Influence of ovarian hormones on development of ingestive responding to alterations in fatty acid oxidation in female rats

Adult male rats have been demonstrated to increase food intake in response to administration of drugs that interfere with oxidation of fatty acids (e.g. methyl palmoxirate and mercaptoacetate [MA]), effects that are larger in animals maintained on a high-fat diet. In contrast, while administration of MA has been reported to stimulate food intake in pre-pubertal female rats, food intake is not stimulated by MA in adult female rats. Instead, administration of MA to adult females results in changes in reproductive behavior and physiology. The present experiments were designed to examine the effects of administration of MA on food intake in adult female rats. The results demonstrated that, as previously reported, food intake was stimulated by MA in adult male rats on low-fat and high-fat diets, but food intake was not stimulated by MA in gonadally-intact adult female rats on either low-fat or high-fat diet. Further, MA did not stimulate food intake in female rats ovariectomized as adults. However, when females were ovariectomized prior to the onset of puberty (postnatal day 25–28), food intake was stimulated by administration of MA in adulthood. Finally, cyclic injections of 17-β-estradiol benzoate given to females ovariectomized prior to the onset of puberty abolished the stimulatory effects of MA on food intake in adult females. Taken together, the data suggest that exposure to estrogens during the time of puberty in female rats can persistently alter adult ingestive responding to signals related to changes in energy utilization.     

Estrogen replacement stimulates fatty acid oxidation and impairs post-ischemic recovery of hearts from ovariectomized female rats

Women less than 50 years of age, the majority of whom are likely premenopausal and exposed to estrogen, are at greater risk of a poor short-term recovery after myocardial ischemia than men and older women. Since estrogen enhances non-cardiac lipid utilization and increased lipid utilization is associated with poor post-ischemic heart function, we determined the effect of estrogen replacement on post-ischemic myocardial function and fatty acid oxidation. Female Sprague-Dawley rats, either intact (n = 15) or ovariectomized and treated with 17beta-estradiol (0.1 mg x kg(-1) x day(-1), s.c., n = 14) or corn oil vehicle (n = 16) for 5 weeks, were compared. Function and fatty acid oxidation of isolated working hearts perfused with 1.2 mM [9,10-3H]palmitate, 5.5 mM glucose, 0.5 mM lactate, and 100 mU/L insulin were measured before and after global no-flow ischemia. Only 36% of hearts from estrogen-treated rats recovered after ischemia compared with 56% from vehicle-treated rats (p > 0.05, not significant), while 93% of hearts from intact rats recovered (p < 0.05). Relative to pre-ischemic values, post-ischemic function of estrogen-treated hearts (26.3 +/- 10.1%) was significantly lower than vehicle-treated hearts (53.4 +/- 11.8%, p < 0.05) and hearts from intact rats (81.9 +/- 7.0%, p < 0.05). Following ischemia, fatty acid oxidation was greater in estrogen-treated hearts than in the other groups. Thus, estrogen replacement stimulates fatty acid oxidation and impairs post-ischemic recovery of isolated working hearts from ovariectomized female rats.     

The effects of inhibition of fatty acid oxidation in suckling newborn rats.

Inhibition of fatty acid oxidation with pent-4-enoate in suckling newborn rats caused a fall in blood [glucose] and blood [ketone bodies] and inhibition of gluconeogenesis from lactate. Glucose utilization was not increased in newborn rats injected with pent-4-enoate. Active fatty acid oxidation appears to be essential to support gluconeogenesis and to maintain normal blood [glucose] in suckling newborn rats.     

Effect of ovarian hormones on glycosyltransferase activities in the endometrium of ovariectomized rats.

