Homomeric and heteromeric ion channels formed from the kainate-type subunits GluR6 and KA2 have very small, but different, unitary conductances.

摘要

1. Patch-clamp methods were used to study recombinant glutamate receptor (GluR) ion channels expressed in human embryonic kidney cells (HEK 293) after transfection of the cells with cDNAs encoding kainate (KA)-type GluR subunits. Cells were transiently or stably transfected with the fully edited (R) version of GluR6 or they were contransfected with GluR6(R) and KA2. 2. Concentration-response data were obtained for KA and glutamate activation of homomeric GluR6(R) channels and fitted with Hill-type equations to give values for the agonist concentration at half-maximal activation (EC50), the Hill coefficient (nH), and the maximum current (Imax). Analysis of results obtained in seven cells with KA gave mean values of 0.47 microM and 1.47 for the EC50 and nH, respectively. The corresponding values for glutamate were 32 microM and 1.21 (n = 5). The Imax values obtained for KA and glutamate in the same cells were similar, suggesting that both KA and glutamate are full agonists at homomeric GluR6(R) channels.3. Spectral density analysis of current noise evoked by KA and glutamate in GluR6(R)-expressing cells, or in outside-out patches from these cells, was used to obtain estimates of the apparent unitary conductance (gamma noise) of homomeric GluR6(R) channels. Results obtained with 10 microM KA in 15 cells gave a mean gamma noise value of 231 fS. The corresponding value from analysis of noise in outside-out patches was 259 fS (n = 4). Spectral density analysis of glutamate-evoked noise in six cells gave a mean gamma noise value of 264 fS. 4. Noise analysis of whole cell currents recorded in GluR6(R)/ KA2-expressing cell gave a mean gamma noise value of 572 fS (n = 9). Unlike homomeric GluR6(R) channels, GluR6(R)/KA2 heteromers were activated by alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid (AMPA) (0.25-5 mM). The mean EC50 for KA activation of GluR6(R)/KA2 channels was 1.62 microM (n = 6). 5. The results indicate that both homomeric GluR6(R) and heteromieric GluR6(R)/KA2 channels have a unitary conductance in the femtosiemen range. The coassembly of KA2 with GluR6(R) results in channels that have a different affinity for AMPA and KA and a two- to threefold larger unitary conductance.