Acid phosphatase and arylsulphatase activities in testes after AET or MEA treatment of adult mice.

Temporal changes of acid phosphatase (E.C. 3.1.3.2) and arylsulphatase (E.C. 3.1.6.1) activities in testes of adult Swiss mice after AET (2-amino-ethylisothiouronium Br. HBr) or MEA (cysteamine HCl) treatment, were studied. The animals were injected intraperitoneally with the S-containing substances in a single dose of 400 mg/kg body weight. The enzyme activities in crude organ homogenates were assessed every four hours during a 24-hour period. Administration of the aminothiol agents to mouse organism caused greater changes in the acid phosphatase activity than in the arylsulphatase activity, and the two chemical compounds AET and MEA given, influenced the enzyme activities in testes in a different way. Treatment of mice with AET resulted in a decrease of the acid phosphatase activity related to 1 g of fresh tissue at 16.00 and the whole organ weight at 24.00 and 16.00 as well as in a decrease of the arylsulphatase activity expressed per the whole weight of testes at 08.00. After MEA injection, the acid phosphatase activity related to 1 mg of protein, 1 g of fresh tissue and the whole organ weight was decreased at 20.00(1), and the enzyme activity expresse per 1 mg of protein and 1 g of fresh tissue was increased at 24.00, but the arylsulphatase activity related to both 1 mg of protein at 08.00, 12.00 and to the whole weight of testes at 08.00, was reduced.     

Quantity of DNA in the organs of adult Swiss male mice after AET and MEA treatment

Adult Swiss male mice were injected intraperitoneally with 2-aminoethylesothiouronium bromide hydrobromide (AET) or cysteamine hydrochloride (MEA) in a dose of 400 mg/kg body weight. In the thirtieth, sixtieth or ninetieth minute after the injection, the animals were killed and the deoxyribonuclein acid content in 100 mg of fresh tissue of testes, spleen and liver, was measured. DNA was extracted from the organs by means of Burton's method, which is based on the estimation of deoxiribose content in the colour reaction with diphenylamine. The injection of AET and MEA did not distinctly influence the DNA content in the organs of mice. Statistically significant differences among the groups of mice were not observed compared to the controls, in mice treated with the compound, a decreasing tendency in the quantity of the DNA in the organs was found only.     

Temporary changes of protein level in liver after AET or MEA treatment prior to 60Co gamma-irradiation of male mice

The adult male Swiss mice were either whole-body gamma-irradiated with a single dose of 10 Gy from 60Co source, always at 19.00 or, 15 minutes before irradiation injected intraperitoneally with AET (2-aminoethylisothiouronium Br. HBr), or MEA (cysteamine HCl), in a dose of 400 mg/kg body weight. The measurements of the protein level in crude homogenates of liver were done in four-hour internals during a 24-hour period, starting at 20.00. The protein concentration in liver was calculated per 1 g of fresh tissue and the whole organ weight. The body and liver weight was also studied. There were no fluctuations in the liver weight and concentration of protein in the control and irradiated only mice. Temporary changes in the liver weight and level of protein expressed in mg per 1 g of fresh tissue and the whole organ weight could be found in the group of males treated with AET, and daily changes in the liver weight and concentration of protein related to mg per 1 g of fresh tissue, in the group of male mice injected with MEA prior to irradiation, could be recorded. Differences in the liver weight at 20.00, 24.00 and 04.00, as well as in the protein level expressed in mg per 1 g fresh tissue at 04.00, 12.00, 16.00, and the whole liver weight at 24.00, 04.00, and 16.00, of between the particular groups of mice, were observed. There were no temporary changes in the body weight in any of the groups and there were no differences in this value between the groups of mice.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of AET and MEA treatment of mice on the first day of pregnancy

Porton female mice were injected intraperitoneally with 2-aminoethylisothiouronium bromide hydrobromide (AET) or cysteamine hydrochloride (MEA) in a dose of 40 mg/kg of body weight on the first day of pregnancy. On the last, nineteenth, day of gestation, taking into consideration females in whose uterus live fetuses were observed, the increase in their body weight throughout pregnancy, the number of fetuses in the uterus, the body weight of fetuses, and placental weight were found smaller in mice treated with AET or MEA, than in control ones. Among the injected compounds, AET appeared to be less toxic than MEA.     

