Abstract
Loss of intracellular calcium homeostasis is an established mechanism associated with neuronal dysfunction and status epilepticus. Sequestration of free cytosolic calcium into endoplasmic reticulum by Mg2+/Ca2+ adenosinetriphosphatase (ATPase) is critical for maintenance of intracellular calcium homeostasis. Exposing hippocampal cultures to low-magnesium media is a well-accepted in vitro model of status epilepticus. Using this model, it was shown that endoplasmic reticulum Ca2+ uptake was significantly inhibited in homogenates from cultures demonstrating electrophysiological seizure phenotypes. Calcium uptake was mainly neuronal. However, glial Ca2+ uptake was also significantly inhibited. Viability of neurons exposed to low magnesium was similar to neurons exposed to control solutions. Finally, it was demonstrated that Ca2+ uptake inhibition and intracellular free Ca2+ levels increased in parallel with increasing incubation in low magnesium. The results suggest that inhibition of Mg2+/Ca2+ ATPase-mediated endoplasmic reticulum Ca2+ sequestration contributes to loss of intracellular Ca2+ homeostasis associated with status epilepticus. This study describes for the first time inhibition of endoplasmic reticulum Mg2+/Ca2+ ATPase in a mixed primary hippocampal model of status epilepticus. In combination with animal models of status epilepticus, the cell culture model provides a powerful tool to further elucidate mechanisms that result in inhibition of Mg2+/Ca2+ ATPase and downstream consequences of decreased enzyme activity.
Highlights
Status epilepticus (SE) is a deleterious neurological seizure disorder attributed to a considerable rate of morbidity and mortality in children and adults [1,2]
Whole-cell patch-clamp studies showed that neurons displayed continuous tonic high-frequency epileptiform discharges, similar to electroencephalographic patterns seen in animal models [16,17], throughout the standard
We revealed that inhibition of sarco/endoplasmic reticulum Mg2+ /Ca2+ ATPase (SERCA)-mediated Ca2+ uptake in cortex microsomes was dependent upon N-methyl-D-aspartate (NMDA) receptor activation [21]
Summary
Status epilepticus (SE) is a deleterious neurological seizure disorder attributed to a considerable rate of morbidity and mortality in children and adults [1,2]. Associated with cellular damage and dysfunction in the hippocampus as a result of prolonged seizure activity is loss of intracellular free Ca2+ ([Ca2+ ]i ) homeostasis. This has been demonstrated in both animal [6,7,8] and cell culture [9,10,11] models of SE. While animal models are important for an in vivo understanding of prolonged seizure disorders and have a closer resemblance to human pathology, neuronal cell cultures provide an important in vitro tool for the elucidation of intercellular and intracellular molecular mechanisms associated with SE. Whole-cell patch-clamp studies showed that neurons displayed continuous tonic high-frequency epileptiform discharges, similar to electroencephalographic patterns seen in animal models [16,17], throughout the standard
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