CONICET | Buscador de Institutos y Recursos Humanos

The muscle nicotinic acetylcholine receptor (AChR) has an extracellular domainwhich contains neurotransmitter-binding sites and a transmembrane domain thatforms the ion channel pore and exhibits extensive contacts with the surroundinglipids. The AChR is present in high-density clusters in the muscle cell membranewhere it localizes mainly in liquid-ordered (Lo) domains enriched in cholesterol andsphingolipids. We studied the relationship between AChR and cholesterol in T.californicaAChR-rich membranes, model membranes containing purified AChR andcells expressing AChR and evaluated different experimental conditions: depletionof cholesterol by methyl-b-cyclodextrin, enrichment of cholesterol or cholesterolhemisuccinate (symmetric or asymmetric situations), and oxidation of cholesterolusing cholesterol oxidase. After having analyzed: i) membrane order by anisotropyand GP, ii) augmentation/diminution of Lo domains by GUVs formation andfluorescence microscopy, iii) AChR location in these domains by detergenttreatment and SDS-PAGE, iv) AChR structural conformation by crystal violetfluorescent probe, and v) AChR functionality by electrophysiology, we canconclude that a change in the amount, distribution or oxidation of cholesterolimpacts not only in the size and location of Lo domains and in the AChR preferencefor them, but also in AChR functionality and AChR structural conformation.