Time course of dysregulation of calcium homeostasis in acutely isolated CA1 hippocampal pyramidal neurons after pilocarpine-induced Status Epilepticus

Abstract

Glutamate induces excitotoxic damage to hippocampal pyramidal neurons in Status Epilepticus (SE) and epilepsy. In this study, we investigated time course of dysregulation of calcium homeostasis at various intervals after an episode of SE in acutely isolated CA1hippocampal pyramidal. For this purpose, male Sprague-Dawley rats (200 g) were subjected to pilocarpine-induced SE. The SE was blocked after 3 h with diazepam and MK-801. The CA1 pyramidal neurons were acutely isolated at 24 h, 48 h, 6 days, 10 days and one-month post-SE. The basal intracellular calcium levels [Ca++]i from individual neurons were determined by high speed image flourometery using Fura-2. Rise in neuronal [Ca++]i in response to 30 ?M glutamate (1 min) exposure was captured by Merlin image acquisition software. Calcium decay response curves were then constructed and compared with neurons from control animals at the same time points. The basal calcium levels in acutely isolated CA1 pyramidal neurons post SE were significantly higher (then control at different time points. The neurons studied at 24 h, 48 h, and 6 days were not able to restore calcium levels to normal after glutamate exposure. The neurons at 10 days and 1 month were gradually able to restore calcium levels to pre-glutamate levels. SE induced excitotoxic damage to the hippocampal pyramidal neurons that leads to neuronal death and disruption of calcium homeostasis. This is evidenced by an increase in basal [Ca++]i levels and inability to restore [Ca++]i to pre-glutamate levels after exposure. The neurons recover gradually over a period of 10 days after acute SE. This is the first direct evidence of dysregulation of calcium homeostasis in pyramidal neurons isolated from animals subjected to SE.