Effects of Membrane Geometry on Voltage-Gated ion Channel Distribution Studied with a Model System

Abstract

Voltage-gated ion channels are inhomogeneously distributed between the highly curved axons and distal dendrites and the relatively flat soma and proximal dendrites. To investigate the effects of membrane geometry on channel distribution and diffusion, we developed a model system based on membrane nano-tubes connected to cell-sized Giant Unilamellar Vesicles (GUVs). KvAP, a bacterial analog of eukaryotic Kv channels [1], was purified, fluorescently labeled and reconstituted into GUVs. Channel density and homogeneity in GUVs were quantified via confocal microscopy while patch-clamp was used to measure the activity of the reconstituted channels. To study the effect of membrane curvature, we pulled a membrane nano-tube from a GUV and could set the tube radius between 10 nm and 200 nm by varying the tension of the GUV membrane. The concentrations of channels in the tube and GUV were measured via confocal microscopy while diffusion was measured by tracking individual channels labeled with quantum dots. As the tube radius decreased, the channel concentration increased while the diffusion coefficient decreased. Results obtained with this model system should give insight into the diffusion of membrane proteins into and out of synaptic boutons.[1] Ruta et al., Nature 2003, 422 : 180-185.