Single nanoparticle tracking of [Formula: see text]-methyl-d-aspartate receptors in cultured and intact brain tissue.
Neurophoton. 2016-07-12; 3(4): 041808
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1. Neurophotonics. 2016 Oct;3(4):041808. doi: 10.1117/1.NPh.3.4.041808. Epub 2016
Single nanoparticle tracking of [Formula: see text]-methyl-d-aspartate receptors
in cultured and intact brain tissue.
Varela JA(1), Ferreira JS(1), Dupuis JP(1), Durand P(1), Bouchet D(1), Groc L(1).
(1)Université de Bordeaux, Interdisciplinary Institute for Neuroscience, UMR
5297, 146 rue Leo Saignat, 33077 Bordeaux, France; Centre National de la
Recherche Scientifique, Interdisciplinary Institute for Neuroscience, UMR 5297,
33000 Bordeaux, France.
Recent developments in single-molecule imaging have revealed many biological
mechanisms, providing high spatial and temporal resolution maps of molecular
events. In neurobiology, these techniques unveiled that plasma membrane
neurotransmitter receptors and transporters laterally diffuse at the surface of
cultured brain cells. The photostability of bright nanoprobes, such as quantum
dots (QDs), has given access to neurotransmitter receptor tracking over long
periods of time with a high spatial resolution. However, our knowledge has been
restricted to cultured systems, i.e., neurons and organotypic slices, therefore
lacking several aspects of the intact brain rheology and connectivity. Here, we
used QDs to track single glutamatergic [Formula: see text]-methyl-d-aspartate
receptors (NMDAR) in acute brain slices. By delivering functionalized
nanoparticles in vivo through intraventricular injections to rats expressing
genetically engineered-tagged NMDAR, we successfully tracked the receptors in
native brain tissue. Comparing NMDAR tracking to different classical brain
preparations (acute brain slices, cultured organotypic brain slices, and cultured
neurons) revealed that the surface diffusion properties shared several features
and are also influenced by the nature of the extracellular environment. Together,
we describe the experimental procedures to track plasma membrane NMDAR in
dissociated and native brain tissue, paving the way for investigations aiming at
characterizing receptor diffusion biophysics in intact tissue and exploring the
physiopathological roles of receptor surface dynamics.