D127 of the human 5-HT3A receptor is a major determinant of ion permeation

Matthew Livesey, Michelle Cooper, Jeremy Lambert, John Peters. Centre for Neuroscience, University of Dundee, Angus, United Kingdom.

The 5-HT3A receptor is a cation-selective member of the Cys-loop receptor family. The single channel conductance and ion selectivity of such channels depends crucially upon the structure of the channel-lining second transmembrane domain and also the large intracellular loop (Barnes et al., 2009). Here, D127, an extracellular residue, is shown to be an additional determinant of the biophysical properties of the 5-HT3A receptor.

Single channel currents evoked by 5-HT (10 μM) were recorded from outside-out membrane patches excised from tsA-201 cells expressing either the high conductance form of the human 5-HT3A receptor (5-HT3A QDA) (Livesey et al., 2008), or a mutant (5-HT3A QDA D127N) thereof. Single channel conductances (γ) were obtained from full current-voltage relationships. The relative permeability of Ca2+ versus Cs+ (PCa/PCs) was calculated from measurements of macroscopic current reversal potential in a solution wherein Ca2+ was the sole permeant cation. Table 1 summarises the data.

Table 1. Data are reported as the mean ± s.e.m. ***Significantly reduced (P = 0.001) in comparison to 5-HT3A QDA (unpaired t-test). Vh = holding potential.

In addition to reducing γ and PCa/PCs, the D127N mutation essentially abolished inhibition by Ca2+ of single channel currents mediated by monovalent cations at the 5-HT3A QDA receptor (Livesey et al., 2008). We suggest that reduced electronegativity within the extracellular vestibule underlies the effects of the D127N mutation.

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