Induction of phosphoenolpyruvate carboxykinase by hydrocortisone in rat liver and brain as a function of age

The activity and induction pattern of phosphoenolpyruvate carboxykinase (PEPCK) in the liver and brain of young (6-), adult (30-) and old (90-weeks) male rats were studied. The activity of this enzyme increases in both tissues until adulthood and decreases gradually thereafter. Further, the activity of PEPCK is higher in the liver than the brain. Adrenalectomy decreases significantly the activity of this enzyme in the liver of rats of all ages. However, this treatment inhibits brain PEPCK in young and adult rats. Administration of hydrocortisone to adrenalectomized rats increases PEPCK in both tissues of young and adult rats. However, the magnitude of induction is higher in the young, as compared to the adult, rats. This hormone-mediated induction of the enzyme is actinomycin D-sensitive.     

Alterations in the activities of subcellular fractions marker enzymes in rat liver and brain by hydrocortisone and corticosterone treatment

The effect of subcutaneous injection of hydrocortisone and corticosterone on the activity values of some subcellular fractions marker enzymes from rat liver and brain was investigated and compared with controls (without treatment with hormones). The following enzymes were studied (subcellular fraction are shown between parentheses): N-acetyl-beta-D-glucosaminidase and beta-glucuronidase (lysosomes); succinate dehydrogenase = SDH (mitochondria); glucose-6-phosphatase (endoplasmic reticulum); 5'-nucleotidase and Na+-K+-Mg2+ ATPase (plasma membrane). The specific activity of lysosomal enzymes from liver showed no change when rats were injected either with hydrocortisone or corticosterone. The same enzymes from brain showed significant increases in their activities with both hydrocortisone or corticosterone except beta-glucuronidase; this enzyme gave activity values remaining between the control levels, after treatment with corticosterone. The activity of mitochondrial SDH was increased after corticosterone injection either in liver or brain. After hydrocortisone injection, its activity rises significantly in brain (72%), but it falls in liver compared to the control values. Glucose-6-phosphatase behaves similarly in brain or liver fractions; its activity increases always after corticosterone treatment and decreases by hydrocortisone. The plasma membrane marker enzymes did not change practically in brain fractions, excepted Na+-K+-Mg2+ ATPase which tends to rise its activity after hydrocortisone injection. In liver fractions, both 5'-nucleotidase and Na+-K+-Mg2+ ATPase activities increase either by corticosterone or hydrocortisone treatment, except 5'-nucleotidase which specific activity decreases in liver after hydrocortisone treatment.     

Regulation of NAD- and NADP-linked isocitrate dehydrogenase by thyroxine in the brain and liver of female rats of various ages

The specific activity of NAD- and NADP-linked isocitrate dehydrogenase and their regulation by thyroxine in the brain and liver of female rats of various ages were studied with the ultimate goal of better understanding the decreased physiological functioning of the brain and liver during old age. Both thyroidectomy and thyroxine treatment have differential age-dependent effects on the activities of these enzymes in both tissues. The activity of NAD-ICDH decreases whereas both cytoplasmic and mitochondrial NADP-ICDH increase simultaneously following thyroidectomy. Thyroxine administration induces NAD-ICDH and depresses NADP-ICDH. The degree of induction and/or repression is lowest in old rats. These effects of thyroxine are actinomycin D sensitive in both the tissues of rats.     

Regulation of NAD- and NADP-linked isocitrate dehydrogenase by hydrocortisone in the brain and liver of male rats of various ages

The activity and hormonal regulation of NAD- and NADP-linked isocitrate dehydrogenase (EC 1.1.1.41 and 1.1.1.42, respectively) in the brain and liver of rats of various ages were investigated. The activity of NAD-linked isocitrate dehydrogenase of the brain was greater than cytoplasmic or mitochondrial NADP-linked isocitrate dehydrogenase. In contrast, the cytoplasmic NADP-isocitrate dehydrogenase of the liver predominates over both NAD- and mitochondrial NADP-isocitrate dehydrogenases at the three ages studied. The activity of NAD-isocitrate dehydrogenase increased in the brain (139%) and liver (17%) of rats upt o 33 weeks of age and decreased (57 and 39%, respectively) in old rats (85-week-old). The activity of cytoplasmic NADP-isocitrate dehydrogenase was maximum in immature (6-week-old) rat brain and decreased as the age of the rats increased; whereas, in liver, the activity of this enzyme was found to be maximum in adult rats (33-week-old). Brain mitochondrial NADP-isocitrate dehydrogenase activity increased (64%) in adult rats, but in liver it decreased (45 and 33% in 33- and 85-week-old rats, respectively). In both tissues, adrenalectomy and hydrocortisone treatment showed differential age-dependent response. Hydrocortisone-mediated induction of the level of enzymes was inhibited by actinomycin D.     

