Abstracts

RATIONALE: In patients and models of temporal lobe epilepsy, granule cells sprout aberrant axon collaterals into the granule cell layer (GCL) and molecular layer (ML) of the dentate gyrus. Previous studies provided ultrastructural and functional evidence that sprouted axons synapse with other granule cells. This sort of synaptic reorganization would form a recurrent excitatory network and may be epileptogenic. However, other studies suggested that sprouted axons synapse with inhibitory interneurons. This sort of synaptic reorganization would enhance recurrent inhibition and may be anti-epileptic. To clarify the effects of granule cell axon sprouting we measured the synaptic density and examined the ultrastructure and neurochemistry of the postsynaptic targets of sprouted mossy fibers in epileptic rats.
METHODS: Granule cells in pilocarpine-induced epileptic rats were biocytin-labeled using [italic]in vivo[/italic] intracellular recording techniques. After rats were perfused, hippocampi were isolated, sectioned (40 [mu]m), and processed by the ABC method. Axon projections were 3-D reconstructed and measured at the light microscopic level. Selected regions that included biocytin-labeled mossy fibers in the GCL and ML were further sectioned (80 nm) and processed for EM. Axon segments were 3-D reconstructed from serial EM sections, and the number of synapses formed per axon length was measured. Post-embedding immunostaining was employed to detect GABA-positive structures.
RESULTS: 8 granule cells were examined in 5 epileptic rats. On average, each granule cell projected a summed length of approximately 1 mm of axon into the GCL and 1 mm of axon into the ML. 7 axon segments and 276 synapses were examined at the EM level. 4 axon segments in the ML (409 [mu]m summed length) and 2 axon segments in the GCL (91 [mu]m summed length) were 3-D reconstructed. In both regions axons formed 1 synapse an average of every 2-7 [mu]m. In the ML 97% of the synapses were with dendritic spines, the rest were with dendritic shafts. In the GCL 77% of the synapses were with dendritic spines, 14% were with dendritic shafts, and 9% were with granule cell somata. In both regions over 97% of the synapses were with GABA-negative structures. Integrating the light and EM data suggests that a granule cell[ssquote]s sprouted mossy fibers in the GCL and ML form 280-1000 new synapses, but [lt]3% are with interneurons.
CONCLUSIONS: These findings suggest that almost all of the synapses formed by mossy fibers in the GCL and ML are with other granule cells, and that axon reorganization forms a recurrent excitatory network that may contribute to temporal lobe epileptogenesis.