Abstracts

Rationale: In order to identify time dependent physiological changes underlying epileptogenesis, we have previously investigated time-course of hippocampal gene expression profiles of postnatal day (P)15 and P30 rats after kainic acid-induced status epilepticus (SE) using high-density oligonucleotide arrays. We selected for differentially regulated genes (P30/P15) by capitalizing on age-dependent physiological responses to KA; KA-SE causes hippocampal neurodegeneration and development of a chronic epileptic state in mature rats (P30), while KA causes neither cell death nor chronic spontaneous seizures in younger animals prior to P21. Genes associated with the mitogen activated protein kinase (MAPK) pathways and inflammation related genes were among the most significantly regulated after KA-SE. Gene expressions were more robust and more sustained in P30 compared to P15. One gene that belonged to both MAPK and inflammation pathway and that displayed a particularly striking gene expression profile after KA-SE, exhibiting an exponentially increasing trend near 240h only in P30 animals was CD74. We performed immunohistochemistry to verify our microarray data and to localize CD74 expression in P15 and P30 rat brains and also in surgically resected cortices from patients with intractable childhood epilepsy. Methods: Long Evans rats were injected with KA at P15 (3 mg/kg, i.p.) or at P30 (10mg/kg). Control littermates were injected with PBS. Animals that showed generalized convulsions for at least one hour were included in the study and perfused at 24h and 240h (n=3/group/time point). Cortices were from two patients (ages 6 and 12 months) who underwent hemispherectomy for drug-resistant multiple daily seizures. Immunohistochemistry was performed using anti-rat and anti-human CD74 antibodies (Sata Cruz). Results: In parallel to microarray data, prominent CD74 expressing cells were detected within the hippocampus only in P30 rats at 240h. In addition, intensely CD74 cells of glia morphology were symmetrically present surrounding white matter tracts including internal capsule, fornix, and anterior commissure and within reuniens thalamic nucleus. Widespread CD74 cells were detected in both white matter and gray matter of resected cortices. They have morphology of both neurons and glia. Conclusions: CD74 immunohistochemistry validated our microarray time-dependent expression data of CD74. The presence of cells expressing CD74 was specifically prominent in P30 KA rats at 240h in agreement with the exponential increase in CD74 mRNA noted at 240h after KA in P30 hippocampus. Also, we observed a distinct path of activation for CD74, beginning around the internal capsule and ending near the anterior commissure. CD 74 is a receptor for macrophage migration inhibitory factor (MIF), a proinflammatory cytokine. Binding of MIF to CD74 is required to activate MIF-mediated signal transduction pathways. Delayed activation of CD 74 after KA-SE in adult animals suggests that CD74 protein may play a functional role to regulate seizure-induced inflammation and immune response.