Treatment Of Adrenalectomized Rats Research Articles

Aldosterone Regulates the Na-K-2Cl Cotransporter in Vascular Smooth Muscle

Aldosterone increases cation transport and contractility of vascular smooth muscle, but the specific transporter involved and how it is linked to smooth muscle tone is unknown. Because the Na-K-2Cl cotransporter (NKCC1) contributes to vascular smooth muscle contraction and is regulated by vasoactive compounds, we sought to determine whether this transporter is a target of aldosterone in rat aorta. Treatment of adrenalectomized rats with aldosterone for 7 days resulted in a 63% increase in NKCC1 activity as measured by bumetanide-sensitive efflux of 86Rb+. Treatment of normal aortas in culture with aldosterone for 3 and 7 days resulted in 29% and 47% increases in NKCC1 activity, respectively. Aldosterone had no acute effect on 86Rb+ efflux. Stimulation of NKCC1 was blocked by spironolactone, a mineralocorticoid receptor antagonist, but not by RU38486, a glucocorticoid receptor antagonist. Aldosterone did not augment the stimulation of NKCC1 by phenylephrine and did not increase NKCC1 mRNA as determined by real-time polymerase chain reaction. We conclude that aldosterone regulates the Na-K-2Cl cotransporter in vascular smooth muscle through classic mineralocorticoid receptors but not through changes in the abundance of NKCC1 mRNA. This could account for the increase in Na+, K+, and Cl- fluxes previously observed in vascular smooth muscle from mineralocorticoid-treated animals and may contribute to increased vascular tone.

Is the mineralocorticoid receptor in Brown Norway rats constitutively active?

In a previous study using corticosterone treatment of adrenalectomized rats, we hypothesized that mineralocorticoid receptor (MR)-related mechanisms are constitutively active and that glucocorticoid receptor (GR)-mediated mechanisms are more efficient in Brown Norway rats compared to Fischer 344 (F344) rats. In order to discriminate the mineralocorticoid from the glucocorticoid actions exerted by corticosterone, F344 and Brown Norway adrenalectomized rats were treated with increasing doses (1, 5 and 25 microg/ml of drinking water) of deoxycorticosterone (DOC, MR-specific ligand) or RU 28362 (GR-specific ligand). These rats were compared with long-term adrenalectomized (ADX) untreated rats and sham-ADX rats. This study confirms our previous results, notably the lack of effect of ADX on body weight and fluid intake in Brown Norway rats. Moreover, DOC treatment had no effect in Brown Norway rats whereas the higher dose restored fluid intake of the F344 ADX group to sham values. These results support the hypothesis of a constitutive activation of the MR and therefore the insensitivity of this receptor to its ligand in Brown Norway rats. Alternatively, RU 28362 treatment induced greater weight loss, decrease in food intake, anxiolysis, thymus involution, and decrease in plasma transcortin concentration and pituitary corticosteroid receptor densities in Brown Norway rats than in F344 rats, which is consistent with greater efficiency of GR mechanisms in Brown Norway rats than in F344 rats. Therefore, these strains are of great utility to disentangle MR and GR effects on complex phenotypes.

Fos immunoreactivity in the lamina terminalis of adrenalectomized rats and effects of angiotension II type 1 receptor blockade or deoxycorticosterone

