Osmotic pump

All surgeries were performed according to the National Institutes of Health guidelines for the humane treatment of laboratory animals according to the “Guide for the Care and Use of Laboratory Animals” (NIH publication 86–23) and with approval of the Institutional Animal Care and Use Committee of Vanderbilt University Medical School. Prior to hepatectomy or for sham operations, mice were anesthetized with 60 mg/kg ketamine (Hospira) and 7 mg/kg xylazine (Phoenix Pharmaceutics) and positioned supine. A transverse incision was made inferior to the xiphoid process, which was excised. The median and left lateral lobes were eviscerated and ligated resulting in 60% liver removal. About 5 mg/kg/day of recombinant human IGF-1 (Cell Sciences) dissolved in water was given continuously with an osmotic pump (DURECT). To quantitate dividing hepatocytes, mice received 1 mg intraperitoneal injection of 5-bromo-2-deoxyuridine (BrdU; BD Pharmingen) 2 h before sacrifice. At tissue recovery, mice were anesthetized and weighed. Livers were excised, rinsed, blotted, and weighed. Sections were fixed in 10% neutral buffered formalin. Mortality after hepatectomy was < 5% and not associated with a particular genotype.

Chronically WHV-infected woodchucks were treated for 28 weeks with the nucleoside analogue entecavir (ETV, Bristol-Myers Squibb, New York, NY). Initially, the drug was administered for 12 weeks by using osmotic pumps (DURECT, Cupertino, CA) which were implanted surgically under the skin of the animals. The pump releases subcutaneously approximately 0.2 mg of ETV per day. Pumps were exchanged every 4 weeks and overall 3 pumps were implanted for each woodchuck. From week 10 to 28, subcutaneous injections of 1.5 mg of ETV were performed weekly.

To assess the long-term central effects of the IL18 system on food intake, body composition and iWAT morphology, 10–11 weeks old Il18r1-KO and WT mice (4–7 mice/genotype/treatment) were infused ICV with saline or recombinant IL18 (20 μg/mouse/7 days, Sino Biological, Inc., Beijing, China)56 (link). For this purpose, brain infusion canulae were placed stereotaxically into the lateral ventricle as previously described56 (link). A catheter tube was connected from the brain infusion cannula to the osmotic minipump (model 1007D, total volume 100 μL, flow rate 0.5 μL/h; Alzet Osmotic Pumps, Durect, CA, USA) flow moderator.

Chronic nicotine exposure was carried out using subcutaneously implanted osmotic pumps (DURECT Corporation, Cupertino, CA, USA), infusing nicotine bis-L-(+)-tartrate dehydrate (4.5 mg/kg b.w./day) over an average period of 14.8 ± 0.2 days (considered as a period of chronic exposure). The dose and regimen of nicotine exposure were selected to maintain the levels of nicotine and cotinine as observed in chronic smokers [53 (link),54 (link),55 (link)].