To determine the response to alteration in site and form of carbohydrate delivery to the digestive tract, in vitro rates of lipogenesis and lipolysis in mesenteric (MESA), omental (OMA), and subcutaneous (SQA) adipose depots were compared. Forty crossbred beef steers (243 +/- 2 kg of BW) were fed 161 (LI) or 214 (HI) kcal of ME/(kg of BW(0.75) x d) or they were fed LI and infused for 35 d into the rumen (R) or abomasum (A) with starch hydrolysate (SH) or into the abomasum with glucose (G). Jugular blood samples were collected, steers were slaughtered, and adipose depots were sampled and prepared for assessment of lipogenesis and lipolysis in vitro. Blood concentrations of glucagon were increased (P = 0.04) in HI-H2O compared with LI-H2O steers, whereas A-SH tended to increase (P = 0.08) circulating IGF-I relative to R-SH, and A-G tended to have elevated (P = 0.09) T3 compared with A-SH. Lipolysis, as assessed by NEFA release, was unaffected by treatment. Glycerol release by the MESA and SQA was increased or tended to be increased (P < or = 0.08) in HI-H2O compared with LI-H2O steers. In A-G compared with A-SH steers, glycerol release from OMA increased (P = 0.008) and from SQA tended to be increased (P = 0.08). Acetate incorporation into total neutral lipids (TNL) increased or tended to increase with ME intake and SH infusion (P < or = 0.09) across all depots. Rates of acetate incorporation into fatty acids (FA) also increased or tended to be increased (P < or = 0.1) by SH infusion across all depots, but only that of SQA was increased with ME intake (HI-H2O vs. LI-H2O; P = 0.02). Rates of acetate incorporation into FA and TNL in MESA were increased (P < or = 0.03) by A-SH compared with R-SH, but site of SH infusion did not affect the rates in SQA or OMA. Glucose incorporation into TNL for MESA and SQA increased or tended to be increased (P < or = 0.1) by dietary and infused energy, whereas for OMA they tended to be increased (P = 0.1) only by SH infusion. In contrast, glucose incorporation into FA was unaffected by energy supply but tended to be increased (P = 0.07) by SH in MESA and tended to be greater (P = 0.08) for A-G than A-SH in OMA. The general across-depot pattern of acetate incorporation rate into FA and TNL was SQA > OMA > MESA, whereas, for glucose incorporation, rates across depots were equivalent. These data provide evidence that the postruminal supply of energy, specifically carbohydrate, stimulates lipogenesis from acetate and glucose and is more pronounced in abdominal depots relative to the subcutaneous depot.