Lowering plasma free fatty acids with Acipimox mimics the antidiabetic effects of the beta 3-adrenergic agonist CL-316243 in obese Zucker diabetic fatty rats.

We previously reported that long-term treatment of Zucker diabetic fatty (ZDF) rats with the selective beta(3) agonist CL-316243 normalizes glycemia, decreases plasma free fatty acids (FFA) concentration, improves insulin responsiveness, and increases glucose uptake, not only in brown and white adipose tissues, but also in skeletal muscles. Because muscles do not express typical beta(3) adrenoceptors, we postulated that the muscle effect was indirect and that it was possibly mediated by an activation of the glucose-fatty acid cycle. To test this hypothesis, we investigated the effects of Acipimox, a potent inhibitor of lipolysis in adipose tissue. Similar to CL-316243, Acipimox (150 mg/kg orally) markedly decreased plasma FFA, glucose, and insulin concentrations and improved glucose tolerance while reducing the insulin response in obese (350 to 400 g) ZDF rats. Plasma FFA concentrations were significantly correlated with plasma glucose and insulin concentrations (r =.72 and.83, respectively; P <.01), indicating strong metabolic relationships between these parameters. Euglycemic-hyperinsulinemic clamps combined with the 2-[(3)H]deoxyglucose method revealed that Acipimox markedly improved insulin responsiveness and significantly increased glucose uptake (Rg') in the diaphragm, the heart, and various skeletal muscles. Unlike CL-316243, Acipimox did not increase glucose use in brown or white adipose tissues. This selectivity shows that it is possible to improve diabetes in obese ZDF rats without necessarily stimulating thermogenesis in adipose tissues. Thus, decreasing plasma FFA with 2 drugs (Acipimox or CL-316243) that act via different mechanisms (acute inhibition of lipolysis or chronic stimulation of FFA oxidation) is associated with increased glucose uptake in muscles and enhanced insulin responsiveness. These observations support the hypothesis that CL-316243 may indirectly stimulate glucose uptake in muscles of type II diabetic rats by first stimulating brown adipose tissue (increasing uncoupling protein content and fatty acid oxidation) and progressively decreasing the levels of circulating FFA, resulting in activation of the glucose-fatty acid cycle or other mechanisms regulating insulin responsiveness in skeletal muscles.

Mechanisms of the antidiabetic effects of the beta 3-adrenergic agonist CL-316243 in obese Zucker-ZDF rats.

Previous studies have demonstrated that chronic cold exposure activates the sympathetic nervous system, increases energy expenditure, improves glucose uptake in peripheral tissues [brown and white adipose tissues (BAT and WAT) and muscles] of normal rats. The goal of the present studies was to test whether the selective beta 3-adrenergic agonist CL-316243 (CL) would mimic the beneficial beneficial effects of cold exposure in lean and obese ZDF/Gmi-fa male (ZDF) rats, a new model of type II diabetes. In obese ZDF rats, chronic infusion of CL (1 mg.kg-1.day-1 for 14 days) significantly decreased body weight gain, food intake, and WAT weight. It also increased total tissue cytochrome oxidase activity, not only in BAT (15 times), but also in WAT (2-4) times, suggesting that it progressively enhanced mitochondriogenesis in adipose tissues. CL treatment normalized hyperglycemia and reduced hyperinsulinemia and circulating free fatty acid (FFA) levels. It also improved glucose tolerance and reduced insulin response during an intravenous glucose tolerance test. In general, the beneficial effects of CL were more pronounced in obese than in lean rats. Hyperinsulinemic-euglycemic glucose clamps combined with the [2-3H]deoxyglucose method revealed that CL markedly improved insulin responsiveness in obese rats (3-4 times) and increased glucose uptake in BAT (21 times), WAT (3 times), skeletal muscles (2-3 times), and in the diaphragm (2.8 times), but not in the heart. It is concluded that chronic CL treatment improves glucose tolerance and insulin responsiveness in obese ZDF rats by a mechanism similar to that induced by chronic cold exposure, i.e., by stimulating facultative thermaogenesis, mitochondriogenesis, and glucose utilization in BAT and WAT. In addition to this mechanism, the reduction in plasma FFA levels induced by chronic CL treatment may further contribute to enhance glucose uptake in skeletal muscles (a tissue that does not express typical beta 3-adrenoceptors) via the "glucose-fatty acid" cycle. The antiobesity and antidiabetic properties of CL suggest that selective beta 3-adrenergic agonists may represent useful agents for the treatment of type II diabetes.

Chronic norepinephrine infusion stimulates glucose uptake in white and brown adipose tissues.

