Antilipolytic activity in vitro of various analogs of nicotinic acid. Structure-activity relationship

The antilipolytic activity of a series of N aryl-nicotinamides and of alpha picolinic acid, has been tested in vitro. Lipolysis was stimulated by epinephrine (20 micrograms/ml of incubation medium) using rat's epididymal adipose tissue slices. Only N(2-carboxy methyl phenyl) nicotinamide showed antilipolytic effect comparable to that of nicotinic acid at similar concentrations (2 X 10(-5) M). Picolinic acid (10(-4) M) showed no antilipolytic effect. These results, together with those of the literature, are discussed in regard to the relations between structure and antilipolytic activity.     

Clearing-factor lipase in adipose tissue. Distinction of different states of the enzyme and the possible role of the fat cell in the maintenance of tissue activity

1. Incubation of intact epididymal adipose tissue from fed rats at 37 degrees in an albumin solution at pH7.4 in vitro results in rapid loss of clearing-factor lipase activity until a low activity, stable to prolonged incubation, is attained. The clearing-factor lipase activity of intact tissue from starved rats, which is initially much less than that of tissue from fed rats, is mainly stable to incubation at 37 degrees . 2. Much of the clearing-factor lipase activity of intact epididymal adipose tissue from fed rats is inactivated by collagenase. The enzyme activity of intact tissue from starved rats is not inactivated by collagenase. 3. The clearing-factor lipase activity of fat cells isolated from the epididymal adipose tissue of fed rats is stable to prolonged incubation at 37 degrees . It represents only a small proportion of the total activity of the intact tissue. In starved rats, the isolated fat cells contain a much higher proportion of the activity of the intact tissue. Their activity is also stable at 37 degrees . 4. Incubation of isolated fat cells in a serum-based medium leads to a progressive rise in clearing-factor lipase activity. Actinomycin increases the extent of this rise in activity. No rise in clearing-factor lipase activity occurs when stromal-vascular cells isolated from epididymal adipose tissue are incubated in the medium. 5. The findings indicate that less than 20% of the activity of intact adipose tissue from fed rats is retained when fat cells are isolated from the tissue by collagenase treatment. The activity that is lost could be that which normally functions in the uptake of triglyceride fatty acids by the tissue.     

Inhibition of adenosine 3':k'-monophosphate accumulation white fat acids, lactate, and beta-hydroxybutyrate.

The large increase in cyclic AMP accumulation by rat white fat cells seen in the presence of lipolytic agents plus methylxanthines and adenosine deaminase was markedly inhibited by lactate. However, lipolysis was unaffected by lactate. Octanoate, hexanoate, heptanoate, and beta-hydroxybutyrate inhibited both cyclic AMP accumulation and lipolysis by rat fat cells. The mechanism by which these acids inhibit lipolysis differs from that for long chain fatty acids such as oleate. Oleate directly inhibited triglyceride lipase activity of homogenized rat adipose tissue. In contrast, octanoate, beta-hydroxybutyrate, and lacatate had no effect on triglyceride lipase activity. Hormone-stimulated adenylate cyclase activity of rat fat cell ghosts was inhibited by oleate and 4mM octanoate but not by 1.6 mM octanoate, heptanoate, hexanoate, beta-hydroxybutyrate or lactate. None of the acids affected the soluble protein kinase activity of rat adipose tissue. There was no stimulation by lactate, butyrate, beta-hydroxybutyrate, or octanoate of the soluble or particulate cyclic AMP antilipolytic action of a short chain acid such as octanoate or hexanoate was not accompanied by any drop in total fat cell ATP. The mechanism by which lactate lowers cyclic AMP but not lipolysis remains to be established.     

Adrenergic lipolysis in human fat cells: properties and physiological role of alpha-adrenergic receptors (author's transl)

