Summary
Several intrinsic properties of acetylcholine receptor-rich membrane vesicles prepared fromElectrophorus electricus, which need to be considered in measurements of receptor-mediated ion flux, have been identified. One of these properties is a slow exchange of inorganic ions in the vesicles. The slow exchange of ions is not related to the receptor-mediated flux of ions and accounts for 30–35% of the efflux observed. A method to separate this process from the receptor-controlled flux has been developed. It has also been shown, using a light-scattering method, that aggregation-disaggregation of the vesicles can interfere with the efflux measurements, and a method to overcome this problem has been developed. The difference in the amplitude of effluxes induced by saturating amounts of carbamylcholine and gramicidin has been investigated and has been shown not to be due to a receptor-controlled process; therefore, the amplitude difference does not need to be considered in understanding the receptor-controlled process.
Similar content being viewed by others
References
Aoshima, H., Cash, D.J., Hess, G.P. 1980. Acetylcholine receptor-controlled ion flux in electroplax membrane vesicles: A minimal mechanism based on rate measurements in the millisecond to minute time region.Biochem. Biophys. Res. Commun. 92:896
Bernhardt, J., Neumann, E. 1978. Kinetic analysis of receptor-controlled tracer efflux from sealed membrane fragments.Proc. Nat. Acad. Sci. USA 75:3756
Cash, D.J., Hess, G.P. 1980. On the molecular mechanism of acetylcholine receptor-controlled ion translocation across cell membranes.Proc. Nat. Acad. Sci. USA 77:842
Cash, D.J., Hess, G.P. 1981. Quenched flow technique with plasma membrane vesicles: Acetylcholine receptor mediated transmembrane ion flux.Anal. Biochem. (in press)
Cass, A., Finkelstein, A., Krespi, Y. 1970. The ion permeability induced in thin lipid membranes by the polyene antibiotics nystatin and amphotericin B.J. Gen. Physiol. 56:100
Doty, P., Edsall, J.T. 1951. Light scattering in protein solution.Adv. Protein Chem. 6:35
Epstein, N., Hess, G.P., Kim, P.S., Noble, R.L. 1980. Inactivation (desensitization) of the acetylcholine receptor inElectrophorus electricus membrane vesicles by carbamylcholine: Comparison between ion flux and α-bungarotoxin binding.J. Membrane Biol. 56:133
Fu, J.-j.L., Donner, D.B., Moore, D.E., Hess, G.P. 1977. Allosteric interactions between the membrane-bound acetylcholine receptor and chemical mediators: Equilibrium measurements.Biochemistry 16:678
Gent, M.P.N., Prestegard, J.H. 1976. Interaction of the polyene antibiotics with lipid bilayer vesicles containing cholesterol.Biochim. Biophys. Acta 426:17
Hanley, M.R. 1978. Crotoxin effects inTorpedo californica, cholinergic excitable vesicles and the role of its phospholipase activity.Biochem. Biophys. Res. Commun. 82:392
Hess, G.P. 1979. Acetylcholine receptor-controlled ion fluxes in microsacs (membrane vesicles) obtained from the electroplax ofElectrophorus electricus.In: The Neurosciences: Fourth Study Program. F.O. Schmitt and F.G. Worden, editors, Chapter 49, p. 847. MIT Press, Cambridge
Hess, G.P., Andrews, J.P. 1977. Functional acetylcholine receptor-electroplax membrane microsacs (vesicles): Purification and characterization.Proc. Nat. Acad. Sci. USA 74:482
Hess, G.P., Andrews, J.P., Struve, G.E. 1976) Apparent cooperative effects in acetylcholine receptor-mediated ion flux in electroplax membrane preparations.Biochem. Biophys. Res. Commun. 69:830
