Fig 2: Antibody decoration of additional epitope-tagged NMDA receptors isolated from the total cellular pool. A and B, schematic illustration of a GluN1/GluN2A NMDA receptor subunit heterodimer, with the location of the tag (HA/His8) used to isolate the receptor indicated by the arrow, and the sites of antibody decoration (A, anti-HA (ABD), and B, anti-GluN2A (CTD)) indicated by the asterisk. Numbers refer to the subunits used (GluN1 and GluN2A). C, frequency distribution of angles between pairs of bound anti-HA antibodies. The curve indicates the fitted Gaussian function. The peak of the distribution is indicated. D, frequency distribution of angles between pairs of bound anti-GluN2A antibodies. E and F, schematic illustration of a GluN1/GluN2A NMDA receptor subunit heterodimer, with the location of the tag (HA/His8) used to isolate the receptor indicated by the arrow, and the sites of antibody decoration (E, anti-Myc (ABD), and F, anti-HA (ABD)) indicated by the asterisk. G, frequency distribution of angles between pairs of bound anti-Myc antibodies. H, frequency distribution of angles between pairs of bound anti-HA antibodies. For each distribution, the subunit arrangement revealed by the data is shown in the inset.

Fig 3: Arrangement of GluN2A subunits in GluN1/GluN2A NMDA receptors isolated from the total cellular pool. A, schematic illustration of a GluN1/GluN2A NMDA receptor subunit heterodimer, with the location of the tag (FLAG/His8) used to isolate the receptor indicated by the arrow, and the site of anti-His antibody decoration (post-TMD) indicated by the asterisk. Numbers refer to the subunits used (GluN1 and GluN2A). B, gallery of enlarged images showing receptors after incubation with anti-His antibody. The gallery shows single- (upper) and double-decorated receptors (lower). Scale bar, 20 nm; height scale, 0–3 nm. C, frequency distribution of molecular volumes of the central particles decorated by anti-His antibodies. The curve indicates the fitted Gaussian function. The peak of the distribution is indicated. D, frequency distribution of angles between pairs of bound antibodies. The subunit arrangement revealed by the data is shown in the inset.

Fig 4: Plantaris tendon tissue proper (TP), peritendinous connective tissue (PT), and cultured tendon cells (TC) stained for NMDAR1 a, b, c, d and pNMDAR1 e, f, g, h: Examples of immunoreactions (arrows) can be observed in slender and rounded tenocytes in the tendon tissue proper a, b, e, f, in cultured tendon cells c, gand in cells in the peritendinous tissue d, h. Nuclei in c and g are stained with DAPI (blue). In i and j, control stainings replacing the primary rabbit i and goat j antibodies with PBS are shown. No positive reactions can be seen. Bars indicate 20 µm length

Fig 5: Results of 2 and 3 days of strain on the gene expression of Gls a and Got1 b, as seen by qPCR, and the protein content of NMDAR1 as seen by densitometry of western blot c. After 2 days, there was a slight increase of Gls and Got1 mRNA a, b. After 3 days this increase is more pronounced a, b. NMDAR1 is reduced after 2 days and slightly, but not significantly, increased after 3 days c. Asterisks mark significance for outcome in relation to the control (CTR). Error bars show standard deviation. (* p < 0.05; ** p < 0.01; *** p < 0.001). n = 3