PATHOLOGICAL AND PATHOPHYSIOLOGICAL ALTERATIONS IN RAT HIPPOCAMPUS IN MTBI

TBI is one of the most important predisposing factors for acquired epilepsy. In 68% of patients suffering from posttraumatic epilepsy, the epileptic focus resides in temporal lobe structures such as the hippocampus and the amygdala. Yet, the current knowledge of the morphological and functional alterations that occur in these brain regions after TBI leading to epileptogenesis is very limited. Here, we utilized the controlled cortical impact (CCI), a widely used experimental form of closed head injury, to investigate path physiological alterations in the amygdala and hippocampus that underlie the development of posttraumatic epilepsy. Whole-cell recordings from CA1 and BLA pyramidal neurons showed a significant reduction in the frequency and amplitude of GABAA-mediated spontaneous inhibitory postsynaptic currents (IPSCs), seven days after CCI. This reduction was accompanied by a decrease in the levels of the ¿1, ¿2/3 and ¿2 subunits of the GABAA receptor expressed in the membrane, as determined by biotinylation assay and Western blots. We have also observed loss of inhibitory neurons in the BLA whereas we have not detected loss of the total number of neurons. Activation of ¿7 containing nicotinic receptors by fast application of acetylcholine or choline showed that the function of ¿7 nicotinic receptors in the CA1 region was unaltered. Accordingly, there is no change in the level of ¿7 containing nicotinic receptors in the hippocampus one week after CCI. However, in the BLA, the function of ¿7 nicotinic receptors was significantly increased seven days after CCI, suggesting that compensatory mechanisms involving the cholinergic system may take place after CCI. Our results show that CCI exposure causes a significant reduction in the inhibitory tonus of the CA1and BLA regions seven days after CCI. This reduction may significantly underlie the behavioral abnormalities that follow TBI. Genome-wide expression patterns for mRNA, and the microarray (miRNA) which epigenetically influences mRNA activity, can reveal long-term effects of brain injury. An Illumina microarray comprising a comprehensive set of rat expressed genes was interrogated with hippocampal tissue from the ipsilateral side of sham-operated or cortically injured male rats at 1, 7 and 30 days post-injury. The chemokine ligands Ccl2 and Ccl7, the blood-brain-barrier regulator Cldn1, the GABA type b2 receptor (Gabrb2), and the Pou4f1, Klf4 and Klf2 transcription factors were identified as early (Ccl2, Ccl7, Cldn1), intermediate (Gabrb2) and late (Klf2, Klf4 and Pou4f1) markers for the hippocampal response to CCI. Illumina Solexa sequencing revealed 31 significantly up- or down-regulated miRNAs. A series of bioinformatic tools identified miRNAs and putative target mRNAs whose expression was altered inversely after CCI, indicating potential miRNA-dependent repression or de-repression of mRNA after brain injury. Several-fold up-regulation of Ccl2 and Ccl7 24 hours after CCI, subsiding by one week after injury, suggests short-term inflammatory responses which resolve quickly and are not under miRNA control.Altered expression of Gabrb2, changes in GABA receptor trafficking after CCI and Pou4f1/Klf4 suppression might all contribute to excitatory/inhibitory neuronal imbalance in hippocampus with long-term behavioral consequences.