Neural activity, as measured immunohistochemically by the presence of Fos protein, was determined in the lamina terminalis, a thin strip of tissue forming the anterior wall of the third brain ventricle, after adrenalectomy. Several weeks after surgery, the adrenalectomized rats were maintained with access to water and a low sodium diet for five days. In addition, hypertonic (0.5 M) NaCl solution was available for the entire five-day period (sodium available) or only during the first four days (sodium unavailable). The number of neurons expressing Fos, determined at the end of the fifth day, was increased in the adrenalectomized rats with or without NaCl solution to drink. Fos activity in the median preoptic nucleus was increased only in adrenalectomized rats without access to NaCl solution. Treatment of adrenalectomized rats with the sodium-retaining mineralocorticoid hormone, deoxycorticosterone, at the end of the fourth day, decreased Fos expression in the subfornical organ and the organum vasculosum of the lamina terminalis when NaCl solution was available but not when the NaCl solution was unavailable. In the adrenalectomized rats with NaCl solution available, mineralocorticoid treatment decreased both urinary sodium excretion and daily sodium intake. Brain nuclei in the lamina terminalis also became activated in intact rats made sodium deplete by treatment with the diuretic, furosemide. Relative to sodium-deplete intact rats, however, sodium-deplete adrenalectomized rats had a greater number of neurons expressing Fos in the organum vasculosum. Treatment of sodium-deplete rats, adrenalectomized or intact, with the angiotensin II-type 1 receptor antagonist, ZD7155, decreased sodium intake and Fos expression in the subfornical organ but not in the organum vasculosum of the lamina terminalis or median preoptic nucleus. In conclusion, the results demonstrated that activation of the brain nuclei located in the lamina terminalis of adrenalectomized rats was primarily related to sodium deficit and not to the absence of the mineralocorticoid hormones, although the adrenal hormones may have a role in limiting the activation of organum vasculosum of the lamina terminalis during sodium depletion. Furthermore, the results obtained with the administration of the angiotensin receptor antagonist are consistent with the proposal that sodium appetite of the sodium-deplete rat, adrenalectomized or intact, is mediated by circulating angiotensin II acting in the subfornical organ.

Glucocorticoids regulate the synthesis of HSP27 in rat brain slices

Using mild heat shock of rat brain slices as a model for cellular insult, corticosteroid-mediated regulation of protein synthesis has been investigated. Following a single in vivo injection of rats with corticosterone or the Type II glucocorticoid receptor agonist, RU-28362, synthesis of a 28 kDa protein is elevated in cerebellar slices which are subsequently incubated in vitro at 39°C for 3 h. Immunoblotting of proteins subsequent to separation by two-dimensional gel electrophoresis has identified this glucocorticoid-sensitive protein to be the small molecular weight heat-shock protein, HSP27. Synthesis of the major heat-shock proteins, HSP70 and HSP90, is not glucocorticoid-sensitive. When animals are sacrificed at either 4 h following an aldosterone injection or at 24 h following a corticosterone injection, the synthesis of HSP27 in cerebellar slices is decreased. Treatment of adrenalectomized rats with either corticosterone, RU-28362 or aldosterone produces increased synthesis of HSP27. With duration of heat shock, there is a transient increase in the synthesis of HSP27 after 2 h at 39°C in slices from the cerebral cortex, with a more sustained synthesis of HSP27 in cerebellar slices. In hippocampal slices, HSP27 is rarely present. The upregulated synthesis of HSP27 in the cerebellum following an acute exposure to stress-like elevations in corticosterone titers may contribute to the relative resistance of this brain region to cellular insults.

The effect of corticosterone on reactivity to spatial novelty is mediated by central mineralocorticosteroid receptors.

Corticosterone, secreted by the adrenal glands, binds to central mineralocorticoid receptors with high affinity and to glucocorticoid receptors with a tenfold lower affinity. In previous studies we have shown that the selective activation of either mineralocorticoid receptors or glucocorticoid receptors exerts distinctly different behavioural effects. In this study we examined in particular the mineralocorticoid receptor-mediated effect of corticosterone on the control of the behavioural response of male Wistar rats to spatial novelty. This analysis was based on our observation that in adrenal-intact rats the presence of an object in the centre of an open field alters the time spent and distance walked in the centre compared to the peripheral area, i.e. the pattern of reactive locomotor activity is changed. Using this paradigm we found that 1 day after removal of the adrenals the rats increased their behavioural reactivity towards the object. Treatment of adrenalectomized rats with a low dose of corticosterone (50 micrograms/kg s.c.) 1 h prior to testing restored the behavioural reactivity to the level of sham-operated, intact rats. Surprisingly, a high dose of corticosterone (1000 micrograms/kg s.c.) also increased the rat's reactivity towards the object. The same high dose of corticosterone given to adrenal-intact rats also increased behavioural reactivity. Pretreatment of these rats with an intracerebroventricular injection of the selective mineralocorticoid receptor antagonist RU28318 (100 ng/microliters) prevented the corticosterone-induced increase in behavioural reactivity, while the blockade of glucocorticoid receptors with the antagonist RU38486 (100 ng/microliters) was not effective. Administration of the mineralocorticoid receptor antagonist without corticosterone to adrenal-intact rats also increased behavioural reactivity, but this increase did not reach statistical significance.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of Maternal Bilateral Adrenalectomy and Betamethasone Administration on Renal Rat Encephalic Development