Lipolytic activity of human isolated fat cells from different fat deposits was studied. The purpose of the present investigations was to determine the epinephrine responsiveness, with regard to alpha- and beta-adrenergic receptor site activity, of omental and subcutaneous adipocytes (abdominal or from the lateral part of the thigh). Adipocytes were obtained from normal subjects or from obese subjects on iso- or hypocaloric diets. The lipolytic effect of epinephrine varied according to the fat deposits, while the beta-lipolytic effect of isoproterenol was more stable (Fig. 1). We explored the possible involvement of adrenergic alpha-receptors, in order to explain these results. The potentiating action of phentolamine on epinephrine-induced lipolysis, and the antilipolytic effect of alpha-agonists on basal or theophylline--induced lipolysis, were found to be a good indication of alpha-adrenergic activity. The alpha-adrenergic antilipolytic effect was most prominent in adipose tissue from the lateral part of the thigh, and less noticeable in omental adipocytes. In conclusion, the inability of epinephrine to induce lipolysis, and the epinephrine-induced inhibition of lipolysis observed when the basal rate of FFA release was spontaneously increased in subcutaneous fat-cells of the thigh, could be explained by an increased alpha adrenergic responsiveness (Fig. 2). Moreover, various alpha-adrenergic agonists (phenylephrine, noradrenaline and adrenaline) showed a clear inhibiting effect on theophylline-stimulated adipocytes from the thigh. The pharmacological study of the antilipolytic effect of epinephrine on theophylline-induced lipolysis showed that the inhibition was linked to a specific stimulation of the alpha-receptors of the subcutaneous adipocytes (Fig. 4). From the different sets of experiments, it is shown that the modifications in the lipolytic effect of epinephrine on adipocytes of different areas could be explained by the occurrence of a variable alpha-adrenergic effect initiated by catecholamine. Furthermore, theophylline stimulation of lipolysis provides an accurate system to investigate the alpha-inhibiting effect of catecholamines. Our study was completed by the investigation of the lipolytic activity of subcutaneous fat cells from obese subjects submitted to a hypocaloric diet (800-1 000 Cal/day). An increased alpha-inhibitory effect of epinephrine was shown on the increased basal lipolytic activity observed in the fat cells of obese subjects on a hypocaloric diet (Fig. 5); a similar effect was observed when these adipocytes were stimulated by theophylline. To conclude, these investigations allow the alpha-adrenergic effect to be considered as a regulator mechanism of the in vitro lipolytic activity in human adipose tissue, since the antilipolytic effect is operative whenever the basal rate of lipolysis is increased (spontaneously, after caloric restriction, or with a lipolytic agent such as theophylline).     

Activation of hormone-sensitive lipase in extracts of adipose tissue

Rat adipose tissue was homogenized in 0.154 m KCl, and the supernatant fluid, obtained after centrifugation at 15,000 g, was extracted with benzene to remove triglycerides. Most of the lipase activity in the extracted fluid was precipitated with ammonium sulfate between 15 and 40% saturation. The specific activity of the lipase in this fraction was about three times that in the benzene-extracted supernatant fluid. The specific activity of the monoglyceride esterase was increased to a lesser extent. Lipase activity in the benzene-extracted fluid and in the ammonium sulfate fraction was increased 15-45% by incubation with 0.3 mm ATP, 10 mm MgCl(2), and 0.03 mm cyclic AMP for 10 min before assay. None of these compounds alone or in combinations of two was as effective as all three together. The specific activity of the 15-40% ammonium sulfate fraction prepared from fat cells exposed to epinephrine and glucagon was greater than that from portions of the same cell pool not exposed to hormones. In addition, the already elevated lipase activity in preparations from hormone-treated cells was not enhanced by incubation with ATP, MgCl(2), and cyclic AMP. Thus, it seems probable that the lipase activity in the ammonium sulfate fractions represents, at least in part, hormone-sensitive lipase.     

Regulation of lipoprotein lipase. Induction by insulin.

Lipoprotein lipase activity in intact epididymal adipose tissue of fasted rats increased rapidly after treatment with insulin in vivo. In contrast, lipoprotein lipase activity in adipocytes isolated from the contralateral fat pads remained essentially unchanged. When adipocytes were incubated for 30 min at ambient temperature in vitro, about 2 times more lipoprotein lipase activity was found in the medium of cells from insulin-treated rats than in medium from cells of control animals. Following insulin treatment, extracts of tissue acetone powders separated by gel chromatography showed increases in both enzyme activity fractions obtained (designated lipoprotein lipase a and b). However, no consistent differences were observed between fractions derived from adipocyte acetone powders of insulin-treated and control animals. All the observed effects of insulin on lipoprotein lipase activity were abolished by cycloheximide treatment in vivo. These data indicate that following insulin treatment, increased lipoprotein lipase activity in adipose tissue results from enhanced enzyme secretion by the fat cell and subsequent accumulation in the tissue, thus implicating the adipocyte secretory mechanism as a major site of regulation of lipoprotein lipase activity in adipose tissue.     

