Datensatz

Zusammenfassung

1. Patch-clamp methods have been used to examine single-channel properties of recombinant GluR5 and GluR6 kainate-preferring glutamate receptors which differ in a single amino acid residue as a result of RNA editing at the Q/R (glutamine/arginine) site. Subunits were expressed alone or in combination with the high-affinity kainate receptor subunit KA - 2 in transfected human embryonic kidney (HEK-293) cells. 2. In outside-out patches, unedited homomeric GluR6(Q) receptors exhibited directly resolved domoate-activated single-channel conductances of 8, 15 and 25 pS. Variance analysis of GluR6(Q) responses gave a mean conductance of 5.4 pS, while the edited isoform GluR6(R) had an unusually low channel conductance (225 fS). 3. Homomeric channels composed of GluR5(Q) subunits exhibited three conductance states of 5, 9 and 14 pS characterized by prolonged burst activations in the presence of domoate. In contrast, the GluR5(R) subunit, which has not previously been reported to form functional homomeric receptors, had an extremely low conductance (< 200 fS). 4. Heteromeric GluR6(Q)/KA-2 kainate receptors gave single-channel events indistinguishible from homomeric GluR6(Q) channels. Conversely, openings produced by GluR5(Q)KA-2 and GluR5(Q) receptors differed from each other in their kinetic properties. The primary effect of co- expression of KA-2 with GluR5(Q) was a dramatic shortening in channel burst length. 5. Spectral and variance analyses were used to estimate mean single-channel conductances of heteromeric edited receptor- channels; channel conductances were 950 fS for GluR5(R)KA-2 receptors and 700 fS for GluR6(R)/KA-2 receptors. Both receptor types had significantly higher conductances than the respective homomeric channels, GluR5(R) and GluR6(R). 6. We conclude that Q/R site editing dramatically reduces single-channel conductance. Furthermore, we find similarity between the kainate receptor-channels described in sensory neurones and the recombinant GluR5(Q) homomeric channel. Characterization of recombinant single-channel properties could therefore aid identification of the native kainate receptors