title: Caveolin-rich lipid rafts of the plasma membrane of mature cerebellar granule neurons are microcompartments for calcium/reactive oxygen and nitrogen species cross-talk signaling.
abstract: In previous works, we have shown that L-type voltage-operated calcium channels, N-methyl-d-aspartate receptors (NMDAr), neuronal nitric oxide synthase (nNOS) and cytochrome b5 reductase (Cb5R) co-localize within the same lipid rafts-associated nanodomains in mature cerebellar granule neurons (CGN). In this work, we show that the calcium transport systems of the plasma membrane extruding calcium from the cytosol, plasma membrane calcium pumps (PMCA) and sodium-calcium exchangers (NCX), are also associated with these nanodomains. All these proteins were found to co-immunoprecipitate with caveolin-1 after treatment with 25mM methyl-β-cyclodextrin, a lipid rafts solubilizing agent. However, the treatment of CGN with methyl-β-cyclodextrin largely attenuated the rise of cytosolic calcium induced by l-glutamate through NMDAr. Fluorescence energy transfer imaging revealed that all of them are present in sub-microdomains of a size smaller than 200nm, with a peripheral distribution of the calcium extrusion systems PMCA and NCX. Fluorescence microscopy images analysis revealed high calcium dynamic sub-microcompartments near the plasma membrane in fura-2-loaded CGN at short times after addition of l-glutamate. In addition, the close proximity between sources of nitric oxide (nNOS) and superoxide anion (Cb5R) suggests that these nanodomains are involved in the fast and efficient cross-talk between calcium and redox signaling in neurons.
authors.affiliation: Department of Biochemistry and Molecular Biology, Faculty of Sciences, University of Extremadura, 06006 Badajoz, Spain.
authors.forename: D
authors.initials: D
authors.lastname: Marques-da-Silva
authors.affiliation: Department of Biochemistry and Molecular Biology, Faculty of Sciences, University of Extremadura, 06006 Badajoz, Spain. Electronic address: carlosgm@unex.es.
authors.forename: C
authors.initials: C
authors.lastname: Gutierrez-Merino
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020746:Calcium Channels, L-Type / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects; D051242:Caveolin 1 / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology; D042966:Cytochrome-B(5) Reductase / Q000378:metabolism; D018698:Glutamic Acid / Q000494:pharmacology; D021962:Membrane Microdomains / Q000378:metabolism*; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D052248:Nitric Oxide Synthase Type I / Q000378:metabolism; D053499:Plasma Membrane Calcium-Transporting ATPases / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D026361:Reactive Nitrogen Species / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D016194:Receptors, N-Methyl-D-Aspartate / Q000378:metabolism; D019831:Sodium-Calcium Exchanger / Q000378:metabolism; D047392:beta-Cyclodextrins / Q000494:pharmacology
chemical_list: D020746:Calcium Channels, L-Type; D051242:Caveolin 1; D026361:Reactive Nitrogen Species; D017382:Reactive Oxygen Species; D016194:Receptors, N-Methyl-D-Aspartate; D019831:Sodium-Calcium Exchanger; D047392:beta-Cyclodextrins; C108732:methyl-beta-cyclodextrin; D018698:Glutamic Acid; D052248:Nitric Oxide Synthase Type I; D042966:Cytochrome-B(5) Reductase; D053499:Plasma Membrane Calcium-Transporting ATPases; D002118:Calcium
keywords: Calcium microcompartments; Calcium signaling; Caveolin-1; Cerebellar granule neurons; Cytochrome b(5) reductase; L-type voltage-operated calcium channels; Lipid rafts; NMDA receptors; Nitric oxide synthase; Plasma membrane calcium pump; Reactive oxygen and nitrogen species; Sodium–calcium exchanger