Relationship of lipoprotein lipase activity to triglyceride uptake in adipose tissue

Fasted rats injected with actinomycin or fed glucose show increased lipoprotein lipase activity of epididymal adipose tissue. Data from the actinomycin-treated animals showed a direct correlation between the lipoprotein lipase activity and the uptake of lipoprotein triglyceride by the epididymal fat pad in vitro and in vivo. Data from the animals fed glucose confirmed these findings in vitro. These data strongly suggest that lipoprotein lipase plays a major role in triglyceride deposition in adipose tissue.     

Dietary fish oil reverse epididymal tissue adiposity, cell hypertrophy and insulin resistance in dyslipemic sucrose fed rat model small star, filled

The present work was designed to assess the possible benefits of (7% w/w) dietary fish oil in reversing the morphological and metabolic changes present in the adipose tissue of rats fed an SRD for a long time. With this purpose, in the epididymal fat tissue, we investigated the effect of dietary fish oil upon: i) the number, size and distribution of cells, ii) the basal and stimulated lipolysis, iii) the lipoprotein lipase (LPL) and the glucose 6-phosphate dehydrogenase activities, and iv) the antilipolytic action of insulin. The study was conducted on rats fed an SRD during 120 days with fish oil being isocaloric substituted for corn oil for 90-120 days in half the animals. Permanent hypertriglyceridemia, insulin resistance and abnormal glucose homeostasis were present in the rats before the source of fat in the diet was replaced. The major new findings of this study are the following: i) Dietary fish oil markedly reduced the fat pads mass, the hypertrophy of fat cells and improved the altered cell size distribution. ii) The presence of fish oil in the diet corrected the inhibitory effect of high sucrose diet upon the antilipolytic action of insulin, reduced the "in vitro" enhanced basal lipolysis and normalized isoproterenol-stimulated lipolysis. Fat pads lipoprotein lipase activity decreased reaching values similar to those observed in age-matched controls fed a control diet (CD). These effects were not accompanied by any change in rat body weight. All these data suggest that the dyslipemic rats fed a moderate amount of dietary fish oil constitute a useful animal model to study diet-regulated insulin action.     

Inhibition of the reversible deactivation of chicken adipose tissue hormone-sensitive lipase by heat-stable proteins from adipose tissue and skeletal muscle.

The reversible deactivation of chicken adipose tissue hormone-sensitive lipase is catalyzed by a lipase phosphatase. Heat-stable protein preparations from rat epididymal fat pads, chicken adipose tissue, and rabbit skeletal muscle inhibited lipase phosphatase activity. Phosphatase inhibitor preparations from rat adipose tissue did not inhibit the protein kinase-catalyzed activation of hormone-sensitive lipase, whereas inhibitor preparations from rabbit skeletal muscle were contaminated with protein kinase inhibitor.     

Enzymes catalyzing the hydrolysis of long-chain monoacyglycerols in rat adipose tissue

Acetone-ether preparations of epididymal fat pads from fasted or fed rats contained two enzymes catalyzing the hydrolysis of long-chain monoacylglycerols. The enzymes were identified as monoacylglycerol lipase (Tornqvist, H. and Belfrage, P., (1976) J. Biol Chem. 251, 813--819) and lipoprotein lipase by their apparent pI values after electrofocusing in non-ionic detergent, selective inhibition properties, substrate specificity and positional specificity. It was estimated that monoacylglycerol lipase accounted for about 90% of the total monoacylglycerol-hydrolyzing activity in acetone-ether preparations from fasted and 70% from fed rats. Its enzyme activity did not change with the nutritional state in contrast to that of lipoprotein lipase. The latter enzyme hydrolyzed 2-monoacylglycerols at a much lower rate than the 1(3)-isomers. Monoacylglycerol lipase was located almost entirely in the adipocytes, thus most of the enzyme activity towards monoacylglycerols in the adipose tissue was found in this site. Fractionated sucrose homogenates of rat epididymal fat pads also contained a third enzyme with monoacylglycerol-hydrolyzing activity, identified as hormone-sensitive lipase by its pI, selective inhibition properties and substrate specificity. It was estimated that hormone-sensitive lipase accounted for less than 20% of the total activity against monoacylglycerols in these tissue preparations from fasted rats. Over-all quantitative estimations emphasized the dominant role of monoacylglycerol lipase over the other two enzymes in the hydrolysis of monoacylglycerols.     

