Microglia and peripheral monocytes contribute to hypersensitivity in rodent models of neuropathic pain. However, the precise respective function of microglia and peripheral monocytes has not been investigated in these models. To address this question, here we combined transgenic mice and pharmacological tools to specifically and temporally control the depletion of microglia and monocytes in a mouse model of spinal nerve transection (SNT). We found that although microglia and monocytes are required during the initiation of mechanical allodynia or thermal hyperalgesia, these cells may not be as important for the maintenance of hypersensitivity. Moreover, we demonstrated that either resident microglia or peripheral monocytes are sufficient in gating neuropathic pain after SNT. We propose that resident microglia and peripheral monocytes act synergistically to initiate hypersensitivity and promote the transition from acute to chronic pain after peripheral nerve injury.

Background and Purpose White matter (WM) ischemic injury, a major neuropathological feature of cerebral small vessel diseases, is an important cause of vascular cognitive impairment in later life. The pathogenesis of demyelination following WM ischemic damage are often accompanied by microglial activation. Fingolimod (FTY720) was approved for the treatment of multiple sclerosis for its immunosuppression property. In this study, we evaluated the neuroprotective potential of FTY720 in a WM ischemia model. Methods Chronic WM ischemic injury model was induced by bilateral carotid artery stenosis (BCAS). Cognitive function, WM integrity, microglial activation and potential pathway involved in microglial polarization were assessed after BCAS. Results Disruption of WM integrity was characterized by demyelination in the corpus callosum and disorganization of Ranvier’s Nodes using Luxol Fast Blue staining, immunofluorescence staining and electron microscopy. In addition, radial maze test demonstrated that working memory performance was decreased at one-month post BCAS-induced injury. Interestingly, FTY720 could reduce cognitive decline and ameliorate the disruption of WM integrity. Mechanistically, cerebral hypoperfusion induced microglial activation, production of associated pro-inflammatory cytokines and priming of microglial polarization toward the M1 phenotype; whereas FTY720 attenuated microglia mediated neuroinflammation after WM ischemia and promoted oligodendrocytogenesis by shifting microglia toward M2 polarization. FTY720’s effect on microglial M2 polarization was largely suppressed by selective STAT3 blockade in vitro, revealing that FTY720-enabled shift of microglia from M1 to M2 polarization state was possibly mediated by STAT3 signaling. Conclusions Our study suggested that FTY720 might be a potential therapeutic drug targeting brain inflammation by skewing microglia toward M2 polarization after chronic cerebral hypoperfusion.

Clinical studies show that chronic pain is accompanied by memory deficits and reduction in hippocampal volume. Experimental studies show that spared nerve injury (SNI) of the sciatic nerve induces long-term potentiation (LTP) at C-fiber synapses in spinal dorsal horn, but impairs LTP in the hippocampus. The opposite changes may contribute to neuropathic pain and memory deficits, respectively. However, the cellular and molecular mechanisms underlying the functional synaptic changes are unclear. Here, we show that the dendrite lengths and spine densities are reduced significantly in hippocampal CA1 pyramidal neurons, but increased in spinal neurokinin-1-positive neurons in mice after SNI, indicating that the excitatory synaptic connectivity is reduced in hippocampus but enhanced in spinal dorsal horn in this neuropathic pain model. Mechanistically, tumor necrosis factor-alpha (TNF-␣) is upregulated in bilateral hippocampus and in ipsilateral spinal dorsal horn, whereas brain-derived neurotrophic factor (BDNF) is decreased in the hippocampus but increased in the ipsilateral spinal dorsal horn after SNI. Importantly, the SNI-induced opposite changes in synaptic connectivity and BDNF expression are prevented by genetic deletion of TNF receptor 1 in vivo and are mimicked by TNF-␣ in cultured slices. Furthermore, SNI activated microglia in both spinal dorsal horn and hippocampus; pharmacological inhibition or genetic ablation of microglia prevented the region-dependent synaptic changes, neuropathic pain, and memory deficits induced by SNI. The data suggest that neuropathic pain involves different structural synaptic alterations in spinal and hippocampal neurons that are mediated by overproduction of TNF-␣ and microglial activation and may underlie chronic pain and memory deficits.Key words: memory deficit; microglia; neuropathic pain; SNI; synaptic plasticity; TNF-␣ Significance StatementChronic pain is often accompanied by memory deficits. Previous studies have shown that peripheral nerve injury produces both neuropathic pain and memory deficits and induces long-term potentiation (LTP) at C-fiber synapses in spinal dorsal horn (SDH) but inhibits LTP in hippocampus. The opposite changes in synaptic plasticity may contribute to chronic pain and memory deficits, respectively. However, the structural and molecular bases of these alterations of synaptic plasticity are unclear. Here, we show that the complexity of excitatory synaptic connectivity and brain-derived neurotrophic factor (BDNF) expression are enhanced in SDH but reduced in the hippocampus in neuropathic pain and the opposite changes depend on tumor necrosis factor-alpha/tumor necrosis factor receptor 1 signaling and microglial activation. The regiondependent synaptic alterations may underlie chronic neuropathic pain and memory deficits induced by peripheral nerve injury.

