A microfluidics platform for studying ion and solute transport by isolated synaptic vesicles.

Loading of synaptic vesicles with neurotransmitters, an essential step in synaptic transmission, is mediated by vesicular neurotransmitter transporters that are fuelled by an electrochemical proton gradient across the vesicle membrane. While advances have recently been made in the structural and mechanistic understanding of these transporters, it is difficult to study transport kinetics and associated ion fluxes, largely due to technical constraints in accessing intracellular transporters. Here we describe a microfluidics platform that enables rapid solution changes, thus resolving solute fluxes across the vesicle membrane. Antibodies directed against general or subclass-specific vesicle proteins are immobilized on functionalized glass slides, followed by immunocapture of synaptic vesicles yielding a vesicle monolayer virtually free of contamination. After mounting in a specially designed microfluidics setup, acidification of vesicles, as measured by fluorescent reporters, can be repetitively switched on and off with a time resolution of seconds, demonstrating the versatility of the system.