Some of the properties of galactosyl- and sialyltransferases present in rat endometrial tissue were investigated. The enzyme activities were found to be partly membrane-bound and partly in soluble form. The galactose enzyme was also present in uterine secretions. The specific activities of both galactosyl- and sialyltransferases were greatly enhanced in endometrium of ovariectomized rats following 17 β-estradiol injections, although the enzyme activities in the liver remained unaffected. Mixing experiments with the homogenates of endometrium from control and estradiol-treated rats failed to suggest the presence of any “activator” or “inhibitor” of the enzymes. Diethylstilbesterol, estrone, and estriol also stimulated galactosyl- and sialyltransferase activities, whereas testosterone stimulated sialyltransferase only. Prolactin administration had no effect on either of the enzymes. The effect of estradiol on both enzymes was shown to be dose-dependent and the specific activities of the enzymes started to increase about 6 hr after hormone administration, reaching a peak around 48 h. Progesterone, on its own, had no effect on the galactosyltransferase in ovariectomized rat endometrium but effectively prevented the stimulatory effect of estradiol. When estradiol-primed rats were treated with progesterone, it was found that very small doses of progesterone resulted in decrease of galactosyltransferase activity. In such animals sialyltransferase activity was stimulated by a low concentration of progesterone which was followed by inhibition at higher concentrations. These effects of ovarian hormones on glycosyltransferase activities in endometrium are compatible with earlier reports on the effects of these hormones on glycoprotein and glycosaminoglycan levels in rabbit uterus after ovariectomy (7). Regulation of glycosyltransferase activities in endometrium induced by estradiol and progesterone may bear some relationship to the “receptive” state of the uterus for blastocyst implantation.     

Differential effects of fatty acids and exercise on body weight regulation and metabolism in female Wistar rats

High-fat diets made with different fats may have distinct effects on body weight regulation and metabolism. In the present study, the metabolic effects of high-fat (HF) diets made with fish oil, palm oil, and soybean oil were compared with a low-fat diet in female Wistar rats that were either exercised (EX, swimming) or that remained sedentary as controls. Each adult rat was exposed to the same diet that their dams consumed during pregnancy and lactation. When they were 9 weeks old, rats began an EX regimen that lasted for 6 weeks. Twenty-four hours after the last EX bout, rats were sacrificed in a fasted state. It was observed that HF feeding of soybean oil induced more body weight and fat gain, as well as insulin resistance, as indicated by insulin/glucose ratios, than other oils. Female rats fed a HF diet made with fish oil had body weight and insulin sensitivity not different from that observed in low fat fed control rats. For rats fed HF diets made with soybean oil or palm oil, EX also exerted beneficial effects by reducing body fat %, blood insulin, triglyceride and leptin levels, as well as improving insulin sensitivity.     

Regulation of sociosexual communication in female Long-Evans rats by ovarian hormones

Two experiments examined the role of the steroid hormones, estradiol (E2), progesterone (P), and testosterone (T), in activating scent marking and 50-kHz ultrasonic vocalizations in ovariectomized Long-Evans rats in response to a devocalized male rat. In Experiment 1, females received, in a counterbalanced order, a series of six hormone treatments consisting of two injections (48-54 and 4 hr before behavioral tests). The six treatments were 8 micrograms E2 followed by 500 micrograms P or oil, 2 micrograms E2 followed by 500 micrograms P or oil, and oil followed by 500 micrograms P or oil. Injections of either the high or low dose of E2 followed by P resulted in high levels of vocalizations. Neither E2 by itself or P by itself were very effective. Surprisingly, none of the hormone treatments were effective in activating marking above the level seen when the females received control injections of oil. Four other hormone treatments were examined in Experiment 2: daily injections of 500 micrograms T, daily injections of 50 micrograms E2, implantation of silastic capsules of E2 (5% E2, 5 mm length) followed by 500-micrograms P injections before behavioral tests, and implantation of silastic capsules of E2 followed by oil injections. Animals receiving E2 by silastic capsule followed by P injection displayed the highest levels of marking and vocalizations across the five weekly tests. These results suggest that while E2 and P synergize for the display of female-typical behaviors similar to the hormonal regulation of lordosis, the mechanism of E2 action may be different for the two signaling behaviors. Scent marking appears to be responsive to the tonic levels of E2 released from silastic capsules.     

