Ann C. Sullivan

Inhibition of lipogenesis in rat liver by (−)-hydroxycitrate

Abstract The purpose of these investigations was to determine the effect of the stereoisomers of hydroxycitrate on the rate of lipogenesis in rat liver. These observations were made under conditions in which there was an induced rate of lipid synthesis. Only (−)-hydroxycitrate significantly decreased the conversion of [14C]citrate into lipid in liver high speed supernatant and the conversion of [14C]alanine into lipid in vivo. The in vivo rate of lipogenesis was markedly decreased for 150 min following the administration of (−)-hydroxycitrate either iv (0.017 mmoles/kg) or ip (0.0263 mmoles/kg). Fatty acid and cholesterol synthesis were significantly inhibited by the oral administration of (−)-hydroxycitrate (5.26, 3.95 and 2.63 mmoles/kg) only when the compound was given before the feeding period.

Effect of (−)-hydroxycitrate upon the accumulation of lipid in the rat: I. Lipogenesis

The purpose of these investigations was to ascertain the effect of (−)-hydroxycitrate on the accumulation of lipid in the meal fed rat by examining the rates of lipogenesis after acute and chronic treatment. Oral administration of (−)-hydroxycitrate depressed significantly the in vivo lipogenic rates in a dose-dependent manner in the liver, adipose tissue, and small intestine. The hepatic inhibition was significant for the 8 hr period, when control animals demonstrated elevated rates of lipid synthesis. The kinetics of this reduction of in vivo hepatic lipogenesis were identical after acute or chronic administration of (−)-hydroxycitrate. However, in vitro rates of lipogenesis were elevated after chronic administration of (−)-hydroxycitrate for 30 days. Rats receiving (−)-hydroxycitrate consumed less food than the untreated controls; however, this decreased caloric intake was not responsible for the drug induced depression of hepatic lipogenesis, as shown by studies using pair fed rats.

Effect of (−)-hydroxycitrate upon the accumulation of lipid in the rat: II. Appetite

These studies were designed to determine the effect of (−)-hydroxycitrate upon the accumulation of lipid in the rat by examining appetite, wt gain, and total body lipid profiles. The chronic oral administration of a nontoxic dose of (−)-hydroxycitrate to growing rats for 11–30 days caused a significant reduction in body wt gain, food consumption, and total body lipid. The administration of equimolar amounts of citrate did not alter wt gain, appetite, or body lipid. No increase in liver size or liver lipid content occurred with either treatment. Pair feeding studies demonstrated that the reduction in food intake accounted for the decrease in wt gain and body lipid observed with (−)-hydroxycitrate treatment.

Abnormal sympatho-adrenal function and plasma catecholamines in obese Zucker rats

The functional integrity of the peripheral sympathetic nervous system and adrenal medulla was assessed in homozygous, lean and obese, 7--8 month old male Zucker rats by the changes in plasma catecholamines during cold and immobilization stresses. Five of eight obese, but no lean rats died during a 24 hr cold stress (4--7 degrees C) from hypothermia. While both lean and obese rats had decreased rectal temperatures after 4 hr of cold stress, the obese had lower temperatures, relatively less of an increase of plasma norepinephrine (NE) and epinephrine (E) than the lean rats, and were unable to consistently maintain their temperatures even during intravenous NE infusions. Obese rats had lower rectal temperatures and higher plasma NE and dopamine levels at 21--22 degrees C ambient temperature, a relative failure to increase plasma NE and E levels after 1 hr of immobilization, but normal or supranormal plasma catecholamine levels after decapitation compared to the lean rats. These results suggest that the obese Zucker rat has abnormalities of both peripheral sympatho-adrenal function and thermoregulation, which may play roles in the development and/or maintenance of many of the physiological and metabolic defects in this animal model of genetic obesity.

Catecholamine levels in discrete brain nuclei of seven month old genetically obese rats.

