Channel gating in the absence of agonist by a homo-oligomeric molluscan GABA receptor expressed in Xenopus oocytes from a cloned cDNA.
We have previously described the isolation of a complementary DNA (cDNA) from the freshwater mollusc Lymnaea stagnalis encoding a polypeptide that exhibits approximately 50% identity to the beta-subunits of vertebrate gamma-aminobutyric acid (GABA) type A (GABAA) receptor. When expressed in Xenopus laevis oocytes from in vitro-transcribed RNA, the snail subunit forms functional homo-oligomeric receptors possessing chloride-selective ion channels. In recordings from voltage-clamped oocytes held at -60 mV, GABA induced an inward current, whereas application of the chloride-channel blocker picrotoxin (in the absence of agonist) elicited an apparent outward current. Single channel recordings obtained from cell-attached patches have revealed a single population of approximately 20 pS channels, with an open probability greater than 90% (at a pipette potential of -100 mV) in the absence of GABA. The relationship between single channel current and pipette potential was linear over the studied range (-100 mV to +60 mV), but the open probability was less for hyperpolarizations than for depolarizations. The spontaneous channel openings were blocked by micromolar concentrations of picrotoxin. Functional hetero-oligomeric receptors were formed when the molluscan subunit was co-expressed in oocytes with the bovine GABAA receptor alpha 1-subunit, but the channels gated by these receptors did not open spontaneously.
Bhandal
N S
NS
Department of Life Science, University of Nottingham, UK.
Ramsey
R L
RL
Harvey
R J
RJ
Darlison
M G
MG
Usherwood
P N
PN
D000818:Animals; D002417:Cattle; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D005260:Female; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology*; D007473:Ion Channels / Q000187:drug effects / Q000502:physiology*; D008195:Lymnaea; D008974:Mollusca / Q000378:metabolism*; D009865:Oocytes / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D018079:Receptors, GABA / Q000737:chemistry / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology; D014982:Xenopus laevis / Q000378:metabolism*; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology
D018076:DNA, Complementary; D007473:Ion Channels; D018079:Receptors, GABA; D010566:Virulence Factors, Bordetella; D005680:gamma-Aminobutyric Acid