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Aims: Adenosine 2A receptor (A 2A R) is widely expressed in the brain and plays important roles in neuroinflammation, and the nucleotide-binding oligomerization domain, leucine-rich repeat, and pyrin domain-containing protein 3 (NLRP3) inflammasome is a crucial component of the innate immune system while the regulation of A 2A R on it in the central nervous system (CNS) has not been clarified., Methods: The effects of microglial A 2A R on NLRP3 inflammasome assembly and activation were investigated in wild-type, A 2A R- or NLRP3-knockout primary microglia with pharmacological treatment. Microglial A 2A R or NLRP3 conditional knockout mice were used to interrogate the effects of this regulation on neuroinflammation posttraumatic brain injury (TBI)., Results: We found that A 2A R directly interacted with NLRP3 and facilitated NLRP3 inflammasome assembly and activation in primary microglia while having no effects on mRNA levels of inflammasome components. Inhibition of the interaction via A 2A R agonist or knockout attenuated inflammasome assembly and activation in vitro. In the TBI model, microglial A 2A R and NLRP3 were co-expressed at high levels in microglia next to the peri-injured cortex, and abrogating of this interaction by microglial NLRP3 or A 2A R conditional knockout attenuated the neurological deficits and neuropathology post-TBI via reducing the NLRP3 inflammasome activation., Conclusion: Our results demonstrated that inhibition of the interaction between A 2A R and NLRP3 in microglia could mitigate the NLRP3 inflammasome assembly and activation and ameliorate the neuroinflammation post-TBI. It provides new insights into the effects of A 2A R on neuroinflammation regulation post-TBI and offers a potential target for the treatment of NLRP3 inflammasome-related CNS diseases., (© 2023 The Authors. CNS Neuroscience & Therapeutics published by John Wiley & Sons Ltd.)

The formation of fear memory is crucial in emotional disorders such as PTSD and anxiety. Traumatic brain injury (TBI) can cause emotional disorders with dysregulated fear memory formation; however, their cross-interaction remains unclear and hurdled the treatment against TBI-related emotional disorders. While adenosine A2A receptor(A2AR) contributes to the physiological regulation of fear memory, this study aimed to evaluate the A2AR role and possible mechanisms in post-TBI fear memory formation using a craniocerebral trauma model, genetically modified A2AR mutant mice, and pharmacological A2AR agonist CGS21680 and antagonist ZM241385. Our finding showed (i) TBI enhanced mice freezing levels (fear memory) at seven days post-TBI; (ii) The A2AR agonist CGS21680 enhanced the post-TBI freezing levels; conversely, the A2AR antagonist ZM241385 reduced mice freezing level; further (iii) Genetic knockdown of neuronal A2AR in the hippocampal CA1, CA3, and DG regions reduced post-TBI freezing levels, while A2AR knockout in DG region yielded the most reduction in fear memory; finally, (iv) AAV-CaMKII-Cre virus-mediated DG deletion of A2AR on excitatory neurons led to a significant decreased freezing levels post-TBI. These findings indicate that brain trauma increases fear memory retrieval post-TBI, and A2AR on DG excitatory neurons plays a crucial role in this process. Importantly, inhibition of A2AR attenuates fear memory enhancement, which provides a new strategy to prevent fear memory formation/enhancement after TBI., Competing Interests: Declaration of Competing Interest The authors declare no competing interests., (Copyright © 2023. Published by Elsevier Inc.)

Traumatic brain injury (TBI) survivors suffer from long-term disability and neuropsychiatric sequelae due to irreparable brain tissue destruction. However, there are still few efficient therapies to promote neurorestoration in damaged brain tissue. This study aimed to investigate whether the pro-oncogenic gene ski can promote neurorestoration after TBI. We established a ski-overexpressing experimental TBI mouse model using adenovirus-mediated overexpression through immediate injection after injury. Hematoxylin-eosin staining, MRI-based 3D lesion volume reconstruction, neurobehavioral tests, and analyses of neuronal regeneration and astrogliosis were used to assess neurorestorative efficiency. The effects of ski overexpression on the proliferation of cultured immature neurons and astrocytes were evaluated using imaging flow cytometry. The Ski protein level increased in the perilesional region at 3 days post injury. ski overexpression further elevated Ski protein levels up to 14 days post injury. Lesion volume was attenuated by approximately 36-55% after ski overexpression, with better neurobehavioral recovery, more newborn immature and mature neurons, and less astrogliosis in the perilesional region. Imaging flow cytometry results showed that ski overexpression elevated the proliferation rate of immature neurons and reduced the proliferation rate of astrocytes. These results show that ski can be considered a novel neurorestoration-related gene that effectively promotes neurorestoration, facilitates neuronal regeneration, and reduces astrogliosis after TBI., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

