Cholesterol depletion by methyl-beta-cyclodextrin alters axonal behavior of retinal ganglion cells

FIORE, LUCIANO; DI NAPOLI, JENNIFER; RAPACIOLI, MELINA; RODRIGUEZ CELIN, ALEJANDRA; CARRI, NESTOR G; SCICOLONE, GABRIEL

Huerta Grande, Cordoba

Congreso; First Joint Meeting of the Argentine Society for Neurosciences (SAN) and the Argentine Workshop in Neurosciences (TAN). IBRO/LARC; 2009

Argentine Society for Neurosciences (SAN) and the Argentine Workshop in Neurosciences (TAN).

Cholesterol depletion by methyl-beta-cyclodextrin alters axonal behavior of retinal ganglion cells. Fiore, Luciano1; Di Napoli, Jennifer1; Rapacioli, Melina3; Rodriguez Celin, Alejandra3; Carri, Nestor G2; Scicolone, Gabriel1. 1Instituto de Biologia Celular y Neurociencia "Prof Eduardo De Robertis", Facultad de Medicina, UBA-CONICET, Buenos Aires. 2Instituto Multidisciplinario de Biologia Celular -IMBICE- (CIC y CONICET), La Plata. 3Departamento de Ciencias Bioestructurales, Universidad Favaloro, Buenos Aires.                                                                            We have previously demonstrated that EphA3 stimulates axons of retinal ganglion cells to grow toward caudal tectum. We suggested that ephrin-As act as receptors for EphA3 and it is known that ephrin-As are located in cholesterol-rich membrane microdomains (rafts). Our objective was to examine if membrane-cholesterol microenvironment is necessary for axon growth and guidance. We used cultures of chicken embryos retinal explants exposed to methyl-beta-cyclodextrin (MCD) at different concentrations and/or aggregated EphA3Fc. We analyzed the axonal length and morphology, and studied  the expression pattern of EphAs and ephrin-As by immunocytochemistry. Explants exposed to MCD presented an early significant increase in the number of  axonal varicosities and of collapsed growth cones followed by a significant decrease in the axonal length. Ephrin-A2 was reduced in the growth cones and was increased in the axonal varicosities. Explants exposed to EphA3Fc presented a significant increase in the number of expanded growth cones. Exposure to EphA3Fc and MCD together produced intermediate results than those produced by them separately. Results obtained with MCD suggest that cholesterol depletion decreases the ephrin-As located in growth cones and the axonal growth by stimulating endocytosis. The substractive effects of EphA3 and MCD suggest: a) axonal growth mediated by EphA3 is reduced by MCD for resting membrane by endocytosis and/or reducing the ephrin-As in the growth cones, and b) the interaction between ephrin-As and EphA3 reduces the endocytosis mediated by MCD. These data suggest that cholesterol environment  is necessary for regulating the axon guidance mediated by EphA/ephrin-A system.   Funded by CONICET and UBA.