Abstracts

Rationale: Recent evidence suggests that immune/inflammatory activation may contribute to the control of neuronal excitability, and may have a role in epileptogenesis and epilepsy (Riazi et al., 2008; Marchi et al., 2009). Microglia have been shown to take part in synapse formation and function with MHC class I and DAP12 (DNAX activating protein of 12 kD) dependent mechanisms (Huh et al., 2000; Boulanger et al., 2001; Bessis et al., 2007; Roumier et al., 2008). Our hypothesis is that similar mechanisms are responsible for the modulation of synapse plasticity during epileptogenesis. Methods: Western blot (WB) and immunocytochemical (ICC) measures of protein expression were performed for activated microglia (CD45, CD67, CD11b) in hippocampus of rats following kainate (KA) status epilepticus (SE) as well as in human tissue harvested from epilepsy surgical resections of adults with temporal lobe epilepsy, and age-matched controls. In addition, changes in microglia-related synaptic factors, including MHC class I antigen, and DAP12 were also evaluated. Results: The microglia activation marker CD45 was upregulated in hippocampus of rats at 48 hours post KA -induced SE, with a 2.5-fold increase (OD CD45/actin = 0.8±0.18; n=4) when compared to controls (OD CD45/actin 0.31±0.16; n=4) (p<0.001). ICC confirmed the presence of activate ameboid looking CD45, CD67 and CD11b positive microglial cells within the dentate gyrus and the CA3 of rat hippocampi at 48-hours post KA-induced SE. A parallel 3.4-fold downregulation of expression of MHCI antigen was observed in post KA-induced SE animals (OD MHCI/actin = 0.24±0.04; n=4) compared to controls (OD MHCI/actin = 0.82±0.012; n=4) (p<0.001). ICC techniques confirmed the loss of MHCI immunoreactivity in the DG and the CA3 of rat hippocampi at 48-hours post KA-induced SE. Furthemore, in the same regions of rat hippocampi, confocal imaging microscopy showed the expression of DAP12 on CD67-positive microglial cells. In human tissue, WB analysis revealed a 1.33-fold increase in the expression of CD45 in the epilepsy specimens (OD CD45/actin = 0.31±0.15; n=5) compared to normal controls (OD CD45/actin 0.231±0.15; n=3) (p<0.05). In human specimens, the presence of ramified CD45-positive microglial cells surrounding NeuN-positive dystrophic neurons was observed. In the same specimens, confocal imaging confirmed the coexpression of CD45 and DAP12 on ramified microglial cells.