Differential effects of hydrocortisone on alanine aminotransferase isoenzymes of the cerebral hemispheres and cerebellum of rats during growth, development, and senescence

The activities and induction patterns of the isoenzymes of alanine aminotransferase (AAT) of the cerebral hemispheres and cerebellum of rats of various ages were studied. The activities of both the soluble (s-) and mitochondrial (m-) isoenzymes of ATT of the cerebral hemispheres and cerebellum were highest in the immature rat and decreased significantly thereafter with increasing age. Adrenalectomy decreased, and hydrocortisone administration increased significantly, the activity of s-AAT in both cerebral hemispheres and cerebellum of immature, adult, and senescent rats. However, these treatments resulted in significant changes in the activity of m-AAT in both tissues of the immature rat only. The hormone-mediated induction of these isoenzymes was actinomycin D-sensitive.     

Hydrocortisone and triiodothyronine regulation of malate-aspartate shuttle enzymes during postnatal development of chicken.

The normal endogenous level of malate-aspartate shuttle enzymes and its regulation by hydrocortisone and triiodothyronine were studied in the liver and kidney of 0-, 30- and 60-day old male Rhode Island Red (RIR) chicken. The endogenous activity of cytosolic malate dehydrogenase (c-MDH) was significantly higher in the liver of day 30 as compared to day 0 and 60. In contrast, mitochondrial malate dehydrogenase (m-MDH) activity decreased at day 60 in the liver. However, both c- and m-MDH had significantly lower activities at day 0, which increased sharply at day 30 and 60 in the kidney. On the other hand, activity of both cytosolic and mitochondrial aspartate aminotransferase (c- and m-AsAT) showed peak value at day 30 in both liver and kidney. Hydrocortisone administration induced c-MDH in the liver at all the ages studied, but did not influence the activity of the isoenzymes in the kidney whereas, it induced m-MDH in the liver at day 0 and in kidney at day 30. Administration of hydrocortisone, however, did not influence AsAT isoenzymes (c- and m-AsAT) in either of the tissues at any of the postnatal ages. Triiodothyronine induced c-MDH in the liver at all the ages whereas kidney isoenzyme was induced only at day 60. In contrast, m-MDH was induced by triiodothyronine in both liver and kidney at day 30 and 60. Administration of triiodothyronine did not influence c-AsAT of liver and kidney at either of the ages, whereas it induced m-AsAT of only liver at day 0 and 60. These findings indicated a tissue- and age-specific expression of the malate-aspartate shuttle enzymes in chicken and difference in the regulation exerted by hydrocortisone and triiodothyronine during postnatal development of chicken.     

Induction of particulate and soluble isoenzymes of tyrosine aminotransferase by hydrocortisone in the liver of rats as a function of age.

The induction of soluble cytoplasmic (c-), and particulate mitochondrial (m-) and nuclear (n-) isoenzymes of tyrosine aminotransferase (TAT) by hydrocortisone in the liver of 6-, 35- and 76- week old rats was studied. In contrast to the earlier reports, both the particulate isoenzymes (m- &; n-TAT) are induced by hydrocortisone. This induction is actinomycin D sensitive. The degree and pattern of induction of the three isoenzymes of TAT vary with age. The possibility of separate regulatory mechanisms for the synthesis of the three isoenzymes is discussed.     

Differential effects of hydrocortisone on aspartate aminotransferase isoenzymes of the liver of rats during growth, development, and senescence

The activity and induction patterns of the isoenzymes of aspartate aminotransferase (AsAT) of the liver of male rats during growth, development, and senescence were studied. The activity of both the cytoplasmic and mitochondrial isoenzymes increased significantly until adulthood and remained constant thereafter. Adrenalectomy decreased, and hydrocortisone administration increased, the activity of the cytoplasmic AsAT of the liver of young-immature, adult, and senescent rats. However, the degree of these responses decreased with increasing age of the animal. The hormone-mediated induction of this isoenzyme is actinomycin D-sensitive. Mitochondrial AsAT on the other hand is irresponsive towards these treatments throughout the life span of the rat.     