Quantity of glycogen in the liver of 19-day-old foetuses after AET, 5-HT, MEA, or GSH treatment of pregnant mice on the first day of gestation

Swiss mice were treated intraperitoneally with AET, 5-HT, MEA, or GSH, in a dose of 80 mg/kg of body weight, on the first day of gestation. On the 19th day of pregnancy, the fresh weight of liver of the foetuses, as well as glycogen content in 1 g of fresh tissue and in the whole organ were analysed. The determination of glycogen content in the foetal liver were made according to the anthrone method. As compared with controls, in the remaining groups of mice a lower fresh weight of foetal liver less glycogen per g of fresh tissue and a smaller total amount of glycogen in the whole organ were found. Among the compounds, AET appeared to be more toxic than 5-HT, MEA, and GSH.     

Glycogen metabolism in white and red muscle or normal and diabetic rats. The glycogen concentration and glycogen synthesis from glucose.

The amount of glycogen and its synthesis from glucose was studied in white muscle (extensor digitorum longus -- EDL) and red muscle (soleus -- SOL) of normal rats and rats with alloxan diabetes by the anthrone method. The amount of glycogen was higher in the white muscle of normal rats, both after a 24 hours' fast (0.37+/-0.02 mg/g as against 0.29+/-0.01 mg/g in the SOL) and with feeding ad libitium (0.72+/-0.05 mg/g as against 0.58+/-0.03 mg/g in the SOL). After a 24 hours' fast, the glycogen content of both muscles was non-significantly higher in alloxan-diabetic rats than in normal animals, whereas in diabetic animals fed ad libitum it was significantly lower than in normal rats fed in the same manner (0.54+/-0.07 mg/g in the EDL and 0.33+/-0.03 mg/g in the SOL). The difference between the glycogen content of the white and red muscle of diabetic rats was also in favour of the white muscle. Muscle glycogenesis from intragastrically administered glucose was higher in the red muscle in all the experimental groups. In normal fed ad libitum the glycogen content of the EDL did not change after glucose administration, but in the SOL it rose from 0.58+/-0.03 to 0.83+/-0.05 mg/g. In fasting (24 hours) normal rats it rose sharply in both muscles, from 0.037+/-0.02 to 0.57+/-0.03 mg/g in the EDL and from 0.29+/-0.01 to 0.87+/-0.06 mg/g in the SOL. In fasting (24 hours) diabetic animals, the glycogen content rose after glucose in the SOL only, from 0.36+/-0.01 to 0.66+/-0.06 mg/g. The differences found in glycogen synthesis in the white and red muscle of normal and diabetic rats are discussed mainly from the aspect of the existence of a relationship between the glycogen concentration and glycogen synthetase activity.     

Quantity of DNA in the organs of 19-day-old foetuses after AET, MEA, or 5-HT treatment of mice on the first day of gestation

On the first day of gestation, Porton mice were injected intraperitoneally with AET (2-aminoethylisothiouronium bromide hydrobromide), MEA (cysteamine hydrochloride,) or 5-HT (serotonin-creatinine sulphate), in a dose of 40 mg/kg of bodyweight. On the nineteenth day of pregnancy, the fresh weight of both heart and kidneys of foetuses, as well as DNA content in 25 mg of fresh tissue and in these whole organs were analysed. DNA was extracted from the foetal organs by means of Burton's method, which is based on the estimation of deoxiribose content in the colour reaction with diphenylamine. As compared to controls, in the remaining groups of mice lower fresh weight of both heart and kidneys of foetuses, greater DNA content in 25 mg of fresh tissue and smaller total amounts of DNA in the whole organs were found. Among the experimental groups of mice, statistically significant differences in the analysed values were observed between the group of animals treated with 5-HT and the remaining groups, with the exception of statistically non-significant difference in the DNA content of the whole kidneys between those injected with 5-HT and MEA.     

