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The interaction of trichloroethanol with murine recombinant 5‐HT 3 receptors
Author(s) -
Downie David L.,
Hope Anthony G.,
Belelli Delia,
Lambert Jeremy J.,
Peters John A.,
Bentley Kirn R.,
Steward Lucinda J.,
Chen ChongYang,
Barnes Nicholas M.
Publication year - 1995
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1111/j.1476-5381.1995.tb14952.x
Subject(s) - chloral hydrate , chemistry , receptor , xenopus , recombinant dna , 5 ht receptor , ethanol , biophysics , biochemistry , serotonin , pharmacology , biology , gene
1 The effects of ethanol, chloral hydrate and trichloroethanol upon the 5‐HT 3 receptor have been investigated by use of electrophysiological techniques applied to recombinant 5‐HT3 receptor subunits (5‐HT 3 R‐A or 5‐HT 3 R‐As) expressed in Xenopus laevis oocytes. Additionally, the influence of trichloroethanol upon the specific binding of [ 3 H]‐granisetron to membrane preparations of HEK 293 cells stably transfected with the murine 5‐HT 3 R‐A s subunit and 5‐HT 3 receptors endogenous to NG 108‐15 cell membranes was assessed. 2 Ethanol (30–300 mM), chloral hydrate (1–30 mM) and trichloroethanol (0.3‐10mM), produced a reversible, concentration‐dependent, enhancement of 5‐HT‐mediated currents recorded from oocytes expressing either the 5‐HT 3 R‐A, or the 5‐HT 3 R‐A s subunit. 3 Trichloroethanol (5 mM) produced a parallel leftward shift of the 5‐HT concentration‐response curve, reducing the EC 50 for 5‐HT from 1 ± 0.04 μm (n = 4) to 0.5 ± 0.01 μm (n = 4) for oocytes expressing the 5‐HT 3 R‐A. A similar shift, from 2.1 ± 0.05 μm (n = 11) to 1.3 ± 0.1 μ m (n = 4), was observed in oocytes expressing the 5‐HT 3 R‐A s subunit. Trichloroethanol (5 mM) had little or no effect upon the maximum current produced by 5‐HT for either recombinant receptor. 4 Trichloroethanol (5 mM) similarly reduced the EC50 for 2‐methyl‐5‐HT from 13 ± 0.4 μm (n = 4) to 4.6 ± 0.2 μ m (n = 4) and from 15 ± 2μ m (n = 4) to 5 ± 0.4μm (n = 4) for oocytes expressing the 5‐HT 3 R‐A and 5‐H 3 R‐A s subunit respectively. Additionally, trichloroethanol (5 mM) produced a clear enhancement of the maximal current to 2‐methyl‐5‐HT (expressed as a percentage of the maximal current to 5‐HT) from 63 ± 0.7% (n = 4) to 101 ± 1.6% (n = 4) and from 9 ± 0.2% (n = 4) to 74 ± 2% (n = 4) for oocytes expressing the 5‐HT 3 R‐A and 5‐HT 3 R‐As subunit respectively. 5 Trichloroethanol (2.5 mM) had no effect upon the K d , or B max , of specific [ 3 H]‐granisetron binding to membrane homogenates of NG 108‐15 cells or HEK 293 cells. Similarly, competition for [ 3 H]‐granisetron binding by the 5‐HT 3 receptor antagonists ondansetron and tropisetron was unaffected. However, competition for [ 3 H]‐granisetron binding by the 5‐HT 3 receptor agonists, 5‐HT, 2‐methyl‐5‐HT and phenylbiguanide was enhanced by trichloroethanol (2.5 mM). 6 Unexpectedly, the competition for [ 3 H]‐granisetron binding by the 5‐HT3 receptor antagonist, quipazine, was enhanced by 2.5 mM trichloroethanol. Quipazine (1 nM‐0.3 μ m ) antagonized 5‐HT‐evoked currents recorded from oocytes expressing the 5‐HT 3 R‐A subunit with an IC 50 of 18 ± 3 nM (n = 4). Additionally, quipazine (30 nM‐0.3 μ m ) produced a small inward current which was greatly enhanced by 5 mM trichloroethanol and antagonized by 100 nM ondansetron. Collectively, these observations suggest that quipazine may act as a partial agonist. 7 The demonstration that a recombinant homo‐oligomeric receptor, expressed in a foreign membrane, retains a sensitivity to alcohols, together with the sequencing of alcohol‐insensitive 5‐HT 3 receptor subunits, may lead to a better definition of the alcohol binding site(s).