Isocitrate dehydrogenase activity and its regulation by estradiol in tissues of rats of various ages

The activity and hormonal regulation of NAD- and NADP-linked isocitrate dehydrogenase (EC.1.1.1.41 and EC.1.1.1.42, respectively) in the brain, liver and kidney cortex of female rats of various ages was investigated. The activity of NAD-ICDH of brain was greater than extramitochondrial (-c) or intramitochondrial (-m) NADP-ICDH. In contrast, liver c-NADP-ICDH was much higher than NAD- or m-NADP-ICDH, whereas in kidney cortex the activity of m-NADP-ICDH is dominant over both NAD- and c-NADP-ICDH in all the age group of rats studied. The activity of the NAD-ICDH of brain and all the enzymes of liver and kidney cortex increases until adulthood (33-weeks) and decreases thereafter in old rats (85-weeks). In brain c-NADP-ICDH was much higher in immature (6-weeks) rats and decreases with increasing age of the animal, whereas m-NADP-ICDH showed no significant change with the age of the rats. Bilateral ovariectomy decreases the level of all the three forms of enzyme in all the tissues of 6-, 13- and 33-week rats but failed to show any significant effect in 85-week old rats. Exogenous administration of estradiol induces all the three forms of enzyme in all the tissues of ovariectomized rats. The degree of response is tissue- and age-specific.     

Tissue-specific differential induction of aspartate transcarbamylase by hydrocortisone in rats of various ages

The activity and induction pattern of aspartate transcarbamylase (ATCase) of the liver (a mitotic tissue) and heart (a post-mitotic tissue) of rats of various ages were studied. The activity of the enzyme of both tissues was highest in the immature rat and decreased significantly thereafter with increasing age of the animal. Adrenalectomy and hydrocortisone treatments altered the activity of liver ATCase in rats of all the ages. However, these treatments altered the heart enzyme of the immature and adult rats, but not of senescent rat. The magnitude of induction of the enzyme by hydrocortisone was highest in the immature rat. Actinomycin D inhibited the hormone-mediated induction of ATCase in both the liver and heart.     

Induction of a Peroxisomal Malate Dehydrogenase Isoform in Liver of Starved Rats

The influence of starvation on malate dehydrogenase (MDH) in rat liver was investigated. Native electrophoresis revealed two MDH isoforms in non-starved rats and three isoenzymes in starved rats. After sucrose density gradient centrifugation of cell organelles from liver, MDH activity was detected in the mitochondrial and cytosolic fractions from non-starved rats. However, additional activity was found in the peroxisomal fraction from starved rats. The latter was identified as the electrophoretically new isoform in starved animals. The three isoforms of malate dehydrogenase from hepatocytes were separated and partially purified by chromatography on DEAE-Toyopearl. Several kinetic and regulatory properties of the three isoforms were rather similar. It is suggested that the newly expressed isoform of MDH operates in the glyoxylate cycle of liver peroxisomes of food-starved animals.     

Lack of inducibility of brain monooxygenase activities including parathion desulfuration

The ability of phenobarbital and beta-naphthoflavone to induce parathion desulfuration, aminopyrine N-demethylation, and NADPH-cytochrome-c reductase activity in the brain and liver of male and female rats was investigated. Activities of all three enzymes were found in similar levels in both the mitochondrial and microsomal fractions of brain. There were no sex differences in brain activities. Liver activities were from 10- to 30-fold higher than brain activities when computed on a tissue-wet-weight-equivalent basis. Although exposure to both inducers increased all three enzyme activities and cytochrome P-450 in liver, neither inducer increased the enzyme activities in mitochondrial or microsomal brain fractions of either sex. Thus, these brain monooxygenase activities appear to be refractory to induction by two classical types of cytochrome P-450 inducers. This lack of inducibility could serve to protect the animal against environmentally enhanced increases in the activation of xenobiotics to neurotoxic metabolites, such as parathion desulfuration to paraoxon.     

