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. 2018 Jul 10;8:10414. doi: 10.1038/s41598-018-28682-6

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© The Author(s) 2018

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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The mGlu₂ receptor is mainly dimeric in cultured hippocampal neurons. (a) Schematic representation of the homodimeric mGlu₂ receptor N-terminally SNAP-tagged (ST-mGlu₂) and labeled with Alexa Fluor 488 (Alexa488). (b) Intensity image by two-photon microscopy of a primary neuron transfected with ST-mGlu₂ labeled with Alexa488. The white dashed square represents the area selected for further imaging and analysis. (c) Average intensity map obtained from a stack of 100 images of the dashed square in (b) after thresholding. (d) Molecular brightness (ε, counts/40 μs/molecule) map of the dashed square in (b). (e) Histogram of molecular brightness values corresponding to the map in (d). The center of the distribution is estimated with a Gaussian fit (red line) giving ε = 0.0798 ± 0.0030 or 1.66 ± 0.06 subunits/complex (expression 98 subunits per effective volume (V_eff = 0.355 fL)). (f) Number of mGlu₂ receptor subunits per complex as a function of subunits per V_eff. The vertical dotted lines indicate three expression regimes with different complex sizes. The horizontal dotted lines indicate the molecular brightness of a dimer ± SD. Data are from 14 independent experiments.