Accumulating evidence has revealed the adenosine 2A receptor is a key tuner for neuropathological and neurobehavioral changes following traumatic brain injury by experimental animal models and a few clinical trials. Here, we highlight recent data involving acute/sub-acute and chronic alterations of adenosine and adenosine 2A receptor-associated signaling in pathological conditions after trauma, with an emphasis of traumatic brain injury, including neuroinflammation, cognitive and psychiatric disorders, and other severe consequences. We expect this would lead to the development of therapeutic strategies for trauma-related disorders with novel mechanisms of action., (Copyright © 2023. Published by Elsevier Inc.)

Effective treatment for cognitive dysfunction after traumatic brain injury (TBI) is lacking in clinical practice. Increased brain-derived neurotrophic factor (BDNF) expression in cognitive circuits can significantly alleviate cognitive dysfunction in animal models of TBI. Selective 5-hydroxytryptamine receptor 6 (5-HT 6 R) agonists significantly increase BDNF expression and improve cognitive function. Therefore, we evaluated the protective effect of a highly selective 5-HT 6 R agonist, WAY-181187, on cognitive dysfunction after TBI. We established a controlled cortical impact model of moderate TBI in rats and performed drug intervention for five consecutive days. Rats had spatial reference memory impairment in the Morris water maze one and four weeks after TBI. BDNF expression in the medial prefrontal cortex (mPFC) and hippocampus decreased two and five weeks after TBI. Additionally, five weeks after TBI, decreases in neuronal dendritic spine density and the proportion of thin, mushroom-shaped dendritic spines and an increased proportion of stubby-type dendritic spines were observed. WAY-181187 administration (3 mg/kg) for five consecutive days after TBI significantly alleviated cognitive dysfunction at one and four weeks (P < 0.001 and P < 0.01), upregulated BDNF expression in the mPFC and hippocampus at two (P < 0.01 and P < 0.05) and five (P < 0.01 and P < 0.001) weeks and increased the dendritic spine density and the proportions of thin, mushroom-shaped dendrites in the mPFC (P < 0.05, P < 0.001 and P < 0.01) and hippocampus (P < 0.05, P < 0.001 and P < 0.05) at five weeks after TBI. Our results confirm that WAY-181187 administration (3 mg/kg) in the acute phase alleviated cognitive dysfunction after TBI, possibly by upregulating BDNF expression in the mPFC and hippocampus, enhancing neuroplasticity., (Copyright © 2022 Elsevier B.V. All rights reserved.)

The purpose of the present study was to investigate the effect of glutamate scavenger oxaloacetate (OA) combined with CGS21680, an adenosine A2A receptor (A2AR) agonist, on acute traumatic brain injury (TBI), and to elucidate the underlying mechanisms. C57BL/6J mice were subjected to moderate-level TBI by controlled cortical impact, and then were treated with OA, CGS21680, or OA combined with CGS21680 at acute stage of TBI. At 24 h post TBI, neurological severity score, brain water content, glutamate concentration in cerebrospinal fluid (CSF), mRNA and protein levels of IL-1β and TNF-α, mRNA level and activity of glutamate oxaloacetate aminotransferase (GOT), and ATP level of brain tissue were detected. The results showed that neurological deficit, brain water content, glutamate concentration in CSF, and the inflammatory cytokine IL-1β and TNF-α production were exacerbated in CGS21680 treated mice. Administrating OA suppressed the rise of both glutamate concentration in CSF and brain water content, and elevated the ATP level of cerebral tissue. More interestingly, neurological deficit, brain edema, glutamate concentration, IL-1β and TNF-α levels were ameliorated significantly in mice treated with OA combined with CGS21680. The combined treatment exhibited better therapeutic effects than single OA treatment. We also observed that GOT activity was enhanced in single CGS21680 treatment group, and both the GOT mRNA level and GOT activity were up-regulated in early-stage combined treatment group. These results suggest that A2AR can improve the efficiency of GOT and potentiate the ability of OA to metabolize glutamate. This may be the mechanism that A2AR activation in combination group augmented the neuroprotective effect of OA rather than aggravated the brain damages. Taken together, the present study provides a new insight for the clinical treatment of TBI with A2AR agonists and OA.