Polyacrylamide gel electrophoresis of rat brain acetylcholinesterase: isoenzymes of normal rat brain

Abstract— Triton-solubilized acetylcholinesterase (EC 3.1.1.7) of rat brain was submitted to vertical flatbed polyacrylamide gel electrophoresis. Three anodally migrating isoenzyme zones with low relative mobilities could be resolved, each of which on quantitative densitometry appeared to consist of more than one subzone. More than 50 per cent of the total AChE activity was exhibited by the isoenzyme zone closest to the origin (isoenzyme zone 3). Regional differences in AChE isoenzyme activity were quantitative only with the caudate-putamen complex, midbrain, pons and medulla oblongata exhibiting relatively high content of the three isoenzymes and the cerebral cortex and olfactory bulb possessing weak isoenzyme activities. Intermediate levels of isoenzyme activities were observed in the cerebellum and hippocampus. In all areas examined, the relative percentage values for each isoenzyme remained constant. AChE isoenzymes from the forebrain, brain stem and cerebellum of 15- and 30-day-old rats appeared to have identical patterns. In brain stem, no quantitative differences could be detected in the isoenzyme activities between 15 and 30 days of age. At both ages, the isoenzymes of male and female rats did not show any qualitative differences. The single cholinesterase (EC 3.1.1.8) isoenzyme which could be identified in brain stem supernatants of 30-day-old rats was weakly reactive and appeared to have the same relative mobility as the major acetylcholinesterase zone, zone 3. Acetylcholinesterase isoenzymes failed to demonstrate any differential response toward varying concentrations of inhibitors and to changes in pH. While there were basic similarities in the acetylcholinesterase and cholinesterase isoenzyme patterns of brain, serum, liver, skeletal muscle and intestine, brain alone exhibited a marked preponderance of the acetylcholinesterase isoenzyme zone 3.     

Rat brain and liver mitochondria develop oxidative stress and lose enzymatic activities on aging

The mitochondrial mass of rat brain and liver remained unchanged on aging in young adults, old adults, and senescent animals (28, 60, and 92 wk of age); the values were 15-17 and 29-31 mg protein/g for brain and liver, respectively. The whole aging process was associated with an increased content of the oxidation products, thiobarbituric acid-reactive substances and protein carbonyls, by 61-69% in brain and 36-45% in liver, respectively. The activities of critical enzymes for mitochondrial function, mitochondrial nitric oxide synthase, Mn-superoxide dismutase, complex I, and complex IV, decreased progressively during aging with activity losses of 73, 37, 29, and 28%, respectively, in the brain and 47, 46, 30, and 24% in the liver of senescent rats compared with young adults. Brain mitochondria isolated from aged rats showed increased mitochondrial fragility, as assayed by mitochondrial marker enzyme activities in the postmitochondrial supernatant, and increased volume and water permeability, as assayed by light scattering. Liver mitochondria isolated from young and old rats did not show differences in fragility and water permeability. A subpopulation of brain mitochondria with increased size and fragility was differentiated in aging rats, whereas liver showed a homogeneous mitochondrial population.     

Aspartate and alanine aminotransferase activity in the tissues of adrenalectomized rabbits following administration of hydrocortisone and corticotropin

Aspartate and alanine aminotransferase (AsT, AlT) activities were studied in tissues of adrenalectomized rabbits which were treated with a single and multiple administrations of hydrocortisone (5 mg/kg) or a single administration of corticotropine (ACTH, 10 units/kg). It is shown that adrenalectomy decreases the AsT activity in homogenate of femoral muscle tissue and decreases the AlT activity in homogenate and supernatant of the liver, spleen and muscle tissue and in blood plasma. A single administration of hydrocortisone increases the AsT activity in supernatant of femoral muscle tissue and in blood plasma and increases AIT activity in the brain, liver, muscle and blood plasma. Parallel with that AsT and AlT activities are decreased in the spleen tissue. Multiple administration of hydrocortisone induces analogous changes in the AsT activity in the muscle and in the AlT activity in the liver, muscle and blood plasma. A single administration of ACTH induces an increase of the AsT activity in the muscle supernatant and in blood plasma. It also causes a rise of the AlT activity in the liver, muscle supernatant and blood plasma. The AlT activity is decreased in the brain supernatant. A question about stability of free amino acids metabolism (especially of alanine and aspartic acid) in the rabbit brain with changes in corticosteroid levels of organism is under discussion.     

