Studying the Dynamics of Ligand–Receptor Complexes by Single-Molecule Techniques - Single Molecule Dynamics in Life Science - page 143

AMPAR containing the

glutamate receptor subunit
GluR2 in cultured hippocam-
pal neurons and tagged with
anti-GluR2 antibody labeled
with Cy5 or Alexa 647

Glutamate SMI Diffusion: Glutamate application increased both receptor

mobility inside synapses and the proportion of mobile
receptors in the juxta-synaptic region.

[154]

[155] a Benzodiazepine receptor in Benzodiazepine Ro7-1986/ FCS Diffusion: Two populations of ligand receptor complexes

with different mobility. A clustering preceding internali-
zation of the GABA A R

-

hippocampal neurons 602 uor-aze-

pam) labeled with Alexa 532

(N-des-diethyl-

ligand complexes could explain the

-

slower diffusion time.

Binding: Saturation experiments yielded B max ¼ 38 nM and

K d ¼ 9.9 nM. Time course displacement with an excess of

non-labeled agonist, midazolam, showed a mono-expo-
nential decrease with k diss ¼ 10 3 s 1 . 1.3 ·

GABA A R in hippocampal Muscimol labeled with Alexa FCS Diffusion: Two populations of ligand receptor complexes [156 158]

- -

neurons 532 with different mobility.

Binding: Binding isotherm yielded one binding site per
receptor with K d ¼ 18.4 nM. Time-de-

pendent displacement experiments showed a mono-ex-
ponential decrease with k diss ¼

3.4 nM and B max ¼

10 2 s 1 . A positive
cooperativity of ligand binding yielding to a selective in-
crease in the fraction of ligand

5.4 ·

receptor complexes with
hindered mobility was observed by co-incubation with
midazolam [156], protoberberine type 2 alkaloids [157], or
xanthohumol [158].

-

[159] b

GABA A R in human cortical
neurons

Kavain labeled with
tetramethylrhodamine

FCS Diffusion: One fraction of bound ligand.

(Continued)