Rompum 2

After AP induction, 20 rats in each group remained under observation for a maximum of 7 days, with access to standard oral diet (AIN-93M) and water ad libitum. The animals were observed individually every 8 h for death registration. Animals that survived until 7 days after AP were sacrificed with an intraperitoneal injection of 80 mg/kg ketamine hydrochloride (Ketamin-S(+)®; Cristália) and 8.0 mg/kg xylazine hydrochloride (Rompum® 2%; Bayer).

All mice were anaesthetized 30 min after surfacing (after grip tests) first with halothane (5% in oxygen, Halothane, Belamont, France) in order to gain time and to minimize stress, and then by intraperitoneal injection of a mixture of 16 mg/kg xylazine (Rompum® 2%, Bayer Pharma) and 100 mg/kg ketamine (Imalgène® 1000, Laboratoire Rhône). Mice in the spad-pool were kept for heart blood sampling for biochemistry and then sacrificed by injecting pentobarbital (200 mg/kg ip, Sanofi Santé, France).

Male C57BL/6 mice were obtained from Janvier (Le Genest Saint Isle, France). All experiments were performed in accordance with protocols approved by the Institutional Review Boards. (Comité d’Ethique NOrmandie en Matière d’EXpérimentation Animale ENOMEXA n°54).
APS was induced in mice as described previously [7 ] by a single i.v. injection of a 300μL (4.2mg) aPL. Control groups received either saline (sham) or 300μL of serum without antiphospholipid activity obtained from healthy donors (NSIg). These injections were given through an insulin syringe (29 G, Myjector U-100 INSULIN). The mice were treated by daily tube-fed with HCQ at 12μg/g for 7 days.
On day 7, mice were anesthetized with 3.6mg/kg xylazine (Rompum 2%, Bayer, France) and 90mg/kg ketamine (Imalgène 1000, Merial, France) intraperitoneally and underwent laparotomy.

(−)-EPI (+)-bitartrate salt, L-(-)-NE (+)-bitartrate salt monohydrate, and ICI 118,551 were purchased from Sigma-Aldrich (St. Louis, MO, USA). Ketamine (Imalgene 1000, Merial, Lisboa, Portugal) and xilazine (Rompum 2%, Bayer, Lisboa, Portugal).

The rats were then anesthetized by an intraperitoneal injection of a mixture of 10 mg.kg⁻¹ xylazine (Rompum^® 2%, Bayer Pharma, Germany) and 100 mg.kg⁻¹ ketamine (Imalgene^®1000, Rhône laboratory, France).
A sample of arterial blood was taken from each rat by direct aortic puncture and blood gases were measured using an i-STAT System (Abbott Laboratories, Illinois, USA).
The rats were sacrificed by injecting pentobarbital (200 mg.kg⁻¹ ip, Sanofi Santé, France).

(±)-Sotalol hydrochloride, (-)-adrenaline, L-(-)-noradrenaline, dopamine hydrochloride, 2,3-dihydroxybenzoic acid, and perchloric acid were purchased from Sigma-Aldrich (St. Louis, United States). Ketamine (Imalgene 1,000, Merial, Lisboa, Portugal) and xylazine (Rompum 2%, Bayer, Lisboa, Portugal) were purchased from Agrofauna (Vila Nova de Gaia, Portugal).