Upregulation of mGluR5 uncovered in postmortem prefrontal cortex of 14 FXS patients
We are proud that our mGluR5 antibody was selected for this important study. The investigators found that mGluR5 binding density and protein expression were increased in the brains of FXS patients or carriers: Talakad G Lohith, Emily K Osterweil, Masahiro Fujita, Kimberly J Jenko, Mark F Bear, Robert B Innis. Is metabotropic glutamate receptor 5 upregulated in prefrontal cortex in fragile X syndrome? Molecular Autism 2013, 4:15 (24 May 2013).
Protocol: Tissue samples were homogenized using a mortar and pestle in fresh ice-cold, 50 mM Tris– HCl buffer (1:10 w/v) completing 3 × 10 passes with cooling on ice between homogenizations. Homogenates were centrifuged at 20,000g and 4°C for 25 minutes,followed by removal of the supernatant. Pellets were then resuspended in fresh ice-cold 50 mM Tris–HCl buffer and centrifuged again at the same settings. Pellets were resuspended in fresh ice-cold 50 mM Tris–HCl buffer at a protein concentration of approximately 1 mg of protein/mL. Aliquots were stored in a freezer at −80°C until further use. Protein concentrations were determined using the Bradford protein assay (Bio-Rad, Hercules, CA), and absorption was read at 595 nm.
Figure: mGluR5 receptor density and expression in prefrontal cortex of FXS individuals and healthy controls. (A) Binding curves from homologous competition binding of 0.48 nM of [3H] MPEPy to membrane preparation from FXS and control subject samples at concentrations of unlabeled MPEPy ranging from 0.1 nM to 1μM. Individual binding curves were obtained from the average of triplicate measurements for e ach unlabeled ligand concentration. Data represent mean ± standard error in the mean from 14 FXS patients or carriers (FX) and 17 healthy controls (HC). At low concentration of unlabeled ligand, specific binding was higher for FXS than control samples. (B) Results of unpaired t-test tocompare the two groups. mGluR5 density tended to be higher (+16%;P = 0.058) in FXS patients than in the control group. Data represent mean ± standard deviation. Solid triangles (▲) in the FX group indicate the location of three FXS carriers; semisolid triang les indicate the location of a carrier with FXTAS. (C) Representative immunoblot for mGluR5. The mGluR5 band intensity was stronger for the FXS than control subject. Total protein stain of the same lanes confirmed equal-protein loading. (D) Average mGluR5: total protein ratio normalized to control subjects. The ratio was high and marginally significant (+32%; P=0.048) for the FXS group compared with controls. Data represent mean ± standard deviation.Solid triangles (▲) in the FX group indicate the location of three FXS carriers; semisolid triangles indicate the location of a carrier with FXTAS.
This study was the first to identify upregulation of mGluR5 density and expression in the prefrontal cortex of FXS patients or carriers compared to an age- and sex-matched control group. This is consistent with several studies in FXS model mice that postulate that the syndromic features of FXS are caused by an upregulated mGluR5 signaling pathway. Although the sample size was relatively small and the results could be secondary to prior medication treatment, these initial findings provide strong rationale for measuring mGluR5 in live patients using PET. Such in-vivo studies could measure mGluR5 in all brain regions; the results could also be correlated with treatment response to mGluR5 negative allosteric modulators.
Scientists grow retina cells from skin-derived stem cells - WASHINGTON - University of Wisconsin-Madison researchers have successfully grown multiple types of retina cells from two types of stem cells, giving new ho...
7 years ago