Traumatic brain injury (TBI) is one of the most common pathological conditions impacting the central nervous system (CNS). A neurological deficit associated with TBI results from a complex of pathogenetic mechanisms including glutamate excitotoxicity, inflammation, demyelination, programmed cell death, or the development of edema. The critical components contributing to CNS response, damage control, and regeneration after TBI are glial cells-in reaction to tissue damage, their activation, hypertrophy, and proliferation occur, followed by the formation of a glial scar. The glial scar creates a barrier in damaged tissue and helps protect the CNS in the acute phase post-injury. However, this process prevents complete tissue recovery in the late/chronic phase by producing permanent scarring, which significantly impacts brain function. Various glial cell types participate in the scar formation, but this process is mostly attributed to reactive astrocytes and microglia, which play important roles in several brain pathologies. Novel technologies including whole-genome transcriptomic and epigenomic analyses, and unbiased proteomics, show that both astrocytes and microglia represent groups of heterogenic cell subpopulations with different genomic and functional characteristics, that are responsible for their role in neurodegeneration, neuroprotection and regeneration. Depending on the representation of distinct glia subpopulations, the tissue damage as well as the regenerative processes or delayed neurodegeneration after TBI may thus differ in nearby or remote areas or in different brain structures. This review summarizes TBI as a complex process, where the resultant effect is severity-, region- and time-dependent and determined by the model of the CNS injury and the distance of the explored area from the lesion site. Here, we also discuss findings concerning intercellular signaling, long-term impacts of TBI and the possibilities of novel therapeutical approaches. We believe that a comprehensive study with an emphasis on glial cells, involved in tissue post-injury processes, may be helpful for further research of TBI and be the decisive factor when choosing a TBI model.

• Klíčová slova
• experimental models, glia, intercellular signaling, neurodegeneration, neuroinflammation, traumatic brain injury,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

INTRODUCTION: Astrocytic Aquaporin 4 (AQP4) and Transient receptor potential vanilloid 4 (TRPV4) channels form a functional complex that likely influences cell volume regulation, the development of brain edema, and the severity of the ischemic injury. However, it remains to be fully elucidated whether blocking these channels can serve as a therapeutic approach to alleviate the consequences of having a stroke. METHODS AND RESULTS: In this study, we used in vivo magnetic resonance imaging (MRI) to quantify the extent of brain lesions one day (D1) and seven days (D7) after permanent middle cerebral artery occlusion (pMCAO) in AQP4 or TRPV4 knockouts and mice with simultaneous deletion of both channels. Our results showed that deletion of AQP4 or TRPV4 channels alone leads to a significant worsening of ischemic brain injury at both time points, whereas their simultaneous deletion results in a smaller brain lesion at D1 but equal tissue damage at D7 when compared with controls. Immunohistochemical analysis 7 days after pMCAO confirmed the MRI data, as the brain lesion was significantly greater in AQP4 or TRPV4 knockouts than in controls and double knockouts. For a closer inspection of the TRPV4 and AQP4 channel complex in the development of brain edema, we applied a real-time iontophoretic method in situ to determine ECS diffusion parameters, namely volume fraction (α) and tortuosity (λ). Changes in these parameters reflect alterations in cell volume, and tissue structure during exposure of acute brain slices to models of ischemic conditions in situ, such as oxygen-glucose deprivation (OGD), hypoosmotic stress, or hyperkalemia. The decrease in α was comparable in double knockouts and controls when exposed to hypoosmotic stress or hyperkalemia. However, during OGD, there was no decrease in α in the double knockouts as observed in the controls, which suggests less swelling of the cellular components of the brain. CONCLUSION: Although simultaneous deletion of AQP4 and TRPV4 did not improve the overall outcome of ischemic brain injury, our data indicate that the interplay between AQP4 and TRPV4 channels plays a critical role during neuronal and non-neuronal swelling in the acute phase of ischemic injury.

• Klíčová slova
• AQP4, ECS diffusion, MRI, TRPV4, brain edema, cerebral ischemia,
• Publikační typ
• časopisecké články MeSH

