title: Early disruption of the actin cytoskeleton in cultured cerebellar granule neurons exposed to 3-morpholinosydnonimine-oxidative stress is linked to alterations of the cytosolic calcium concentration.
abstract: Cytoskeleton damage is a frequent feature in neuronal cell death and one of the early events in oxidant-induced cell injury. This work addresses whether actin cytoskeleton reorganization is an early event of SIN-1-induced extracellular nitrosative/oxidative stress in cultured cerebellar granule neurons (CGN). The actin polymerization state, i.e. the relative levels of G-/F-actin, was quantitatively assessed by the ratio of the fluorescence intensities of microscopy images obtained from CGN double-labelled with Alexa594-DNase-I (for actin monomers) and Bodipy-FL-phallacidin (for actin filaments). Exposure of CGN to a flux of peroxynitrite as low as 0.5-1μM/min during 30min (achieved with 0.1mM SIN-1) was found to promote alterations of the actin cytoskeleton dynamics as it increases the G-actin/F-actin ratio. Because L-type voltage-operated Ca(2+) channels (L-VOCC) are primary targets in CGN exposed to SIN-1, the possible role of Ca(2+) dynamics on the perturbation of the actin cytoskeleton was also assessed from the cytosolic Ca(2+) concentration response to the L-VOCC's agonist FPL-64176 and to the L-VOCC's blocker nifedipine. The results showed that SIN-1 induced changes in the actin polymerization state correlated with its ability to decrease Ca(2+) influx through L-VOCC. Combined analysis of cytosolic Ca(2+) concentration and G-actin/F-actin ratio alterations by SIN-1, cytochalasin D, latrunculin B and jasplakinolide support that disruption of the actin cytoskeleton is linked to cytosolic calcium concentration changes.
authors.affiliation: Dept. Biochemistry and Molecular Biology, Faculty of Sciences, University of Extremadura, 06006-Badajoz, Spain.
authors.forename: Teresa
authors.initials: T
authors.lastname: Tiago
authors.forename: Dorinda
authors.initials: D
authors.lastname: Marques-da-Silva
authors.forename: Alejandro K
authors.initials: AK
authors.lastname: Samhan-Arias
authors.forename: Manuel
authors.initials: M
authors.lastname: Aureliano
authors.forename: Carlos
authors.initials: C
authors.lastname: Gutierrez-Merino
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002120:Calcium Channel Agonists / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000737:chemistry / Q000378:metabolism; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology; D008981:Molsidomine / Q000031:analogs & derivatives* / Q000494:pharmacology; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D009543:Nifedipine / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects*; D030421:Peroxynitrous Acid / Q000494:pharmacology; D011758:Pyrroles / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar
chemical_list: D002120:Calcium Channel Agonists; D020746:Calcium Channels, L-Type; D011758:Pyrroles; C071658:FPL 64176; D030421:Peroxynitrous Acid; C002385:linsidomine; D008981:Molsidomine; D009543:Nifedipine; D002118:Calcium