Selective antilipolytic effect of bacitracin in the isolated fat cell

Bacitracin, an antibiotic which decreases extracellular degradation, has been used to study peptide hormone degradation $\text{in}\text{vitro}$. The biologic effectiveness of these hormones in the presence of bacitracin has received minimal attention. This study demonstrates inhibition of lipolysis induced by both epinephrine and glucagon in the isolated fat cell (IFC). IFC from epididymal tissue were incubated with 0.5 μM epinephrine and increasing concentrations of bacitracin. Lipolysis was inhibited in a dose-dependent fashion, with a concentration of 5.7 × 10^?4$\text{M}$ bacitracin suppressing lipolysis 50%. Increasing the concentration of epinephrine in the presence of a constant dose of bacitracin overcame the antilipolytic effect. Bacitracin did not increase oxidation of glucose-U-C¹⁴ over basal. In the perifusion system, acute exposure to 5.7 × 10^?4$\text{M}$ bacitracin plus 5 × 10^?9$\text{M}$ glucagon suppressed lipolysis below unstimulated basal levels. Constant bacitracin perifusion produced no change in basal lipolysis but blunted the response to glucagon. ¹²⁵I-glucagon degradation was decreased in the presence of bacitracin. Additional studies with dibutyryl cyclic AMP demonstrated that the antilipolytic effect of bacitracin is exerted at a step beyon the second messenger. Bacitracin exerts a direct antilipolytic effect in isolated fat cells without stimulating glucose uptake and may afford a means of studying antilipolysis in the absence of other insulin-like effects.     

The role of beta and alpha adrenergic receptors in the lipolytic effect of catecholamines on dog adipocytes (author's transl)]

Pharmacological properties of adrenergic receptors have been investigated in fat cells isolated from omental adipose tissue of the Dog. From the results, the following points can be stated. 1. Lipolysis is markedly enhanced by isoproterenol. This effect is competitively inhibited by propranolol (a beta-adrenoceptor blocking agent). (Fig 1). 2. The beta 2-sympathomimetic salbutamol is found to have only a slight effect on lipolysis rate (Fig. 2). 3. The epinephrine-induced lipolysis is potentiated by phentolamine (an alpha-adrenoceptor blocking agent) only on large sized adipose cells (mean fat cell size 96.7 +/- 5.3 micrometer; Fig. 5). 4. The isoproterenol-induced lipolysis is partially inhibited by phenylephrine (an alpha-adrenomimetic drug) (Table I). These findings show that beta 1 nature of the receptors involved in the adrenergic control of lipolysis in Dog adipose tissue. Moreover an antilipolytic effect of epinephrine, via alpha-adrenergic receptors, is observed, especially in large adipose cells.     

Inhibition of rat and human adipocyte adenylate cyclase in the antilipolytic action of insulin, clofibrate, and nicotinic acid.

Clofibrate (Atromid-S), nicotinic acid, and insulin are known to be potent hypolipidemic and antilipolytic agents. The present study was undertaken to define the mechanism of action of this latter effect on isolated rat and human fat cells. Sodium clofibrate (0.42 mM), nicotinic acid (0.42 mM), and insulin (100 microU/mL) were shown to inhibit norepinephrine-stimulated lipolysis in rat and human adipose cells and this inhibition was associated with a reduction in intracellular 3',5'-cyclic AMP levels. A similar cyclic AMP lowering effect was demonstrated with insulin in the presence of procaine-HCL, which uncouples the adenylate cyclase system from lipolysis. This insulin effect was attributed to inhibition of adenylate cyclase. A direct and significant inhibition of adenylate cyclase in membrane fractions obtained from isolated human adipocytes was demonstrated for all three antilipolytic agents. The common membrane site of action of these agents whereby adenylate cyclase activity is depressed, thus decreasing cyclic AMP production and free fatty acid (FFA) mobilization from adipose stores, implies a central role for the adenylate cyclase system. These findings are consistent with the view that the hypotriglyceridemic effects of clofibrate, nicotinic acid, and insulin may be partly explained by deprivation of FFA substrate for hepatic very low density lipoprotein synthesis.     

