The Journal of Neurophysiology Vol. 78 No. 2 August 1997, pp. 1135-1143
Copyright ©1997 The American Physiological Society

N-Methyl-D-Aspartate Evokes Rapid Net Depolymerization of Filamentous Actin in Cultured Rat Cerebellar Granule Cells

Spencer L. Shorte

Institut National de la Santé et de la Recherche Médicale (INSERM), Unité 29, Laboratoire de Neurobiologie et Physiopathologie du Développement, Hôpital de Port-Royal, 75014 Paris; and INSERM Unité 261, Institut Pasteur, 75015 Paris, France

Shorte, Spencer L. N-methyl-D-aspartate evokes rapid net depolymerization of filamentous actin in cultured rat cerebellar granule cells. J. Neurophysiol. 78: 1135-1143, 1997. Filamentous actin(F-actin) was measured in cultured rat cerebellum granule neurons with the use of fluorescently labeled phallotoxin as a site-specific probe for F-actin, and fluorescence microscopy. The averaged apparent intensity of soma-associated F-actin-derived fluorescence (F_app) was measured from fixed cells after incubation in either 1) normal Krebs solution containing 2 mM extracellular calcium ([Ca²⁺]ex) or 2) normal Krebs solution plus N-methyl-D-aspartate (NMDA) for 2 min immediately before fixation. NMDA (10, 50, and 100 µM) decreased F_app to 63 ± 5% (mean ± SE), 53 ± 4%, and 47 ± 2%, respectively, of that measured from control cells. This effect was mimicked by treatment of cells with ionomycin. The ability of NMDA to reduce the F_app in the presence of [Ca²⁺]ex was abolished when cells were maintained in [Ca²⁺]ex-free medium. Cells first treated with NMDA for 2 min and then left in normal medium for 30 min before fixation gave F_app fluorescence similar to control values (91 ± 12%). However, if the F-actin polymerization inhibitor cytochalasin D was added to cells immediately after NMDA was removed, the F_app did not recover with time (36 ± 3%). Cells treated for 30 min with cytochalasin D alone showed a small reduction in staining (~20%). It is concluded that the actin polymerization state of rat cerebellar granule neurons is sensitive to changes in intracellular calcium, and that NMDA receptor activation evokes an initial rapid depolymerization of F-actin.

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