The present study examined the effects of maternal bilateral adrenalectomy and betamethasone treatment on fetal encephalic development, in terms of fetal body weight, brain weight, DNA, protein and lipid content and morphological development. Both influenced the developmental time patterns of fetal brain and cerebellum. Fetuses of adrenalectomized rats had decreased body weights, whereas brain weight was not affected. Maternal adrenalectomy produces in fetal brain a decreased number of cells and increased cell size, while betamethasone treatment of adrenalectomized rats increased cell number, which was not different from control values; cell size remained lower than in control fetuses. Lipid content was increased in the fetuses of betamethasone-treated rats. In terms of morphological development, laminated structures (hippocampus and brain and cerebellar cortex) were the ones most affected.

Adipocyte G-proteins and adenylate cyclase. Effects of adrenalectomy

Steroid hormones modulate the ability of cells to respond to hormones that act via cyclic AMP. In adipocytes of adrenalectomized rats, cyclic AMP accumulation and lipolysis in response to adrenaline are attenuated. However, the mechanism(s) of these effects are poorly understood. The effects of altered glucocorticoid status in vivo on the steady-state amounts of components of the hormone-sensitive adenylate cyclase were analysed in rat adipocytes. beta-Adrenergic receptors were analysed by using radioligand binding and immunoblotting with an anti-receptor antiserum. Neither the amount of radioligand binding nor the amount of beta-adrenergic-receptor peptide (Mr 67,000) was altered by adrenalectomy, whereas treatment of adrenalectomized rats with dexamethasone was found to increase both parameters by more than 25% with respect to the control. Forskolin-stimulated adenylated cyclase activity was unchanged in membranes isolated from adipocytes of adrenalectomized rats, but was decreased (50%) in those from dexamethasone-treated rats. The alpha-subunit of Gs was probed by using cholera-toxin-catalysed ADP-ribosylation. Immunoblotting was used to analyse the steady-state amounts of G-protein beta-subunits (beta-G35/36). Adrenalectomy was associated with decreases in the steady-state amounts of alpha-Gs (30%) and beta-G35/36 (50%). Dexamethasone treatment of adrenalectomized animals partially restored the lipolytic response of adipocytes to adrenaline and the amounts of alpha-Gs, increased the amounts of beta-G35/36 subunits from 50% to 150% of control values, increased beta-adrenergic receptors by more than 25% and decreased adenylate cyclase activity (50%). These results suggest that the steady-state amounts of components of hormone-sensitive adenylate cyclase are differentially regulated by glucocorticoids.

Relationship between the cytoplasmic activator of adenylate cyclase and glycogen metabolism in rat lung

The role of cytoplasmic activator of adenylate cyclase in rat lung metabolism was investigated. Mouse adrenal tumor (MAT) cells undergo differentiation in response to choleratoxin which acts through cyclic AMP. The activator of adenylate cyclase from rat lung also produced cyclic AMP in a disrupted MAT cell preparation. However, unlike choleratoxin, it did not induce MAT cell differentiation in whole cells. These results suggest impermeability of MAT cells, and possibly other cells, to the activator. Thus, means of altering activator activity in lung cytoplasm were sought, and changes in activator activity were related to lung glycogen. Adrenalectomy (ADX) in rats led to a reduction in activator activity that was accompanied by an elevation in lung glycogen. Dexamethasone treatment of adrenalectomized rats reversed both of these effects. Streptozotocin-induced diabetes in rats elevated activator activity and lowered lung glycogen. Insulin treatment of the diabetic rats restored activator activity to the normal control values. Preweaning of rats on day 16 instead of day 22 increased activator activity on the 19th day over the controls and there was a concomitant decrease in lung glycogen. Feeding the separated pups with homogenized milk restored glycogen and activator activity to the control values. These results indicate that activator activity in rat lung cytoplasm was dependent on the circulating levels of cortisol and insulin, and that there appeared to be an inverse relationship between activator activity and glycogen level in rat lungs.

