Central sensitization and network hyperexcitability of the nociceptive system is a basic mechanism of neuropathic pain. We hypothesize that development of cortical hyperexcitability underlying neuropathic pain may involve homeostatic plasticity in response to lesion-induced somatosensory deprivation and activity loss, and can be controlled by enhancing cortical activity. In a mouse model of neuropathic pain, in vivo two-photon imaging and patch clamp recording showed initial loss and subsequent recovery and enhancement of spontaneous firings of somatosensory cortical pyramidal neurons. Unilateral optogenetic stimulation of cortical pyramidal neurons both prevented and reduced pain-like behavior as detected by bilateral mechanical hypersensitivity of hindlimbs, but corpus callosotomy eliminated the analgesic effect that was ipsilateral, but not contralateral, to optogenetic stimulation, suggesting involvement of inter-hemispheric excitatory drive in this effect. Enhancing activity by focally blocking cortical GABAergic inhibition had a similar relieving effect on the pain-like behavior. Patch clamp recordings from layer V pyramidal neurons showed that optogenetic stimulation normalized cortical hyperexcitability through changing neuronal membrane properties and reducing frequency of excitatory postsynaptic events. We conclude that development of neuropathic pain involves abnormal homeostatic activity regulation of somatosensory cortex, and that enhancing cortical excitatory activity may be a novel strategy for preventing and controlling neuropathic pain.

HIV-1 invades CNS in the early course of infection, which can lead to the cascade of neuroinflammation. NADPH oxidases (NOXs) are the major producers of reactive oxygen species (ROS), which play important roles during pathogenic insults. The molecular mechanism of ROS generation via microRNA-mediated pathway in human microglial cells in response to HIV-1 Tat protein has been demonstrated in this study. Over-expression and knockdown of microRNAs, luciferase reporter assay, and site-directed mutagenesis are main molecular techniques used in this study. A significant reduction in miR-17 levels and increased NOX2, NOX4 expression levels along with ROS production were observed in human microglial cells upon HIV-1 Tat C exposure. The validation of NOX2 and NOX4 as direct targets of miR-17 was done by luciferase reporter assay. The over-expression and knockdown of miR-17 in human microglial cells showed the direct role of miR-17 in regulation of NOX2, NOX4 expression and intracellular ROS generation. We demonstrated the regulatory role of cellular miR-17 in ROS generation through over-expression and knockdown of miR-17 in human microglial cells exposed to HIV-1 Tat C protein. (Tat) is a regulatory protein and among the early HIV proteins to be produced after infection. HIV Tat protein has been reported in serum of HIV-1-infected patients (Westendorp et al. 1995;Goldstein 1996;Xiao et al. 2000), where its concentration can reach up to 300-500 ng/mL (Poggi et al. 2004). HIV Tat protein is released from HIV-1-infected cells (Ensoli et al. 1990)

Background Despite its identification as a key checkpoint regulator of microglial activation in Alzheimer’s disease, the overarching role of CX3CR1 signaling in modulating mechanisms of Aβ driven neurodegeneration, including accumulation of hyperphosphorylated tau is not well understood. Methodology Accumulation of soluble and insoluble Aβ species, microglial activation, synaptic dysregulation, and neurodegeneration is investigated in 4- and 6-month old 5xFAD;Cx3cr1+/+ and 5xFAD;Cx3cr1−/− mice using immunohistochemistry, western blotting, transcriptomic and quantitative real time PCR analyses of purified microglia. Flow cytometry based, in-vivo Aβ uptake assays are used for characterization of the effects of CX3CR1-signaling on microglial phagocytosis and lysosomal acidification as indicators of clearance of methoxy-X-04+ fibrillar Aβ. Lastly, we use Y-maze testing to analyze the effects of Cx3cr1 deficiency on working memory. Results Disease progression in 5xFAD;Cx3cr1−/− mice is characterized by increased deposition of filamentous plaques that display defective microglial plaque engagement. Microglial Aβ phagocytosis and lysosomal acidification in 5xFAD;Cx3cr1−/− mice is impaired in-vivo. Interestingly, Cx3cr1 deficiency results in heighted accumulation of neurotoxic, oligomeric Aβ, along with severe neuritic dystrophy, preferential loss of post-synaptic densities, exacerbated tau pathology, neuronal loss and cognitive impairment. Transcriptomic analyses using cortical RNA, coupled with qRT-PCR using purified microglia from 6 month-old mice indicate dysregulated TGFβ-signaling and heightened ROS metabolism in 5xFAD;Cx3cr1−/− mice. Lastly, microglia in 6 month-old 5xFAD;Cx3cr1−/− mice express a ‘degenerative’ phenotype characterized by increased levels of Ccl2, Ccl5, Il-1β, Pten and Cybb along with reduced Tnf, Il-6 and Tgfβ1 mRNA. Conclusions Cx3cr1 deficiency impairs microglial uptake and degradation of fibrillar Aβ, thereby triggering increased accumulation of neurotoxic Aβ species. Furthermore, loss of Cx3cr1 results in microglial dysfunction typified by dampened TGFβ-signaling, increased oxidative stress responses and dysregulated pro-inflammatory activation. Our results indicate that Aβ-driven microglial dysfunction in Cx3cr1−/− mice aggravates tau hyperphosphorylation, neurodegeneration, synaptic dysregulation and impairs working memory.