fármacos
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