Long-term alteration of calcium homeostatic mechanisms in the pilocarpine model of temporal lobe epilepsy.
The pilocarpine model of temporal lobe epilepsy is an animal model that shares many of the clinical and pathophysiological characteristics of temporal lobe or limbic epilepsy in humans. This model of acquired epilepsy produces spontaneous recurrent seizure discharges following an initial brain injury produced by pilocarpine-induced status epilepticus. Understanding the molecular mechanisms mediating these long lasting changes in neuronal excitability would provide an important insight into developing new strategies for the treatment and possible prevention of this condition. Our laboratory has been studying the role of alterations in calcium and calcium-dependent systems in mediating some of the long-term neuroplasticity changes associated with epileptogenesis. In this study, [Ca(2+)](i) imaging fluorescence microscopy was performed on CA1 hippocampal neurons acutely isolated from control and chronically epileptic animals at 1 year after the induction of epileptogenesis with two different fluorescent dyes (Fura-2 and Fura-FF) having high and low affinities for [Ca(2+)](i). The high affinity Ca(2+) indicator Fura-2 was utilized to evaluate [Ca(2+)](i) levels up to 900 nM and the low affinity indicator Fura-FF was employed for evaluating [Ca(2+)](i) levels above this range. Baseline [Ca(2+)](i) levels and the ability to restore resting [Ca(2+)](i) levels after a brief exposure to several glutamate concentrations in control and epileptic neurons were evaluated. Epileptic neurons demonstrated a statistically significantly higher baseline [Ca(2+)](i) level in comparison to age-matched control animals. This alteration in basal [Ca(2+)](i) levels persisted up to 1 year after the induction of epileptogenesis. In addition, the epileptic neurons were unable to rapidly restore [Ca(2+)](i) levels to baseline following the glutamate-induced [Ca(2+)](i) loads. These changes in Ca(2+) regulation were not produced by a single seizure and were not normalized by controlling the seizures in the epileptic animals with anticonvulsant treatment. Peak [Ca(2+)](i) levels in response to different concentrations of glutamate were the same in both epileptic and control neurons. Thus, glutamate produced the same initial [Ca(2+)](i) load in both epileptic and control neurons. Characterization of the viability of acutely isolated neurons from control and epileptic animals utilizing standard techniques to identify apoptotic or necrotic neurons demonstrated that epileptic neurons had no statistically significant difference in viability compared to age-matched controls. These results provide the first direct measurement of [Ca(2+)](i) levels in an intact model of epilepsy and indicate that epileptogenesis in this model produced long-lasting alterations in [Ca(2+)](i) homeostatic mechanisms that persist for up to 1 year after induction of epileptogenesis. These observations suggest that altered [Ca(2+)](i) homeostatic mechanisms may underlie some aspects of the epileptic phenotype and contribute to the persistent neuroplasticity changes associated with epilepsy.
Raza
M
M
Department of Neurology, Medical College of Virginia, Virginia Commonwealth University, P.O. Box 980599, Richmond, VA 23298-0599, USA.
Pal
S
S
Rafiq
A
A
DeLorenzo
R J
RJ
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002470:Cell Survival / Q000502:physiology; D004195:Disease Models, Animal; D004833:Epilepsy, Temporal Lobe / Q000139:chemically induced / Q000378:metabolism*; D005456:Fluorescent Dyes; D016257:Fura-2; D018698:Glutamic Acid / Q000378:metabolism; D006624:Hippocampus / Q000166:cytology / Q000378:metabolism; D006706:Homeostasis / Q000502:physiology*; D008297:Male; D008856:Microscopy, Fluorescence; D018721:Muscarinic Agonists; D009473:Neuronal Plasticity / Q000502:physiology; D010862:Pilocarpine; D051381:Rats; D013226:Status Epilepticus / Q000139:chemically induced / Q000378:metabolism; D013997:Time Factors
D005456:Fluorescent Dyes; D018721:Muscarinic Agonists; D010862:Pilocarpine; D018698:Glutamic Acid; D002118:Calcium; D016257:Fura-2