Daily exercise increases hepatic fatty acid oxidation and prevents steatosis in Otsuka Long-Evans Tokushima Fatty rats

Exercise training is commonly prescribed for treatment of nonalcoholic fatty liver disease (NAFLD). We sought to determine whether exercise training prevents the development of NAFLD in Otsuka Long-Evans Tokushima Fatty (OLETF) rats and to elucidate the molecular mechanisms underlying the effects of exercise on hepatic steatosis. Four-week-old OLETF rats were randomly assigned to either a sedentary control group (Sed) or a group given access to voluntary running wheels for 16 wk (Ex). Wheels were locked 2 days before euthanasia in the Ex animals, and both groups were euthanized at 20 wk old. Voluntary wheel running attenuated weight gain and reduced serum glucose, insulin, free fatty acids, and triglycerides in Ex animals compared with Sed (P < 0.001). Ex animals exhibited significantly reduced hepatic triglyceride levels and displayed fewer lipid droplets (Oil Red O staining) and reduced lipid droplet size compared with Sed. Wheel running increased by threefold the percent of palmitate oxidized completely to CO(2) in the Ex animals but did not alter AMP-activated protein kinase-alpha (AMPKalpha) or AMPK phosphorylation status. However, fatty acid synthase and acetyl-coenzyme A carboxylase (ACC) content were significantly reduced (approximately 70 and approximately 35%, respectively), and ACC phosphorylation and cytochrome c content were significantly elevated (approximately 35 and approximately 30%, respectively) in the Ex animals. These results unequivocally demonstrate that daily physical activity attenuates hepatic steatosis and NAFLD in an obese rodent model and suggest that this effect is likely mediated, in part, through enhancement of hepatic fatty acid oxidation and reductions in key protein intermediates of fatty acid synthesis.     

Age-associated blood glucose levels during exercise in female swimmers

The purpose of this study was to determine whether age-associated alterations in blood glucose levels occur during exercise. In addition, blood lactate and fitness levels (VO2max) were examined to ascertain if these factors influenced the age-related responses. Sixty-four female masters swimmers (25-75 years) were classified into either a well trained (WT) or recreational trained (RT) group and exercised on a treadmill to VO2max X VO2max data confirmed our classification of WT and RT swimmers based on activity levels. There were no differences in post-absorptive blood glucose and lactate levels across age and fitness. Significant age-related effects on blood glucose levels during exercise (p less than 0.01) but no fitness effect were revealed by ANOVA. Within the first or second exercise stage all age groups demonstrated a significant decline in blood glucose (6.3 to 14.1%). A hyperglycemic response was observed during recovery in all age groups with the exception of the over 60 (60+) group. The 60+ group exhibited lower blood glucose levels compared to all other age groups during exercise commencing with the second exercise stage. There were no significant differences in glucose levels among any of the other age groups during exercise or recovery. There were no age-related differences in maximal or recovery lactates. These data indicate that there is an alteration of blood glucose homeostasis during exercise in females over 60 years of age.     

Cardioprotective effects of exercise training on myofilament calcium activation in ovariectomized rats.

The risks associated with hormone replacement therapy, especially cardiac diseases in postmenopausal women, have prompted extensive studies for other preventive or therapeutic alternatives. We investigated the cardioprotective effects of exercise training on the changes in cardiac myofilament Ca2+ activation in 10-wk-old ovariectomized rats. The exercise groups were subjected to a 9-wk running program on a motor-driven treadmill 1 wk after surgery. The relationship between pCa (-log molar free Ca2+ concentration) and myofibrillar MgATPase activity of exercise-sham myofibrils or exercise-ovariectomized myofibrils was the same and could not be distinguished from that of sedentary-sham control hearts. In contrast, a significant suppression in maximum MgATPase activity and a leftward shift of pCa50 (half-maximally activating pCa) in the pCa-ATPase activity relationship were detected in sedentary-ovariectomized rats. Exercise training also prevented the shift in myosin heavy chain (MHC) isoforms toward beta-MHC in ovariectomized hearts. The upregulation of beta1-adrenergic receptors in the left ventricular membranes of ovariectomized rat hearts, as measured by receptor binding and immunoblot analyses, was no longer observed in exercise-ovariectomized hearts. Immunoblot analyses of heat shock protein (HSP) 72, an inducible form of HSP70, demonstrated a significant downregulation in ovariectomized hearts. Exercise training in ovariectomized rats completely reversed the expression of HSP72 to the same level as sham controls. Our results clearly indicate the cardioprotective effects of exercise training on changes in cardiac myofilament Ca2+ activation in ovariectomized rats. Alterations in expression of beta1-adrenergic receptors and HSP72 may, in part, play a mechanistic role in the cardioprotective effects.     