Hess, G.P., Andrews, J.P., Struve, G.E., Coombs, S.E. 1975. Acetylcholine receptor-mediated ion flux in electroplax membrane preparations.Proc. Nat. Acad. Sci. USA 72:4371
Hess, G.P., Cash, D.J., Aoshima, H. 1979. Acetylcholine receptor-controlled ion fluxes in membrane vesicles investigated by fast reaction techniques.Nature (London) 282:329
Hess, G.P., Lipkowitz, S., Struve, G.E. 1978. Acetylcholine-receptor-mediated ion flux in electroplax membrane microsacs (vesicles): Change in mechanism produced by asymmetrical distribution of sodium and potassium ions.Proc. Nat. Acad. Sci. USA 75:1703
Hsiao-Ping, H., Moore, H., Hartig, P.R., Raftery, M.A. 1979. Correlation of polypeptide composition with functional events in acetylcholine receptor-enriched membranes fromTorpedo californica.Proc. Nat. Acad. Sci. USA 76:6265
Kasai, M., Changeux, J.-P. 1971a.In vitro excitation of purified membrane fragments by cholinergic agonists. I. Pharmacological properties of the excitable membrane fragments.J. Membrane Biol. 6:1
Kasai, M., Changeux, J.-P. 1971b.In vitro excitation of purified membrane fragments by cholinergic agonists. II. The permeability change caused by cholinergic agonists.J. Membrane Biol. 6:24
Kasai, M., Changeux, J.-P., 1971c.In vitro excitation of purified membrane fragments by cholinergic agonists. III. Comparison of the dose-response curves to decamethonium with the corresponding binding curves of decamethonium to the cholinergic receptor.J. Membrane Biol. 6:58
Katz, B., Thesleff, S. 1957. A study of the “desensitization” produced by acetylcholine at the motor endplate.J. Physiol. (London) 138:64
Keynes, R.D., Martins-Ferreira, H. 1953. Membrane potentials in the electroplates of the electric eel.J. Physiol. (London) 119:315
Kim, P.S. 1979. Acetylcholine receptor rich electroplax membrane microsacs (vesicles): Identification and characterization of membrane properties that interfere with measurements of receptor-mediated ion flux. A.B. Honors Thesis. Cornell University, Ithaca
Lowry, O.H., Rosebrough, N.J., Farr, A.L., Randall, R.J. 1951. Protein measurements with the Folin phenol reagent.J. Biol. Chem. 193:265
Palfrey, C., Littauer, V.Z. 1976. Sodium-dependent efflux of K+ and Rb+ through the activated sodium channel of neuroblastoma cells.Biochem. Biophys. Res. Commun. 72:209
Popot, J.L., Sugiyama, H., Changeux, J.-P. 1976. Studies on the electrogenic action of acetylcholine withTorpedo marmorata electric organ. II. The permeability response of the receptor-rich membrane fragments to cholinergic agonistsin vitro.J. Mol. Biol. 106:469
Pressman, B. 1976. Biological applications of ionophores.Annu. Rev. Biochem. 45:501
Schiebler, W., Hucho, F. 1978. Membranes rich in acetylcholine receptor: Characterization and reconstitution to excitable membranes from exogenous lipids.Eur. J. Biochem. 85:55
Selwyn, M.J., Dawson, A.P., Stockdale, M., Gains, N. 1970. Chloride-hydroxide exchange across mitochondrial erythrocyte and artifiecal lipid membranes mediated by trialkyl- andtriphenyltin compounds.Eur. J. Biochem. 14:120
Sillen, L.G., Martell, A.E. (Compilers) 1964. Stability Constants of Metal Ion Complexes. Special Publication. No.17. The Chemical Society, London
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Kim, P.S., Hess, G.P. Acetylcholine receptor-controlled ion flux in electroplax membrane vesicles: Identification and characterization of membrane properties that affect ion flux measurements. J. Membrain Biol. 58, 203–211 (1981). https://doi.org/10.1007/BF01870906
Received:
Revised:
Issue Date:
DOI: https://doi.org/10.1007/BF01870906