Background: Cognitive impairment in the late stage of traumatic brain injury (TBI) is associated with the NOD-, LRR and pyrin domain-containing protein 3 (NLRP3) inflammasome, which plays an important role in neuroinflammation. Although classical inflammatory pathways have been well-documented in the late stage of TBI (4-8 weeks post-injury), the mechanism by which the NLRP3 inflammasome impairs cognition is still unclear., Methods: Mice lacking the gene encoding for NLRP3 (NLRP3-knockout mice) and their wild-type littermates were used in a controlled cortical impact model of TBI. Levels of NLRP3 inflammasome and inflammatory factors such as IL-1β and HMGB1 were detected in post-injury hippocampal tissue, as well as long-term potentiation. Behaviors were assessed by T-maze test, novel object recognition, and nesting tests. Glycyrrhizin was used to antagonize HMGB1. Calcium imaging were performed on primary neuronal cultures., Results: By using the NLRP3-knockout TBI model, we found that the continuous activation of the NLRP3 inflammasome and high mobility group box 1 (HMGB1) release were closely related to cognitive impairment. We also found that inhibition of HMGB1 improved LTP reduction and cognitive function by increasing the phosphorylation level of the NMDAR1 subunit at serine 896 while reducing NLRP3 inflammasome activation., Conclusion: NLRP3 inflammasome damages memory in the late stage of TBI primarily through HMGB1 upregulation and provides an explanation for the long-term progression of cognitive dysfunction., (© 2021. The Author(s).)

Spatial recognition memory impairment is an important complication after traumatic brain injury (TBI). We previously found that spatial recognition memory impairment can be alleviated in adenosine A 2A receptor knockout (A 2A R KO) mice after TBI, but the mechanism remains unclear. In the current study, we used manganese-enhanced magnetic resonance imaging and the Y-maze test to determine whether the electrical activity of neurons in the retrosplenial cortex (RSC) was reduced and spatial recognition memory was impaired in wild-type (WT) mice after moderate TBI. Furthermore, spatial recognition memory was damaged by optogenetically inhibiting the electrical activity of RSC neurons in WT mice. Additionally, the electrical activity of RSC neurons was significantly increased and spatial recognition memory impairment was reduced in A 2A R KO mice after moderate TBI. Specific inhibition of A 2A R in the ipsilateral RSC alleviated the impairment in spatial recognition memory in WT mice. In addition, A 2A R KO improved autophagic flux in the ipsilateral RSC after injury. In primary cultured neurons, activation of A 2A R reduced lysosomal-associated membrane protein 1 and cathepsin D (CTSD) levels, increased phosphorylated protein kinase A and phosphorylated extracellular signal-regulated kinase 2 levels, reduced transcription factor EB (TFEB) nuclear localization and impaired autophagic flux. These results suggest that the impairment of spatial recognition memory after TBI may be associated with impaired autophagic flux in the RSC and that A 2A R activation may reduce lysosomal biogenesis through the PKA/ERK2/TFEB pathway to impair autophagic flux., (© 2020 The Authors. Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)

Traumatic brain injury-induced acute lung injury (TBI-ALI) is a serious complication of traumatic brain injury (TBI). Our previous clinical study found that high levels of blood glutamate after TBI were closely related to the occurrence and severity of TBI-ALI, while it remains unknown whether a high concentration of blood glutamate directly causes or aggravates TBI-ALI. We found that inhibition of the adenosine A2A receptor (A2AR) after brain injury alleviated the TBI-ALI; however, it is unknown whether lowering blood glutamate levels in combination with inhibiting the A2AR would lead to better effects. Using mouse models of moderate and severe TBI, we found that intravenous administration of L-glutamate greatly increased the lung water content, lung-body index, level of inflammatory markers in bronchoalveolar lavage fluid and acute lung injury score and significantly decreased the PaO2/FiO2 ratio. Moreover, the incidence of TBI-ALI and the mortality rate were significantly increased, and the combined administration of A2AR activator and exogenous glutamate further exacerbated the above damaging effects. Conversely, lowering the blood glutamate level through peritoneal dialysis or intravenous administration of oxaloacetate notably improved the above parameters, and a further improvement was seen with concurrent A2AR genetic inactivation. These data suggest that A2AR activation aggravates the damaging effect of high blood glutamate concentrations on the lung and that combined treatment targeting both A2AR and blood glutamate may be an effective way to prevent and treat TBI-ALI.

