Veränderte Ca²⁺-abhängige Inaktivierung von spannungsabhängigen Kalziumkanälen bei humaner Epilepsie

Zusammenfassung

Spannungsabhängige Ca²⁺-Kanäle sind eine der Haupteintrittsrouten, durch die Ca²⁺ während Aktionspotenzialen in neuronale Zellen eintreten kann und koppeln hierdurch Ca²⁺-Einstrom an neuronale Aktivität. Neurone können die Menge des Ca²⁺-Einstromes durch Expression eines großen Repertoires von Ca²⁺-Kanälen mit unterschiedlichen biophysikalischen Eigenschaften regulieren. Zusätzlich wird der Ca²⁺-Eintritt durch Ca²⁺-abhängige Inaktivierung von Ca²⁺-Kanälen begrenzt. Experimente in Epilepsietiermodellen sowie in reseziertem Hirngewebe von Epilepsiepatienten legen bisher nahe, dass in hippokampalen Körnerzellen die Dichte von Ca²⁺-Kanälen erhöht ist. Zusätzlich wurde eine zweite funktionell wichtige Veränderung identifiziert. Das Ca²⁺-bindende Protein Calbindin-D_28k ist in Körnerzellen von Patienten mit läsionsassoziierter Epilepsie in normalen Mengen enthalten, während es bei Ammonshornsklerose nahezu völlig verloren geht. Der Verlust von Calbindin-D_28k führt zu einer stark vermehrten Ca²⁺-abhängigen Inaktivierung spannungsabhängiger Ca²⁺-Kanäle. Die intrazelluläre Applikation von aufgereinigtem Calbindin-D_28k verminderte Ca²⁺-abhängige Inaktivierung in diesen Zellen bis zu Werten, die von Patienten mit läsionsassoziierter Epilepsie und normalem Calbindin-D_28k-Gehalt nicht unterscheidbar waren. Diese Experimente führen zu der ungewöhnlichen Hypothese, dass der Verlust von Calbindin-D_28k den Ca²⁺-Einstrom in Neurone während längerer Depolarisationen durch erhöhte Ca²⁺-abhängige Inaktivierung vermindert. Dieser Mechanismus könnte neuroprotektiv sein und zum Überleben von Körnerzellen bei chronischer Epilepsie beitragen. Zusätzlich erlauben Messungen an humanen nativen Zellen grundsätzliche Aussagen über die biophysikalischen Eigenschaften und Modulation humaner Ionenleitfähigkeiten in situ.

Abstract

Voltage-dependent Ca²⁺ channels are one of the main routes of Ca²⁺ entry into neurons during action potentials, thus coupling neuronal activity to Ca²⁺ influx. For fine-tuning the Ca²⁺ entry through voltage-dependent Ca²⁺ channels, neurons can rely on a large repertoire of Ca²⁺ channel subunits and splice variants from which channels with different biophysical properties may be assembled. Furthermore, Ca²⁺ entry is limited by Ca²⁺-dependent inactivation of Ca²⁺ channels. So far, the data from epilepsy patients and experimental models of epilepsy suggest that, in hippocampal granule cells, Ca²⁺ current density is augmented. The underlying mechanisms for this change are not clear. As a second important functional change, the Ca²⁺-binding protein Calbindin-D_28k is lost from dentate granule cells in epilepsy patients with hippocampal sclerosis. In contrast, patients with lesionassociated epilepsy retained normal Calbindin-D_28k levels. A loss of Calbindin-D_28k was accompanied by a markedly augmented Ca²⁺-dependent inactivation of voltage-dependent Ca²⁺ channels. Introducing purified Calbindin-D_28k into granule cells restored Ca²⁺-dependent inactivation to levels observed in cells with normal Calbindin-D_28k content obtained from patients with lesion-associated epilepsy. Thus, by limiting Ca²⁺ influx through an enhanced Ca²⁺-dependent inactivation of voltage-dependent Ca²⁺ channels during prolonged neuronal discharges, the loss of Calbindin-D_28k may constitute a neuroprotective mechanism contributing to survival of dentate granule cells. In addition to such information relevant to the pathophysiology of epilepsy, recordings on human neurons allow to obtain information about the biophysical properties and modulation of human ion channels in native neurons.

Key words

Ca²⁺ channel - Ca²⁺-dependent inactivation - patch-clamp - calbindin-D_28k

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