Abstract Catecholamine levels were measured in microdissected nuclear groups of seven month old, obese and lean, male and female Zucker rats. By analysis of variance, obese rats showed significantly reduced levels of norepinephrine and epinephrine in the paraventricular nucleus and of epinephrine and dopamine in the dorsomedial hypothalamic nucleus. Norepinephrine levels were increased in the median forebrain bundle and caudate nucleus. When compared by sex and genotype (t-test), the female obese rats had significant decreases compared to the leans of norepinephrine and epinephrine levels in the paraventricular and dorsomedial hypothalamic nuclei, while norepinephrine was increased in the median forebrain bundle and caudate nucleus. Dopamine was decreased in the dorsomedial and increased in the C2 nuclei. Male obese Zucker rats showed changes only in the dorsomedial and C2 nuclei where dopamine levels were decreased. In general, female obese and lean rats tended to have lower levels of catecholamines in various brain areas than males of the same genotype. Comparisons of these data to previous studies in younger Zucker rats [9,10] suggested that changes also occurred in various nuclei with aging. It is postulated that catecholamine deficits in certain hypothalamic nuclei of the Zucker obese rat may be a contributing factor to the development and/or maintenance of obesity, possibly in association with abnormalities in thermal regulation.

Effects of diet and obesity on brain α1- and α2-noradrenergic receptors in the rat

Abstract The chronic feeding of a sweetened condensed milk/corn oil diet (CM diet) to adult male rats produced significant increases in body weight and levels of plasma insulin in 34% of the rats fed this diet with respect to chow-fed controls. Levels of α 1 -noradrenergic receptor binding were lower (32%) in the hypothalamic ventromedial nucleus (VMN) of only those rats which became obese (DIO rats) with respect to both chow-fed controls and those rats which resisted the development of obesity on the CM diet (DR rats). Also, α 1 -noradrenergic binding was inversely proportional to body weight gain in the VMN ( r = −0.831). α 2 -Noradrenergic receptors were 30–37% lower in both the DIO and DR rats in the dorsomedial nucleus and dorsal area of the hypothalamus, and the medial dorsal area and nucleus reuniens of the thalamus. The similar decreases in α 2 -noradrenergic receptors in both the DIO and DR rats in these areas suggested that dietary factors alone were responsible for these changes. There were no significant differences from chow-fed rats for hypothalamic dopamine (D 2 ) or β-noradrenergic ( β 1 - and β 2 -) receptors in either DR or DIO rats. These results indicate that VMN α 1 -noradrenergic receptors co-vary with body weight and implicate a role for α 1 -receptors in the VMN in the central neuronal regulation of body weight.

Hypolipidemic activity of (−)-hydroxycitrate

The influence of (−)-hydroxycitrate, a potent competitive inhibitor of adenosine triphosphate (ATP) citrate lyase, on serum triglyceride and cholesterol levels, and in vitro and in vivo rates of hepatic fatty acid and cholesterol synthesis was investigated in normal and hyperlipidemic rat model systems. (−)-Hydroxycitrate reduced equivalently the biosynthesis of triglycerides, phospholipids, cholesterol, diglycerides, cholesteryl esters, and free fatty acids in isolated liver cells. In vivo hepatic rates of fatty acid and cholesterol synthesis determined in meal-fed normolipidemic rats were suppressed significantly by the oral administration of (−)-hydroxycitrate for 6 hr, when control animals exhibited maximal rates of lipid synthesis; serum triglyceride and cholesterol levels were significantly reduced by (−)-hydroxycitrate. In two hypertryglyceridemic models—the genetically obese Zucker rat and the fructose-treated rat—elevated triglyceride levels were due, in part, to enhanced hepatic rates of fatty acid synthesis. (−)-Hydroxycitrate significantly reduced the hypertriglyceridemia and hyperlipogenesis in both models. The marked hypertriglyceridemia exhibited by the triton-treated rat was only minimally due to increased hepatic lipogenesis; (−)-hydroxycitrate significantly inhibited both serum triglyceride levels and lipogenesis in this model.

