U73122

Thapsigargin (Sigma, T9033), Tunicamycin (Sigma, T7765), Maprotiline (Sigma, M9651), Amoxapine (MedChemExpress, HY-B0991), Desloratadine (MedChemExpress, HY-B0539), Desipramine (Sigma, D3900), Trifluoperazine (Sigma, T8516), Clomipramine (Sigma, C7291), Amitriptyline (Sigma, A8404), Quetiapine (Sigma, Q3638), Olanzapine (MedChemExpress, HY-14541), Doxepin (MedChemExpress, B078), Loxapine (Sigma, L106), NorQuetiapine (Sigma, 07849), dimethyl sulfoxide (DMSO) (Sigma, D8418), 2-APB (MedChemExpress, HYW009724), U-73122 (MedChemExpress, HY13419), Go 6983 (MedChemExpress, HY13689), GSK2606414 (Sigma, 516535), ISRIB (MedChemExpress, HY-12495).

Ca²⁺ was measured by a Ca²⁺ fluorescent probe fluo‐4‐AM kit following the manufacturer's instructions. C2C12 cells were incubated with ryanodine (100 nM, MedChemExpress) or U73122 (1 μM, MedChemExpress, Monmouth Junction, USA) for 1 h to block the endoplasmic reticulum Ca²⁺ channel. Then, the cells were washed 3 times with Hank's balanced salt solution and incubated with 10 μM fluo‐4‐AM at 37℃ for 1 h. After incubation, cells were then rewashed 3 times. Nikon Eclipse Ti‐s microscopy (Nikon) was used to observe fluorescence. Fluorescent data were acquired after excitation at 490 nm and intensity at 525 nm.

All salts were from Sangon Biotech in Shanghai. Acetylcholine, N^ω-nitro-L-arginine and indomethacin were offered by Sigma-Aldrich (St Louis, USA). NE was purchased from GRANDPHARMA (China) co. LTD. 17β-Estradiol, LPS, Propyl pyrazole triol (PPT), MPP hydrochloride (MPP), G-1, G-15, Diarylpropionitrile (DPN), PHTPP, ouabain, TRAM-34, apamin, U73122, U73343, 2-APB and SKA-31 were supplied by MedChemExpress (MCE; New Jersey, USA), SN-6 was provided by Tocris (Bristol, UK). LiCl was supplied by Macklin (China). The human phospholipase C Beta 1 Polyclonal antibody (Cat No. 26551-1-AP) and human ITPR1-specific Polyclonal antibody (Cat No. 19962-1-AP) were from proteintech. The commercial kits of serum ALT, AST, LD and BUN were from Neobioscience (Jiancheng Bioengineering Institute, Nanjing, China). The commercial kit of serum DAO was from Solarbio. The commercial kit of serum Cr was from Beijing Leagene Biotechnology co. LTD.

Ca²⁺ was measured by a Ca²⁺ fluorescent probe fluo-4-AM kit following the manufacturer's instructions. 3T3-L1 cells were incubated with ryanodine (100 nM, MedChemExpress, Monmouth Junction, USA) or U73122 (1 μM, MedChemExpress, Monmouth Junction, USA) for 1 h to block the endoplasmic reticulum Ca²⁺ channel. Then, the cells were washed 3 times with Hank's Balanced Salt Solution and incubated with 10 μM fluo-4-AM at 37°C for 1 h. After incubation, cells were then rewashed 3 times. Nikon Eclipse Ti-s microscopy (Nikon, Tokyo, Japan) was used to observe fluorescence. Fluorescent data were acquired after excitation at 490 nm and intensity at 525 nm.

Dimethyl sulfoxide (DMSO) and platelet-derived growth factor (PDGF) were purchased from Beyotime Biotechnology; 2-amino-ethoxydiphenyl borate (2-APB) and nifedipine were from Sigma-Aldrich; thapsigargin and U73122 were from MedChemExpress (Shanghai, China); Ruthenium red (RuR) was from Macklin (Shanghai, China); fibronectin and the calcium-free culture medium were from Thermo Fisher Scientific.

Rat pancreatic INS-1 cells were kindly provided by Prof. Hai Qian (China Pharmaceutical University, Nanjing, China) [21 (link)]. The cells were routinely reseeded every 2-3 days and cultured in RPMI 1640 medium containing 11 mmol/L (mM) glucose, 2 mM L-glutamine supplemented with 10% heat-inactivated fetal bovine serum (FBS), 1 mM sodium pyruvate, 50 μmol/L (μΜ) β-mercaptoethanol, 10 mM N-2-hydroxyethyl piperazine-N-2-ethane-sulphonic acid (HEPES), 100 IU/mL penicillin, and 100 mg/mL streptomycin in a humidified cell incubator at 37°C in a humidified atmosphere (5% CO₂ and 95% air). Lek Pharmaceuticals d.d. generously provided us with atorvastatin (Ljubljana, Slovenija). Final concentrations of atorvastatin ranged from 0.2 μM to 20 μM. Pioglitazone hydrochloride, U-73122, and GW1100 were purchased from MedChem Express (NJ, USA). INS-1 cells were incubated with atorvastatin, pioglitazone, GW1100, FFA1 siRNA, and/or U-73122 for 24 h prior to insulin secretion and other experiments. atorvastatin, pioglitazone, GW1100, and U-73122 were dissolved in dimethyl sulfoxide (DMSO), and the final concentration of DMSO was adjusted to 0.1% (v/v). The medium containing the same amount of DMSO was used as the control.