Abstracts

Rationale: Posttraumatic epilepsy (PTE) is a common and serious complication of acute traumatic brain injury (TBI), especially after dural penetration. Seizures occur after a latent period of months to years. This delay in epileptic development suggests the initiation of a slow process after injury leading to a permanently epileptic brain. We have modeled a penetrating TBI by transecting the Schaffer collateral (SC) pathway in vivo and in vitro. These lesions have been shown previously to result in delayed axonal sprouting of CA3 pyramidal cells in vitro and an increase in the probability that CA3 cells are connected by excitatory synapses. The extent of this axonal sprouting is correlated with hyperexcitability and transgenic mice with reduced trkB expression are less likely to exhibit axonal sprouting after SC transection. We now extend these findings using mice in which a single amino acid mutation in the trkB receptor (F616A) has been knocked-in rendering it susceptible to pharmacological blockade by 1NMPP1 (Chen et al., 2005). TrkBF616A receptors are fully functional without the drug present and allow for full pharmacological blockade in the presence of the drug. Methods: SC pathway transection was performed in hippocampal slice cultures derived from trkBF616A mice at day in vitro 14; cultures were treated with 1NMPP1 or normal media. Cultures were processed for immunohistochemical and Western blot (WB) analysis of GAP43, a marker for growing axons. In order to determine the contribution of trkB receptors and axonal sprouting under more physiological conditions, we performed SC transections in vivo in trkBF616A mice using a microknife mounted on a stereotaxic carrier. Extracellular recordings of acute physiology slices were carried out to determine hyperexcitability. WB analysis was carried out to determine GAP43 levels after lesion. Results: Our in vitro model revealed that the number of GAP43 immunoreactive fibers in the vicinity of the lesion was significantly reduced in cultures treated with 1NMPP1, compared to untreated cultures. Blockade of the trkB receptor with 1NMPP1 prevented the increase in GAP43 protein levels that were observed after the lesion (n=4, p=0.001, ANOVA).Extracellular recording in area CA3 from acute hippocampal brain slices obtained the in vivo model showed a marked increase in their coastline bursting index indicating they were hyperexcitable (n=6, p=0.001, ANOVA). WB analysis of GAP43 levels indicated an increase GAP43 protein following the lesion as compared to sham controls (n=4, p=0.001, ANOVA). Conclusions: We confirm our previous suggestion that lesion induced neurotrophin-trkB signaling is a critical promoter of axonal sprouting after injury. We are currently treating mice with 1NMPP1 to test the hypothesis further that trkB receptor activation is required for injury-induced axonal sprouting and hyperexcitability. These data will provide a better understanding of the role of trkB receptor signaling and axonal sprouting after TBI and PTE.