Changes in Lipid Metabolism in Diet-Induced Obesity

Mature male Sprague-Dawley rats fed a powdered Purina Chow diet containing corn oil and condensed milk (CM) were compared to rats fed a Purina Chow diet (control). CM rats gained more weight and consumed more calories over a 73-day period than the control rats. The increased weight gain and body fat in CM rats was accompanied by increased cell number in retroperitoneal and inguinal but not epididymal fat pads while cell size was unchanged in all three pads. After obesity had developed there was an increase in insulin levels, lipolysis, hepatic fatty acid synthesis, and fatty acid oxidation. While CM rats demonstrated hyperinsulinemia and hyperglycerolemia, they maintained normal glucagon and glucose levels. They demonstrated higher rates of fatty acid synthesis in isolated hepatocytes but not in vivo, suggesting that a greater potential for fatty acid synthesis in CM rats was masked in vivo by the inhibitory action of dietary lipids. Beta-oxidation of (1-14C) palmitate in vivo and in vitro, and in vivo ketogenesis were greater in CM than in chow fed rats. These studies demonstrate that, after the development of obesity, CM rats, like genetically obese Zucker rats, are hyperinsulinemic and have elevated levels of fatty acid synthesis. However, unlike obese Zucker rats, CM rats displayed an increase in beta-oxidation. These studies suggest that increased insulin levels and hepatic fatty acid synthesis may contribute to dietary obesity (as they do to genetic obesity), whereas increased fatty acid oxidation in dietary obesity may be a compensatory response to maintain a lower body weight.

Defective catecholamine metabolism in peripheral organs of genetically obese zucker rats

Catecholamine (CA) metabolism in peripheral organs of lean and obese, 3-4 and 7-8 month (mo) old male Zucker rats was studied to define further the known abnormalities of peripheral sympatho-adrenal functions in the obese rat. Norepinephrine (NE) levels in all sympathetically innervated organs from obese rats (aorta, heart, pancreas, brown adipose tissue and white adipose tissue) were decreased to 9-55% of those in lean rats at 3-4 mo. NE turnover, measured by inhibition of tyrosine hydroxylase (TH) with a-methyl-p-tyrosine, was also decreased in these same organs. NE levels and turnover were also decreased (by 50-95%) in many, but not all organs assayed of 7-8 mo old obese rats, while there were inconsistent changes in organ dopamine and epinephrine levels at both ages. Decreased NE metabolism was associated with decreased dopamine-beta-hydroxylase (D beta H) activity in every organ assayed from obese rats at 3-4 mo and 7-8 mo of age except in the superior cervical ganglia of 7-8 mo old rats. There were no consistent changes in TH or phenylethanolamine-N-methyltransferase activities. In 3-4 mo old obese rats, decreased D beta H activity was associated with decreased immunoprecipitable enzyme protein in the hearts but not in the adrenal glands, where differences in the affinity for substrate appeared to explain the activity differences. These results suggest that the previously reported defect in stress-induced plasma NE levels in obese rats could be explained by decreased D beta H activity in nerve terminals of their sympathetic nervous system and that, in this case, D beta H may play an important role in the regulation of NE synthesis.

Metabolic consequences of fasting in old lean and obese Zucker rats.

The effects of fasting on lipid and carbohydrate metabolism and plasma insulin and glucagon levels were compared in lean and obese Zucker rats. Sixteen-month-old female and male rats were fasted for periods of 2, 4, 6 and 12 days. Fasting produced significant decreases in hepatic rates of lipid, cholesterol, and glycogen synthesis, as well as circulating levels of triglycerides, cholesterol, phospholipids, and insulin. Significant increases in hepatic lipid levels and serum free fatty acids were noted. When compared to lean rats, obese rats had elevated rates of hepatic lipid and glycogen synthesis, hepatic lipid and glycogen stores, serum triglycerides, cholesterol, phospholipids, and plasma insulin. Lean rats had higher plasma glucagon levels. Sex differences in several parameters were observed. Females demonstrated higher levels of lipid and cholesterol synthesis and serum free fatty acids, whereas serum cholesterol levels and hepatic glycogen stores were higher in males. Following a 12-day fast, carcass fat and protein content were decreased in both lean and obese rats, but the obese animals maintained an obese body composition. It is concluded that fasting results in qualitatively similar metabolic and hormonal changes in both lean and obese rats, but that abnormalities in carbohydrate and lipid metabolism persist in obese rats even after a 12-day fast.