Interaction of androgens and prolactin on prostatic enzymes of the pyruvate-malate cycle involved in lipogenesis in castrated mature monkey, Macaca radiata.

Effects of androgens, prolactin (Prl) and bromocriptine (Br) on the specific activities of prostatic (caudal and cranial) enzymes of the pyruvate-malate cycle were studied in castrated mature bonnet monkeys. Castration decreased the activity of NADP+ isocitrate dehydrogenase (ICDH), ATP citrate lyase, malate dehydrogenase (MDH), malic enzyme and fatty acid synthase (FAS). Administration of testosterone propionate (TP)/dihydrotestosterone (DHT) increased the activities of all these enzymes in both lobes. Malate dehydrogenase maintained normal activity. Prl also had a stimulatory effect on the enzymes and was further enhanced when Prl was given in combination with TP/DHT. Unlike Prl, bromocriptine treatment inhibited all the enzymes in both lobes. Thus, prolactin was found to have a direct as well as a synergistic effect with androgens on enzymes of the pyruvate-malate cycle in the prostate of castrated mature monkeys.     

Alpha-Glycerophosphate dehydrogenase (insoluble) and lactic dehydrogenase activities in the skeletal muscles of two insects.

The flight muscle preparations of the dragonfly Pantala flavescens and the aquatic beetle Cybister confusus showed extremely low levels of lactic dehydrogenase activity and high levels of alpha-glycerophosphate dehydrogenase (insoluble) activity. The activities of these two enzymes in the leg muscle of the beetle were approximately the same (1:1), but lactic dehydrogenase activity was several times higher than that in the flight muscles of both Insects. These results have been interpreted as indicating the high energy-yielding demands of the flight muscles during continuous sustained activity, while the leg muscles of the beetle which are involved in swimming activity derive their energy predominantly through anaerobic glycolysis.     

Activity patterns of phosphofructokinase,glyceraldehydephosphate dehydrogenase,lactate dehydrogenase and malate dehydrogenase in microdissected fast and slow fibres from rabbit psoas and soleus muscle

Summary Methods for standardized determination of phosphofructokinase (PFK), glyceraldehydephosphate dehydrogenase (GAPDH), lactate dehydrogenase (LDH) and malate dehydrogenase (MDH) activities in nanogram samples of microdissected single fibres of rabbit psoas and soleus muscle are described. Fast and slow fibres in soleus muscle show lower absolute activities of these enzymes than the respective fibre types in psoas muscle. Slow fibres represent a more uniform population in the two muscles according to absolute and relative activities of the enzymes investigated. Slow fibres are characterized by high activities of MDH and relatively low activities of glycolytic enzymes. Fast fibres in the soleus muscle represent a population with high activities of MDH and glycolytic enzymes. Fast fibres in psoas muscle represent a heterogeneous population with high activities of glycolytic enzymes and extremely variable activity of MDH. More than 10-fold differences exist in the MDH activities of the extreme types of this fibre population. Differences in the activity levels of MDH in single fast type fibres but also in the activities of glycolytic enzymes between fast and slow fibres are greater than those reported between extreme white and red rabbit muscles.     

Phosphamidon, methylparathion and dichlorvos impact on tissue oxidative metabolism in penaeid prawn, Metapenaeus monoceros

Changes in oxidative metabolism of hepatopancreas and muscle tissues of penaeid prawn, Metapenaeus monoceros was studied, following its exposure to selected organophosphorous insecticides phosphamidon, dichlorovos and methylparathion. The OPI are found to inhibit the activity levels of acetylcholinesterase, succinate dehydrogenase, isocitrate dehydrogenase, pyruvate dehydrogenase, lactate dehydrogenase and cytochrome-c-oxidase and cause accumulation of acetylcholine in the hepatopancreas and muscle tissues. These changes in the activity levels of selected oxidative enzymes during insecticide exposure in these tissues of prawn indicates the shift in the metabolic emphasis from aerobic to anaerobic conditions and is interpreted as a functional adaptation to insecticide induced metabolic stress. These observed changes at cellular level pave way for successful survival of prawns in insecticide polluted environ.     

Activity patterns of phosphofructokinase, glyceraldehydephosphate dehydrogenase, lactate dehydrogenase and malate dehydrogenase in microdissected fast and slow fibres from rabbit psoas and soleus muscle.

