Drosophila GABA-gated chloride channel

Modified [3H]EBOB binding site associated with Ala → Ser or Gly mutants of Rdl subunit

Loretta M. Cole, Richard T. Roush, John E. Casida

Research output: Contribution to journalArticle

63 Citations (Scopus)

Abstract

The non-competitive blocker site of the GABA-gated chloride ion channel in normal susceptible strains of Drosophila melanogaster and simulans binds 4-n [3H]propyl-4′-ethynylbicycloorthobenzoate ([3H]EBOB) at specific sites with Kds of 1.6-1.9 nM and Bmaxs of 171-181 fmol/mg protein. This specific binding of [3H]EBOB is strongly inhibited by: a large number and variety of insecticidal channel blockers at 20 nM (lindane, α-endosulfan, dieldrin, 12-ketoendrin, fipronil, and a representative bicycloorthobenzoate and dithiane) or 200 nM (picrotoxinin); the insecticidal channel activators avermectin and moxidectin at 20 nM; muscimol at 30 μM and GABA at 300 μM. Cyclodiene resistance in D. melanogaster has been attributed to a mutation resulting in an Ala302 → Ser replacement in the Rdl GABA receptor subunit and in D. simulans to an homologous Ala → Ser or Gly replacement. These mutations are shown here to greatly reduce [3H]EBOB binding, i.e. lower affinity and apparent number of binding sites. The Ala → Ser replacement with both melanogaster and simulans almost always reduces the potency in inhibiting [3H]EBOB binding of each of eight channel blockers and of muscimol and GABA. The Ala → Gly replacement in D. simulans is generally less effective than the Ala → Ser modification in reducing sensitivity to the channel blockers and to muscimol and GABA. The channel activators avermectin and moxidectin usually retain their inhibitory potency in the Rdl subunit mutants. Thus, it appears that replacement of Ala by Ser generally modifies the non-competitive blocker site and its coupling to the GABA-recognition site with less effect on the channel activator site. In contrast, the Ala → Gly replacement has less impact in protecting the chloride channel from the action of insecticidal blockers. Each of the resistant strains has the same level of resistance to the lethal action of the five channel blockers examined but none to avermectins and muscimol.

Original languageEnglish (US)
Pages (from-to)757-765
Number of pages9
JournalLife Sciences
Volume56
Issue number10
DOIs
StatePublished - Jan 27 1995

Fingerprint

Chloride Channels
Muscimol
gamma-Aminobutyric Acid
Drosophila
Binding Sites
Drosophila melanogaster
Endosulfan
Dieldrin
Lindane
Mutation
GABA Receptors
1-(4-ethynylphenyl)-4-propyl-2,6,7-trioxabicyclo(2.2.2)octane
avermectin
Proteins

All Science Journal Classification (ASJC) codes

  • Medicine(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Pharmacology, Toxicology and Pharmaceutics(all)

Cite this

@article{ecf55820582341c48f3c595019026fd7,
title = "Drosophila GABA-gated chloride channel: Modified [3H]EBOB binding site associated with Ala → Ser or Gly mutants of Rdl subunit",
abstract = "The non-competitive blocker site of the GABA-gated chloride ion channel in normal susceptible strains of Drosophila melanogaster and simulans binds 4-n [3H]propyl-4′-ethynylbicycloorthobenzoate ([3H]EBOB) at specific sites with Kds of 1.6-1.9 nM and Bmaxs of 171-181 fmol/mg protein. This specific binding of [3H]EBOB is strongly inhibited by: a large number and variety of insecticidal channel blockers at 20 nM (lindane, α-endosulfan, dieldrin, 12-ketoendrin, fipronil, and a representative bicycloorthobenzoate and dithiane) or 200 nM (picrotoxinin); the insecticidal channel activators avermectin and moxidectin at 20 nM; muscimol at 30 μM and GABA at 300 μM. Cyclodiene resistance in D. melanogaster has been attributed to a mutation resulting in an Ala302 → Ser replacement in the Rdl GABA receptor subunit and in D. simulans to an homologous Ala → Ser or Gly replacement. These mutations are shown here to greatly reduce [3H]EBOB binding, i.e. lower affinity and apparent number of binding sites. The Ala → Ser replacement with both melanogaster and simulans almost always reduces the potency in inhibiting [3H]EBOB binding of each of eight channel blockers and of muscimol and GABA. The Ala → Gly replacement in D. simulans is generally less effective than the Ala → Ser modification in reducing sensitivity to the channel blockers and to muscimol and GABA. The channel activators avermectin and moxidectin usually retain their inhibitory potency in the Rdl subunit mutants. Thus, it appears that replacement of Ala by Ser generally modifies the non-competitive blocker site and its coupling to the GABA-recognition site with less effect on the channel activator site. In contrast, the Ala → Gly replacement has less impact in protecting the chloride channel from the action of insecticidal blockers. Each of the resistant strains has the same level of resistance to the lethal action of the five channel blockers examined but none to avermectins and muscimol.",
author = "Cole, {Loretta M.} and Roush, {Richard T.} and Casida, {John E.}",
year = "1995",
month = "1",
day = "27",
doi = "10.1016/0024-3205(95)00006-R",
language = "English (US)",
volume = "56",
pages = "757--765",
journal = "Life Sciences",
issn = "0024-3205",
publisher = "Elsevier Inc.",
number = "10",

}

Drosophila GABA-gated chloride channel : Modified [3H]EBOB binding site associated with Ala → Ser or Gly mutants of Rdl subunit. / Cole, Loretta M.; Roush, Richard T.; Casida, John E.