Embryonic survival in mice treated with AET, MEA or 5-HT on the first day of pregnancy

The embryotoxicity of AET, MEA, and 5-HT was investigated in Porton mice. Female mice on the first day of gestation were injected intraperitoneally with 2-amino-ethylisothiouronium bromide hydrobromide (AET), cysteamine hydrochloride (MEA) or serotonin-creatinine sulphate (5-HT) in a dose of 40 mg/kg body weight. Uterine contents were examined on the nineteenth day of pregnancy. As compared with controls, in mice treated with AET, MEA or 5-HT, a smaller number of live fetuses and a greater number of non-implanted embryos, resorptions, and dead fetuses were found. Not all females which were injected with these compounds had live fetuses. Among the compounds, MEA appeared to be more toxic than AET and 5-HT.     

Levodopa with carbidopa diminishes glycogen concentration, glycogen synthase activity, and insulin-stimulated glucose transport in rat skeletal muscle.

We hypothesized that levodopa with carbidopa, a common therapy for patients with Parkinson's disease, might contribute to the high prevalence of insulin resistance reported in patients with Parkinson's disease. We examined the effects of levodopa-carbidopa on glycogen concentration, glycogen synthase activity, and insulin-stimulated glucose transport in skeletal muscle, the predominant insulin-responsive tissue. In isolated muscle, levodopa-carbidopa completely prevented insulin-stimulated glycogen accumulation and glucose transport. The levodopa-carbidopa effects were blocked by propranolol, a beta-adrenergic antagonist. Levodopa-carbidopa also inhibited the insulin-stimulated increase in glycogen synthase activity, whereas propranolol attenuated this effect. Insulin-stimulated tyrosine phosphorylation of insulin receptor substrate (IRS)-1 was reduced by levodopa-carbidopa, although Akt phosphorylation was unaffected by levodopa-carbidopa. A single in vivo dose of levodopa-carbidopa increased skeletal muscle cAMP concentrations, diminished glycogen synthase activity, and reduced tyrosine phosphorylation of IRS-1. A separate set of rats was treated intragastrically twice daily for 4 wk with levodopa-carbidopa. After 4 wk of treatment, oral glucose tolerance was reduced in rats treated with drugs compared with control animals. Muscles from drug-treated rats contained at least 15% less glycogen and approximately 50% lower glycogen synthase activity compared with muscles from control rats. The data demonstrate beta-adrenergic-dependent inhibition of insulin action by levodopa-carbidopa and suggest that unrecognized insulin resistance may exist in chronically treated patients with Parkinson's disease.     

Liver glycogen synthase but not the muscle isoform differentiates between glucose 6-phosphate produced by glucokinase or hexokinase

Using adenovirus-mediated gene transfer into FTO-2B cells, a rat hepatoma cell line, we have overexpressed hexokinase I (HK I), glucokinase (GK), liver glycogen synthase (LGS), muscle glycogen synthase (MGS), and combinations of each of the two glucose-phosphorylating enzymes with each one of the GS isoforms. FTO-2B cells do not synthesize glycogen even when incubated with high doses of glucose. Adenovirus-induced overexpression of HK I and/or LGS, two enzymes endogenously expressed by these cells, did not produce a significant increase in the levels of active GS and the total glycogen content. In contrast, GK overexpression led to the glucose-dependent activation of endogenous or overexpressed LGS and to the accumulation of glycogen. Similarly overexpressed MGS was efficiently activated by the glucose-6-phosphate (Glc-6-P) produced by either endogenous or overexpressed HK I and by overexpressed GK. These results indicate the existence of at least two pools of Glc-6-P in the cell, one of them is accessible to both isoforms of GS and is replenished by the action of GK, whereas LGS is excluded from the cellular compartment where the Glc-6-P produced by HK I is directed. These findings are interpreted in terms of the metabolic role that the two pairs of enzymes, HK I-MGS in the muscle and GK-LGS in the hepatocyte, perform in their respective tissues.     

