Nicotinic acetylcholine receptor transmembrane mutations convert ivermectin from a positive to a negative allosteric modulator.
Ivermectin is a macrocyclic lactone that acts as a positive allosteric modulator of alpha7 nicotinic acetylcholine receptors (nAChRs) but has no modulatory activity on 5-hydroxytryptamine (5-HT) type 3 (5-HT(3)) receptors. By examining the influence of ivermectin on subunit chimeras containing domains from the nAChR alpha7 subunit and the 5-HT3A subunit, we have concluded that the transmembrane domains play a critical role in influencing allosteric modulation by ivermectin. A series of mutations located within the alpha-helical transmembrane domains of the alpha7 subunit were examined, and seven were found to have significant effects on allosteric modulation by ivermectin. Four mutations (A225D, Q272V, T456Y, and C459Y) caused a significant reduction in the potency of ivermectin as an allosteric potentiator. Compared with wild-type alpha7 nAChRs, potentiation by ivermectin was reduced dramatically (by 89-97%) by these mutations. Somewhat unexpectedly, three mutations (S222M, M253L, and S276V located in TM1, TM2, and TM3) converted ivermectin from a positive allosteric modulator into an antagonist. Levels of inhibition of 56, 84, and 89% were observed on M253L, S276V, and S222M, respectively. Antagonism by ivermectin was insurmountable and had no effect on EC(50) of acetylcholine, indicating that it is acting noncompetitively. The seven mutations that influence allosteric modulation by ivermectin are located near a predicted intrasubunit transmembrane cavity. Computer docking simulations provide support for the hypothesis that ivermectin binds in close proximity to this cavity. We conclude that transmembrane mutations in alpha7 nAChRs are able to convert ivermectin from a positive to a negative allosteric modulator.
Collins
Toby
T
Department of Neuroscience, Physiology & Pharmacology, University College London, London, United Kingdom.
Millar
Neil S
NS
D000494:Allosteric Regulation; D000818:Animals; D003198:Computer Simulation; D006801:Humans; D007559:Ivermectin / Q000494:pharmacology*; D017354:Point Mutation; D011978:Receptors, Nicotinic / Q000235:genetics*; D014982:Xenopus laevis
D011978:Receptors, Nicotinic; D007559:Ivermectin
Eur J Pharmacol. 2000 Apr 14;394(2-3):163-70
10771281
Vet Hum Toxicol. 2000 Feb;42(1):30-5
10670084
J Neurosci. 2005 Apr 27;25(17):4396-405
15858066
Neuron. 2007 Apr 19;54(2):263-74
17442247
J Pharmacol Exp Ther. 2007 Oct;323(1):294-307
17625074
J Mol Biol. 2006 Jan 13;355(2):310-24
16307758
Neuroscience. 2000;101(3):785-91
11113327
J Parasitol. 1995 Apr;81(2):286-94
7707209
J Biol Chem. 1998 Feb 6;273(6):3314-9
9452448
J Biol Chem. 2002 Jan 18;277(3):2000-5
11714703
Br J Pharmacol. 2008 Mar;153 Suppl 1:S177-83
18246096
Pflugers Arch. 2008 Aug;456(5):939-50
18427835
Neuropharmacology. 2006 Feb;50(2):174-81
16225893
J Mol Biol. 2007 Jun 15;369(4):895-901
17481657
Eur J Pharmacol. 1994 Jul 1;259(2):121-8
7957605
J Neurochem. 1998 Jun;70(6):2585-93
9603225
J Biol Chem. 2001 Apr 20;276(16):12556-64
11278873
J Neurophysiol. 1994 Sep;72(3):1317-26
7807214
Nat Rev Drug Discov. 2002 Jun;1(6):427-36
12119744
Mol Pharmacol. 2003 Feb;63(2):419-28
12527814
Mol Pharmacol. 1987 Dec;32(6):749-52
2447480
Br J Pharmacol. 2007 Oct;152(4):501-12
17721553
Mol Pharmacol. 2005 Nov;68(5):1431-8
16120769
Mol Pharmacol. 2007 Feb;71(2):389-97
17065235
J Mol Biol. 2005 Mar 4;346(4):967-89
15701510
J Pharmacol Exp Ther. 2003 Jun;305(3):1024-36
12649296
Nature. 2003 Jun 26;423(6943):949-55
12827192
Biochem Pharmacol. 2007 Oct 15;74(8):1155-63
17707779
Proc Natl Acad Sci U S A. 2008 Sep 23;105(38):14686-91
18791069
Neuropharmacology. 2002 Sep;43(3):374-84
12243767
J Neurosci. 1999 Sep 1;19(17):7289-99
10460235
Nature. 1997 Sep 25;389(6649):385-9
9311780
Mol Pharmacol. 2008 Nov;74(5):1407-16
18678621
Biochem Pharmacol. 2009 Oct 1;78(7):836-43
19549506
Nature. 2006 Nov 23;444(7118):486-9
17108970
Med Res Rev. 2007 Sep;27(5):723-53
17238157
Hum Mol Genet. 2003 Nov 15;12(22):3017-24
14506129
J Pharmacol Exp Ther. 2006 Sep;318(3):1108-17
16738207
Mol Pharmacol. 1998 Feb;53(2):283-94
9463487