In vitro Radiopharmaceutical Evidence for MCHR1 Binding Sites in Murine Brown Adipocytes

Figures

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  Figure 1. Overlay of kinetic binding toward CHO-K1-ADRB3 and displacement curves of [125I]Iodocyanopindolol using the MCHR1 ligands SNAP-7941 (red line) and FE@SNAP (blue line) and ethanol (black line) as the respective vehicle control. Arrows indicate the addition time points of the MCHR1 ligands and vehicle, respectively.

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  Figure 2. Histological staining of lipids in mature brown adipocytes was performed using Oil Red O and haematoxylin for counterstaining. Lipids appear in red and chromatin in bluish.

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  Figure 3. Representative, background-corrected time-activity curves of [18F]FDG uptake by mature brown adipocytes (black) and pre-adipocytes (red) are shown as overlay.

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  Figure 4. Baseline refers to 60 min [18F]FDG uptake in the presence of vehicle control and was normalized to 100% uptake/well. Data is presented as mean ± standard deviation (SD) from at least three independent experiments.

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  Figure 5. [11C]SNAP-7941 binding to brown adipocytes in the presence of vehicle control (ethanol or DMSO) represents baseline (100% binding/well). SNAP-7941 (2 μM) significantly reduced [11C]SNAP-7941 binding by 22.9 ± 5.0%. The effect of the ADRB3 agonist CL 316,243 was less pronounced and the non-selective adrenergic receptor beta antagonist propranolol did not alter [11C]SNAP-7941 binding. Data is presented as mean ± standard deviation (SD) from at least three independent experiments.

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  Figure 6. Gene expression of Mchr1 in mouse lung, brain and murine brown adipocytes normalized to the respective housekeeping gene (Rpl27 n = 2, ß-actin n = 2). Values were calculated with respect to mouse spleen as negative control using the ΔΔCt method.

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  Figure 7. Western Blot analysis of MCHR1 expression (A) in murine brown adipocyte lysates and reference tissues. Data is presented as cropped image format. Corresponding Ponceau S staining as loading control (B) and calculated relative density of MCHR1 bands normalized to Ponceau S (C).