We conducted two experiments to evaluate the flooding dose method for measuring intestinal and liver protein synthesis in sheep. Experiment 1 showed that large doses of phenylalanine did not cause marked metabolic disturbances. Experiment 2 examined the effectiveness of flooding with phenylalanine and the time dependency of the protein synthesis calculation. Rams were injected with 1.2 MBq L-[ring 2,6-3H]phenylalanine/kg body wt and slaughtered 20, 40 or 60 min later. Plasma specific radioactivity reached a plateau within 2.5 min and did not change significantly (P > 0.05) throughout the experiment. Tissue intracellular free pool specific radioactivity also remained constant from 20 to 60 min postinfusion. Flooding conditions were achieved in the intracellular free pool of intestinal tissues (specific radioactivity 70-96% of plasma specific radioactivity), although liver flooding was less successful (57-67%). Protein synthesis rates measured after 20 min were significantly (P < 0.05) higher in the liver, jejunum and ileum than those measured at 60 min. Protein synthesis rates also tended to decline with time in the duodenum and colon (P > 0.05). There were no significant differences (P > 0.05) between protein synthesis rates calculated using the intracellular specific radioactivity vs. plasma specific radioactivity in the duodenum, ileum or colon. Therefore, this method represents an improvement over continuous infusion methods for measurements of protein synthesis in visceral tissues.

The effect of corticosterone treatment on the sensitivity of muscle protein synthesis to insulin infusion was assessed in postabsorptive young rats. To select the optimal time period for corticosterone treatment, protein synthesis was measured by injection of L-[2,6-3H]phenylalanine (1.5 mmol/kg body weight) 1, 4, 12 or 24 h after injection of corticosterone (5 mg/kg body wt.). Muscle protein synthesis was significantly decreased at 4 h and the effect was maximal by 12 h; liver protein synthesis was elevated at 12 h and 24 h. The dose-response of muscle protein synthesis to a 30 min infusion with 0-150 munits of insulin/h was then compared in rats pretreated with corticosterone (10 mg/100 g body wt.) or vehicle alone. When no insulin was infused, corticosterone inhibited protein synthesis in gastrocnemius muscle. High doses of insulin stimulated protein synthesis, but the inhibition by corticosterone was similar to that in the absence of insulin. At intermediate doses of insulin there was an increased requirement for insulin to elicit an equivalent response in muscle protein synthesis. Plantaris muscle responded in a manner similar to that of gastrocnemius, but neither soleus muscle nor liver responded significantly to insulin. These data suggest that corticosterone has two modes of action; one which is independent from and opposite to that of insulin, and a second which causes insulin-resistance through a decrease in sensitivity rather than a change in responsiveness.

The early time course (1, 3, 9, 24 h) of changes in rates of protein synthesis (ks) in liver and three different muscles (gastrocnemius, soleus and heart) was investigated after injection of saline, interleukin-1 beta (IL-1) or turpentine in rats. IL-1 injection induced a consistent increase in body temperature of about 3 degrees C between 3 and 5 h, but thereafter a hypothermic response occurred. With turpentine, a delayed fever response with a peak value by 9 h was observed. Both IL-1 and turpentine had no effect on protein synthesis in the small intestine, but produced a significant increase in ks in the liver at 9 h. By 24 h in IL-1-treated animals, liver ks had returned back to control values, whereas the turpentine-treated group showed a progressive rise in ks. Gastrocnemius and soleus muscles exhibited a significant fall in ks at 9 h after IL-1 and turpentine injection compared with the control. In contrast, the ks of heart muscle increased at 3-9 h after IL-1 injection, but there was no effect of turpentine. Thus for the first time a marked decrease of protein synthesis in skeletal muscle in response to IL-1 could be demonstrated.

of in vitro and in vivo energy metabolism of the gastrointestinal tract of fed or fasted sheep. Can

Insulin at a concentration close to the physiological range (100 mu-units/ml) stimulated protein synthesis in L6 myoblasts by 17%. Pre-treatment with the phospholipase A2 inhibitors mepacrine or dexamethasone prevented this stimulation and decreased the release of prostaglandin F2 alpha, implicating the action of phospholipase A2 and the subsequent metabolism of arachidonic acid to prostaglandins in the stimulation of protein synthesis by physiological doses of insulin. Higher concentrations of insulin (500-1000 mu-units/ml) stimulated protein synthesis in the presence of mepacrine or dexamethasone, suggesting that an alternative pathway may become important in insulin action when phospholipase A2 is inhibited.