Lh 21

AM251, SR141716A (Rimonabant), URB447, LH21, AM630, O-2545, and CP55,940 were obtained from Cayman Chemical (Ann Arbor, MI) and Gp1a was obtained from Tocris Bioscience (Minneapolis, MN).

8-week-old female C57BL/6J mice (NC mice) and spontaneous diabetic KKAy mice were purchased from the Experimental Animal Center, Chinese Academy of Medical Sciences, Beijing. Mice were housed in a room under controlled temperature (23 ± 1°C) and 12-h light–dark cycle. The KKAy mice were fed with a high-fat diet and water ad libitum. At the beginning of the study, the KKAy mice were randomized into the vehicle-treated model control group (MC) or the two LH-21 (1 and 3 mg/kg) treatment groups based on their initial body weight and blood glucose levels (n = 8). LH-21 (Cayman Chemical, Ann Harbor, MI, USA) was administered by IP injection daily for 3 weeks, while mice in NC and MC group were IP injected with vehicle (5% Tween 80 and 1% ethanol) for 3 weeks. The dosages of LH-21 were chosen according to previously described (12 (link), 30 (link)). Individual body weight and food consumption were measured every 2 days. At the last day of the experiment, overnight fasted mice were sacrificed by decapitation. Plasma was collected for immediate assessment of serum biochemical parameters. The intraperitoneal WAT and interscapular brown adipose tissue (BAT) were excised and weighed, and then the WAT and the isolated aortae were rapidly frozen in liquid nitrogen for subsequent gene expression and western blot assay.

LH-21 was purchased from Cayman (Cayman Chemical, Ann Harbor, MI), dissolved in 100% ethanol and stored in 30 mg/ml aliquots. For daily injections, fresh dilutions were prepared in vehicle (5% Tween 80 in saline; final ethanol concentration 1%). Either vehicle or LH-21 were i.p. injected daily for two weeks in the obese pre-diabetic mice that had been fed a HFD for 15 weeks. An effective dosage of 3 mg/Kgday⁻¹ LH-21 was used to treat animals as previously described^{12 (link)}. For the pharmacological experiments in naïve healthy mice LH-21 was prepared and administered as above. CID16020046 was purchased from Tocris (Tocris Bioscience, Bristol, UK), dissolved in 100% DMSO and stored in 30 mg/ml aliquots. In the experiments, CID16020046 was dissolved in saline and i.p. injected 15 min before LH-21 injections.