Postexercise glucose uptake and glycogen synthesis in skeletal muscle from GLUT4-deficient mice.

To determine the role of GLUT4 on postexercise glucose transport and glycogen resynthesis in skeletal muscle, GLUT4-deficient and wild-type mice were studied after a 3 h swim exercise. In wild-type mice, insulin and swimming each increased 2-deoxyglucose uptake by twofold in extensor digitorum longus muscle. In contrast, insulin did not increase 2-deoxyglucose glucose uptake in muscle from GLUT4-null mice. Swimming increased glucose transport twofold in muscle from fed GLUT4-null mice, with no effect noted in fasted GLUT4-null mice. This exercise-associated 2-deoxyglucose glucose uptake was not accompanied by increased cell surface GLUT1 content. Glucose transport in GLUT4-null muscle was increased 1.6-fold over basal levels after electrical stimulation. Contraction-induced glucose transport activity was fourfold greater in wild-type vs. GLUT4-null muscle. Glycogen content in gastrocnemius muscle was similar between wild-type and GLUT4-null mice and was reduced approximately 50% after exercise. After 5 h carbohydrate refeeding, muscle glycogen content was fully restored in wild-type, with no change in GLUT4-null mice. After 24 h carbohydrate refeeding, muscle glycogen in GLUT4-null mice was restored to fed levels. In conclusion, GLUT4 is the major transporter responsible for exercise-induced glucose transport. Also, postexercise glycogen resynthesis in muscle was greatly delayed; unlike wild-type mice, glycogen supercompensation was not found. GLUT4 it is not essential for glycogen repletion since muscle glycogen levels in previously exercised GLUT4-null mice were totally restored after 24 h carbohydrate refeeding.-Ryder, J. W., Kawano, Y., Galuska, D., Fahlman, R., Wallberg-Henriksson, H., Charron, M. J., Zierath, J. R. Postexercise glucose uptake and glycogen synthesis in skeletal muscle from GLUT4-deficient mice.     

Post-exercise glucose uptake and glycogen synthesis in human muscle during oral or IV glucose intake

Post-exercise muscle uptake and the intracellular fate of glucose was studied after oral or intravenous glucose administrations which caused similar plasma glucose concentrations, but high and moderate plasma insulin concentrations, respectively. Five male subjects participated in two experiments with 6-16 weeks in between. In the first experiment, the oral glucose experiment (OG), 1.4, 0.7 and 0.7 g.kg-1 body mass of glucose was given as oral loads at 0, 1 and 2 h of a 3-h post-exercise observation period (3hOP). In the second experiment, the glucose clamp experiment (GC), a glucose infusion clamp technique was employed. Based on repetitive, immediate plasma glucose measurements performed every 5th min, the rate of glucose infusion was adjusted to obtain the same temporal pattern of the plasma glucose concentration as in OG. The average plasma glucose concentrations during 3hOP were 9.2 +/- 1.1 and 9.3 +/- 1.2 mmol.1(-1) in OG and GC, respectively. The average arterio-femoral venous (a-v)f glucose differences were 1.0 +/- 0.3 and 0.5 +/- 0.2 mmol.1(-1) (p less than 0.001), while the average plasma insulin concentrations were 56 +/- 12 and 26 +/- 5 microU.ml-1 (p less than 0.001) for the two experiments. Increases in muscle glycogen concentrations were 28 +/- 4 and 25 +/- 3 mmol.kg-1 (NS) during 3hOP in OG and GC, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of ginger extract ingestion on skeletal muscle glycogen contents and endurance exercise in male rats

