Abstracts

Rationale: The kindling phenomenon is a generally accepted experimental model of epilepsy and epileptogenesis. The PTZ-induced chemical kindling is generally accepted as a model for a generalized epilepsy. Vitamin D has been shown to have neuroprotective effect in neurons, possibly by down-regulating L-type voltage-dependent Ca²⁺-channels. We hypothesized that vitamin D inhibits epileptogenesis and prevents seizure-induced neuronal damage. We used PTZ- kindling model to test the hypothesis. To investigate the effect of vitamin D on PTZ kindling and the resulting neuronal damage in mouse hippocampus. Methods: The CD-1 stain mice were used for the experiments. For kindling, 35 mg/kg PTZ was injected intraperitoneally (IP) once every 48h. After each injection the animals were observed for 20 min and the convulsive behavior was scored (stage 0-5). Vitamin D (1a,25-Dihydroxyvitamin D3; 1 µg/kg IP) or vehicle (ethanol, 2.4 µl/ml/kg IP) was administered every 24h for 5 days before PTZ kindling and then 15 min prior to each PTZ injection (N=16, vitamin D; N=20, vehicle). The animals received 10 PTZ injections during the kindling procedure. Once fully kindled (N=6, vitamin D; N=6, vehicle) regardless of numbers of PTZ injection, animals were examined histologically using cresyl violet stain after 6 consecutive fully kindled seizures. Cresyl violet stains were analyzed to assess neuronal damage by consensus among 3 blinded observers. Hippocampus was divided into 3 sectors for damage analysis. Histological damage score was assigned on a 0-3 grading scale (0= no damage, 1= mild, 2= moderate and 3= severe neuronal damage). The data were statistically analyzed using the Fisher's-exact test for evaluating the differences in survival rate during kindling and the Mann-Whitney U-test for differences between both groups. Results: The vitamin D treated group showed significant inhibition of kindling development from the 3rd kindling injection to the 10th kindling injection (Mann-Whitney U-test P<0.05 or P<0.01) and a higher survival rate (vitamin D 88% vs vehicle 50% at the end of kindling, Fisher's exact test, P<0.01). Cresyl violet staining demonstrated hippocampal neuronal damage in CA1, CA3, hilus, and dentate gyrus in PTZ-kindled animals pretreated with vehicle (N=6), but significantly decreased neuronal damage in PTZ-kindled animals pretreated with vitamin D (N=6) (Mann-Whitney U-test, P<0.05). Conclusions: These results indicate that vitamin D inhibits epileptogenesis in PTZ-kindling and that vitamin D protects hippocampal neurons from damages induced by PTZ-kindled seizures. Better mechanistic understanding of vitamin D effect in kindling could provide novel approaches for more effective therapy in epilepsy to prevent epileptogenesis and to protect from seizure induced neuronal injury.