Summary
Proteoliposomes made by a butanol-sonication technique from electric organ presynaptic membranes showed choline transport activity. In contrast to intact nerve terminals, the uptake of choline was dissociated from its conversion to acetylcholine in this preparation. The kinetics of choline uptake by proteoliposomes was best described by two Michaelis-Menten components. At a low concentration of choline, uptake was inhibited by hemicholinium-3 and required external Na+ and, thus, closely resembled high-affinity choline uptake by intact cholinergic nerve terminals. Choline transport could be driven by the Na+ gradient and by the transmembrane potential (inside negative) but did not directly require ATP. External Cl−, but not a Cl− gradient, was needed for choline transport activity. It is suggested that internal K+ plays a role in the retention of choline inside the proteoliposome. Proteoliposomes should prove a useful tool for both biochemical and functional studies of the highaffinity choline carrier.
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Abbreviations
- ACh:
-
acetylcholine
- HC-3:
-
hemicholinium-3
- ChAT:
-
choline acetyltransferase
References
Adam-Vizi, V., Marchbanks, R.M. 1983. Studies on the osmotic disruption and resealing of synaptosomes.J. Neurochem. 41:780–785
Beach, R.L., Vaca, K., Pilar, G. 1980. Ionic and metabolic requirements for high affinity choline uptake and acetylcholine synthesis in nerve terminals at a neuromuscular junction.J. Neurochem. 34:1387–1398
Bradford, M. 1976. A new technique for the determination of protein in small samples.Anal. Biochem. 72:248–254
Breer, H., Lueken, W. 1983. Transport of choline by membrane vesicles prepared from synaptosomes of insect nervous tissue.Neurochem. Int. 5:713–720
Carroll, P.T., Goldberg, A.M. 1975. Relative importance of choline transport to spontaneous and potassium depolarized release of ACh.J. Neurochem. 25:523–527
Eytan, G.D. 1982. Use of liposomes for reconstitution of biological functions.Biochim. Biophys. Acta 694:185–202
Fonnum, F. 1975. A rapid radiochemical method for the determination of choline acetyltransferase.J. Neurochem. 24:407–409
Gulik-Krzywicki, T., Costello, M.J. 1978. The use of low temperature X-ray diffraction to evaluate freezing methods used in freeze-fracture electron microscopy.J. Microsc. (Oxford) 112:103–113
Israël, M., Lesbats, B. 1981. Chemiluminescent determination of acetylcholine, and continuous detection of its release from Torpedo electric organ synapses and synaptosomes.Neurochem. Int. 3:81–90
Israël, M., Lesbats, B., Manaranche, R., Morel, N. 1983. Acetylcholine release from proteoliposomes equipped with synaptosomal membrane constituents.Biochim. Biophys. Acta 728:438–448
Israël, M., Lesbats, B., Morel, N., Manaranche, R., Gulik-Krzywicki, T., Dedieu, J.C. 1984. Reconstitution of a functional synaptosomal membrane possessing the protein constituents involved in acetylcholine translocation.Proc. Natl. Acad. Sci. USA 81:277–281
Johnson, M.R. 1960. The intracellular distribution of glycolytic and other enzymes in rat brain homogenates and mitochondrial preparations.Biochem. J. 77:610–618
Jope, R.S. 1979. High affinity choline transport and acetyl-CoA production in brain and their roles in the regulation of acetylcholine synthesis.Brain Res. Rev. 1:313–344
Jope, R., Weiler, M.H., Jenden, D.J. 1978. Regulation of acetylcholine synthesis: Control of choline transport and acetylation in synaptosomes.J. Neurochem. 30:949–954
King, R.G., Marchbanks, R.M. 1982a. The incorporation of solubilized choline-transport activity into liposomes.Biochem. J. 204:565–576
Kuhar, M.J., Murrin, L.C. 1978. Sodium dependent, high affinity choline uptake.J. Neurochem. 30:15–21
Kuhar, M.J., Zarbin, M.A. 1978. Synaptosomal transport: A chloride dependence for choline, GABA, glycine and several other compounds.J. Neurochem. 31:251–256
Marchbanks, R.M., Wonnacott, S.P., Rubio, M.A. 1981. The effect of acetylcholine release on choline fluxes in isolated synaptic terminals.J. Neurochem. 36:379–393
Martin, K. 1972. Extracellular cations and the movement of choline across the erythrocyte membrane.J. Physiol. (London) 224:207–230
Massarelli, R., Gorio, A., Dreyfus, H. 1982. Influx, metabolism and efflux of choline in nerve cells and synaptosomes; Role of sialocompounds and glycoconjugates.J. Physiol. (Paris) 78:392–398
Meyer, E.M., Cooper, J.R. 1983. High affinity choline uptake and calcium-dependent acetylcholine release in proteoliposomes derived from rat cortical synaptosomes.J. Neurosci.3:987–994
Morel, N., Israël, M., Manaranche, R., Lesbats, B. 1979. Stimulation of cholinergic synaptosomes isolated fromTorpedo electric organ.Prog. Brain Res. 49:191–202
Morel, N., Israël, M., Manaranche, R., Mastour-Franchon, P. 1977. Isolation of pure cholinergic nerve endings fromTorpedo electric organ. Evaluation of their metabolic properties.J. Cell Biol. 75:43–55
Murrin, L.C., Kuhar, M.J. 1976. Activation of high affinity choline uptakein vitro by depolarizing agents.Mol. Pharmacol. 12:1082–1090
O'Regan, S. 1982. The synthesis, storage and release of propionylcholine by the electric organ ofTorpedo marmorata.J. Neurochem. 39:764–772
O'Regan, S., Collier, B. 1981. Factors affecting choline transport by the cat superior cervical ganglion during and following stimulation, and the relationship between choline uptake and acetylcholine synthesis.Neuroscience 6:511–520
Pressman, B.C. 1976. Biological applications of ionophores.Annu. Rev. Biochem. 45:501–530
Rothlein, J.E., Parsons, S.M. 1979. Characterization of high affinity choline uptake byTorpedo californica T-sacs.J. Neurochem. 33:1189–1194
Schultz, S.G., Curran, P.F. 1970. Coupled transport of sodium and organic solutes.Physiol. Rev. 50:637–718
Wheeler, D.D. 1979. A model of high affinity choline transport in rat cortical synaptosomes.J. Neurochem. 32:1197–1213
Yamamura, H.I., Snyder, S.H. 1973. High affinity transport of choline into synaptosomes of rat brain.J. Neurochem. 21:1355–1374
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Vyas, S., O'Regan, S. Reconstitution of carrier-mediated choline transport in proteoliposomes prepared from presynaptic membranes oftorpedo electric organ, and its internal and external ionic requirements. J. Membrain Biol. 85, 111–119 (1985). https://doi.org/10.1007/BF01871264
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DOI: https://doi.org/10.1007/BF01871264