Abstracts

RATIONALE: During limbic epileptogenesis in vivo, the dentate granule cells (DGC) exhibit increased expression of brain derived neurotrophic factor (BDNF) followed by striking morphologic plasticities, namely formation of basal dendrites and sprouting of mossy fibers. These morphological plasticities may underlie epileptogenesis. Since BDNF and its cognate receptor TrkB exert powerful morphoregulatory effects on diverse types of neurons, we hypothesized that increased expression of BDNF intrinsic to DGC is sufficient to induce these plasticities. By assessing the role played by BDNF in inducing mossy fiber sprouting and basal dendrite formation, we hope to gain insight into the development of epilepsy in patient populations and to elucidate potential drug targets for therapeutic intervention.
METHODS: DGC in hippocampal slice cultures made from P10 rat pups were transfected with BDNF + green fluorescent protein (GFP), nerve growth factor (NGF) + GFP, or GFP + empty vector control using particle-mediated gene transfer. All procedures conformed to NIH and Institutional guidelines for the care and use of animals. NGF served as a negative control as DGC do not express the NGF recptor TrkA. Twenty-four hours after transfection the slice cultures were fixed in paraformaldehyde and neuronal processes were visualized using a Biorad MRC 600 confocal microscope. Confocal images were reconstructed using the Neurolucida system and multiple neuronal parameters were assessed, including dendritic and axonal branch number and dendritic length.
RESULTS: Transfection with BDNF produced significant increases in axonal branch and basal dendrite number relative to NGF or empty vector controls. BDNF-transfection also increased basal dendrite length. Increases in axonal and dendritic branch number were restricted to a region within 50 microns of the soma. Significantly, structural changes were prevented by the tyrosine kinase inhibitor, K252a, indicating that the BDNF effects are likely mediated by the BDNF receptor, TrkB.
CONCLUSIONS: Repeated and/or prolonged focal hippocampal seizures promote limbic epileptogenesis, the process by which a normal brain becomes epileptic. The cascade of gene expression activated by focal hippocampal seizures includes marked increases of BDNF content in multiple neuronal populations including the DGC. Pharmacological and genetic interventions implicate a causal role for BDNF in epileptogenesis. Here we show that increasing expression of BDNF in DGC is sufficient to induce the formation of basal dendrites and axonal sprouting. In conclusion, these morphological consequences of increased BDNF expression may underlie the recurrent excitatory synapses demonstrated among DGC in epileptic animals and thus may constitute one mechanism by which seizure-induced BDNF expression promotes limbic epileptogenesis.
[Supported by: This work was supported by NIH grants NS07370, NS32334 and NINDS grant NS17771. SCD was supported by an NIH NRSA grant and the PhRMA Foundation.]