Adaptation to a high protein, carbohydrate-free diet induces a marked reduction of fatty acid synthesis and lipogenic enzymes in rat adipose tissue that is rapidly reverted by a balanced diet.

We have previously shown that in vivo lipogenesis is markedly reduced in liver, carcass, and in 4 different depots of adipose tissue of rats adapted to a high protein, carbohydrate-free (HP) diet. In the present work, we investigate the activity of enzymes involved in lipogenesis in the epididymal adipose tissue (EPI) of rats adapted to an HP diet before and 12 h after a balanced diet was introduced. Rats fed an HP diet for 15 days showed a 60% reduction of EPI fatty acid synthesis in vivo that was accompanied by 45%-55% decreases in the activities of pyruvate dehydrogenase complex, ATP-citrate lyase, acetyl-CoA carboxylase, glucose-6-phosphate dehydrogenase, and malic enzyme. Reversion to a balanced diet for 12 h resulted in a normalization of in vivo EPI lipogenesis, and in a restoration of acetyl-CoA carboxylase activity to levels that did not differ significantly from control values. The activities of ATP-citrate lyase and pyruvate dehydrogenase complex increased to about 75%-86% of control values, but the activities of glucose-6-phosphate dehydrogenase and malic enzyme remained unchanged 12 h after diet reversion. The data indicate that in rats, the adjustment of adipose tissue lipogenic activity is an important component of the metabolic adaptation to different nutritional conditions.

Dietary sodium restriction exacerbates age-related changes in rat adipose tissue and liver lipogenesis.

To investigate the effects of prolonged dietary sodium restriction on lipid metabolism, male rats weighing 35 to 40 g (just weaned) were fed either a low-salt (LSD) or a normal salt diet (NSD) and used in metabolic experiments after 1, 2, or 3 months of diet consumption. After 2 and 3 months on the diet, LSD rats showed increased amounts of lipid in carcass and retroperitoneal tissue. In both LSD and NSD, extending the feeding period from 2 to 3 months resulted in a marked reduction in the in vivo rates of adipose tissue fatty acid synthesis that was accompanied by increases in liver lipogenesis and in the activity of adipose tissue lipoprotein lipase (LPL). However, these increases were more marked in LSD rats. Thus, in vivo rates of liver fatty synthesis and LPL activity in LSD rats, which were already higher (by about 35% and 20%, respectively) than in controls after 2 months, attained levels 50% higher than those in NSD animals after another month on the diet. Brown adipose tissue (BAT) thermogenic capacity, estimated after 2 and 3 months by the tissue temperature response to norepinephrine (NE) injection and by guanosine diphosphate (GDP) binding to BAT mitochondria, did not change in controls, but was significantly reduced in LSD rats. This raises the possibility that a decrease in overall energy expenditure, together with an LPL-induced increased uptake of preformed fatty acids from the circulation, may account for the excessive lipid accumulation in LSD rats. Taken together, the data indicate that prolonged dietary sodium restriction exacerbates normal, age-related changes in white and BAT metabolism.

Control of glyceroneogenic activity in rat brown adipose tissue.

Brown adipose tissue (BAT) glyceroneogenesis was evaluated in rats either fasted for 48 h or with streptozotocin-diabetes induced 3 days previously or adapted for 20 days to a high-protein, carbohydrate-free (HP) diet, conditions in which BAT glucose utilization is reduced. The three treatments induced an increase in BAT glyceroneogenic activity, evidenced by increased rates of incorporation of [1-14C]pyruvate into triacylglycerol (TAG)-glycerol in vitro and a marked, threefold increase in the activity of BAT phosphoenolpyruvate carboxykinase (PEPCK). BAT glycerokinase activity was not significantly affected by fasting or diabetes. After unilateral BAT denervation of rats fed either the HP or a balanced diet, glyceroneogenesis activity increased in denervated pads, evidenced by increased rates of nonglucose carbon incorporation into TAG-glycerol in vivo (difference between 3H2O and [14C]glucose incorporations) and of [1-14C]pyruvate in vitro. PEPCK activity was not significantly affected by denervation. The data suggest that BAT glyceroneogenesis is not under sympathetic control but is sensitive to hormonal/metabolic factors. In situations of reduced glucose use there is an increase in BAT glyceroneogenesis that may compensate the decreased generation of glycerol-3-phosphate from the hexose.

Expression of glycerokinase in brown adipose tissue is stimulated by the sympathetic nervous system.

The effect of cold exposure (4 degrees C) or prolonged norepinephrine infusion on the activity and mRNA levels of glycerokinase (GyK) was investigated in rat interscapular brown adipose tissue (BAT). Cold exposure for 12 and 24 h induced increases of 30% and 100%, respectively, in the activity of BAT GyK, which was paralleled by twofold and fourfold increase in enzyme mRNA levels. BAT hemidenervation resulted in reductions of 50% and 30% in GyK activity and in mRNA levels, respectively, in denervated pads from rats kept at 25 degrees C, and suppressed in these pads the cold-induced increases in both GyK activity and mRNA levels. The increase in GyK activity induced by cold exposure was not affected by phenoxybenzamine, but was markedly inhibited by previous administration of propranolol or actinomycin D. BAT GyK activity did not change significantly after 6 h of continuous subcutaneous infusion of norepinephrine (20 microg/h), but increased twofold and fourfold after 12 and 24 h, with no further increase after 72 h of infusion. Norepinephrine infusion also activated mRNA production, but the effect was comparatively smaller than that on enzyme activity. beta-Adrenergic agonists also stimulated GyK activity with the following relative magnitude of response: CL316243 (beta(3)) > isoproterenol (non-selective) > dobutamine (beta(1)). In vitro rates of incorporation of glycerol into glyceride-glycerol were increased in BAT from rats exposed to cold. The data suggest that in conditions of a sustained increase in BAT sympathetic flow there is a stimulation of GyK gene expression at the pretranslational level, with increased enzyme activity, mediated by beta-adrenoreceptors, mainly beta(3).

