, 2010) supports the notion that ET pores are maintained open in a long lasting manner. Cell-attached recordings, during which ET has been applied inside the recording patch-clamp Pexidartinib nmr pipette, have shown that ET induces large transmembrane unitary currents on granule cells in organotypic cerebellar slices (Lonchamp et al., 2010). The corresponding unitary conductance of which has been estimated around ∼270 pS. Such a conductance is larger than that of most endogenous channels in neuron, except the Ca2+-dependent K channels (also termed big K) that may reach 150 up to 250 pS.

However, at variance of most endogenous ionic channels, no voltage dependence has been detected in ET-induced currents (Lonchamp et al., 2010). The conductance of ∼270 pS induced by ET in granule cell is compatible with that determined in bilayers membrane (∼480 pS, Nestorovich et al., 2010; ∼550 pS, Petit et al., 2001). Similar as for many cytolysins of bacterial origin, lipidic environment in plasma membrane impacts GW-572016 chemical structure the effects of ET. Overall, the integrity of the plasma membrane is needed for ET to exert its effects (Dorca-Arévalo et al., 2012; Nagahama and Sakurai, 1992; Petit et al., 1997). Studies made using

liposomes devoid of specific receptor have suggested that membrane fluidity plays an important role in the interaction of ET with liposomes, insertion in the membrane bilayer, and assembly into complex process in the bilayer (Nagahama et al., 2006; Petit et al., 2001). Reminiscent of data obtained using renal cells (Chassin et al., 2007; Miyata et al., 2002; Petit et al., 1997) the cholesterol sequestration by methyl-β-cyclodextrin (mβCD) does not prevent ET binding onto target neural cells as assessed by immuno-staining

of ET on cerebellum slices or cultured granule cells (Lonchamp et al., 2010). Note, however, that a decrease in 35S-ET binding on rat synaptosomes has been reported (Miyata et al., 2002). These results are consistent with single-molecule Florfenicol tracking experiments made on ET at the apical membrane of MDCK cells, which have shown that the ET binding onto plasma membranes does not require presence of cholesterol (Türkcan et al., 2012). Therefore, the cholesterol is dispensable for ET binding to its receptor. This is not the case for the subsequent steps. In the one hand, pre-incubation of renal cells with mβCD prevents ET-oligomerization and ET-induced cytotoxicity (reviewed by Popoff, 2011a), and mβCD prevents ET-oligomerization in synaptosomal membranes fractions (Miyata et al., 2002). In the other hand, the oligomerization process and the pore formation (see below) can occur in artificial membrane in absence of cholesterol (Nagahama et al., 2006; Petit et al., 2001). The contradiction between these different insights is only apparent, and has recently received an explanation.