[Purpose]Skeletal muscle glycogen is a determinant of endurance capacity for some athletes. Ginger is well known to possess nutritional effects, such as anti-diabetic effects. We hypothesized that ginger extract (GE) ingestion increases skeletal muscle glycogen by enhancing fat oxidation. Thus, we investigated the effect of GE ingestion on exercise capacity, skeletal muscle glycogen, and certain blood metabolites in exercised rats. [Methods]First, we evaluated the influence of GE ingestion on body weight and elevation of exercise performance in rats fed with different volumes of GE. Next, we measured the skeletal muscle glycogen content and free fatty acid (FFA) levels in GE-fed rats. Finally, we demonstrated that GE ingestion contributes to endurance capacity during intermittent exercise to exhaustion. [Results]We confirmed that GE ingestion increased exercise performance (p<0.05) and elevated the skeletal muscle glycogen content compared to the non-GE-fed (CE, control exercise) group before exercise (Soleus: p<0.01, Plantaris: p<0.01, Gastrocnemius: p<0.05). Blood FFA levels in the GE group were significantly higher than those in the CE group after exercise (p<0.05). Moreover, we demonstrated that exercise capacity was maintained in the CE group during intermittent exercise (p<0.05). [Conclusion]These findings indicate that GE ingestion increases skeletal muscle glycogen content and exercise performance through the upregulation of fat oxidation.     

Crowding or ACTH treatment of pregnant mice affects adult copulatory behavior of male offspring

Sexual behavior of male offspring from female mice chronically crowded during Days 12-17 of pregnancy was investigated. In an 80-min test pairing with a sexually experienced female primed with estradiol and progesterone injections, males from crowded mothers displayed poorer copulation than controls: mount and intromission latencies were longer, number of mounts and intromissions lower, and ejaculations within the test period were abolished. Daily injections of 500 micrograms testosterone propionate improved copulatory vigor in offspring from crowded mothers. A second series of experiments investigated the effects of ACTH treatment of females during the same period of pregnancy. A low dose rate (1 IU injected daily) had little effect but male offspring from females injected daily with 8 IU displayed longer intromission latency and fewer mounts and intromissions than controls. Daily injections of 500 micrograms testosterone propionate improved copulatory vigor, although mount frequency remained depressed. The similarity of the effects on male offspring copulation of crowding their mothers during pregnancy or ACTH treatment during pregnancy suggest mediation by similar mechanisms, implicating involvement of maternal pituitary-adrenocortical secretions during pregnancy in the production of these behavioral deficits. Postnatal influences were minimized by fostering all litters at birth to untreated dams.     

Effect of chronic metformin treatment of hepatic and muscle glycogen metabolism in KK mice.

Noninsulin-dependent diabetic KK mice, aged 90-100 days, with hyperinsulinemia and insulin resistance were treated with either metformin (N = 13) or water (control, N = 10) orally at a concentration of 50 mg/kg twice daily for 28 weeks. Age-matched nondiabetic Swiss Webster (SW) mice were also similarly treated. Liver and skeletal muscle glycogen synthase and phosphorylase enzymes were determined in all groups of mice. Both enzymes were significantly lower in control KK than in control SW mice. Metformin did not influence either of these enzymes in nondiabetic SW mice. However, it significantly increased the active form of glycogen synthase (a form) in both the liver and muscle of KK mice. Metformin also increased the active form of phosphorylase (a form) in the liver but not in the muscle of these mice. Hepatic glycogen content was similar in both control and metformin-treated KK mice. However, the muscle glycogen content was significantly higher in metformin-treated than in control KK mice. These data suggest that metformin preferentially stimulates glycogen synthesis in skeletal muscle, and this seems to be responsible for the observed improvement in fasting glucose and glucose response to an oral glucose load in KK mice.     

Effects of AET and MEA on the lysosomal hydrolase activities in the mouse organs.

The adult male Swiss mice were injected intraperitoneally with AET (2-aminoethylisothiouronium Br.HBr) or MEA (cysteamine HCl), in a toxic dose of 400 mg/kg body weight. The acid phosphatase (E.C. 3.1.3.2) and arylsulphatase (E. C. 3.1.6.1) activities in crude homogenates of liver and kidneys were assessed every fourth hour throughout a 24-h period. Different patterns of temporal changes in the acid phosphatase and arylsulphatase activities in liver and kidneys expressed in nkat per 1 mg of protein, 1 g of fresh tissue and per the whole organ weight, were found. The extent and timing of the alterations in the activity of each of the lysosomal hydrolases were dependent on the particular organ chosen and aminothiol compound given.