Age-related differential induction of tryptophan pyrrolase by hydrocortisone in the liver of male rats

The activity and the regulatory pattern of tryptophan pyrrolase of the liver of male rats during various phases of the life span were studied with a view to investigate the differential effectiveness of hydrocortisone in relation to growth, development, and senescence of an organism. The level of this enzyme shows no significant change till adulthood but decreases significantly thereafter with increasing age. Adrenalectomy and hydrocortisone treatments decrease and increase, respectively the activity of this enzyme significantly in rats of all the ages. However, the effects of these treatments are highest in the mature rat. Induction of the enzyme by hydrocortisone is actinomycin D-sensitive.     

Anti-oxidant defences and peroxidation in liver and brain of aged rats.

Old rats (28 months), when compared with young adults (9 months), did not show differences in activities of superoxide dismutase (SOD) or selenium-dependent and -independent glutathione peroxidases (GPx), or in levels of GSH, GSSG, GSSG/GSH and endogenous peroxidation in liver and brain. Rates of stimulated peroxidation in vitro were decreased in the livers of old rats. Old animals showed decreased levels of hepatic catalase and glutathione reductase. Nevertheless, when enzyme activities were referred to cytochrome oxidase activity these decreases disappeared, and GPx and SOD (brain) were even increased in old rats.     

Glycolytic enzymes in polyamine-treated bovine retina

The retina is characterized by glycolysis under aerobic conditions, mediated by lactate dehydrogenase isoenzyme-5 (LDH-5) as well as by the soluble isoenzyme of malate dehydrogenase. Bovine retina LDH and MDH isoenzymes and their activities were studied after polyamine treatment. Our results showed that LDH-5 isoenzyme presented the highest activity in untreated as well as in putrescine-treated retina. Decreased activity was present when the retina was treated with spermidine or spermine. It was demonstrated that retinic LDH-5 had a high affinity for lactate which enabled the isoenzyme to be more effective than the other LDH isoenzymes in the conversion of NADH to NAD. Therefore, the putrescine enhancing LDH-5 activity appeared to be capable of stimulating NAD-mediated rhodopsin regeneration. Putrescine induced a marked increase of both MDH isoenzymes--soluble (s-MDH) and mitochondrial (m-MDH), while spermine and spermidine mostly affected the soluble form of the enzyme. Putrescine induced a three-fold increase in s-MDH and m-MDH activities, while spermine and spermidine induced a four to five-fold increase in s-MDH. These results document the differential effects of polyamine treatment on LDH and MDH isoenzyme activities.     

The Induction of Glyoxylate Cycle Enzymes in Tissues of Starving Rats

The induction of glyoxylate cycle enzyme activities was revealed in the liver and other organs of starving rats. A five day deprivation of food was followed by the appearance of isocitrate lyase (ICL) and malate synthase activities and the increase of malate dehydrogenase (MDH) and citrate synthase activities. The induction of MDH was associated with the appearance of its new isoform with Rf 0.52. ICL activity was revealed in the liver, blood, pancreas, kidney, lungs, heart, and skeletal muscles of starving rats, reaching a peak on day 5 of food deprivation. No significant changes of blood glucose level in starving rats were revealed until day 9. A homogeneous ICL preparation with a specific activity of 12.4 IU per mg protein was obtained as the result of a five-stage purification procedure.     

Effect of a short-term fasting and altitude hypoxia on the cytochrome oxidase activity of rat brain mitochondria

The effect of short-term fasting and thirst, prolonged fasting and hypoxic hypoxia upon the activity of cytochrome oxidase was studied in mitochondrial fractions obtained from the brain and the liver. The investigation was carried out in two groups of rats, 5 and 60 days old. a) The activity of cytochrome oxidase in mitochondria isolated from the brain cortex, subcortical regions and the medulla oblongata rises, while the changes in liver mitochondrial fractions are reverse. b) A significant increase of mitochondrial cytochrome oxidase was found in 5-day-old rats after both types of fasting and hypoxia in all regions of the brain, as well as in the liver. c) The cytochrome oxidase activity in brain and liver mitochondria of 60-day-old rats was not affected appreciably after 24 h nutritional deprivation, with the exception of a significant rise of activity in the medulla oblongata. Prolonged fasting and hypoxia again markedly increased the activity of this enzyme in all regions of the brain and in the liver.