CNS-resident macrophages (microglia and CNS-associated macrophages) are the main immunocompetent cells of the central nervous system (CNS) and respond by rapid activation to brain injury. Molecular events occurring during IFNgamma-activation and identification of potential markers of the CNS-resident macrophage subsets were investigated using microglial-derived clones (EOC) differing in their morphology and their antigen presenting activities for CD4+ and CD8+ T-cells. By applying the subtractive process of cDNA representational difference analysis (cRDA), 16 differentially expressed mRNAs were isolated and sequenced, revealing 8 known and 8 novel molecules; 15 of these messages were unpreviously reported in microglia. Two markers of all activated microglial EOC cells were identified (iNOS; IRG-1) and specific subpopulation markers were highlighted, including molecules known to be closely expressed in perivascular spaces. Moreover, some messages could support the distinct morphology, adhesive characteristics, and potential functions of the different clones.