BBS tag inserted into the R1a subunit is able to bind BTX without
altering receptor pharmacology. A, schematic of R1a subunit
showing relative positions for the myc tag (blue) and the BBS
(red) and the primary sequence. B, images of fixed GIRK
cells transfected with R1aBBSR2 + GFP, α7/5HT3a +
GFP and GFP alone after treatment with 3 μg/ml BTX-Rhd. GFP (FITC) and
rhodamine channels are shown separately and then merged. C,
concentration response curves from GIRK cells transfected with either R1aR2
(▪; n = 8–11) or R1aBBSR2 with (▴;
n = 5) or without (•; n = 5) an application of 3
μg/ml BTX-Rhd. D, CGP55845 inhibition curves for R1aR2 (▪)
and R1aBBSR2 (•) using an EC50 GABA concentration
(n = 5). E, time course of responses to submaximal GABA
concentration (10 μm) for R1aR2 (▪; n =
5–8), and R1aBBSR2 with either BTX-Rhd (3 μg/ml)
pre-applied (•; n = 6–7) or applied after the first and
second GABA applications (○; n = 5–6).