Abstracts

Rationale: Basket cells in the dentate gyrus normally provide inhibitory control of granule cells. Changes in excitatory synaptic input to basket cells could affect their level of activity and inhibitory influence. We previously found a tendency for decreased frequency of miniature excitatory postsynaptic currents (mEPSC) in basket cells of epileptic pilocarpine-treated rats. Possible mechanisms include decreased probability of glutamate release and loss of synaptic contacts. Synapse loss might occur if basket cells lose or retract dendritic branches after epileptogenic injuries. To test this possibility we reconstructed and measured dendritic arbors of biocytin-labeled dentate basket cells in control and epileptic rats. Methods: Pilocarpine (380 mg/kg) was administered to young adult male Sprague-Dawley rats to induce status epilepticus, which was treated after 2 h with diazepam. Beginning 10 days later rats were video-monitored 40 hr/week for motor seizures and used after at least one spontaneous seizure was observed. Horizontal slices (300 μm thick) were prepared, and putative basket cells recorded and labeled at 32°C using patch pipettes filled with K-gluconate-based internal solution containing 20 mM biocytin. During an experiment basket cells were tentatively identified based on their short action potential duration (0.69 ms ± 0.06, mean ± SEM) and nonadapting high-frequency spike firing. Slices were fixed in 4% paraformaldehyde for at least 24 hrs and processed for biocytin-labeling using a whole-mount protocol. Labeled cells were scanned with a confocal microscope at a magnification just large enough to include the entire dendritic arbor. Stack height was adjusted to include all dendritic processes, and the optical section interval was 3 μm. Cells were reconstructed three dimensionally using a Neurolucida system confocal module. Results: All basket cells (n=6 controls; n=6 epileptics) had a soma located at the hilar border of the dentate gyrus with axon collaterals densely concentrated in the granule cell layer. Most had pyramidal-shaped cell bodies, but some were multipolar. All basket cells had basal dendritic projections into the hilus and apical dendritic projections into the molecular layer. Basket cells in control and epileptic rats had similar soma areas (350 ± 18 vs 343 ± 26 μm²) and a similar number of primary dendrites (4.3 ± 0.3 vs 4.5 ± 0.5) and dendritic ends (37 ± 4 vs 41 ± 5). Total dendritic length was 1.3 times longer in epileptic rats (5104 ± 556 μm) compared to controls (3900 ± 152 μm), but the difference was not statistically significant (p = 0.18). Conclusions: These findings suggest that dendritic length of dentate basket cells is similar or slightly longer in epileptic pilocarpine-treated rats compared to controls. Therefore, reduced frequency of miniature EPSCs in basket cells of epileptic rats is unlikely to be caused by synapse loss after dendritic loss or retraction.