Ca2+ effects on glucose transport and fatty acid oxidation in L6 skeletal muscle cell cultures

We examined the effect of Ca²⁺ on skeletal muscle glucose transport and fatty acid oxidation using L6 cell cultures. Ca²⁺ stimulation of glucose transport is controversial. We found that caffeine (a Ca²⁺ secretagogue) stimulation of glucose transport was only evident in a two-part incubation protocol (“post-incubation”). Caffeine was present in the first incubation, the media removed, and labeled glucose added for the second. Caffeine elicited a rise in Ca²⁺ in the first incubation that was dissipated by the second. This post-incubation procedure was insensitive to glucose concentrations in the first incubation. With a single, direct incubation system (all components present together) caffeine caused a slight inhibition of glucose transport. This was likely due to caffeine induced inhibition of phosphatidylinositol 3-kinase (PI3K), since nanomolar concentrations of wortmannin, a selective PI3K inhibitor, also inhibited glucose transport, and previous investigators have also found this action.We did find a Ca²⁺ stimulation (using either caffeine or ionomycin) of fatty acid oxidation. This was observed in the absence (but not the presence) of added glucose. We conclude that Ca²⁺ stimulates fatty acid oxidation at a mitochondrial site, secondary to malonyl CoA inhibition (represented by the presence of glucose in our experiments). In summary, the experiments resolve a controversy on Ca²⁺ stimulation of glucose transport by skeletal muscle, introduce an important experimental consideration for the measurement of glucose transport, and uncover a new site of action for Ca²⁺ stimulation of fatty acid oxidation.     

Myocardial fatty acid oxidation during ischemia and reperfusion

Inhibition of fatty acid oxidation is an early event in myocardial ischemia that most likely contributes to tissue injury by the accumulation of potentially toxic intermediates such as acylCoA and acylcarnitine. After reperfusion both myocardial oxygen consumption and fatty acid oxidation may rapidly recover to preischemic levels, even when contractile function remains depressed. The mechanisms underlying the apparent dissociation between contractile function and oxidative metabolism early during reperfusion are still controversial. In isolated rat hearts subjected to 60 min of no-flow ischemia myocardial oxygen consumption and oxidation of palmitate were lowered during reperfusion by 3 mM of NiCl₂ and by 6 µM of ruthenium red. The results provide indirect evidence for the hypothesis that intracellular calcium transport may be involved in the mechanisms responsible for the high oxidative metabolic rate early after reperfusion     

Suppressive effects of genistein dosage and resistance exercise on bone loss in ovariectomized rats.

This study was designed to determine whether combined treatments with genistein dosage and moderate resistance exercise would exhibit synergistically preventive effects on bone loss following the onset of menopause. Forty-one 12 wk-old female SD rats were assigned to five groups: 1) Sham operated (Sham); 2) ovariectomized (OVX-Cont); 3) OVX received genistein (OVX-GEN); 4) OVX exercised (OVX-EXE); and 5) OVX treated with both genistein and exercise (OVX-GEN-EXE). All rats were fed a low Ca (0.1%) diet ad libitum. Daily genistein dosage was 12 mg/kg body weight. Exercising rats took 40 sets of 1-min run interspersed with 1-min rest with a 100 g weight on the back on an uphill treadmill at 20 m/min. The experimental duration consisted of the adaptation and treatment periods of 4 weeks each. Uterine weight in OVX-Cont, OVX-GEN, OVX-EXE and OVX-GEN-EXE decreased to about 15% of that in Sham (p < 0.001). The femoral BMD (mg/cm2; mean +/- SE), assessed by DEXA (Lunar), of OVX-Cont was significantly lowered to 206 +/- 5 by -9%, as compared to 226 +/- 2 of Sham (p < 0.001). The BMD of OVX-GEN, OVX-EXE and OVX-GEN-EXE were 217 +/- 2, 217 +/- 2 and 222 +/- 2, respectively, and genistein dosage and resistance exercise equally increased the BMD of OVX rats by 5% (p < 0.01). Combined treatment of genistein and exercise more successfully recovered their decreased BMD by 8% (p < 0.001). BMD of the fourth lumbar vertebrae in OVX-Cont was declined to 191 +/- 7 by -15%, as compared to 225 +/- 4 in Sham (p < 0.001). OVX-EXE and OVX-GEN-EXE gained the BMD by 6% to 205 +/- 4 and 203 +/- 3, respectively, as compared to that of OVX-Cont (p < 0.01). These results suggest the possibility that the combined treatment of genistein dosage and resistance exercise have more beneficial effects by acting rather independently than their separate trials on the prevention of ovx-induced bone loss in femurs.     

