Replacement of asparagine with arginine at the extracellular end of the second transmembrane (M2) region of insect GABA receptors increases sensitivity to penicillin G.
The actions of penicillin-G (PCG) on wild-type and mutant Drosophila GABA receptor (RDL) subunits expressed in Xenopus oocytes were studied under two-electrode voltage-clamp. PCG was found to be a non-competitive antagonist of homomeric Drosophila RDL receptors with an IC(50) of 20.41 +/- 1.66 mM at EC(50) GABA. Substitution of a single amino acid (N318R) at the extracellular end of the channel lining region of the RDL subunit increased the potency of GABA approximately four fold, and increased the IC(50) of PCG to 5.09 +/- 0.38 mM. Although the antagonism by PCG on wild-type RDL receptors was independent of membrane potential, PCG action on the N318R mutant showed pronounced voltage-dependency, being much more effective at positive membrane potentials. Thus, in RDL homomers, the replacement of N318 by R318, a residue present at the equivalent position in vertebrate GABA(A) receptors, confers a vertebrate-like PCG pharmacology to the N318R mutant receptor. The A301S mutation that confers resistance to dieldrin did not significantly affect the antagonism by PCG.
Hosie
Alastair M
AM
Department of Pharmacology, University College, Gower St, London WC1E 6BT, UK.
Buckingham
Steven D
SD
Hamon
Alain
A
Sattelle
David B
DB
D000818:Animals; D001120:Arginine / Q000235:genetics / Q000378:metabolism*; D001216:Asparagine / Q000235:genetics / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004330:Drosophila; D018756:GABA Antagonists / Q000494:pharmacology; D018757:GABA Modulators / Q000494:pharmacology*; D020128:Inhibitory Concentration 50; D019476:Insect Proteins / Q000737:chemistry / Q000502:physiology; D007313:Insecta; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology / Q000528:radiation effects; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed / Q000379:methods; D009154:Mutation / Q000502:physiology; D009865:Oocytes; D018408:Patch-Clamp Techniques / Q000379:methods; D010400:Penicillin G / Q000494:pharmacology*; D018079:Receptors, GABA / Q000737:chemistry / Q000187:drug effects* / Q000502:physiology; D014981:Xenopus
D018756:GABA Antagonists; D018757:GABA Modulators; D019476:Insect Proteins; D018079:Receptors, GABA; D001216:Asparagine; D001120:Arginine; D010400:Penicillin G