Voltage fluctuations at the frog sartorius motor endplate produced by a covalently attached activator

Abstract

The depolarization that develops after covalent attachment of trimethylammonium benzoyl to the dithiothreitol-reduced frog sartorius acetylcholine receptor is accompanied by a small increase in voltage fluctuations. The amplitude of the elementary voltage event produced by the covalently attached activator is about 0.04 microV, almost an order of magnitude below the acetylcholine shot-effect amplitude in the control preparation, and about one-fourth the acetylcholine shot amplitude after disulfide-bond reduction. Spectral density plots of trimethylammonium-benzoyl noise can be analyzed in terms of two relaxation rates that bracket the single rate observed in response to acetylcholine.

Optochemical control of genetically engineered neuronal nicotinic acetylcholine receptors

“…Once attached, QBr activated the channel in its trans -form, that is at 500 nm or in the dark, but could be switched to an inactive cis -form by irradiation with 330 nm light. The resulting light-activated nAChR could be used to stimulate reversibly Electrophorus electroplaques, frog muscles and rat myoballs with light 28,29,31,32 . At the time these studies were carried out, however, receptors could not be manipulated genetically and expressed heterologously, so the impact of synthetic photo-switches on neuroscience research remained limited.…”

Physiological and Pharmacological Manipulations with Light Flashes

“…(b) Because the unreacted molecules are washed out of the preparation after the tethering reaction, there is no chance that they can also function as competitive antagonists or "open-channel blockers." Their disadvantage is that, like all known agonists, they eventually desensitize receptors, so that the number of open channels decreases with time (31,32). This shortcoming is avoided by the photoisomerizable tethered agonist, QBr (13, 65; Figure 1).…”

Activation of acetylcholine receptor channels by covalently bound agonists in cultured rat myoballs.

“…We have therefore combined whole-cell and single-channel measurements on cultured rat myoballs in order to extend earlier experiments with tethered agonists at the ACh receptor of frog muscle (Cox et al 1979) and ofElectrophorus electroplaques (Lester et al 1980). As found previously, the ACh channel can be activated by a single tethered agonist, although other channel properties, such as the conductance, kinetics, and voltage dependence, are very similar to those seen with reversibly bound agonists.…”

“…They eliminate the first step in the agonist-receptor interaction, the diffusion-limited formation of an initial complex (Burgen, 1966). The tethered agonist is present at a very high effective local concentration: Cox, Kawai, Karlin & Brandt (1979) estimated -1 M. If a reversibly TETHERED NICOTINIC AGONISTS AT RAT MYOBALLS bound agonist molecule were studied at such concentrations, channel block by the agonist itself would intolerably complicate the current records. In addition, by comparing the fractional conductance with the fraction of active (trans) tethered QBr, it has been shown that the open state of the receptor is controlled by the configuration of a single tethered QBr molecule (Lester et al 1980).…”