The three-ion test for the identity of the ionophores mediating the hyperpolarizing response to cholinomimetics in neurones of Helix aspersa.

1. The hyperpolarizing response investigated is mediated by Cl- ions. Additional anions NO-3 and BrO-3 were introduced into the perfusate to produce a system with 3 permeant anions. 2. Under such circumstances the reversal potential (Er) becomes a means of discriminating between the molecular mechanisms invoked by different agonists that produce an increase in anionic conductance. 3. A statistically significant difference of 0.49 mV was detected between the Er for acetylcholine and those for acetyl-beta-methyl choline and tetramethylammonium (acetylcholine more negative). 4. This difference was not significantly affected by changing to a different 3-ion solution. 5. It is argued from theoretical grounds that a change in this difference is to be expected when molecular mechanisms differ; alternative explanations for the difference are suggested.