In vitro reconstitution of neurotransmitter release.

The vesicular hypothesis has stimulated fruitful investigations on many secreting systems. In the case of rapid synaptic transmission, however, the hypothesis has been found difficult to reconcile with a number of well established observations. Brief impulses of transmitter molecules (quanta) are emitted from nerve terminals at the arrival of an action potential by a mechanism which is under the control of multiple regulations. It is therefore not surprising that quantal release could be disrupted by experimental manipulation of a variety of cellular processes, such as a) transmitter uptake, synthesis, or transport, b) energy supply, c) calcium entry, sequestration and extrusion, d) exo- or endocytosis, e) expression of vesicular and plasmalemmal proteins, f) modulatory systems and second messengers, g) cytoskeleton integrity, etc. Hence, the approaches by "ablation strategy" do not provide unequivocal information on the final step of the release process since there are so many ways to stop the release. We propose an alternate approach: the "reconstitution strategy". To this end, we developed several preparations for determining the minimal system supporting Ca2+-dependent transmitter release. Release was reconstituted in proteoliposomes, Xenopus oocytes and transfected cell lines. Using these systems, it appears that a presynaptic plasmalemmal proteolipid, that we called mediatophore should be considered as a key molecule for the generation of transmitter quanta in natural synapses.