Methods for standardized determination of phosphofructokinase (PFK), glyceraldehydephosphate dehydrogenase (GAPDH), lactate dehydrogenase (LDH) and malate dehydrogenase (MDH) activities in nanogram samples of microdissected single fibres of rabbit psoas and soleus muscle are described. Fast and slow fibres in soleus muscle show lower absolute activities of these enzymes than the respective fibre types in psoas muscle. Slow fibres represent a more uniform population in the two muscles according to absolute and relative activities of the enzymes investigated. Slow fibres are characterized by high activities of MDH and relatively low activities of glycolytic enzymes. Fast fibres in the soleus muscle represent a population with high activities of MDH and glycolytic enzymes. Fast fibres in psoas muscle represent a heterogeneous population with high activities of glycolytic enzymes and extremely variable activity of MDH. More than 10-fold differences exist in the MDH activities of the extreme types of this fibre population. Differences in the activity levels of MDH in single fast type fibres but also in the activities of glycolytic enzymes between fast and slow fibres are greater than those reported between extreme white and red rabbit muscles.     

Cyclic AMP-dependent phosphorylation, Ca2+ and calmodulin: possible influences on acyltransferases and pyruvate dehydrogenase activities of rat mammary gland

Specific activities of diacylglycerol acyltransferase, glycerol 3-phosphate acyltransferase and pyruvate dehydrogenase were studied in virgin, pregnant, lactating and involuting rat mammary glands. An inverse relationship was evident between cAMP binding to protein kinase(s) and the activities of the above enzymes in lactating rat mammary glands. Results suggested that free Ca2+ concentration may also contribute to control of the activity of pyruvate dehydrogenase in these glands. However, no consistent change was observed between the activities of these enzymes and cAMP binding in young, pregnant and involuting rat mammary glands. Calmodulin levels paralleled bound Ca2+ except in lactating rats. Almost all parameters studied peaked on day 8 of lactation.     

Malate-aspartate shuttle enzymes in rat brain regions, liver and heart during alloxan diabetes and insulin replacement

Regionally selective and time-dependent variations were observed in the activity of brain aspartate aminotransferase at early phases of diabetes. Malate dehydrogenase activity showed an opposite pattern of changes in soluble and particulate fractions of cerebral hemispheres and brain stem, with cerebellum showing consistent increase in the activity. The activity of both the enzymes increased significantly in liver, in contrast to heart where malate dehydrogenase activity decreased in particulate fraction. Insulin treatment to diabetic animals restored the enzymes to near control levels at early stages of diabetes, except in liver. The results indicate that malate-aspartate shuttle is probably stimulated under diabetic conditions to enable glycolysis to continue and ATP levels to be restored partially, particularly in cerebellum and liver.     

Effect of prostaglandins in vivo on enzymes involved in thyroid carbohydrate metabolism.

Treatment of adult guinea pigs with prostaglandins produces changes in the levels of enzymes involved in carbohydrate metabolism of the thyroid gland. A decrease in glucose-6-phosphate dehydrogenase activity is observed with a concomitant increase in 6-phosphogluconic dehydrogenase; the glycolytic enzymes appear unaffected by the same treatment. The results indicate that prostaglandins do not have the biochemical effects obtained with thyrotropin and cAMP administration, showing that these compounds play an antagonistic role in comparison with the above mentioned stimulating agents.     

Regulation of asparaginase, glutamine synthetase, and glutamate dehydrogenase in response to medium nitrogen concentrations in a euryhaline chlamydomonas species

The ammonium assimilatory enzymes glutamine synthetase (EC 6.3.1.2) and glutamate dehydrogenase (EC 1.4.1.3) were investigated for a possible role in the regulation of asparaginase (EC 3.5.1.1) in a Chlamydomonas species isolated from a marine environment. Cells grown under nitrogen limitation (0.1 millimolar NH(4) (+), NO(3) (-), or l-asparagine) possessed 6 times the asparaginase activity and approximately one-half the protein of cells grown at high nitrogen levels (1.5 to 2.5 millimolar). Biosynthetic glutamine synthetase activity was 1.5 to 1.8 times greater in nitrogen-limited cells than cells grown at high levels of the three nitrogen sources.Conversely, glutamate dehydrogenase (both NADH- and NADPH-dependent activities) was greatest in cells grown at high levels of asparagine or ammonium, while nitrate-grown cells possessed little activity at all concentrations employed. For all three nitrogen sources, glutamate dehydrogenase activity was correlated to the residual ammonium concentration of the media after growth (r = 0.88 and 0.94 for NADH- and NADPH-dependent activities, respectively).These results suggest that glutamate dehydrogenase is regulated in response to ambient ammonium levels via a mechanism distinct from asparaginase or glutamine synthetase. Glutamine synthetase and asparaginase, apparently repressed by high levels of all three nitrogen sources, are perhaps regulated by a common mechanism responding to intracellular nitrogen depletion, as evidenced by low cellular protein content.     

