Functional properties of a cloned 5-hydroxytryptamine ionotropic receptor subunit: comparison with native mouse receptors.
1. A comparative study of the whole-cell and single-channel properties of cloned and native mouse 5-hydroxytryptamine ionotropic receptors (5-HT3) was undertaken using mammalian cell lines expressing the cloned 5-HT3 receptor subunit A (5-HT3R-A), superior cervical ganglia (SCG) neurones and N1E-115 cells. 2. No pharmacological difference was found in the sensitivity to the agonists 5-HT and 2-methyl-5-HT, or to the antagonists d-tubocurare and 3-tropanyl-3,5-dichlorobenzoate (MDL-72222). 3. Current-voltage (I-V) relationships of whole-cell currents showed inward rectification in the three preparations. Rectification was stronger both in cells expressing the 5-HT3R-A subunit and in N1E-115 cells when compared with SCG neurones. 4. No clear openings could be resolved in 5-HT-activated currents in patches excised from cells expressing the 5-HT3R-A subunit or N1E-115 cells. Current fluctuation analysis of whole-cell and excised-patch records revealed a slope conductance of 0.4-0.6 pS in both preparations. Current-voltage relationships of these channels showed strong rectification that fully accounted for the whole-cell voltage dependence. 5. In contrast, single channels of about 10 pS were activated by 5-HT in patches excised from SCG neurones. The weak voltage dependence of their conductance did not account completely for the rectification of whole-cell currents. A lower unitary conductance (3.4 pS) was inferred from whole-cell noise analysis. 6. We conclude that the receptor expressed from the cloned cDNA is indistinguishable from the 5-HT3 receptor of N1E-115 cells, suggesting an identical structure for these two receptors. The higher conductance and different voltage dependence of the 5-HT3 receptor in SCG neurones might indicate the participation of an additional subunit in the structure of native ganglionic 5-HT3 receptors. Homo-oligomeric 5-HT3R-A channels may also be present as suggested by the lower conductance estimated by whole-cell noise analysis.
Hussy
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N
Glaxo Institute for Molecular Biology, Geneva, Switzerland.
Lukas
W
W
Jones
K A
KA
D000818:Animals; D003001:Cloning, Molecular; D004594:Electrophysiology; D005822:Genetic Vectors; D007473:Ion Channels / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D009447:Neuroblastoma / Q000378:metabolism; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D018408:Patch-Clamp Techniques; D011985:Receptors, Serotonin / Q000187:drug effects / Q000378:metabolism*; D012672:Semliki forest virus; D012702:Serotonin Antagonists / Q000494:pharmacology; D017366:Serotonin Receptor Agonists / Q000494:pharmacology; D017783:Superior Cervical Ganglion / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D014407:Tumor Cells, Cultured
D007473:Ion Channels; D011985:Receptors, Serotonin; D012702:Serotonin Antagonists; D017366:Serotonin Receptor Agonists