PurposeMicroglia represent the primary resident immune cells in the CNS, and have been implicated in the pathology of neurodegenerative diseases. Under basal or “resting” conditions, microglia possess ramified morphologies and exhibit dynamic surveying movements in their processes. Despite the prominence of this phenomenon, the function and regulation of microglial morphology and dynamic behavior are incompletely understood. We investigate here whether and how neurotransmission regulates “resting” microglial morphology and behavior.MethodsWe employed an ex vivo mouse retinal explant system in which endogenous neurotransmission and dynamic microglial behavior are present. We utilized live-cell time-lapse confocal imaging to study the morphology and behavior of GFP-labeled retinal microglia in response to neurotransmitter agonists and antagonists. Patch clamp electrophysiology and immunohistochemical localization of glutamate receptors were also used to investigate direct-versus-indirect effects of neurotransmission by microglia.ResultsRetinal microglial morphology and dynamic behavior were not cell-autonomously regulated but are instead modulated by endogenous neurotransmission. Morphological parameters and process motility were differentially regulated by different modes of neurotransmission and were increased by ionotropic glutamatergic neurotransmission and decreased by ionotropic GABAergic neurotransmission. These neurotransmitter influences on retinal microglia were however unlikely to be directly mediated; local applications of neurotransmitters were unable to elicit electrical responses on microglia patch-clamp recordings and ionotropic glutamatergic receptors were not located on microglial cell bodies or processes by immunofluorescent labeling. Instead, these influences were mediated indirectly via extracellular ATP, released in response to glutamatergic neurotransmission through probenecid-sensitive pannexin hemichannels.ConclusionsOur results demonstrate that neurotransmission plays an endogenous role in regulating the morphology and behavior of “resting” microglia in the retina. These findings illustrate a mode of constitutive signaling between the neural and immune compartments of the CNS through which immune cells may be regulated in concert with levels of neural activity.
Retinal microglia normally occupying uninjured tissue display a continuous, dynamic behavior that suggests functions of tissue surveillance and intercellular communication. Microglial behavior is highly regulated by, and immediately responsive to, focal tissue injury and may constitute a therapeutic cellular response to focal laser photocoagulation. Ex vivo live imaging in the retina is an experimental approach well suited to the study of dynamic aspects of microglial physiology.
PURPOSE Microglia in the central nervous system display a marked structural dynamism in their processes in the resting state. This dynamic behavior, which may play a constitutive surveying role in the uninjured neural parenchyma, is also highly responsive to tissue injury. The role of CX3CR1, a chemokine receptor expressed in microglia, in regulating microglia morphology and dynamic behavior in the resting state and after laser-induced focal injury was examined. METHODS Time-lapse confocal imaging of retinal explants was used to evaluate the dynamic behavior of retinal microglia labeled with green fluorescent protein (GFP). Transgenic mice in which CX3CR1 signaling was ablated (CX3CR1GFP/GFP/CX3CR1−/−) and preserved (CX3CR1+/GFP/CX3CR1+/−) were used. RESULTS Retinal microglial density, distribution, cellular morphology, and overall retinal tissue anatomy were not altered in young CX3CR1−/− animals. In the absence of CX3CR1, retinal microglia continued to exhibit dynamic motility in their processes. However, rates of process movement were significantly decreased, both under resting conditions and in response to tissue injury. In addition, microglia migration occurring in response to focal laser injury was also significantly slowed in microglia lacking CX3CR1. CONCLUSIONS CX3CR1 signaling in retinal microglia, though not absolutely required for the presence of microglial dynamism, plays a role in potentiating the rate of retinal microglial process dynamism and cellular migration. CX3CL1 signaling from retinal neurons and endothelial cells likely modulates dynamic microglia behavior so as to influence the level of microglial surveillance under basal conditions and the rate of dynamic behavior in response to tissue injury.
Purpose-To evaluate the effect of intravitreal ranibizumab on retinal capillary hemangioblastomas (RCHs) associated with von Hippel-Lindau (VHL) disease that are not amenable or responsive to standard therapy.Design-Prospective, non-comparative, interventional case series.Participants-Five patients with VHL associated RCH with exudative changes and visual loss.Methods-Monthly intravitreal injections of ranibizumab (0.5mg) were given over a course of 6 months for a total of 7 injections, with additional injections considered until week 52. The final study visit was designated as 8 weeks after the final study injection.Main Outcome Measures-The primary outcome was the change in best-corrected visual acuity (BCVA) of ≥ 15 letters at the final visit compared to baseline. Secondary outcomes included change in lesion size, exudation as assessed clinically and by fluorescein angiography, change in retinal thickness as evaluated by optical coherence tomography, and adverse event assessments.Results-Patients received an average of 10.0±3.1 injections over an average period of 47±14 weeks including follow-up. Mean change in BCVA was a decrease of 9±20 letters, with 1 patient gaining ≥ 15 letters, and 2 patients losing ≥ 15 letters. Changes in both lesion size and exudation were variable.Conclusions-Intravitreal ranibizumab, delivered as monotherapy every 4 weeks had minimal beneficial effects on most VHL-related RCHs. Possible treatment efficacy was demonstrated in the patient with the smallest lesion with less exudation. Future prospective studies are needed to determine the potential role of an anti-angiogenic agent possibly in combination with other therapies for the treatment of such advanced ocular tumors associated with VHL.
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