Regulation of plasma long-chain fatty acid oxidation in relation to uptake in human skeletal muscle during exercise

In the present study, we investigated possible sites of regulation of long-chain fatty acid (LCFA) oxidation in contracting human skeletal muscle. Leg plasma LCFA kinetics were determined in eight healthy men during bicycling (60 min, 65% peak oxygen uptake) with either high (H-FOX) or low (L-FOX) leg fat oxidation (H-FOX: 1,098 +/- 140; L-FOX: 494 +/- 84 micromol FA/min, P < 0.001), which was achieved by manipulating preexercise muscle glycogen (H-FOX: 197 +/- 21; L-FOX: 504 +/- 25 mmol/kg dry wt, P < 0.001). Several blood metabolites and hormones were kept nearly similar between trials by allocating a preexercise meal and infusing glucose intravenously during exercise. During exercise, leg plasma LCFA fractional extraction was identical between trials (H-FOX: 17.8 +/- 1.6; L-FOX: 18.2 +/- 1.8%, not significant), suggesting similar LCFA transport capacity in muscle. On the contrary, leg plasma LCFA oxidation was 99% higher in H-FOX than in L-FOX (421 +/- 47 vs. 212 +/- 37 micromol/min, P < 0.001). Probably due to the slightly higher (P < 0.01) plasma LCFA concentration in H-FOX than in L-FOX, leg plasma LCFA uptake was nonsignificantly (P = 0.17) higher (25%) in H-FOX than in L-FOX, yet the fraction of plasma LCFA uptake oxidized was 61% higher (P < 0.05) in H-FOX than in L-FOX. Accordingly, the muscle content of several lipid-binding proteins did not differ significantly between trials, although fatty acid translocase/CD36 and caveolin-1 were elevated (P < 0.05) by the high-intensity exercise and dietary manipulation allocated on the day before the experimental trial. The present data suggest that, in contracting human skeletal muscle with different fat oxidation rates achieved by manipulating preexercise glycogen content, transsarcolemmal transport is not limiting plasma LCFA oxidation. Rather, the latter seems to be limited by intracellular regulatory mechanisms.     

Effect of glucose on plasma glucagon and free fatty acids during prolonged exercise

The effects of glucose ingestion on the changes in blood glucose, FFA, insulin and glucagon levels induced by a prolonged exercise at about 50% of maximal oxygen uptake were investigated. Healthy volunteers were submitted to the following procedures: 1. a control test at rest consisting of the ingestion of 100 g glucose, 2. an exercise test without, or 3. with ingestion of 100 g of glucose. Exercise without glucose induced a progressive decrease in blood glucose and plasma insulin; plasma glucagon rose significantly from the 60th min onward (+45 pg/ml), the maximal increase being recorded during the 4th h of exercise (+135 pg/ml); plasma FFA rose significantly from the 60th min onward and reached their maximal values during the 4th h of exercise (2177 +/- 144 muEq/l, m +/- SE). Exercise with glucose ingestion blunted almost completely the normal insulin response to glucose. Under these conditions, exercise did not increase plasma glucagon before the 210th min; similarly, the exercise-induced increase in plasma FFA was markedly delayed and reduced by about 60%. It is suggested that glucose availability reduces exercise-induced glucagon secretion and, possibly consequently, FFA mobilization.