Binding of Amyloid ?(1-42)-Calmodulin Complexes to Plasma Membrane Lipid Rafts in Cerebellar Granule Neurons Alters Resting Cytosolic Calcium Homeostasis.

ABSTRACT: Lipid rafts are a primary target in studies of amyloid ? (A?) cytotoxicity in neurons. Exogenous A? peptides bind to lipid rafts, which in turn play a key role in A? uptake, leading to the formation of neurotoxic intracellular A? aggregates. On the other hand, dysregulation of intracellular calcium homeostasis in neurons has been observed in Alzheimer's disease (AD). In a previous work, we showed that A?(1-42), the prevalent A? peptide found in the amyloid plaques of AD patients, binds with high affinity to purified calmodulin (CaM), with a dissociation constant ?1 nM. In this work, to experimentally assess the A?(1-42) binding capacity to intracellular CaM, we used primary cultures of mature cerebellar granule neurons (CGN) as a neuronal model. Our results showed a large complexation of submicromolar concentrations of A?(1-42) dimers by CaM in CGN, up to 120 ± 13 picomoles of A?(1-42) /2.5 × 10⁶ cells. Using fluorescence microscopy imaging, we showed an extensive co-localization of CaM and A?(1-42) in lipid rafts in CGN stained with up to 100 picomoles of A?(1-42)-HiLyteTM-Fluor555 monomers. Intracellular A?(1-42) concentration in this range was achieved by 2 h incubation of CGN with 2 ?M A?(1-42), and this treatment lowered the resting cytosolic calcium of mature CGN in partially depolarizing 25 mM potassium medium. We conclude that the primary cause of the resting cytosolic calcium decrease is the inhibition of L-type calcium channels of CGN by A?(1-42) dimers, whose activity is inhibited by CaM:A?(1-42) complexes bound to lipid rafts.

SUBMITTER: Poejo J 

PROVIDER: S-EPMC7923178 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature