Chemogenetic Regulation of CX3CR1-Expressing Microglia Using Gi-DREADD Exerts Sex-Dependent Anti-Allodynic Effects in Mouse Models of Neuropathic Pain

Abstract

Despite growing evidence suggesting that spinal microglia play an important role in the molecular mechanism underlying experimental neuropathic pain (NP) in male rodents, evidence regarding the sex-dependent role of these microglia in NP is insufficient. In this study, we evaluated the effects of microglial regulation on NP using Gi-designer receptors exclusively activated by designer drugs (Gi-DREADD) driven by the microglia-specific Cx3cr1 promoter. For the Cre-dependent expression of human Gi-coupled M4 muscarinic receptors (hM4Di) in CX3C chemokine receptor 1-expressing (CX3CR1 +) cells, R26-LSL-hM4Di-DREADD mice were crossed with CX3CR1-Cre mice. Mouse models of NP were generated by partial sciatic nerve ligation (PSL) and treatment with anti-cancer agent paclitaxel (PTX) or oxaliplatin (OXA), and mechanical allodynia was evaluated using the von Frey test. Immunohistochemistry revealed that hM4Di was specifically expressed on Iba1 + microglia, but not on astrocytes or neurons in the spinal dorsal horn of CX3CR1-hM4Di mice. PSL-induced mechanical allodynia was significantly attenuated by systemic (intraperitoneal, i.p.) administration of 10 mg/kg of clozapine Noxide (CNO), a hM4Di-selective ligand, in male CX3CR1-hM4Di mice. The mechanical threshold in naive CX3CR1-hM4Di mice was not altered by i.p. administration of CNO. Consistently, local (intrathecal, i.t.) administration of CNO (20 nmol) significantly relieved PSL-induced mechanical allodynia in male CX3CR1-hM4Di mice. However, neither i.p. nor i.t. administration of CNO affected PSL-induced mechanical allodynia in female CX3CR1-hM4Di mice. Both i.p. and i.t. administration of CNO relieved PTX-induced mechanical allodynia in male CX3CR1-hM4Di mice, and a limited effect of i.p. CNO was observed in female CX3CR1-hM4Di mice. Unlike PTX-induced allodynia, OXA-induced mechanical allodynia was slightly improved, but not significantly relieved, by i.p. administration of CNO in both male and female CX3CR1-hM4Di mice. These results suggest that spinal microglia can be regulated by Gi-DREADD and support the notion that

Transcriptome profiling of long noncoding RNAs and mRNAs in spinal cord of a rat model of paclitaxel-induced peripheral neuropathy identifies potential mechanisms mediating neuroinflammation and pain

“…Recently, mounting evidence indicates that spinal microglia is clearly activated after paclitaxel treatment, although different dosages of paclitaxel or treatment regime may vary in these studies [ 44 , 50 – 52 ]. More importantly, inhibition of microglia activity further alleviated paclitaxel-induced neuropathic pain [ 53 ], indicating a critical role of spinal microglia in mediating PIPN. At this moment, we have no clues why the study by Zheng et al found no microglial activation in the spinal cord.…”

Chemogenetic Activation of CX3CR1-Expressing Spinal Microglia Using Gq-DREADD Elicits Mechanical Allodynia in Male Mice

“…To evaluate mechanical allodynia, the 50% paw withdrawal threshold was determined using the von Frey test, in accordance with a previously described method [ 18 , 22 ]. Briefly, the mice were individually placed on a metal mesh (5 × 5 mm) grid floor and covered with an acrylic box.…”

“…Recently, Grace et al demonstrated that chemogenetic inhibition of spinal microglia by hM4Di attenuates mechanical allodynia following peripheral nerve injury, while chemogenetic activation of spinal microglia by hM3Dq induces mechanical allodynia in naïve rats using viral gene transfer [ 17 ]. Moreover, we also found that chemogenetic regulation of CX3C chemokine receptor 1 (CX3CR1)-expressing microglia using hM4Di exerts sex-dependent anti-allodynic effects in nerve injury—or paclitaxel-induced NP [ 18 ]. Nevertheless, current knowledge is not sufficient to fully understand the comprehensive roles of microglia leading to NP, and further investigations focusing on sex differences in microglial function would reveal the underlying mechanisms of NP.…”

Optogenetic activation of spinal microglia triggers chronic pain in mice

“…In addition, many studies have shown that genetic knockout of microglial signaling molecules or depletion of microglia partially reverse neuropathic pain [ 6 , 7 ], suggesting microglia as a key player in chronic pain pathogenesis. Indeed, chemogenetic activation of microglia is sufficient to trigger pain hypersensitivity [ 8 ], while chemogenetic inhibition of microglia attenuates chronic pain in rodents [ 9 , 10 ]. Given the complex interactions between neurons, glia, and immune cells in pain modulation, dissecting the specific role of microglia requires the continuous development of new tools.…”