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On the voltage‐dependent Ca 2+ block of serotonin 5‐HT 3 receptors: a critical role of intracellular phosphates
Author(s) -
Noam Yoav,
Wadman Wytse J.,
Van Hooft Johannes A.
Publication year - 2008
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1113/jphysiol.2008.153486
Subject(s) - homomeric , receptor , biophysics , chemistry , ion channel , protein subunit , biochemistry , stereochemistry , biology , gene
Natively expressed serotonin 5‐HT 3 receptors typically possess a negative‐slope conductance region in their I–V curve, due to a voltage‐dependent block by external Ca 2+ ions. However, in almost all studies performed with heterologously expressed 5‐HT 3 receptors, this feature was not observed. Here we show that mere addition of ATP to the pipette solution is sufficient to reliably observe a voltage‐dependent block in homomeric (h5‐HT 3A ) and heteromeric (h5‐HT 3AB ) receptors expressed in HEK293 cells. A similar block was observed with a plethora of molecules containing a phosphate moiety, thus excluding a role of phosphorylation. A substitution of three arginines in the intracellular vestibule of 5‐HT 3A with their counterpart residues from the 5‐HT 3B subunit (RRR‐QDA) was previously shown to dramatically increase single channel conductance. We find this mutant to have a linear I–V curve that is unaffected by the presence of ATP, with a fractional Ca 2+ current (Pf%) that is reduced (1.8 ± 0.2%) compared to that of the homomeric receptor (4.1 ± 0.2%), and similar to that of the heteromeric form (2.0 ± 0.3%). Moreover, whereas ATP decreased the Pf% of the homomeric receptor, this was not observed with the RRR‐QDA mutant. Finally, ATP was found to be critical for voltage‐dependent channel block also in hippocampal interneurons that natively express 5‐HT 3 receptors. Taken together, our results indicate a novel mechanism by which ATP, and similar molecules, modulate 5‐HT 3 receptors via interactions with the intracellular vestibule of the receptor.