Stimulation of lipoprotein lipase activity of rat adipose tissue and post-heparin plasma by N6-(phenylisopropyl)adenosine

Abstract

Adenosine has profound effects on adipose tissue metabolism. It is an inhibitor of 13-adrenergic stimulation of cyclic AMP accumulation [1,2] and lipolysis [2-101 in tire isolated rat fat cell. It also opposes the effects of adrenaline on phosphate uptake [ 11 ]. In vivo, tile nucleoside has been reported to lower plasma free fatty acid and glycerol concentrations [8]. The effects of adenosine are thought to be mediated by two membrane receptors, one requiring an intact ribose (R-site) and tile other one an intact purine moiety [ 12-14] (P-site). Adenosine is rapidly metabolized but its effects on the R-site [12-14] are shared by purine-substituted analogs such as N6-(phenylisopro pyl)adenosine that are not deaminated by the enzyme adenosine deaminase (EC 3.5.4.4) [15]. Lipoprotein lipase (EC 3.1.1.3.4) is the enzyme responsible for the hydrolysis and uptake of triglycerides from circulating chylomicrons and very low density lipoproteins [16-19] . The enzyme is thought to be located on the endothelial surface of the capillary bed of extrahepatic tissues and it can be released into the circulation by intravenous injection of heparin [ 16-20] . Another heparin-releasable ectoenzyme is probably located on the outer surface of the endothelial cells of the liver and it is therefore called hepatic lipase [20]. This enzyme has been suggested to hydrolyze high density lipoprotein phospholipids and triglycerides [21 ]. This work was undertaken to find out ifN6-(phenyl isopropyl)adenosine has effects on lipoprotein lipase activity and, consequently, on the uptake of circulating triglycerides into tissues. Male Lewis albino rats ( 150 -250 g body wt) were used. They were fed ad libitum with a standard chow until injected intraperitoneally with 50-300/al isotonic NaC1 with/without N6-(phenylisopropyl)adenosine, at which time food was withheld. At the times indicated, the animals were anaesthetized with ether and blood was drawn by cardiac puncture into heparinized syringes. For post-heparin plasma lipase assays, blood was drawn 2 min after intravenous injection ofheparin (2000 units/kg body wt). The blood was centrifuged and the resulting plasma was stored at 2 0 ° C until assayed. Free glycerol was estimated enzymatically by using the commercial kit no. 125 032 of BoehringerMannheim GmbH. Tile assays for porst-heparin plasma lipases and the antibody to rat hepatic lipase have been described in [22]. The method used to measure heparin-releasable lipoprotein lipase activity of adipose tissue will be presented in detail in the text. N6-(phenylisopropyl)Adenosine was a gift from Dr Harald Stork of Boehringer-Mannheim GmbH. Glyceroltri-[14C] oleate (49 mCi/mmol) was from the Radiochemical Centre (Amersham). Heparin was from Medica (ttelsinki). All other reagents were from Sigma (St Louis MO).