5 alpha-reductase activity in rat adipose tissue

We measured the 5 alpha-reductase activity in isolated cell preparations of rat adipose tissue using the formation of [3H]dihydrotestosterone from [3H]testosterone as an endpoint. Stromal cells were prepared from the epididymal fat pad, perinephric fat, and subcutaneous fat of male rats and from perinephric fat of female rats. Adipocytes were prepared from the epididymal fat pad and perinephric fat of male rats. Stromal cells from the epididymal fat pad and perinephric fat contained greater 5 alpha-reductase activity than did the adipocytes from these depots. Stromal cells from the epididymal fat pad contained greater activity than those from perinephric and subcutaneous depots. Perinephric stromal cells from female rats were slightly more active than those from male rats. Estradiol (10(-8) M), when added to the medium, caused a 90% decrease in 5 alpha-reductase activity. Aromatase activity was minimal, several orders of magnitude less than 5 alpha-reductase activity in each tissue studied.     

Acute exposure to long-chain fatty acids impairs {alpha}2-adrenergic receptor-mediated antilipolysis in human adipose tissue

The acute in vitro and in vivo effects of long-chain fatty acids (LCFAs) on the regulation of adrenergic lipolysis were investigated in human adipose tissue. The effect of a 2 h incubation, without or with LCFA (200 mumol/l), on basal and hormonally induced lipolysis was tested in vitro on isolated fat cells. The lipolytic response to epinephrine was enhanced by suppression of the antilipolytic alpha(2)-adrenergic effect. Then, healthy lean and obese male subjects performed a 45 min exercise bout at 50% of their heart rate reserve either after an overnight fast or 3 h after a high-fat meal (HFM: 95% fat, 5% carbohydrates). Subcutaneous adipose tissue lipolysis was measured by microdialysis in the presence or absence of an alpha-antagonist (phentolamine). In vivo, a HFM increased plasma levels of nonesterified fatty acids in lean and obese subjects. In both groups, the HFM did not alter hormonal responses to exercise. Under fasting conditions, the alpha(2)-adrenergic antilipolytic effect was more pronounced in obese than in lean subjects. The HFM totally suppressed the alpha(2)-adrenergic antilipolytic effect in lean and obese subjects during exercise. LCFAs per se, in vitro as well as in vivo, suppress alpha(2)-adrenergic-mediated antilipolysis in adipose tissue. LCFA-mediated suppression of antilipolytic pathways represents another mechanism whereby a high fat content in the diet might increase adipose tissue lipolysis.     

Effects of dietary cholesterol on adipose tissue lipoprotein lipase in the baboon

The effects of infant diet (breast milk or formula containing 2, 30 or 60 mg/dl cholesterol) and subsequent dietary cholesterol (0.02, 1.0 or 1.7 mg/kcal) and fat (saturated or unsaturated) on heparin-releasable lipolytic activity from omental adipose tissue was estimated from 99 baboons of 5-8 years of age. This lipase activity was characterized as lipoprotein lipase based on salt inhibition and apolipoprotein C-II activation. Lipoprotein lipase activity released from adipose tissue by heparin was significantly (P less than 0.002) lower in high cholesterol-fed baboons than in those fed low cholesterol. Most of this difference was due to impaired long-term heparin release of lipoprotein lipase. Adipose tissue lipoprotein lipase increased with increasing fat cell size regardless of diet, but there was no effect of diet on adipocyte size. There were no significant effects of infant cholesterol intake nor adult saturated or unsaturated fat on lipoprotein lipase activity. Adult baboons breast fed as infants had lower adipose tissue lipoprotein lipase activity (P less than 0.07) than adults fed formula as infants.     

Inhibition of adipose S-100 protein release by insulin

The release of S-100 protein brought about in rat epididymal fat pads by 10 microM epinephrine was inhibited by about 50% in the presence of more than 8 nM insulin. The inhibitory effect of insulin was also observed in the release of S-100 protein induced by isoproterenol or adrenocorticotropin (ACTH), but not in the release induced by a high concentration (5 mM) of dibutyryl cyclic AMP. Since insulin suppressed (to about 50%) the increase in cyclic AMP content induced by epinephrine under the same conditions, it is suggested that the inhibitory mechanism is mediated by the cyclic AMP levels in adipocytes. The S-100 protein release induced by catecholamine was significantly decreased (to about 50%) in the fat pads obtained from insulin-injected rats. In contrast, in the fat pads obtained from diabetic or long-term starved rats, the S-100 protein release was greatly enhanced, showing several-fold higher levels of basal release in the absence of hormones, and S-100 protein contents in the epididymal adipose tissues of these rats were significantly lower than those of the control rats. These results suggest that the S-100 protein content in adipocytes is regulated by insulin as well as the lipolytic hormones.     

