Lymphocyte infiltration of neocortex and hippocampus after a single brief seizure in mice

Various immune responses have been described in epileptic patients and animal models of epilepsy, but immune responses in brain after a single seizure are poorly understood. We studied immune responses in brain after a single brief generalized tonic–clonic seizure in mice. C57bl/6 mice, either unanesthetized or anesthetized (pentobarbital, ethyl chloride) received either electrical (15–30 mA, 100 Hz, 1 s) or sham stimulation (subcutaneous electrodes over frontal lobe, no current). Electrical stimulation of unanesthetized mice resulted in tonic–clonic convulsions with hind-limb extension (maximal seizure), tonic–clonic convulsions without hind-limb extension (submaximal seizure), or no seizure. In contrast, such stimulation of anesthetized mice did not result in seizure. Mice were killed at 1 h–7 days after seizure. Brains or regions dissected from brain (neocortex, hippocampus, midbrain, cerebellum) of each group were pooled, single cell suspensions prepared, and cells separated according to density. CD4+ (CD3+CD45Hi) and CD8+ (CD3+CD45Hi) T cell and CD45R+ (CD45Hi) B cell numbers were determined by flow cytometry. At 24 h after a maximal seizure, CD4+ and CD8+ T cells and CD45R+ B cells appeared in brain, reaching peak numbers at 48 h, but were no longer detected at 7 days. CD4+ T cells and CD45R+ B cells were preferentially found in neocortex compared with hippocampus, whereas CD8+ T cells were preferentially found in hippocampus at 24 h after a maximal seizure. In contrast, virtually no lymphocytes were detected in brains of unstimulated or sham stimulated mice, unanesthetized stimulated mice after submaximal or no seizure, and anesthetized stimulated mice at 1 h–7 day. Neither Ly6-G+ neutrophils nor erythrocytes were detected in brains of any animals, nor was there any detectable increase of blood–brain barrier permeability by uptake of Evans Blue dye. The results indicate that lymphocyte entry into brain after a single brief seizure is due to a selective process of recruitment into cortical regions.