In: Life Sciences, Vol. 56, No. 10, 27.01.1995, p. 757-765.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Drosophila GABA-gated chloride channel

T2 - Modified [3H]EBOB binding site associated with Ala → Ser or Gly mutants of Rdl subunit

AU - Cole, Loretta M.

AU - Roush, Richard T.

AU - Casida, John E.

PY - 1995/1/27

Y1 - 1995/1/27

N2 - The non-competitive blocker site of the GABA-gated chloride ion channel in normal susceptible strains of Drosophila melanogaster and simulans binds 4-n [3H]propyl-4′-ethynylbicycloorthobenzoate ([3H]EBOB) at specific sites with Kds of 1.6-1.9 nM and Bmaxs of 171-181 fmol/mg protein. This specific binding of [3H]EBOB is strongly inhibited by: a large number and variety of insecticidal channel blockers at 20 nM (lindane, α-endosulfan, dieldrin, 12-ketoendrin, fipronil, and a representative bicycloorthobenzoate and dithiane) or 200 nM (picrotoxinin); the insecticidal channel activators avermectin and moxidectin at 20 nM; muscimol at 30 μM and GABA at 300 μM. Cyclodiene resistance in D. melanogaster has been attributed to a mutation resulting in an Ala302 → Ser replacement in the Rdl GABA receptor subunit and in D. simulans to an homologous Ala → Ser or Gly replacement. These mutations are shown here to greatly reduce [3H]EBOB binding, i.e. lower affinity and apparent number of binding sites. The Ala → Ser replacement with both melanogaster and simulans almost always reduces the potency in inhibiting [3H]EBOB binding of each of eight channel blockers and of muscimol and GABA. The Ala → Gly replacement in D. simulans is generally less effective than the Ala → Ser modification in reducing sensitivity to the channel blockers and to muscimol and GABA. The channel activators avermectin and moxidectin usually retain their inhibitory potency in the Rdl subunit mutants. Thus, it appears that replacement of Ala by Ser generally modifies the non-competitive blocker site and its coupling to the GABA-recognition site with less effect on the channel activator site. In contrast, the Ala → Gly replacement has less impact in protecting the chloride channel from the action of insecticidal blockers. Each of the resistant strains has the same level of resistance to the lethal action of the five channel blockers examined but none to avermectins and muscimol.

AB - The non-competitive blocker site of the GABA-gated chloride ion channel in normal susceptible strains of Drosophila melanogaster and simulans binds 4-n [3H]propyl-4′-ethynylbicycloorthobenzoate ([3H]EBOB) at specific sites with Kds of 1.6-1.9 nM and Bmaxs of 171-181 fmol/mg protein. This specific binding of [3H]EBOB is strongly inhibited by: a large number and variety of insecticidal channel blockers at 20 nM (lindane, α-endosulfan, dieldrin, 12-ketoendrin, fipronil, and a representative bicycloorthobenzoate and dithiane) or 200 nM (picrotoxinin); the insecticidal channel activators avermectin and moxidectin at 20 nM; muscimol at 30 μM and GABA at 300 μM. Cyclodiene resistance in D. melanogaster has been attributed to a mutation resulting in an Ala302 → Ser replacement in the Rdl GABA receptor subunit and in D. simulans to an homologous Ala → Ser or Gly replacement. These mutations are shown here to greatly reduce [3H]EBOB binding, i.e. lower affinity and apparent number of binding sites. The Ala → Ser replacement with both melanogaster and simulans almost always reduces the potency in inhibiting [3H]EBOB binding of each of eight channel blockers and of muscimol and GABA. The Ala → Gly replacement in D. simulans is generally less effective than the Ala → Ser modification in reducing sensitivity to the channel blockers and to muscimol and GABA. The channel activators avermectin and moxidectin usually retain their inhibitory potency in the Rdl subunit mutants. Thus, it appears that replacement of Ala by Ser generally modifies the non-competitive blocker site and its coupling to the GABA-recognition site with less effect on the channel activator site. In contrast, the Ala → Gly replacement has less impact in protecting the chloride channel from the action of insecticidal blockers. Each of the resistant strains has the same level of resistance to the lethal action of the five channel blockers examined but none to avermectins and muscimol.

UR - http://www.scopus.com/inward/record.url?scp=0028816129&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0028816129&partnerID=8YFLogxK

U2 - 10.1016/0024-3205(95)00006-R

DO - 10.1016/0024-3205(95)00006-R

M3 - Article

VL - 56

SP - 757

EP - 765

JO - Life Sciences

JF - Life Sciences

SN - 0024-3205

IS - 10

ER -