Glucose uptake, glucose transporter GLUT4, and glycolytic enzymes in brown adipose tissue from rats adapted to a high-protein diet.

In vivo rates of glucose uptake, insulin-responsive glucose transporter (GLUT4) content, and activities of glycolytic enzymes were determined in brown adipose tissue (BAT) from rats adapted to a high-protein, carbohydrate-free (HP) diet. Adaptation to the HP diet resulted in marked decreases in BAT glucose uptake and in GLUT4 content. Replacement of the HP diet by a balanced control diet for 24 hours restored BAT glucose uptake to levels above those in rats fed the control diet, with no changes in GLUT4 levels in 4 of 5 animals examined. BAT denervation of rats fed the control diet induced a 50% reduction in glucose uptake, but did not significantly affect the already markedly reduced BAT hexose uptake in HP diet-fed rats. It is suggested that the pronounced decrease in BAT glucose uptake in these animals is due to the combined effects of the HP diet-induced reductions in plasma insulin levels and in BAT sympathetic activity. Adaptation to the HP diet was accompanied by decreased activities of hexokinase, phosphofructo-1-kinase, and pyruvate kinase (PK). The activity of BAT PK in HP diet-fed rats was reduced to about 50% of controls, and approached normal levels 24 hours after diet reversion. BAT denervation induced a small (15%) decrease in BAT PK activity in control rats, but did not affect the activity of the enzyme in HP diet-adapted rats. Also, denervation did not interfere with the restoration of PK activity induced by diet substitution. Treatment with anti-insulin serum resulted in an almost 50% reduction in PK activity in both innervated and denervated BAT from rats fed the control diet, but caused a much smaller ( thick approximate 20%) decrease in BAT from HP diet-fed rats. Furthermore, anti-insulin serum administration completely suppressed the restoration of BAT PK activity induced by diet reversion. These data suggest that, differently from glucose uptake, BAT PK activity is predominantly controlled by hormonal/metabolic factors.

Relative importance of sympathetic outflow and insulin in the reactivation of brown adipose tissue lipogenesis in rats adapted to a high-protein diet.

The effect of denervation or acute insulin deficiency on brown adipose tissue lipogenesis was investigated in rats adapted to a high-protein diet before and after diet reversion to a balanced diet. Denervation of rats fed the balanced diet induced a 50% reduction in in vivo rates of brown adipose tissue fatty acid synthesis, with decreased activities of acetyl-CoA carboxylase and adenosine triphosphate (ATP)-citrate lyase. The markedly (80%) reduced fatty acid synthesis and enzyme activities in brown adipose tissue from rats adapted to the high-protein diet were not affected by denervation. Replacement of the high-protein diet by the balanced diet for 24 hours restored fatty acid synthesis to normal levels, but recovery of enzyme activities was only partial. Lipogenesis restoration and partial recovery of enzyme activities were impaired in denervated tissue from high-protein diet-fed rats. In all experimental conditions, the activities of acetyl-CoA carboxylase and ATP-citrate lyase showed a better correlation with brown adipose tissue lipogenesis than the generators of H(+), glucose-6-P dehydrogenase, and malic enzyme. Anti-insulin serum administration during the 12- to 24-hour period after diet reversion completely blocked lipogenesis recovery in innervated and denervated tissues and drastically reduced brown adipose tissue lipogenesis of concomitantly injected rats fed the balanced diet. The data suggest that efficient and rapid adjustments of brown adipose tissue lipogenesis require sympathetic activation, and that this tissue can maintain significant, albeit reduced, rates of lipogenesis in the absence of sympathetic activation, but not in the absence of insulin.

Glycerokinase activity in brown adipose tissue: a sympathetic regulation?

The effect of brown adipose tissue (BAT) sympathetic hemidenervation on the activity of glycerokinase (GyK) was investigated in different physiological conditions. In rats fed a balanced diet, the activity of the enzyme was approximately 50% lower in BAT-denervated pads than in intact, innervated pads. In rats adapted to a high-protein, carbohydrate-free diet, norepinephrine turnover rates and BAT GyK activity were already reduced, and BAT denervation resulted in a further decrease in the activity of the enzyme. Cold acclimation of normally fed rats at 4 degrees C for 10 days markedly increased the activity of the enzyme. Cold exposure (4 degrees C) for 6 h was insufficient to stimulate BAT GyK, but the activity of the enzyme was already increased after 12 h of cold exposure. The cold-induced BAT GyK stimulation was completely blocked in BAT-denervated pads. The data indicate that an adequate sympathetic flow to BAT is required for the maintenance of normal levels of GyK activity and for the enzyme response to situations, such as cold exposure, which markedly increase BAT sympathetic flow.