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Proteoliposomes were prepared from rat cortical synaptic plasma membranes that retained high and low affinity choline transport. High affinity transport was inhibited by hemicholinium-3 in a competitive manner and was apparently dependent on membrane potential or ion gradients. Proteoliposomes supplemented with an acetylcholine- generating system were able to synthesize [3H]ACh de novo from [3H]choline taken up by high or low affinity transport; however, we were able to specifically load proteoliposomes derived from cholinergic membranes by using a low choline concentration. [3H]ACh, but not [3H]choline, was subsequently released in a calcium-dependent manner by K+ depolarization. This preparation offers a simplified membrane system for studying functional high affinity choline transport and nonvesicular ACh release.

High affinity choline uptake and calcium-dependent acetylcholine release in proteoliposomes derived from rat cortical synaptosomes