Brain edema accompanying ischemic or traumatic brain injuries, originates from a disruption of ionic/neurotransmitter homeostasis that leads to accumulation of K(+) and glutamate in the extracellular space. Their increased uptake, predominantly provided by astrocytes, is associated with water influx via aquaporin-4 (AQP4). As the removal of perivascular AQP4 via the deletion of α-syntrophin was shown to delay edema formation and K(+) clearance, we aimed to elucidate the impact of α-syntrophin knockout on volume changes in individual astrocytes in situ evoked by pathological stimuli using three dimensional confocal morphometry and changes in the extracellular space volume fraction (α) in situ and in vivo in the mouse cortex employing the real-time iontophoretic method. RT-qPCR profiling was used to reveal possible differences in the expression of ion channels/transporters that participate in maintaining ionic/neurotransmitter homeostasis. To visualize individual astrocytes in mice lacking α-syntrophin we crossbred GFAP/EGFP mice, in which the astrocytes are labeled by the enhanced green fluorescent protein under the human glial fibrillary acidic protein promoter, with α-syntrophin knockout mice. Three-dimensional confocal morphometry revealed that α-syntrophin deletion results in significantly smaller astrocyte swelling when induced by severe hypoosmotic stress, oxygen glucose deprivation (OGD) or 50 mM K(+). As for the mild stimuli, such as mild hypoosmotic or hyperosmotic stress or 10 mM K(+), α-syntrophin deletion had no effect on astrocyte swelling. Similarly, evaluation of relative α changes showed a significantly smaller decrease in α-syntrophin knockout mice only during severe pathological conditions, but not during mild stimuli. In summary, the deletion of α-syntrophin markedly alters astrocyte swelling during severe hypoosmotic stress, OGD or high K(+).

• MeSH
• akvaporin 4 genetika   metabolismus   MeSH
• astrocyty metabolismus   patologie   MeSH
• biologický transport MeSH
• draslík metabolismus   MeSH
• draslíkové kanály genetika   metabolismus   MeSH
• edém mozku genetika   metabolismus   patologie   MeSH
• gliový fibrilární kyselý protein MeSH
• glukosa nedostatek   MeSH
• konfokální mikroskopie MeSH
• membránové proteiny nedostatek   genetika   MeSH
• mikrotomie MeSH
• mozková kůra metabolismus   patologie   MeSH
• myši transgenní MeSH
• myši MeSH
• osmolární koncentrace MeSH
• osmotický tlak MeSH
• promotorové oblasti (genetika) MeSH
• proteiny nervové tkáně genetika   metabolismus   MeSH
• proteiny vázající vápník nedostatek   genetika   MeSH
• regulace genové exprese MeSH
• signální transdukce MeSH
• stereotaktické techniky MeSH
• svalové proteiny nedostatek   genetika   MeSH
• techniky tkáňových kultur MeSH
• zelené fluorescenční proteiny genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• akvaporin 4 MeSH
• Aqp4 protein, mouse MeSH Prohlížeč
• draslík MeSH
• draslíkové kanály MeSH
• enhanced green fluorescent protein MeSH Prohlížeč
• glial fibrillary astrocytic protein, mouse MeSH Prohlížeč
• gliový fibrilární kyselý protein MeSH
• glukosa MeSH
• membránové proteiny MeSH
• proteiny nervové tkáně MeSH
• proteiny vázající vápník MeSH
• svalové proteiny MeSH
• syntrophin alpha1 MeSH Prohlížeč
• zelené fluorescenční proteiny MeSH

Aquaporin-4 (AQP4) is the primary cellular water channel in the brain and is abundantly expressed by astrocytes along the blood-brain barrier and brain-cerebrospinal fluid interfaces. Water transport via AQP4 contributes to the activity-dependent volume changes of the extracellular space (ECS), which affect extracellular solute concentrations and neuronal excitability. AQP4 is anchored by α-syntrophin (α-syn), the deletion of which leads to reduced AQP4 levels in perivascular and subpial membranes. We used the real-time iontophoretic method and/or diffusion-weighted magnetic resonance imaging to clarify the impact of α-syn deletion on astrocyte morphology and changes in extracellular diffusion associated with cell swelling in vitro and in vivo. In mice lacking α-syn, we found higher resting values of the apparent diffusion coefficient of water (ADCW) and the extracellular volume fraction (α). No significant differences in tortuosity (λ) or non-specific uptake (k'), were found between α-syn-negative (α-syn -/-) and α-syn-positive (α-syn +/+) mice. The deletion of α-syn resulted in a significantly smaller relative decrease in α observed during elevated K(+) (10 mM) and severe hypotonic stress (-100 mOsmol/l), but not during mild hypotonic stress (-50 mOsmol/l). After the induction of terminal ischemia/anoxia, the final values of ADCW as well as of the ECS volume fraction α indicate milder cell swelling in α-syn -/- in comparison with α-syn +/+ mice. Shortly after terminal ischemia/anoxia induction, the onset of a steep rise in the extracellular potassium concentration and an increase in λ was faster in α-syn -/- mice, but the final values did not differ between α-syn -/- and α-syn +/+ mice. This study reveals that water transport through AQP4 channels enhances and accelerates astrocyte swelling. The substantially altered ECS diffusion parameters will likely affect the movement of neuroactive substances and/or trophic factors, which in turn may modulate the extent of tissue damage and/or drug distribution.