Effect of tumor necrosis factor administration in vivo on lipoprotein lipase activity in various tissues of the rat

When added to murine adipocytes in culture, tumor necrosis factor (TNF) decreases the levels of lipoprotein lipase (LPL). Semb et al (1987. J. Biol Chem. 262: 8390-8394) have shown that administration of murine TNF to rats decreases lipoprotein lipase (LPL) in the epididymal fat pad with maximal inhibition requiring several hours. We have now tested the effects of treatment of rats with TNF on LPL activity in a variety of tissues and find that few show decreases in LPL under conditions that acutely increase serum triglycerides. Ninety minutes after treatment of male rats with human TNF (25 micrograms/200 g, i.v.), serum triglycerides rose 2.2-fold but there was no decrease in LPL activity in epididymal fat. Sixteen hours after TNF treatment LPL activity had decreased by 44% in epididymal fat, consistent with the previously reported data. In contrast, in female rats, no significant decrease was seen in LPL activity in parametrial adipose tissue at either 90 min or 16 hr after TNF administration despite increases in serum triglycerides (1.8-fold and 1.5-fold, respectively). There was little change in LPL activity in most other adipose tissue sites of male or female rats at either time after TNF treatment. No effect of TNF was seen on heart or diaphragm muscle LPL at any time. TNF treatment of both male and female rats produces consistent increases in de novo hepatic lipogenesis in vivo under conditions that increase serum triglycerides. It is unlikely that the limited effects of TNF on LPL in vivo can account for the rapid and sustained increase in serum triglycerides.     

Lack of feedback regulation of cyclic 3':5'-AMP accumulation by free fatty acids in chicken fat cells.

Fat cells isolated from the mesenteric adipose tissue of chickens (pullets) responded to glucagon with an increase in lipolysis and a sustained rise in cyclic adenosine 3':5'-monophosphate (cyclic AMP) over a 30-min incubation. The prolonged accumulation of cyclic AMP due to glucagon in chicken fat cells was primarily intracellular. In addition, there was little increase in cyclic AMP accumulation due to theophylline alone or potentiation of the increase due to glucagon. These data indicate that chicken fat cells, unlike rat fat cells, are relatively insensitive to theophylline. Neither lipolysis nor cyclic AMP accumulation by chicken fat cells was inhibited by free fatty acid to albumin ratios (3 to 7) which markedly reduced both events in rat fat cells. However, in the absence of albumin from the medium, lipolysis in chicken fat cells was reduced, but not to the same extent as in rat fat cells. Chicken fat cells did accumulate more intracellular free fatty acids in response to lipolytic agents than did rat fat cells. The uptake of oleate by rat and chicken fat cells was identical. Glucagon-induced accumulation of cyclic AMP by chicken fat cell ghosts was unaffected by added oleate. Under identical conditions glucagon-induced adenylate cyclase activity of rat fat cell ghosts was markedly inhibited by added oleate. Triglyceride lipase activity of the pH 5.2 precipitate from a 40,000 x g infranatant of homogenized fat cells from chickens was less sensitive than that from rat fat cells to the ratio of oleate to albumin. These results suggest that the maintenance of cyclic AMP levels in chicken fat cells incubated with lipolytic agents results from the relative insensitivity of chicken fat cells to free fatty acid inhibition of cyclic AMP accumulation.     

Effects of repeated injections of epinephrine on lipolysis in rats

The effects of repeated injections of epinephrine on lipolysis in rats were investigated in vivo and in vitro. By daily subcutaneous injections of epinephrine (100 micrograms/kg) for 20 days, the weight of epididymal adipose pads were significantly decreased. By repeated injections of epinephrine for 20 days, the extent of increase in plasma nonesterified fatty acid (NEFA) levels after epinephrine injection became smaller, and the amount of epinephrine-induced NEFA and cyclic AMP release from epididymal adipose tissue were decreased. These results suggested that the lipolytic response to epinephrine in rats was decreased by repeated injections of epinephrine.