Cold exposure activates the sympathetic nervous system and markedly stimulates glucose uptake in rat peripheral tissues [A. L. Vallerand, F. Pérusse, and L. J. Bukowiecki. Am. J. Physiol 259 (Regulatory Integrative Comp. Physiol. 28): R1043-R1049, 1990]. To test whether norepinephrine (NE) mimics the effects of cold exposure, we estimated the effects of chronic NE treatment on tissue glucose uptake by determining the glucose metabolic index using the 2-[1,2-3H(N)]deoxy-D-glucose method. NE was administered in conscious rats at various doses (ranging from 1.9 to 25.1 nmol.kg-1.min-1) during 4 days via minipumps implanted subcutaneously. At doses > 10 nmol.kg-1.min-1, NE maximally stimulated glucose uptake in interscapular brown adipose tissue (approximately 50 times above controls) and epididymal white adipose tissue (approximately 3 times above controls). NE infusion (18.8 nmol.kg-1.min-1) increased the circulating levels of NE from 1.1 +/- 0.1 to 19.2 +/- 0.4 nM (P < 0.001), which is in the range of concentrations for the stimulatory effects of NE on glucose uptake in isolated brown adipocytes. At all concentrations tested, NE infusion did not stimulate glucose uptake in the heart and skeletal muscles. NE treatment did not significantly alter plasma insulin or glucose levels but increased the concentration of circulating free fatty acids. The capacity of brown adipose tissue for NE stimulation of glucose uptake (expressed per g of tissue) was much higher than that of white adipose tissue (100 times), various types of white or red skeletal muscles (10-80 times), or the heart (3-4 times).(ABSTRACT TRUNCATED AT 250 WORDS)

Stimulatory effects of cold exposure and cold acclimation on glucose uptake in rat peripheral tissues.

The effects of cold exposure on the net rates of 2-[3H]deoxy-D-glucose uptake (Ki) in rat peripheral tissues were investigated comparatively in warm- and cold-acclimated animals to determine whether cold acclimation induces regulatory alterations in glucose metabolism. Acute exposure of warm-acclimated (25 degrees C) rats to cold (48 h at 5 degrees C) markedly increased the Ki values in red and white skeletal muscles (2-5 times), in the heart (8 times), in several white adipose tissue (WAT) depots (4-20 times), and in brown adipose tissue (BAT) (110 times). After cold acclimation (3 wk at 5 degrees C), the Ki values further increased in the heart (15 times) and WAT (up to 29 times) but decreased in BAT (36 times). Remarkably, glucose uptake was still increased in muscles of cold-exposed/cold-acclimated animals (that do not shiver), demonstrating that enhanced glucose uptake may occur in muscles in the absence of shivering thermogenesis (or contractile activity). When cold-acclimated rats were returned to the warm for 18 h, the Ki values of all tissues, except WAT, returned to control levels. Cold exposure synergistically potentiated the stimulation of tissue glucose uptake induced by a maximal effective dose of insulin (0.5 U/kg iv) in warm- as well as in cold-acclimated animals.(ABSTRACT TRUNCATED AT 250 WORDS)

Cold exposure reverses inhibitory effects of fasting on peripheral glucose uptake in rats.

The effects of fasting and cold exposure on glucose uptake in skeletal muscles (tibialis anterior, quadriceps, and soleus), heart, and brown adipose tissue (BAT) were studied in conscious rats. Glucose uptake was estimated by determining the glucose metabolic index of individual tissues using the 2-[3H]deoxyglucose method. Fasting for 18 h at 25 degrees C decreased plasma glucose levels (-40%) and glucose uptake in heart (-95%) and skeletal muscles (-64-90%) but did not significantly affect glucose uptake in BAT. Fasting for 48 h did not further decrease these parameters. On the other hand, cold exposure (48 h at 5 degrees C) of fed animals did not alter plasma glucose levels but increased glucose uptake in heart (73%), skeletal muscles (126-326%), and particularly in BAT (95-fold). Remarkably, cold exposure stimulated glucose uptake in BAT and skeletal muscles of 18-h fasted rats by the same order of magnitude as in fed animals (percentagewise), thereby indicating that glucose represents an essential metabolite for shivering (muscles) and nonshivering (BAT) thermogeneses. In the heart of starved animals, the cold-induced increase in glucose uptake was even more important (8-fold) than in fed animals. Considering that cold exposure of fasted rats results in a severe insulinopenia, it is suggested that cold exposure stimulates glucose uptake in peripheral tissues primarily by enhancing glucose oxidation via insulin-independent pathways.

Cold exposure potentiates the effect of insulin on in vivo glucose uptake.