Summary Peripheral nerve injury causes neuropathic pain accompanied by remarkable microgliosis in the spinal cord dorsal horn. However, it is still debated whether infiltrated monocytes contribute to injury-induced expansion of the microglial population. Here we found that spinal microgliosis predominantly results from local proliferation of resident microglia but not from infiltrating monocytes after spinal nerve transection (SNT), using two genetic mouse models (CCR2RFP/+:CX3CR1GFP/+ and CX3CR1creER/+:R26tdTomato/+ mice) as well as specific staining of microglia and macrophages. Pharmacological inhibition of SNT-induced microglial proliferation correlated with attenuated neuropathic pain hypersensitivities. Microglial proliferation is partially controlled by purinergic and fractalkine signaling, as CX3CR1−/− and P2Y12−/− mice show reduced spinal microglial proliferation and neuropathic pain. These results suggest that local microglial proliferation is the sole source of spinal microgliosis, which represents a potential therapeutic target for neuropathic pain management.

Epilepsy has remained a significant social concern and financial burden globally. Current therapeutic strategies are based primarily on neurocentric mechanisms that have not proven successful in at least a third of patients, raising the need for novel alternative and complementary approaches. Recent evidence implicates glial cells and neuroinflammation in pathogenesis of epilepsy with the promise of targeting these cells to complement existing strategies. Specifically, microglial involvement, as a major inflammatory cell in the epileptic brain, has been poorly studied. In this review, we highlight microglial reaction to experimental seizures, discuss microglial control of neuronal activities, and propose the functions of microglia during acute epileptic phenotypes, delayed neurodegeneration and aberrant neurogenesis. Future research that would help fill in the current gaps in our knowledge including epilepsy-induced alterations in basic microglial functions, neuro-microglial interactions during chronic epilepsy, and microglial contributions to developmental seizures. Studying the role of microglia in epilepsy could inform therapies to better alleviate the disease.