Studies on the mechanism of action of ionizing radiations; inhibition of sulfhydryl enzymes by alpha,beta, and gamma rays

The activity of crystalline phosphoglyceraldehyde dehydrogenase and urease was decreased when dilute solutions of these sulfhydryl enzymes were irradiated with small doses of alpha rays from Po, beta rays from Si(89), and gamma rays from Ra. Partial reactivation of the enzyme by addition of glutathione was obtained after inhibition with alpha rays. Evidence that these inhibitions are due to oxidation of the -SH groups of the enzymes was given by the irradiation of the mercury-mercaptide urease with gamma rays. This irradiated complex was completely reactivated by glutathione as was the non-irradiated enzyme. The ionic efficiency of all these ionizing radiations on inhibition of phosphoglyceraldehyde dehydrogenase was similar (ionic yield around 1). The sulfhydryl groups of crystalline phosphoglyceraldehyde dehydrogenase were titrated by enzyme activity measurements and by ferricyanide oxidation.     

Enzyme levels in cultured astrocytes,oligodendrocytes and Schwann cells,and neurons from the cerebral cortex and superior cervical ganglia of the rat

Data are presented for 16 enzymes from 8 metabolic systems in cell cultures consisting of approximately 95% astrocytes and 5% oligodendrocytes. Nine of these enzymes were also measured in cultures of oligodendrocytes, Schwann cells, and neurons prepared from both cerebral cortex and superior cervical ganglia. Activities, in mature astrocyte cultures, expressed as percentage of their activity in brain, ranged from 9% for glycerol-3-phosphate dehydrogenase to over 300% for glucose-6-phosphate dehydrogenase. Creatine phosphokinase activity in astrocytes was about the same as in brain, half as high in oligodendrocytes, but 7% or less of the brain level in Schwann cells and superior cervical ganglion neurons and only 16% of brain in cortical neurons. Three enzymes which generate NADPH, the dehydrogenases for glucose-6-phosphate and 6-phosphogluconate, and the NADP-requiring isocitrate dehydrogenase, were present in astrocytes at levels at least twice that of brain. Oligodendrocytes had enzyme levels only 30% to 70% of those of astrocytes. Schwann cells had much higher lactate dehydrogenase and 6-phosphogluconate dehydrogenase activities than oligodendrocytes, but showed a remarkable similarity in enzyme pattern to those of cortical and superior cervical ganglion neurons.Special issue dedicated to Dr. Lewis Sokoloff.     

Correlation of enzymatic,metabolic, and behavioral deficits in thiamin deficiency and its reversal

To clarify the enzymatic mechanisms of brain damage inthiamin deficiency, glucose oxidation, acetylcholine synthesis, and the activities of the three major thiamin pyrophosphate (TPP) dependent brain enzymes were compared in untreated controls, in symptomatic pyrithiamin-induced thiamin-deficient rats, and in animals in which the symptoms had been reversed by treatment with thiamin. Although brain slices from symptomatic animals produced¹⁴CO₂ and¹⁴C-acetylcholine from [U-¹⁴C]glucose at rates similar to controls under resting conditions, their K⁺-induced-increase declined by 50 and 75%, respectively. In brain homogenates from these same animals, the activities of two TPP-dependent enzymes transketolase (EC 2.2.1.1) and 2-oxoglutarate dehydrogenase complex (EC 1.2.4.2, EC 2.3.1.61, EC 1.6.4.3) decreased 60–65% and 36%, respectively. The activity of the third TPP-dependent enzyme, pyruvate dehydrogenase complex (EC 1.2.4.1, EC 2.3.1.12, EC 1.6.4.3.) did not change nor did the activity of its activator pyruvate dehydrogenase phosphate phosphatase (EC 3.1.3.43). Although treatment with thiamin for seven days reversed the neurological symptoms and restored glucose oxidation, acetylcholine synthesis and 2-oxoglutarate dehydrogenase activity to normal, transketolase activity remained 30–32% lower than controls. The activities of other TPP-independent enzymes (hexokinase, phosphofructokinase, and glutamate dehydrogenase) were normal in both deficient and reversed animals.Thus, changes in the neurological signs during pyrithiamin-induced thiamin deficiency and in recovery paralleled the reversible damage to a mitochondrial enzyme and impairment of glucose oxidation and acetylcholine synthesis. A more sustained deficit in the pentose pathway enzyme, transketolase, may relate to the anatomical abnormalities that accompany thiamin deficiency.Dedicated to Henry McIlwain.     

