Cholesterol Depletion from the Plasma Membrane Impairs Proton and Glutamate Storage in Synaptic Vesicles of Nerve Terminals

We report that cholesterol depletion with methyl-beta-cyclodextrin (M beta CD) acutely applied to rat brain synaptosomes is accompanied by an immediate increase in transporter-mediated glutamate release and decrease in exocytotic release. To clarify the possible mechanisms underlying these phenomena, we investigated the influence of M beta CD on synaptic vesicle acidification and exo/endocytotic process in nerve terminals. As shown by acridine orange fluorescence measurements, the application of M beta CD to synaptosomes, as well as to isolated synaptic vesicles, led to the gradual leakage of the protons from the vesicles, whereas the application of M beta CD complexed with cholesterol stimulated additional vesicle acidification and an increase in Ca2+-dependent exocytotic response. It was found that the treatment of nerve terminals with M beta CD did not block Ca2+-triggered vesicle recycling. We suggest that cholesterol depletion of the plasma membrane with M beta CD induces the removal of cholesterol from the membrane of synaptic vesicles resulting in immediate dissipation of synaptic vesicle proton gradient and redistribution of the neurotransmitter between the vesicular and cytosolic pools. The latter appears to be the main cause of a dramatic decrease in exocytotic and considerable increase in transporter-mediated release of l-[C-14]glutamate.