Pharmascan

All MRI measurements were acquired utilizing a 7 Tesla Bruker scanner with a maximum gradient of 360 mT/m (70/16 PharmaScan, Bruker Biospin GmbH, Germany). Under inhaled isoflurane anesthesia (3% induction; 1.5% maintenance) and circulating warm water, animals were imaged using a receive-only surface coil. Scout T2-weighted images were first acquired in the coronal, transverse and sagittal planes with a rapid-acquisition-with-relaxation-enhancement (RARE) pulse sequence to position the subsequent MR images along standard anatomical orientations in a reproducible manner. For DTI, diffusion weighted images were acquired using the multi-shot spin-echo echo-planar imaging sequence, with repetition time (TR)/echo time (TE) = 3000/30 ms, field-of-view (FOV) = 32 x 32 mm², matrix resolution = 128 x 128, in-plane resolution = 250 x 250 μm², slice thickness = 1 mm, diffusion weighting factor (b) = 0 and 1000 s/mm², diffusion time (Δ) = 15 ms, diffusion gradient duration (δ) = 5 ms, 4 shots and 30 diffusion directions. Cr-enhanced MRI and Mn-enhanced MRI were performed using 2D T1-weighted RARE sequences covering the eye and the brain with TR/TE = 475/8.8 ms, RARE factor = 4, number of averages = 26, FOV = 32 x 32 mm², voxel resolution = 125 x 125 μm² and slice thickness = 0.8 mm.

All MRI measurements were acquired using a 7-T Bruker scanner with a maximum gradient of 360 mT/m (70/16 PharmaScan, Bruker Biospin GmbH, Germany), a 72 mm birdcage transmit-only radiofrequency coil and an actively decoupled receive-only quadrature surface coil. Under inhaled isoflurane anaesthesia (3% induction and 1–1.5% maintenance), T2-weighted images were acquired with a spin-echo rapid-acquisition-with-relaxation-enhancement (RARE) pulse sequence covering the eye and the brain with repetition time (TR) = 4200 ms, echo time (TE) = 36 ms, RARE factor = 8, number of averages = 1, field of view (FOV) = 20 mm × 20 mm (for P1, P5, and P10) and 40 mm × 40 mm (for P15, P21, P28, and P60), in-plane acquisition resolution = 78 μm × 78 μm (for P1, P5, and P10) and 156 μm × 156 μm (for P15, P21, P28, and P60), and slice thickness = 0.8 mm. Chromium-enhanced MRI was performed using a T1-weighted RARE sequence with TR/TE = 475/8.8 ms, RARE factor = 4, number of averages = 26, FOV = 32 mm × 32 mm, in-plane acquisition resolution = 125 μm × 125 μm, and slice thickness = 0.8 mm.

Proton magnetic resonance spectroscopy (¹HMRS) experiments were performed on a MR 7 T horizontal bore magnet 70/16 PharmaScan, ParaVision 6.0.1 (Bruker BioSpin GmbH, Rheinstetten, Germany) using a volume coil with 72 mm inner diameter for transmission and 20 mm surface loop coil for reception. During this stage of the experiments, the animals were anesthetized with an isoflurane and oxygen mixture (3.5% isoflurane for induction and 1.7–2.2% for maintenance). Respiration rate was monitored throughout scanning and isoflurane concentration was adjusted to maintain respirations within a specified target zone (35–45 rpm). Body temperature was controlled by a rectal thermal probe and maintained at physiological values (about 37 °C) using a warm water circulation system. The number of breaths and body temperature were monitored throughout this part of the study using the MR-compatible Small Animal Monitoring System (SA Instruments, Inc., Stony Brook, NY, USA). Rats were scanned in sessions of ~2.5 h each. Four groups of Wistar rats were examined in the study: separated females, non-separated females, separated males and non-separated males.

MRI is used in the rat C6 glioma model for confirmation of the volume of tumor on day 14 after implantation. MRI is undertaken on a 7.0 T animal MRI scanner (70/16 PharmaScan, Bruker Biospin GmbH, Germany) using a 38 mm birdcage rat brain quadrature resonator for radiofrequency transmission and reception. Briefly, rats are anesthetized using inhaled isoflurane/O₂ (3% for induction and 1.5–2% for maintenance). During the MRI scan, the rats are prostrated on a custom made holder to minimize head motion while respiration is maintained at a rate of 50 breaths/min. Scout T₂-weighted imaging (T₂WI) in three planes with a fast spin echo pulse sequence is first acquired to control rat head positioning. Next, a coronal T₂WI scan is acquired using a rapid-acquisition relaxation-enhancement pulse sequence with the following parameters: field of view = 3 × 3 cm, matrix size = 256 × 256, repetition time = 2500 ms, echo time = 33 ms, slice thickness = 1.0 mm, slice gap = 1.0 mm, and acquisition time = 1 min 20 s. [35 (link)]

MRI was examined by the 7.0 T Bruker PharmaScan MRI scanner (70/16 PharmaScan, Bruker BioSpin GmbH, Germany). A quadrature volume resonator (inner diameter 72 mm) was used for radio frequency transmission, and a 4-element surface coil array was used for signal reception. For anesthesia induction, a mixture of isoflurane (5% for induction and 0.3%-0.5% for maintenance) and dexmedetomidine sedation was used [11 (link)]. A nose mask with a bite bar was used to deliver the isoflurane mixture on the MRI bed and to fix the animal in prone position. During the MRI scan, the rat was prostrated on a custom-made holder to minimize head motion, while the respiration rate was controlled between 60 and 80 breaths/min. An echo-planar imaging (EPI) sequence was performed: matrix size = 64 × 64, flip angle = 30°, resolution = 0.5 mm × 0.5 mm, slice thickness = 1.0 mm, slice gap = 0, repetition time (TR) = 2 s, echo time (TE) = 18 ms, and volume = 180. A coplanar T2-weighted scan was also acquired.

Eight-week male FGF21KO mice and WT littermates fed with HFD for 8 weeks were replenished with rmFGF21 or vehicle for 4 weeks. MRI measurements of mice were performed utilizing a 7 T MRI scanner (70/16 PharmaScan, Bruker) by a radiologist who was blinded to group allocation. The mice were anesthetized with 3% isoflurane. During MRI, the animals were placed on a plastic cradle with the head fixed with a tooth bar and plastic screws in the ear canals. Continuous physiological monitoring was performed using an MRI-compatible system (SA Instruments). Spin echo sequences: TR/TE = 150/5.8 ms, FOV = 31.50 × 20.25 mm², matrix size = 210 × 135. The whole abdomen of each mouse were covered with axial slices (thickness 1 mm, no spacing). Acquired images underwent measurement of SAT and VAT using a semiautomated segmentation method. According to the signal intensity of adipose tissue, SFA and VFA outline was manually traced with a graphic user interface. The area inside the outline was automatically labeled and calculated.