• MeSH
• akvaporin 4 metabolismus   MeSH
• astrocyty metabolismus   MeSH
• delece genu * MeSH
• difuze MeSH
• draslík metabolismus   MeSH
• extracelulární prostor metabolismus   MeSH
• genotyp MeSH
• genový knockout MeSH
• ischemie genetika   MeSH
• membránové proteiny genetika   metabolismus   MeSH
• myši knockoutované MeSH
• myši MeSH
• osmotický tlak MeSH
• proteiny vázající vápník genetika   metabolismus   MeSH
• somatosenzorické korové centrum metabolismus   MeSH
• srdeční zástava genetika   metabolismus   MeSH
• svalové proteiny genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• akvaporin 4 MeSH
• draslík MeSH
• membránové proteiny MeSH
• proteiny vázající vápník MeSH
• svalové proteiny MeSH
• syntrophin alpha1 MeSH Prohlížeč

Recently, we have identified two astrocytic subpopulations in the cortex of GFAP-EGFP mice, in which the astrocytes are visualized by the enhanced green-fluorescent protein (EGFP) under the control of the human glial fibrillary acidic protein (GFAP) promotor. These astrocytic subpopulations, termed high response- (HR-) and low response- (LR-) astrocytes, differed in the extent of their swelling during oxygen-glucose deprivation (OGD). In the present study we focused on identifying the ion channels or transporters that might underlie the different capabilities of these two astrocytic subpopulations to regulate their volume during OGD. Using three-dimensional confocal morphometry, which enables quantification of the total astrocytic volume, the effects of selected inhibitors of K⁺ and Cl⁻ channels/transporters or glutamate transporters on astrocyte volume changes were determined during 20 minute-OGD in situ. The inhibition of volume regulated anion channels (VRACs) and two-pore domain potassium channels (K(2P)) highlighted their distinct contributions to volume regulation in HR-/LR-astrocytes. While the inhibition of VRACs or K(2P) channels revealed their contribution to the swelling of HR-astrocytes, in LR-astrocytes they were both involved in anion/K⁺ effluxes. Additionally, the inhibition of Na⁺-K⁺-Cl⁻ co-transporters in HR-astrocytes led to a reduction of cell swelling, but it had no effect on LR-astrocyte volume. Moreover, employing real-time single-cell quantitative polymerase chain reaction (PCR), we characterized the expression profiles of EGFP-positive astrocytes with a focus on those ion channels and transporters participating in astrocyte swelling and volume regulation. The PCR data revealed the existence of two astrocytic subpopulations markedly differing in their gene expression levels for inwardly rectifying K⁺ channels (Kir4.1), K(2P) channels (TREK-1 and TWIK-1) and Cl⁻ channels (ClC2). Thus, we propose that the diverse volume changes displayed by cortical astrocytes during OGD mainly result from their distinct expression patterns of ClC2 and K(2P) channels.

• MeSH
• astrocyty cytologie   účinky léků   metabolismus   MeSH
• biologické modely MeSH
• chloridové kanály metabolismus   MeSH
• draslíkové kanály metabolismus   MeSH
• gliový fibrilární kyselý protein metabolismus   MeSH
• glukosa nedostatek   MeSH
• kotransportéry pro draslík a chloridy MeSH
• kyslík MeSH
• lidé MeSH
• modulátory membránového transportu farmakologie   MeSH
• mozková kůra cytologie   MeSH
• myši transgenní MeSH
• myši MeSH
• pohlavní dimorfismus MeSH
• regulace genové exprese účinky léků   MeSH
• sodík-draslík-chloridové symportéry metabolismus   MeSH
• stanovení celkové genové exprese MeSH
• symportéry metabolismus   MeSH
• techniky in vitro MeSH
• velikost buňky účinky léků   MeSH
• vezikulární transportní proteiny pro glutamát metabolismus   MeSH
• zelené fluorescenční proteiny metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• chloridové kanály MeSH
• draslíkové kanály MeSH
• enhanced green fluorescent protein MeSH Prohlížeč
• gliový fibrilární kyselý protein MeSH
• glukosa MeSH
• kyslík MeSH
• modulátory membránového transportu MeSH
• sodík-draslík-chloridové symportéry MeSH
• symportéry MeSH
• vezikulární transportní proteiny pro glutamát MeSH
• zelené fluorescenční proteiny MeSH