Caffeine is a substance that is consumed worldwide, and it may exert neuroprotective effects against various cerebral insults, including neurotrauma, which is the most prevalent injury among military personnel. To investigate the effects of caffeine on high-intensity blast wave-induced severe blast injury in mice, three different paradigms of caffeine were applied to male C57BL/6 mice with severe whole body blast injury (WBBI). The results demonstrated that chronic caffeine treatment alleviated blast-induced traumatic brain injury (bTBI); however, both chronic and acute caffeine treatments exacerbated blast-induced lung injuries and, more importantly, increased both the cumulative and time-segmented mortalities postinjury. Interestingly, withdrawing caffeine intake preinjury resulted in favorable outcomes in mortality and lung injury, similar to the findings in water-treated mice, and had the trend to attenuate brain injury. These findings demonstrated that although drinking coffee or caffeine preparations attenuated blast-induced brain trauma, these beverages may place personnel in the battlefield at high risk of casualties, which will help us re-evaluate the therapeutic strategy of caffeine application, particularly in multiple-organ-trauma settings. Furthermore, these findings provided possible strategies for reducing the risk of casualties with caffeine consumption, which may help to change the coffee-drinking habits of military personnel., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Excitatory amino acid transporters (EAATs) on cerebral vascular endothelial cells play an important role in maintaining glutamate homeostasis in the brain. The dysfunction of endothelial EAATs is an important reason for the dramatically elevated brain glutamate levels after brain injury, such as traumatic brain injury (TBI). The adenosine A 2A receptor (A 2A R) plays an important role in regulating the brain glutamate level after brain injury; however, researchers have not clearly determined whether this role was related to its ability to regulate endothelial EAATs. Activation of A 2A R in vitro not only decreased the PKA- and glutamate level-dependent strengthening of the interaction between NKA-α1 and the FXYD1 subunit and the subsequent decrease in the activity of Na + /K + -ATPases (NKAs) but also enhanced its interaction with EAATs and ultimately aggravated the reverse transport function of endothelial EAATs under oxygen-glucose deprivation (OGD) conditions. Conversely, inhibition of A 2A R restored the normal transport of EAAT. Moreover, A 2A R inhibition increased NKA activity and decreased its interaction with EAATs in isolated brain capillaries after TBI, further confirming its role in endothelial EAATs in vivo. Based on our results, A 2A R played an important role in regulating endothelial EAAT function, and strategies that restore the normal transport of endothelial EAATs through the inhibition of A 2A R might serve as an effective treatment for brain injury., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Recent studies have shown that after traumatic brain injury (TBI), the number of autophagosomes is markedly increased in brain cells surrounding the wound; however, whether autophagy is enhanced or suppressed by TBI remains controversial. In our study, we used a controlled cortical impact system to establish models of mild, moderate and severe TBI. In the mild TBI model, the levels of autophagy-related protein 6 (Beclin1) and autophagy-related protein 12 (ATG12)-autophagy-related protein 5 (ATG5) conjugates were increased, indicating the enhanced initiation of autophagy. Furthermore, the level of the autophagic substrate sequestosome 1 (SQSTM1) was decreased in the ipsilateral cortex. This result, together with the results observed in tandem mRFP-GFP-LC3 adeno-associated virus (AAV)-infected mice, indicates that autophagosome clearance was also increased after mild TBI. Conversely, following moderate and severe TBI, there was no change in the initiation of autophagy, and autophagosome accumulation was observed. Next, we used chloroquine (CQ) to artificially impair autophagic flux in the injured cortex of the mild TBI model and found that the severity of trauma was obviously exacerbated. In addition, autophagic flux and trauma severity were significantly improved in adenosine A 2A receptor (A 2A R) knockout (KO) mice subjected to moderate TBI. Thus, A 2A R may be involved in regulating the impairment of autophagic flux in response to brain injury. Our findings suggest that whether autophagy is increased after TBI is associated with whether autophagic flux is impaired, and the impairment of autophagic flux exacerbates the severity of trauma. Furthermore, A 2A R may be a target for alleviating the impairment in autophagic flux after TBI.

