Small Fluorescent Probes Show Iglurs Are In The Synapses Of Transfected Neurons Under Basal Conditions
BIOPHYSICAL JOURNAL(2017)
摘要
We imaged transiently transfected live glutamate receptors (iGluRs), namely AMPAR and NMDAR, with super-resolution in three-dimension, labeled with differently sized fluorophores. We used small organic fluorescent dyes (∼ 4 nm), small quantum dots (sQD, ∼10 nm in diameter), or big (commercial) quantum dots (bQD, ∼ 20 nm in diameter). The iGluRs were imaged along with a synaptic protein, Homer1c, using fiducial markers that eliminate stage drift, and under conditions where the receptor cross-linking is monitored or eliminated. With small probes under basal conditions, we find that both AMPAR and NMDAR are predominantly within the synapse (∼84-95%), in contrast to bQDs, which show only 5-10% within the synapse. The results can likely be explained by cross-linking and inhibited mobility of the iGluRs labeled with bQDs, but not with sQDs or organic fluorophores. Hence, this data indicates that there is not a highly diffusible pool of extrasynaptic iGluRs in the plasma membrane, as has been widely reported. In addition, within a synapse, the distribution is non-homogeneous, perhaps due to the non-homogeneous distribution of glutamate.
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