The effects of cold exposure (48 h at 4 degrees C) and insulin injection (0.5 U/kg iv) on the rates of net 2-[3H]deoxyglucose uptake (Ki) in peripheral tissues were investigated in warm-acclimated rats (25 degrees C). Cold exposure and insulin treatment independently increased Ki values in skeletal muscles (soleus, extensor digitorum longus, and vastus lateralis), heart, white adipose tissue (subcutaneous, gonadal, and retroperitoneal), and brown adipose tissue (P less than 0.01). The effects of cold exposure were particularly evident in brown adipose tissue where the Ki increased greater than 100 times. When the two treatments were combined (insulin injection in cold-exposed rats), it was found that cold exposure synergistically enhanced the maximal insulin responses for glucose uptake in brown adipose tissue, all white adipose tissue depots, and skeletal muscles investigated. The results indicate that cold exposure induces an "insulin-like" effect on Ki that does not appear to be specifically associated with shivering thermogenesis in skeletal muscles, because that effect was observed in all insulin-sensitive tissues. The data also demonstrate that cold exposure significantly potentiates the maximal insulin responses for glucose uptake in the same tissues. This potentialization may result from an enhanced responsiveness of peripheral tissues to insulin, possibly occurring at metabolic steps lying beyond the insulin receptor and an increased tissue blood flow augmenting glucose and insulin availability and thereby amplifying glucose uptake.

The role of insulin in nonshivering thermogenesis.

The regulation of nonshivering thermogenesis by insulin was studied in cold-acclimated rats (2 weeks at 5 degrees C) made diabetic after injection with streptozotocin (75 mg/kg, i.p.) and maintained in the warm (25 degrees C) for 2-7 days. To investigate whether thermogenesis was activated in brown adipose tissue (BAT) of diabetic rats under physiological conditions, conscious rats were briefly exposed to cold (2 h at 5 degrees C) and the temperature of interscapular BAT (Tbat) was compared with the colonic temperature (Tcol). It was found that Tbat, Tcol, and Tbat-Tcol (an index of thermogenesis activation in BAT) were significantly reduced in 7-day diabetic rats (P less than 0.01) but not in 2-day diabetic animals, suggesting that diabetes progressively decreases BAT thermogenic capacity. To further assess whether the maximal capacity of BAT for nonshivering thermogenesis was affected by the lack of insulin, the calorigenic response to noradrenaline (0.4 mg/kg, i.m.) was determined at 25 degrees C in anesthetized animals using an open circuit respirometer. The results showed that the calorigenic response to noradrenaline was inhibited by 50 and 70% in 2- and 7-day diabetic rats, respectively. Significantly, tissue weight, protein content, and cytochrome oxidase activity of interscapular BAT were also decreased by 30-40% and 50-70% in the same animals. Insulin treatment of 3-day diabetic rats for 4 days (6.5 U/day per rat) restored their calorigenic response to noradrenaline to control levels.(ABSTRACT TRUNCATED AT 250 WORDS)

An improved glyoxylic acid technique for the histochemical localization of catecholamines in brown adipose tissue.

A glyoxylic acid method using cryostat sections to demonstrate catecholaminergic fibres of the central nervous system was modified to show the extent of the adrenergic innervation in rat brown adipose tissue. It revealed prominent interlacing fluorescent parenchymal fibres surrounding individual adipocytes. The density of this network of fine fibres was not evident using earlier techniques. The new method also confirmed the dense networks of adrenergic fibres associated with arterial vessels. Its specificity was verified by simultaneously performing radioenzymatic determinations of tissue catecholamine levels and histochemical studies of brown adipose tissue from normal and sympathectomized rats. Chemical sympathectomy with 6-hydroxydopamine resulted in a pronounced decrease in brown adipose tissue and heart catecholamine (noradrenalin and dopamine) levels. Significantly, in brown adipose tissue of sympathectomized animals no fluorescence could be detected in terminal nerves of either the parenchyma or those of vascular smooth muscles. Nevertheless, some intense fluorescence was seen in axon bundles. The findings suggest that catecholamines of the parenchymal innervation form a larger proportion of the total catecholamine content of brown adipose tissue than was previously believed, provide stronger support for direct control of the function of multilocular adipocytes, and also confirm unpublished data reporting considerable dopamine content in brown adipose tissue.

Temporal variations in the elimination of antipyrine in normal and adrenalectomized rats.

An oral dose of antipyrine was administered at 08:00 and 20:00 hours to groups of sham operated and adrenalectomized rats standardized in an alternating 12-hour-light-dark cycle for 7 days prior the experiments. Temporal variations in the rate of elimination of antipyrine were obtained in both groups. Adrenalectomy decreased the serum elimination of antipyrine but had no effect on the temporal variations.