Probes for GFAP

Paralleling the activation of dorsal horn microglia after peripheral nerve injury is a significant expansion and proliferation of macrophages around injured sensory neurons in dorsal root ganglia (DRG). Here we demonstrate a critical contribution of DRG macrophages, but not those at the nerve injury site, to both the initiation and maintenance of the mechanical hypersensitivity that characterizes the neuropathic pain phenotype. In contrast to the reported sexual dimorphism in the microglial contribution to neuropathic pain, depletion of DRG macrophages reduces nerve injury-induced mechanical hypersensitivity and expansion of DRG macrophages in both male and female mice. However, fewer macrophages are induced in the female mice and deletion of colony-stimulating factor 1 from sensory neurons, which prevents nerve injury-induced microglial activation and proliferation, only reduces macrophage expansion in male mice. Finally, we demonstrate molecular cross-talk between axotomized sensory neurons and macrophages, revealing potential peripheral DRG targets for neuropathic pain management

Spinal astrocytes contribute to chronic itch via sensitization of itch-specific neurons expressing gastrin-releasing peptide receptor (GRPR). However, whether microglia-neuron interactions contribute to itch remains unclear. In this study, we aimed to explore how microglia interact with GRPR+ neurons and promote chronic itch.RNA sequencing, quantitative real-time PCR, western blot, immunohistochemistry, RNAscope ISH, pharmacologic and genetic approaches were performed to examine the roles of spinal NLRP3 (The NOD-like receptor family, pyrin-containing domain 3) inflammasome activation and IL-1β-IL1R1 signaling in chronic itch. Grpr-eGFP and Grpr KO mice were used to investigate microglia-GRPR+ neuron interactions.We observed NLRP3 inflammasome activation and IL-1β production in spinal microglia under chronic itch conditions. Blockade of microglial activation and the NLRP3/caspase-1/IL-1β axis attenuated chronic itch and neuronal activation. Type 1 IL-1 receptor (IL-1R1) was expressed in GRPR+ neurons, which are essential for the development of chronic itch. Our studies also find that IL-1β+ microglia are localized in close proximity to GRPR+ neurons. Consistently, intrathecal injection of IL1R1 antagonist or exogenous IL-1β indicate that the IL-1β-IL-1R1 signaling pathway enhanced the activation of GRPR+ neurons. Furthermore, our results demonstrate that the microglial NLRP3/caspase-1/IL-1β axis contributes to several different chronic itches triggered by small molecules and protein allergens from the environment and drugs.Our findings reveal a previously unknown mechanism in which microglia enhances the activation of GRPR+ neurons through the NLRP3/caspase-1/IL-1β/IL1R1 axis. These results will provide new insights into the pathophysiology of pruritus and novel therapeutic strategies for patients with chronic itch.

Nonarteritic anterior ischemic optic neuropathy (NAION) is a common acute optic neuropathy and cause of irreversible vision loss in those older than 50 years of age. There is currently no effective treatment for NAION and yet the biological mechanisms leading to neuronal loss are not fully understood. Glial cells activation and intercommunication mediated by molecules such as gap junction protein Connexin 43 (Cx43) is thought to modulate neuronal fate in central nervous system disorders. In this study, we investigated retinal glial changes and neuronal loss following a novel NAION animal model using a 577 nm laser. We induced unilateral photochemical thrombosis using rose bengal at the optic nerve head vasculature in adult C57BL/6 mice using a 577 nm laser and performed morphometric analysis of the retinal structure using serial in vivo optical coherence tomography (OCT) and histology for glial and neuronal markers. OCT imaging revealed peripapillary thickening of the retinal ganglion cell complex (GCC, baseline: 79.5 ± 1.0 μm, n = 8; NAION: 93.0 ± 2.5 μm, n = 8, P < 0.01) and total retina (baseline: 202.9 ± 2.4 μm, n = 8; NAION: 228.1 ± 6.8 μm, n = 8, P < 0.01) at day 1 after NAION, and significant GCC thinning (baseline 78.3 ± 2.1 μm, n = 6; NAION: 72.2 ± 1.9 μm, n = 5, P < 0.05) at day 21. NAION induced a significant increase in retinal VEGF levels at day 1 (control: 2319 ± 195, n = 5; NAION: 4549 ± 683 gray mean value, n = 5, P < 0.05), which correlated with retinal thickness (r = 0.89, P < 0.05). NAION led to increased mRNA levels for Cx43 (Gj1a) at day 1 (control: 1.291 ± 0.38; NAION: 3.360 ± 0.58 puncta/mm2, n = 5, P < 0.05), which was not associated with changes in mRNA levels of glial fibrillary acidic protein (Gfap) at the same time (control: 2800 ± 0.59; NAION: 4690 ± 0.90 puncta/mm2 n = 5, P = 0.19). Retinal ganglion cell loss at day 21 was confirmed by a 30% decrease in Brn3a+ cells (control: 2844 ± 235; NAION: 2001 ± 264 cells/mm2, n = 4, P < 0.05). We described a novel protocol of NAION induction by photochemical thrombosis using a 577 nm laser, leading to retinal edema and VEGF increase at day 1 and RGCs loss at day 21 after injury, consistent with the pathophysiology of human NAION. Early changes in glial cells intercommunication revealed by increased Cx43+ gap junctions are consistent with a retinal glial role in mediating cell-to-cell signaling after an ischemic insult. Our study demonstrates an early glial response in a novel NAION animal model and reveals glial intercommunication molecules such as Cx43 as a promising therapeutic target in acute NAION.