Traumatic brain injury-induced acute lung injury (TBI-ALI) is a serious complication after brain injury for which predictive factors are lacking. In this study, we found significantly elevated blood glutamate concentrations in patients with TBI or multiple peripheral trauma (MPT), and patients with more severe injuries showed higher blood glutamate concentrations and longer durations of elevated levels. Although the increase in amplitude was similar between the two groups, the duration was longer in the patients with TBI. There were no significant differences in blood glutamate concentrations in the patients with MPT with regard to ALI status, but the blood glutamate levels were significantly higher in the patients with TBI-ALI than in those without ALI. Moreover, compared to patients without ALI, patients with TBI showed a clearly enhanced inflammatory response that was closely correlated with the blood glutamate levels. The blood glutamate concentration was also found to be a risk factor (adjusted odds ratio, 2.229; 95% CI, 1.082-2.634) and was a better predictor of TBI-ALI than the Glasgow Coma Scale (GCS) score. These results indicated that dramatically increased blood glutamate concentrations were closely related to the occurrence of TBI-ALI and could be used as a predictive marker for "at-risk" patients.

Traumatic brain injury (TBI) can induce cognitive dysfunction due to the regional accumulation of hyperphosphorylated tau protein (p-tau). However, the factors that cause p-tau to concentrate in specific brain regions remain unclear. Here, we show that AQP4 polarization in the perivascular astrocytic end feet was impaired after TBI, which was most prominent in the ipsilateral brain tissue surrounding the directly impacted region and the contralateral hippocampal CA1 area and was accompanied by increased local p-tau, changes in dendritic spine density and morphology, and upregulation of the adenosine A 2A receptor (A 2A R). The critical role of the A 2A R signaling in these pathological changes was confirmed by alleviation of the impairment of AQP4 polarity and accumulation of p-tau in the contralateral CA1 area in A 2A R knockout mice. Given that p-tau can be released to the extracellular space and that the astroglial water transport via AQP4 is involved in tau clearance from the brain interstitium, our results suggest that regional disruption of AQP4 polarity following TBI may reduce the clearance of the toxic interstitial solutes such as p-tau and lead to changes in dendritic spine density and morphology. This may explain why TBI patients are more vulnerable to cognitive dysfunction.

A series of neurological and psychiatric symptoms occur after traumatic brain injury (TBI), with cognitive dysfunction being one of the most prominent sequela. Given that tau hyperphosphorylation is an important cause of cognitive impairment in patients of Alzheimer's disease, our present study detected the presence of hyperphosphorylated tau (p-tau), mainly at Ser404, in multiple brain regions, including the ipsilateral parietal cortex, contralateral hippocampus and prefrontal cortex, immediately after the injury in a mouse TBI model; these changes lasted for at least 4w. All of these brain regions play important roles in working memory. Hyperphosphorylated tau protein was primarily located in neurons and was accompanied by axonal injury and dendritic spine degeneration. Our study demonstrated that p-tau spreads gradually and selectively from the injured cortex to other brain regions after TBI and that all of the affected regions are part of the working memory circuit. These findings provide experimental support for the role of p-tau in cognitive impairment in the early phase after TBI., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Objective: To investigate the effects of three different ways of chronic caffeine administration on blast- induced memory dysfunction and to explore the underlying mechanisms., Methods: Adult male C57BL/6 mice were used and randomly divided into five groups: control: without blast exposure, con-water: administrated with water continuously before and after blast-induced traumatic brain injury (bTBI), con-caffeine: administrated with caffeine continuously for 1 month before and after bTBI, pre-caffeine: chronically administrated with caffeine for 1 month before bTBI and withdrawal after bTBI, post-caffeine: chronically administrated with caffeine after bTBI. After being subjected to moderate intensity of blast injury, mice were recorded for learning and memory performance using Morris water maze (MWM) paradigms at 1, 4, and 8 weeks post-blast injury. Neurological deficit scoring, glutamate concentration, proinflammatory cytokines production, and neuropathological changes at 24 h, 1, 4, and 8 weeks post-bTBI were examined to evaluate the brain injury in early and prolonged stages. Adenosine A1 receptor expression was detected using qPCR., Results: All of the three ways of chronic caffeine exposure ameliorated blast-induced memory deficit, which is correlated with the neuroprotective effects against excitotoxicity, inflammation, astrogliosis and neuronal loss at different stages of injury. Continuous caffeine treatment played positive roles in both early and prolonged stages of bTBI; pre-bTBI and post-bTBI treatment of caffeine tended to exert neuroprotective effects at early and prolonged stages of bTBI respectively. Up-regulation of adenosine A1 receptor expression might contribute to the favorable effects of chronic caffeine consumption., Conclusion: Since caffeinated beverages are widely consumed in both civilian and military personnel and are convenient to get, the results may provide a promising prophylactic strategy for blast-induced neurotrauma and the consequent cognitive impairment.