Temporal profile of clinical signs, histopathologic changes, gene expression and canonical pathways in a kainic acid induced rat model of mesial temporal lobe epilepsy

Mesial temporal lobe epilepsy (MTLE) is a severe neurological condition of unknown pathogenesis for which several animal models have been developed. To attain a better understanding of the underlying molecular mechanisms, and identify potential biomarkers of lesion progression, we used a rat kainic acid (KA) treatment model of MTLE coupled with global gene expression analysis to examine temporal (4hrs, Days 3,14, or 28) gene regulation relative to development of clinical signs and hippocampal histopathological changes.

Following a single KA subcutaneous dose of 9 mg/kg to young adult male rats, 95% survived, 93% exhibited status epilepticus, and 80% eventually developed spontaneous motor seizures, Histopathology included H&E, autofluorescence, Fluoro-Jade B, Timm's, ED-1/CD68, GFAP, doublecortin, and Ki-67. Neuronal degeneration occurred on Day 3 in the hippocampal CA1, CA3, and dentate hilar regions; amygdaloid and thalamic nuclei, and frontoparietotemporal, entorhinal and piriform cortices. Degeneration severity peaked on Day 6 and persisted until Day 28. Aberrant mossy fiber (MF) sprouting was present in the inner molecular layer of dentate gyrus on Days 6 - 28. Microliosis and astrogliosis peaked on Day 28 and generally colocalized with the distribution of neuronal degeneration. Important correlates to human MTLE included induction of spontaneous seizures, more severe neuronal damage of CA1 than CA3 (in contrast to other animal models but similar to humans), hilar neuronal loss, activated microgliosis and astrogliosis, aberrant MF sprouting and dentate granule cell neurogenesis. Aberrant MF sprouting prior to spontaneous motor seizures and reduced seizure frequency with a decrease in aberrant MF sprouting support the hypothesis that MF sprouts are necessary for spontaneous seizure generation and maintenance.

Analysis of filtered gene expression data using Ingenuity Pathways Analysis (Ingenuity Systems®, www.ingenuity.com) revealed that a number of genes pertaining to neuronal plasticity (RhoA, Racl, Cdc42, BDNF, and Trk), neurodegeneration (Caspase3, Calpain l,Bax, a Cytochromec, and Smac/Diablo), and inflammation/immune-response pathways (TNF-a, CCL2, Cox2) were modulated in a temporal fashion after KA treatment. Expression changes for selected genes known to have a role in neuronal plasticity were subsequently validated by quantitative PCR (qPCR). Notably, canonical pathway analysis revealed that a number of genes within the axon guidance signaling canonical pathway were up-regulated from Days 3 to 28 which correlated with aberrant mossy fiber (MF) sprouting observed histologically beginning at Day 6. Importantly, analysis of the gene expression data also identified potential biomarkers for monitoring neurodegeneration (Cox2) and neuronal/synaptic plasticity (Kalrn).