122 Chapter 5 microglial marker), as well as CD31 (an endothelial marker), as revealed by flow cytometry. Ischemia One of the most popular animal models of ischemia is achieved by a unilateral occlusion of the middle cerebral artery (MCAO)155. This intervention induces a progressive inflammatory reaction that is associated with an increase in TSPO at the mRNA and protein levels138,139. To determine the cellular origin of TSPO in this experimental model, 1 week after a 60-min intraluminal occlusion of MCA, TSPO+ labelling was reported in microglial cells (as shown by lectin immunoreactivity) at the site of the core of the ischemia139. At the periphery of the ischemic core, some of the GFAP+ cells were also TSPO+, compared with the contralateral side. Using a similar protocol but with an occlusion time of 90 min, there was a strong TSPO+CD11b+ colocalization indicative of a microglial origin of TSPO138. However, these data are qualitative and an assessment of the colocalization between TSPO and other cell-type markers was not performed. A third study showed an increase in the number of TSPO+ cells in response to the occlusion of MCA as demonstrated by cytometry140. More precisely, the TSPO+ cells expressed microglial markers (Cd11b+CD45int or IBA1+). Interestingly, pretreatment with the TSPO agonist etifoxine helped to contain the size of ischemia, decreased neurological symptoms, and reduced cytokine release in response to the MCAO. These effects were abolished in a model of ischemia combined with a chemical inactivation of microglia140. Thus, it is probable that TSPO from microglia plays a role in inflammation in MCAO models. However, the search for TSPO in other cell types is not constant and will therefore require further investigations to better define, for example, the role of TSPO of astrocytic origin. Multiple sclerosis Experimental autoimmune encephalomyelitis (EAE) and cuprizone (CPZ) intoxication models induce demyelination and proliferation of microglial and astrocytic cells and are thus useful animal models of multiple sclerosis (MS)156,157. An increase in TSPO density has been reported in such animal models, using TSPO radioligands ([3H](R)-PK11195,[18F]DPA714,[18F]GE180)141,144,145. By assessing the colocalization between TSPO and IBA1 or GFAP after exposure to CPZ (administered per os in the animals’ food) for 1, 3 or 5 weeks, virtually all IBA1+ cells expressed TSPO in both control and CPZ-treated animal. In contrast, only a small portion of GFAP+ were TSPO+ positive in control animals, whereas about 35% of GFAP+ cells become TSPO+ after 5 weeks of treatment. These changes were mainly localized in the corpus callosum and, to a lesser extent, in the grey matter cortex144. The CPZ models make it possible to investigate the expression of TSPO during the demyelinating phase (during CPZ treatment), as well as during the remyelination phase (after CPZ treatment). Thus, Zinnhardt et al. (2019) used a 4- and a 6-week CPZ treatment to explore both phases of the CPZ treatment145. They observed increases in TSPO binding in both states compared with the controls. TSPO levels in the demyelination phase are higher than during the remyelination phase. TSPO expression was mainly microglial during the demyelination phase and both microglial and astrocytic during the remyelination phase. These results were based on the colocalization of TSPO+IBA1+ and TSPO+GFAP+, a qualitative finding, without precise quantitative information regarding the relative contribution of these two glial cell types in the alterations in TSPO binding. In a mouse EAE model, photoemulsion of the in vitro binding of[3H](R)-PK11195 on brain sections141 showed that the TSPO radioligand binding was colocalized with OX-42 (a marker of microglia). The authors reported no co-staining between[3H](R)-PK11195 and GFAP. However, in a mouse model of specific TSPO deficiency in astrocytes (hGFAP-driven conditional TSPO
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