Expression of transcripts for beta 1-, beta 2-, and beta 3-adrenergic receptors (beta 1-AR, beta 2-AR, and beta 3-AR, respectively) was investigated in porcine tissues. Northern blot analysis of total RNA indicated hybridization of the rat beta 1-AR probe to transcripts of approximately 3.0-kb in heart and lung and 2.9-kb in perirenal, omental, and subcutaneous adipose tissue. Hybridization of the hamster beta 2-AR probe with total RNA revealed transcripts of approximately 1.8-kb in the heart, 2.0-kb in the lung, and 2.0-kb in perirenal, omental, and subcutaneous adipose tissue; a minor transcript of 1.8-kb was also observed in the perirenal tissue. Northern blot analysis of poly (A+) RNA from porcine adipose tissue with the mouse beta 3-AR probe revealed no detectable signals in heart and lung but detected a major 2.2-kb transcript and a minor 1.9-kb transcript in subcutaneous adipose tissue. The expression of porcine beta 1-AR and beta 2-AR transcripts was compared in perirenal, omental, and subcutaneous adipose depots. The expression of beta 1-AR transcripts was four to five times greater in the perirenal depot than in the omental depot and two to three times greater in the subcutaneous depot than in the omental depot. The expression of beta 2-AR mRNA seemed to be the same in all three depots.

Young postweaning pigs were fed a high fat diet containing beef tallow (saturated fat) or corn oil (unsaturated fat). Adipose tissue was used to measure adipocyte size and number of cells per gram of tissue, ligand binding by beta-adrenergic receptors and lipolytic and palmitate esterification rates. Pigs fed the saturated fat diet had more saturated and monounsaturated fatty acids and less polyunsaturated fatty acid in the crude membrane fraction. Adipocytes were larger in pigs fed the saturated fat diet. There was no difference in the binding affinities of the receptors; more binding sites were expressed on a protein or cell basis and fewer sites were expressed per unit surface area in adipocyte ghosts isolated from pigs fed the saturated fat diet. Fatty acid esterification was greater in pigs fed saturated fat diets. Isoproterenol inhibition was marginal in both groups but tended to be greater in pigs fed saturated fat diets. The beta-adrenergic receptor-mediated lipolytic rates were not different; only the theophylline-stimulated rates tended to be greater in the saturated fat-fed group. Thus, a large increase in saturated fatty acid concentration of porcine adipose tissue membranes caused an increase in beta-adrenergic receptor number without any change in receptor affinity. These receptor changes were at best only marginally reflected in beta-adrenergic agonist-mediated functions.

Methods have been detailed to prepare a crude membrane fraction from isolated porcine adipose tissue cells. Adipocytes were obtained after incubation of 5 g of adipose tissue slices with 4,500 units of a selected lot of collagenase in a total volume of 15 mL at 37 degrees C for 90 min. There was no bovine serum albumin present during cell isolation because albumin did not enhance cell yield or yield of lipolytic activity. Isolated cells were lysed by exposure to hypotonic conditions in the presence of 7.5 mM ethylene glycol tetraacetic acid (EGTA) and .8 mM phenylmethylsulfonyl fluoride (PMSF). A 30,000 x g centrifugal pellet was used as the crude membrane preparation. Binding of tritiated dihydroalprenolol (DHA), a beta-adrenergic antagonist, was measured in the presence of 7.5 mM EGTA and .2 mM PMSF, because these protease inhibitors improved specific binding by approximately 50% to greater than 150 fmol/mg of protein and decreased non-specific binding to less than 10% at 2.5 nM DHA.