Cholesterol, a very good ally of nicotinic receptors

CAMILA FABIANI; DAÑEL ALEJANDRO PEÑALVA; JEREMÍAS CORRADI; SILVIA SUSANA ANTOLLINI

Congreso; XLVII Reunión Anual de la Sociedad Argentina de Biofísica; 2018

The muscle nicotinic acetylcholine receptor (AChR) has an extracellular domain which contains neurotransmitter-binding sites and a transmembrane domain that forms the ion channel pore and exhibits extensive contacts with the surrounding lipids. The AChR is present in high-density clusters in the muscle cell membrane where it localizes mainly in liquid-ordered (Lo) domains enriched in cholesterol and sphingolipids. We studied the relationship between AChR and cholesterol in T. californica AChR-rich membranes, model membranes containing purified AChR and cells expressing AChR and evaluated different experimental conditions: depletion of cholesterol by methyl--cyclodextrin, enrichment of cholesterol or cholesterol-hemisuccinate (symmetric or asymmetric situations), and oxidation of cholesterol using cholesterol oxidase. After having analyzed: i) membrane order by anisotropy and GP, ii) augmentation/diminution of Lo domains by GUVs formation and fluorescence microscopy, iii) AChR location in these domains by detergent treatment and SDS-PAGE, iv) AChR structural conformation by crystal violet fluorescent probe, and v) AChR functionality by electrophysiology, we can conclude that a change in the amount, distribution or oxidation of cholesterol impacts not only in the size and location of Lo domains and in the AChR preference for them, but also in AChR functionality and AChR structural conformation