Hitoshi Aoshima

Acetylcholine receptor-controlled ion flux in electroplax membrane vesicles: A minimal mechanism based on rate measurements in the millisecond to minute time region

Abstract The dependence of acetylcholine receptor-controlled transmembrane ion flux on carbamylcholine concentration was measured in the msec time region, using membrane vesicles and a quench flow technique. 4 Measurements were made: (1) transmembrane ion influx, (2) rate of inactivation of the receptor by carbamylcholine, (3) rate of recovery, and (4) ion influx mediated by “inactivated” receptor. The minimal model, based on the measurements, accounts for the time dependence of receptor-controlled ion flux over a 200-fold carbamylcholine concentration range. The maximum flux rate of 84 sec−1 indicates that we have succeeded in measuring the receptor-controlled processes which give rise to electrical signals in cells.

Vidicon flame emission spectroscopy of Li+, Na+, and K+ fluxes mediated by acetylcholine receptor in Electrophorus electricus membrane vesicles.

Abstract A vidicon flame spectroscopic method was developed to determine Li+, Na+, and K+ fluxes mediated by the acetylcholine receptor protein in vesicles prepared from Electrophorus electricus electroplax. Although analysis by the vidicon system is susceptible to trace element contamination, the problem can be overcome by taking adequate precautions. The results obtained by flame emission spectroscopy were directly compared to those obtained by using radioactive tracers. Also, the acetylcholine receptor-formed channels were shown to be non-selective for the monovalent metal ions tested.

Acetylcholine-induced receptor-controlled ion flux investigated by flow quench techniques.

Abstract Using a quench flow technique with membrane vesicles, the acetylcholine receptor-controlled transmembrane ion flux and the inactivation of the receptor with acetylcholine were measured in the msec time region. The ion flux was followed by influx of radioactive tracer ion and the inactivation was followed by an ion flux assay of receptor pre-incubated with ligand. The measurements covered a concentration range to complete saturation of the active state of the receptor with ligand, and were consistent with a minimal model previously proposed on the basis of experiments with carbamylcholine. The ion translocation rate at saturation with acetylcholine is about twice that at saturation with carbamylcholine and this reflects a more favored channel opening equilibrium for acetylcholine.