Unraveling mechanisms underlying partial agonism in 5-HT3A receptors

Abstract

Partial agonists are unable to elicit full maximal responses and they have emerged as attractive therapeutic molecules since they can act both as agonists or antagonists depending on the concentration of neurotransmitter. The characterization of a genuine partial agonist is complex because other mechanisms, such as channel block, may also limit maximum open probability. Taking advantage of the high conductance form of the 5-HT3A receptor, we here studied activation by two typical partial agonists: 2-Me-5HT and tryptamine at the single-channel level. For all ligands, activation appears as openings in quick succession grouped in clusters showing high open probability (Popen>0.9). Open time distributions show three components; the slowest one is 6.5- and 3.5-fold briefer for 2-Me-5HT and tryptamine, respectively, than for 5-HT. The duration of this component decreases as a function of agonist concentration due to open-channel block. For 2-Me-5HT, the forward blocking rate is 10-fold faster than for tryptamine and 5-HT. Our single-channel kinetic analysis shows that 2-Me-5HT is actually a full agonist, its maximum response being limited by channel block. In contrast, tryptamine is a genuine partial agonist and its low efficacy is mainly due to a slow transition from the fully-liganded closed state to a pre- open state. After reaching the latter state, activation proceeds similarly as in the presence of 5-HT. Molecular docking shows that interactions at the binding site are similar for 2-Me-5HT and 5-HT. In contrast, the potential to form the cation-Pi interaction with W183 seems to be reduced for tryptamine. Unraveling the mechanism underlying partial responses elicited by 5-HT3 ligands has implications in the design for novel therapeutic compounds. In addition, our results reveal the existence of multiple pre-open states associated to partial agonism in a yet unexplored member of the Cys-loop receptor family