Super-resolution Imaging of Synaptic and Extra-synaptic Pools of AMPA Receptors with Different-sized Fluorescent Probes
Super-resolution Imaging of Synaptic and Extra-synaptic Pools of AMPA Receptors with Different-sized Fluorescent Probes
复制标题
使用不同尺寸荧光探针对 AMPA 受体突触和突触外池进行超分辨率成像
DOI：
10.1101/096966
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发表时间：
2016
期刊：
影响因子：
--
通讯作者：
P. Selvin
中科院分区：
文献类型：
--
作者：
Sang Hak Lee;Chaoyi Jin;En Cai;Pinghua Ge;Y. Ishitsuka;K. Teng;A. Thomaz;Duncan L Nall;M. Baday;Okunola Jeyifous;D. Demonte;Christopher M. Dundas;Sheldon Park;W. N. Green;P. Selvin
AMPA受体（AMPAR）是否进入神经元突触，从内部池的胞吐作用，或从外部膜结合池的扩散，是激烈的争议。3D超分辨率荧光纳米显微镜测量AMPAR的动态和位置是解决这个问题的一个强大的方法。然而，探针尺寸和对紧密填充空间的可接近性可能是有限的。因此，我们用不同尺寸的荧光团标记AMPAR：小有机荧光染料（~ 4 nm），小量子点（sQD，直径~10 nm）或大（商业）量子点（bQD，直径~ 20 nm）。然后，我们比较了它们的扩散率，轨迹和位置相对于突触后密度（PSD）蛋白，荷马1C。用sQD或有机荧光团的小探针标记，我们发现AMPAR主要位于PSD内（约73-93%），并且通常驻留在具有受限扩散的“纳米域”中。相反，当用bQD标记时，只有5-10%的AMPAR在PSD内。结果可以解释为相对自由的访问，或缺乏，突触裂缝的AMPAR标记时，分别与小或大的探针。这意味着AMPAR主要进入PSD后不久，他们的胞吐，而不是从一个大的扩散池突触外AMPAR。
Whether AMPA receptors (AMPARs) enter into neuronal synapses, by exocytosis from an internal pool, or by diffusion from an external membrane-bound pool, is hotly contested. 3D super-resolution fluorescent nanoscopy to measure the dynamics and placement of AMPAR is a powerful method for addressing this issue. However, probe size and accessibility to tightly packed spaces can be limiting. We have therefore labeled AMPARs with differently sized fluorophores: small organic fluorescent dyes (~ 4 nm), small quantum dots (sQD, ~10 nm in diameter), or big (commercial) quantum dots (bQD, ~ 20 nm in diameter). We then compared their diffusion rate, trajectories, and placement with respect to a postsynaptic density (PSD) protein, Homer 1c. Labeled with the small probes of sQDs or organic fluorophores, we find that AMPARs are located largely within PSDs (~73-93%), and generally reside in “nanodomains” with constrained diffusion. In contrast, when labeled with bQDs, only 5-10% of AMPARs are within PSDs. The results can be explained by relatively free access, or lack thereof, to synaptic clefts of the AMPARs when labeled with small or big probes, respectively. This implies that AMPARs primarily enter PSDs soon after their exocytosis and not from a large diffusive pool of extrasynaptic AMPARs.
影响因子：
16.6
作者：
Cai, En;Ge, Pinghua;Lee, Sang Hak;Jeyifous, Okunola;Wang, Yong;Liu, Yanxin;Wilson, Katie M.;Lim, Sung Jun;Baird, Michelle A.;Stone, John E.;Lee, Kwan Young;Davidson, Michael W.;Chung, Hee Jung;Schulten, Klaus;Smith, Andrew M.;Green, William N.;Selvin, Paul R.
通讯作者：
Selvin, Paul R.
影响因子：
25
作者：
Frischknecht, Renato;Heine, Martin;Gundelfinger, Eckart D.
通讯作者：
Gundelfinger, Eckart D.
影响因子：
33.6
作者：
Syková E;Nicholson C
通讯作者：
Nicholson C