Glycolysis in the eye tissues of the rabbit in ontogeny. I. The enzymes of glycolysis and hexosemonophosphate shunt]

The activity of the enzymes of glycolysis (phosphofructokinase, aldolase, pyruvate kinase, lactate dehydrogenase) and hexose monophosphate shunt (glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase) was determined in the eye tissues of the rabbit at different stages of ontogenesis. The activity of these enzymes in the retina was shown to be higher than in other eye tissues. In the uveal tract (iris, ciliary bodies, uvea) the activity of glycolytic enzymes changes with the age. The greatest changes in the activity of enzymes were found during the period of the opening of eyelids. The activity of the enzymes of hexose monophosphate shunt in the eye tissues increases with the age. The relative activity of dehydrogenases of the hexose monophosphate shunt after the establishment of visual function is, however, not high and does not exceed that of phosphofructokinase and pyruvate kinase in the eye tissues of the rabbit.     

Soluble and Bound Apoplastic Activity for Peroxidase, beta-d-Glucosidase, Malate Dehydrogenase, and Nonspecific Arylesterase, in Barley (Hordeum vulgare L.) and Oat (Avena sativa L.) Primary Leaves

An intercellular washing solution containing about 1% of the soluble protein, 0.3% or less of the glucose-6-phosphate dehydrogenase activity, but up to 20% of the peroxidase and β-d-glucosidase activity of barley (Hordeum vulgare L.) or oat (Avena sativa L.) primary leaves was obtained by vacuum infiltrating peeled leaves with pH 6.9 buffered 200 millimolar NaCl. After this wash, segments were homogenized in buffer, centrifuged, and the supernatant was assayed for soluble cytoplasmic enzymes. The pellet was washed and resuspended in 1 molar NaCl to solubilize enzymes strongly ionically bound to the cell wall. The final pellet was assayed for enzyme activity covalently bound in the cell wall. Apoplastic (intercellular washing solution, ionically bound, and covalently bound) fractions contained up to 76% of the β-d-glucosidase activity, 36% of the peroxidase activity, 11% of the nonspecific arylesterase activity, 4% of the malate dehydrogenase activity, but less than 2% of the glucose-6-phosphate dehydrogenase activity of peeled leaf segments. The partitioning and salt-solubility of the enzymes between the apoplast and symplast differed considerably between these two species. Intercellular washing fluid prepared by centrifuging unpeeled leaves had higher activity for glucose-6-phosphate dehydrogenase, less soluble protein, and less peroxidase activity per leaf than intercellular washing solution obtained by our peeling-infiltration-washing technique. The results are discussed in relation to the roles of these enzymes in phenolic metabolism in the cell wall.     

NADPH generating enzymes in Leydig cells from diabetic rats.

Impaired testosterone biosynthesis in Leydig cells from streptozotocin treated rats is correlated with the reduced activity of glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase and isocitrate dehydrogenase. The results shown demonstrate that in the diabetic state the activity of these enzymes is reduced by almost 50 to 59% from normal levels. Insulin treatment restored their activities to normal levels. The diminished supply of NADPH in diabetic interstitial tissue is not the unique factor in the control of steroidogenesis, since the availability of large amounts of exogenous NADPH in the incubations of Leydig cell did not reduce the differences in testosterone synthesis observed when compared with normal cells.     

The early effects of chemical carcinogens on adult rat hepatocytes in primary culture: I. Quantitative changes in intracellular enzyme activities following a single dose of carcinogen.

The effects of exposure of adult rat hepatocytes to chemical carcinogens have been studied using a short-term maintenance culture system. Scanning microdensitometry was used to quantitate the observed changes in enzyme activity. The dose-response curves showed a biphasic response for all 4 enzymes studied (glucose-6-phosphate dehydrogenase, succinate dehydrogenase, NADPH oxidase and gamma-glutamyl transpeptidase) there being decreased enzyme activities at the higher dose levels used, possibly indicating cytotoxicity. The enhancement of enzyme activity at low dose levels was due to generalised increases occurring in every cell, rather than to selection of a cell species particularly high in enzyme activity. A culture period of 24 h was necessary for the complete adaptation of the cells to the culture environment as evidenced by the response of intracellular glucose-6-phosphate dehydrogenase activity to carcinogen treatment. These findings are discussed in relation to previously reported in vivo studies.     