Elevated levels of chemokine C-C motif ligand 2 (CCL2) and its receptor CCR2 have been reported in patients with temporal lobe epilepsy and in experimental seizures. However, the functional significance and molecular mechanism underlying CCL2-CCR2 signaling in epileptic brain remains largely unknown. In this study, we found that the upregulated CCL2 was mainly expressed in hippocampal neurons and activated microglia from mice 1 d after kainic acid (KA)-induced seizures. Taking advantage of double-transgenic mice, we demonstrated that CCL2-CCR2 signaling has a role in resident microglial activation and blood-derived monocyte infiltration. Moreover, seizure-induced degeneration of neurons in the hippocampal CA3 region was attenuated in mice lacking CCL2 or CCR2. We further showed that CCR2 activation induced STAT3 (signal transducer and activator of transcription 3) phosphorylation and IL-1β production, which are critical for promoting neuronal cell death after status epilepticus. Consistently, pharmacological inhibition of STAT3 by WP1066 reduced seizure-induced IL-1β production and subsequent neuronal death. Two weeks after KA-induced seizures, CCR2 deficiency not only reduced neuronal loss, but also attenuated seizure-induced behavioral impairments, including anxiety, memory decline, and recurrent seizure severity. Together, we demonstrated that CCL2-CCR2 signaling contributes to neurodegeneration via STAT3 activation and IL-1β production after status epilepticus, providing potential therapeutic targets for the treatment of epilepsy. Epilepsy is a global concern and epileptic seizures occur in many neurological conditions. Neuroinflammation associated with microglial activation and monocyte infiltration are characteristic of epileptic brains. However, molecular mechanisms underlying neuroinflammation in neuronal death following epilepsy remain to be elucidated. Here we demonstrate that CCL2-CCR2 signaling is required for monocyte infiltration, which in turn contributes to kainic acid (KA)-induced neuronal cell death. The downstream of CCR2 activation involves STAT3 (signal transducer and activator of transcription 3) phosphorylation and IL-1β production. Two weeks after KA-induced seizures, CCR2 deficiency not only reduced neuronal loss, but also attenuated seizure-induced behavioral impairments, including anxiety, memory decline, and recurrent seizure severity. The current study provides a novel insight on the function and mechanisms of CCL2-CCR2 signaling in KA-induced neurodegeneration and behavioral deficits.

Graphical AbstractHighlights d HFS triggers synaptic plasticity of CGRP afferents and chronic pain d LTP-inducible HFS activates spinal microglia through CSF1 signaling d Microglial BDNF is essential for HFS-induced spinal LTP and chronic pain SUMMARY Spinal long-term potentiation (LTP) at C-fiber synapses is hypothesized to underlie chronic pain. However, a causal link between spinal LTP and chronic pain is still lacking. Here, we report that high-frequency stimulation (HFS; 100 Hz, 10 V) of the mouse sciatic nerve reliably induces spinal LTP without causing nerve injury. LTP-inducible stimulation triggers chronic pain lasting for more than 35 days and increases the number of calcitonin gene-related peptide (CGRP) terminals in the spinal dorsal horn. The behavioral and morphological changes can be prevented by blocking NMDA receptors, ablating spinal microglia, or conditionally deleting microglial brain-derived neurotrophic factor (BDNF). HFS-induced spinal LTP, microglial activation, and upregulation of BDNF are inhibited by antibodies against colony-stimulating factor 1 (CSF-1). Together, our results show that microglial CSF1 and BDNF signaling are indispensable for spinal LTP and chronic pain. The microglia-dependent transition of synaptic potentiation to structural alterations in pain pathways may underlie pain chronicity.

BackgroundChronic pain is often accompanied by short-term memory deficit and depression. Currently, it is believed that short-term memory deficit and depression are consequences of chronic pain. Here, we test the hypothesis that the symptoms might be caused by overproduction of interleukin-1beta (IL-1β) in the injured nerve independent of neuropathic pain following spared nerve injury in rats and mice.ResultsMechanical allodynia, a behavioral sign of neuropathic pain, was not correlated with short-term memory deficit and depressive behavior in spared nerve injury rats. Spared nerve injury upregulated IL-1β in the injured sciatic nerve, plasma, and the regions in central nervous system closely associated with pain, memory and emotion, including spinal dorsal horn, hippocampus, prefrontal cortex, nucleus accumbens, and amygdala. Importantly, the spared nerve injury-induced memory deficits, depressive, and pain behaviors were substantially prevented by peri-sciatic administration of IL-1β neutralizing antibody in rats or deletion of IL-1 receptor type 1 in mice. Furthermore, the behavioral abnormalities induced by spared nerve injury were mimicked in naïve rats by repetitive intravenous injection of re combinant rat IL-1β (rrIL-1β) at a pathological concentration as determined from spared nerve injury rats. In addition, microglia were activated by both spared nerve injury and intravenous injection of rrIL-1β and the effect of spared nerve injury was substantially reversed by peri-sciatic administration of anti-IL-1β.ConclusionsNeuropathic pain was not necessary for the development of cognitive and emotional disorders, while the overproduction of IL-1β in the injured sciatic nerve following peripheral nerve injury may be a common mechanism underlying the generation of neuropathic pain, memory deficit, and depression.