2.1 Animals
Dahl Salt Sensitive (DSS) male rats were provided by Beijing Vital River
Laboratory Animal Technology Co., Ltd. The animals were free to access
sterilized standard laboratory food and water. The animal room
environment was controlled (target conditions: temperature 20 to 26°C,
relative humidity 30 to 70%, 12 h of artificial light, and 12 h of
dark). Temperature and relative humidity were monitored twice daily. The
study was conducted in accordance with the Institutional Animal Care and
Use Committee at WuXi AppTec.
2.2 Reagents
All the test drugs were supplied by Huizhou Salubris Co. Ltd (Storage
condition: 25°C nitrogen cabinet free of light): S086 (Lot: ZXM18002,
Purity: 95.51%), EXP3174 (Lot: SXMA18002, Purity: 91.47%), sacubitril
calcium salt (Lot: SXMA18001, Purity: 95.36%), and LCZ696 (Lot:
DYF17006, Purity: 94.37%).
The study used 0.5% CMC-Na as the vehicle for all the compounds. This
vehicle was prepared by weighing and dissolving CMC-Na in deionized
water at a concentration of 0.5 g CMC-Na/100 ml water. All the compounds
were dissolved in this 0.5% CMC-Na solution at a concentration
calculated based on their anhydrous-free acid weight. The theoretical
weight of each compound was multiplied by a corresponding conversion
factor (S086: 1.12; sacubitril calcium salt: 1.10; EXP3174: 1.09; and
LCZ696: 1.14) to determine the actual weight to be used in the solution.
The prepared solutions were stored at 2-8°C and were used within
specific timeframes. S086 and EXP3174 solutions were prepared every 3
days, while sacubitril calcium salt and LCZ696 solutions were prepared
daily. Prior to dosing, the solutions were allowed to reach room
temperature for 10-15 minutes. The rats were each given a dosage of 5
ml/kg of body weight, which was well-mixed with their respective
solutions.
Pentobarbital,
supplier: Alfasan International B.V.; lot: 1709296-02; property: white
powder; storage: light-free at room temperature. Na+test kit, supplier: MedicalSystem; lot: 180908101, 180921101; storage:
2-8℃. 8% salt forage (blue
particles), supplier: Jiangsu Medicience; storage: 2-8℃. 0.3% salt
forage (yellow particles), supplier: Jiangsu Medicience; storage: 2-8℃.
Meloxicam injection, supplier: Qilu Animal Health Products CO., LTD.
lot: 1805001. Gentamicin, supplier: Yichang Humanwell Pharma CO., LTD.
lot: 20180139.