TY - JOUR
T1 - A mutation in transmembrane domain II of the 5-hydroxytryptamine3A receptor stabilizes channel opening and alters alcohol modulatory actions
AU - Sessoms-Sikes, J. S.
AU - Hamilton, Margaret E.
AU - Liu, Li Xin
AU - Lovinger, David M.
AU - Machu, Tina K.
PY - 2003/8/1
Y1 - 2003/8/1
N2 - Mutant 5-hydroxytryptamine (5-HT)3A receptors, in which changes were made at Ile294, position 16′, of the second transmembrane domain, were assessed for alterations in macroscopic response kinetics and modulation by alcohols. Function of heterologously expressed receptors was measured in Xenopus oocytes in the two-electrode voltage clamp configuration and in human embryonic kidney 293 cells using whole cell patch-clamp electrophysiological recordings with rapid drug application. Compared with the wild-type receptor, a decrease in the 5-HT EC50 value in the Ile294Thr mutant was observed, whereas an increase in the 5-HT EC50 value in the Ile294Leu mutant was measured. Ile294Thr receptors showed a marked reduction in the extent of desensitization. Ethanol and 2,2,2-trichloroethanol (TCEt) enhanced 5-HT-mediated currents in wild-type and Ile294Leu receptors, but inhibited or had little stimulatory effect in the Ile294Thr mutant. Kinetic analysis revealed that in the presence of TCEt, the slope of activation was unchanged in the Ile294Thr mutant and increased in the wild-type receptor. Alcohol cutoff was altered with wild-type = heptanol and Ile294Leu = hexanol. Kinetic changes in the Ile294Thr mutant that favor the open channel state, as well as reduction in the rate of channel activation in the presence of TCEt, likely underlie this mutant's altered response to n-chain alcohols.
AB - Mutant 5-hydroxytryptamine (5-HT)3A receptors, in which changes were made at Ile294, position 16′, of the second transmembrane domain, were assessed for alterations in macroscopic response kinetics and modulation by alcohols. Function of heterologously expressed receptors was measured in Xenopus oocytes in the two-electrode voltage clamp configuration and in human embryonic kidney 293 cells using whole cell patch-clamp electrophysiological recordings with rapid drug application. Compared with the wild-type receptor, a decrease in the 5-HT EC50 value in the Ile294Thr mutant was observed, whereas an increase in the 5-HT EC50 value in the Ile294Leu mutant was measured. Ile294Thr receptors showed a marked reduction in the extent of desensitization. Ethanol and 2,2,2-trichloroethanol (TCEt) enhanced 5-HT-mediated currents in wild-type and Ile294Leu receptors, but inhibited or had little stimulatory effect in the Ile294Thr mutant. Kinetic analysis revealed that in the presence of TCEt, the slope of activation was unchanged in the Ile294Thr mutant and increased in the wild-type receptor. Alcohol cutoff was altered with wild-type = heptanol and Ile294Leu = hexanol. Kinetic changes in the Ile294Thr mutant that favor the open channel state, as well as reduction in the rate of channel activation in the presence of TCEt, likely underlie this mutant's altered response to n-chain alcohols.
UR - http://www.scopus.com/inward/record.url?scp=0038637173&partnerID=8YFLogxK
U2 - 10.1124/jpet.103.050542
DO - 10.1124/jpet.103.050542
M3 - Article
C2 - 12730353
AN - SCOPUS:0038637173
VL - 306
SP - 595
EP - 604
JO - Journal of Pharmacology and Experimental Therapeutics
JF - Journal of Pharmacology and Experimental Therapeutics
SN - 0022-3565
IS - 2
ER -