Functional characterization of Musca glutamate- and GABA-gated chloride channels expressed independently and coexpressed in Xenopus oocytes

Insect Mol Biol. 2006 Dec;15(6):773-83. doi: 10.1111/j.1365-2583.2006.00680.x.

Abstract

Ligand-gated chloride channels (LGICs) are important targets for insecticides and parasiticides. Genes encoding subunits of two LGICs, a glutamate-gated chloride channel (MdGluCl-alpha) and a gamma-aminobutyric acid (GABA)-gated chloride channel (MdRdl), were cloned from house-flies (Musca domestica L.). These genes were first expressed independently in Xenopus laevis oocytes by cRNA injection in order to investigate the pharmacology of these ligand-gated channels using two-electrode voltage-clamp electrophysiology. It was found that L-glutamate and GABA activated the MdGluCl-alpha homo-oligomers with an EC(50) value of 30 microM and the MdRdl homo-oligomers with an EC(50) value of 101 microM, respectively. Both channels were chloride ion-permeable, and the MdRdl channel was more sensitive to chloride channel blockers, such as gamma-hexachlorocyclohexane (gamma-HCH), fipronil and picrotoxinin, than the MdGluCl-alpha channel. MdGluCl-alpha required only 1-2 days of incubation after cRNA injection to be expressed in oocytes, whereas 4-7 days of incubation was necessary to achieve MdRdl expression. However, when the cRNA of MdGluCl-alpha was injected at a dose of 1% (w/w) 1 day after the injection of the cRNA of MdRdl, a significant increase in the current amplitude of responses to GABA was observed, and the incubation period necessary for MdRdl expression became shorter. These results suggest that MdGluCl-alpha assists in the expression of MdRdl when the two are coexpressed.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antiparasitic Agents / pharmacology
  • Chloride Channel Agonists
  • Chloride Channels / antagonists & inhibitors
  • Chloride Channels / chemistry
  • Chloride Channels / metabolism*
  • Cloning, Molecular
  • DNA, Complementary / metabolism
  • Dose-Response Relationship, Drug
  • GABA Agonists / pharmacology
  • GABA Antagonists / pharmacology
  • Gene Expression*
  • Glutamates / pharmacology
  • Hexachlorocyclohexane / pharmacology
  • Ion Channel Gating* / drug effects
  • Muscidae / metabolism*
  • Oocytes / drug effects
  • Oocytes / metabolism*
  • Picrotoxin / analogs & derivatives
  • Picrotoxin / pharmacology
  • Protein Structure, Quaternary / drug effects
  • Pyrazoles / pharmacology
  • RNA, Complementary / metabolism
  • Sesterterpenes
  • Xenopus laevis*
  • gamma-Aminobutyric Acid / metabolism*
  • gamma-Aminobutyric Acid / pharmacology

Substances

  • Antiparasitic Agents
  • Chloride Channel Agonists
  • Chloride Channels
  • DNA, Complementary
  • GABA Agonists
  • GABA Antagonists
  • Glutamates
  • Pyrazoles
  • RNA, Complementary
  • Sesterterpenes
  • glutamate-gated chloride channels
  • Picrotoxin
  • gamma-Aminobutyric Acid
  • Hexachlorocyclohexane
  • picrotoxinin
  • fipronil