Changes in the activities of enzymes involved in amino acid metabolism during the senescence of detached wheat leaves

The activities of several enzymes related to amino acid metabolism were investigated in senescing detached wheat leaves ( Triticum aestivum L. cv. Diplomat) in light and darkness and after kinetin treatment. Glutamine synthetase and glutamate synthase activities rapidly declined in darkness. In light, the decline of glutamate synthase activity was retarded, while the activity of glutamine synthetase remained high and even increased transitorily. Kinetin treatment counteracted the decline of the activities of both enzymes. The activity of glutamate dehydrogenase markedly increased during senescence, particularly in light, and kinetin treatment lowered its activity. The activities of glutamate-oxaloacetate and glutamate-pyruvate amino-transferases and of NADP-dependent isocitrate dehydrogenase also increased in detached wheat leaves in light. Kinetin treatment prevented the rise of these enzyme activities. In darkness, the activities of glutamate-oxaloacetate aminotransferase and NADP-dependent isocitrate dehydrogenase decreased slowly while the decline of glutamate-pyruvate aminotransferase activity was more rapid. The activity of NAD-dependent malate dehydrogenase decreased both in light and, more rapidly, in darkness. The pattern of changes of the enzyme activities provides an explanation for the amino acid transformations and the flow of amino nitrogen into transport metabolites in senescing leaves.     

The inhibitory effect of actinomycin D and cycloheximide on the increase in activity of glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase in experimentally induced diseased skeletal muscles

Summary The myotoxic effect of the subcutaneous administration ofN,N ¹-dimethyl-p-phenylenediamine (DPPD) in rats was enhanced by the simultaneous administration of hyaluronidase. The resulting myopathy was associated with an early and dramatic increase in activity of glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase. Administration of actinomycin D or cycloheximide prior to the combined DPPD and hyaluronidase treatment prevented the increase in activity of both pentose phosphate pathway enzymes, indicating that the increase in activity requires RNA synthesis and protein synthesis. The possibility that the increase in activity of both NADPH-regenerating enzymes results from the modification by effectors of existing less active forms of these enzymes leading to more highly active forms was refuted.     

Effect of temperature on the activity and synthesis of glucose-catabolizing enzymes in Pseudomonas fluorescens.

The activity of the enzymes of the oxidative non-phosphorylated pathway, glucose and gluconate dehydrogenases, were not significantly affected by changes in the assay temperature. Both enzymes demonstrated only a threefold difference in activity when compared at assay temperatures of 30 degrees C and 5 degrees C. In contrast, the enzymes involved in the direct phosphorylation and catabolism of glucose or its oxidation products, gluconate and 2-ketogluconate, exhibited a more pronounced response to decreasing assay temperatures. At least one enzyme in each pathway, involved in the direct phosphorylation and catabolism of glucose or 2-ketogluconate (2KG), demonstrated an eightfold decrease in activity with a decrease in assay temperature from 30 degrees C to 5 degrees C. A similar decrease in assay temperature resulted in a fivefold decrease in activity of the enzymes involved in the direct phosphorylation and catabolism of gluconate. The observed differential effect of temperature on the activity of the enzymes of glucose catabolism and on the accumulation of direct oxidation products during growth with glucose in P. fluorescens E-20 is discussed. Growth with glucose at 5 or 20 degrees C resulted in high induced levels of all glucose-catabolizing enzymes examined when compared with the levels of these same enzymes in pyruvate-grown cells. However, only low levels of glucose dehydrogenase were detected during growth at 30 degrees C with glucose, gluconate, or 2-KG. Similarly, only low levels of gluconate dehydrogenase were detected during growth with glucose at 30 degrees C, although a weak induction was observed during growth with gluconate or 2-KG at 30 degrees C. The levels of 2-KG kinase plus KPG reductase during growth at 30 degrees C were undetectable with glucose, weakly induced with gluconate, and fully induced with 2-KG. High induced levels of glucose dehydrogenase, gluconate dehydrogenase, and 2-KG kinase plus KPG reductase were present during growth at 20 degrees C with glucose or 2-KG. The low levels of glucose and gluconate dehydrogenases present at a growth temperature of 30 degrees C was not due to heat lability of the enzymes at this temperature. The low amounts of these two enzymes during growth with glucose at 30 degrees C probably prevented sufficient inducer(s) formation from glucose to allow induction of enzymes of 2-KG catabolism. The results demonstrated that temperature may regulate the pathways of glucose dissimilation by regulating, either directly or indirectly, the activity and synthesis of the enzymes involved in these pathways.