Potassium secretion by neonatal rat distal colon.

Electrogenic K+ secretion across the distal colon of young rats was investigated by measuring the sensitivity of the short-circuit current to Ba2+ added to the mucosal side of the tissue. Ba2+-sensitive short-circuit current (IBasc) was high during the suckling and weaning periods but very low in adult animals. Increasing the mucosal K+ concentration was accompanied by the inhibition of the serosa-to-mucosa IBasc and the induction of the mucosa-to-serosa IBasc. The IBasc was decreased by serosal omission of either Na+ or Cl- as well as by serosal addition of furosemide or ouabain. Mucosal omission of Na+ did not change IBasc. By increasing the plasma level of aldosterone (low-sodium diet) IBasc rose by 95% whereas treatment decreasing this level (high-sodium diet) reduced IBasc by 76%. Bilateral adrenalectomy lowered IBasc by 59% and treatment of adrenalectomized rats with deoxycorticosterone acetate prevented the reduction of IBasc. Tetraethylammonium and quinidine had similar effects on Isc as Ba2+. These data are consistent with the presence of a high level of K+ secretion in the distal colon of neonatal rats. This secretory pathway is electrogenic and independent of Na+ absorption. It appears to be mediated by the Na-K-ATPase as well as a furosemide-sensitive Na-Cl or Na-Cl-K cotransport on the basolateral side and by Ba2+-sensitive K+ conductive pathways on the mucosal side. The results suggest that this K+ secretion can be regulated by mineralocorticoids. The mineralocorticoids are necessary for "stimulated" K+ secretion but they are not essential for maintaining "basal" K+ secretion.

Regulation of parotid salivary proteins by glucocorticoids.

Previous studies have indicated that adrenal-intact rats treated for one week with pharmacological doses of the synthetic glucocorticoid, dexamethasone, show a significant reduction in the proportion of proline-rich proteins and an increase in the proportion of amylase in rat parotid saliva (Johnson et al., 1987). In order to understand more fully the role of glucocorticoids in the regulation of salivary proteins, we performed bilateral adrenalectomies on groups of rats. Some of the adrenalectomized rats were treated with replacement-level doses of the synthetic glucocorticoid, dexamethasone. The food intake was monitored daily for both groups, and sham-operated pair-fed controls were included so that the effects of alterations of food intake could be separated from those of the experimental procedures. After eight to 12 days, uniformly stimulated parotid saliva was collected from these animals as well as from sham-operated controls fed ad libitum. The volume of saliva collected in 30 min was recorded, and the saliva samples were analyzed for concentration and composition of protein. Although the volume of saliva was not affected, parotid saliva collected from adrenalectomized rats exhibited a two-fold greater proportion of proline-rich proteins and reductions in other major secretory proteins: DNase, Fraction I, and Fraction V. The parotid gland secretory granules of adrenalectomized rats were more electron-lucent than in the ad libitum-fed controls. Treatment of adrenalectomized rats with dexamethasone largely prevented the changes in salivary protein composition as well as the alterations in secretory granule morphology.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of adrenalectomy before weaning and short- or long-term glucocorticoid administration on the genetically obese Zucker rat.

Intact obese rats were hyperinsulinaemic, had higher rates of whole-body fatty acid synthesis, higher activities of hepatic acetyl-CoA carboxylase and tyrosine aminotransferase and a higher hepatic glycogen concentration than intact lean animals. Adrenalectomy abolished all these factors of the obese phenotype. Treatment of adrenalectomized rats with corticosterone for 24 h increased the rate of whole-body fatty acid synthesis to the same extent in both phenotypes, but caused a larger increase in glycogen concentration, tyrosine aminotransferase activity and plasma insulin concentration in obese rats.

Microsomal phosphorylase in rat heart: depletion following adrenalectomy and restoration by in vivo administration of dexamethasone.

Phosphorylase activities were determined in homogenates, particulate and soluble fractions of heart muscle from control, adrenalectomized, and adrenalectomized dexamethasone-treated rats so as to assess the influence, if any, of glucocorticoids on the subcellular distribution of this enzyme. The specific activities (enzyme activity expressed as units per mg protein) and recoveries (enzyme activity expressed as units per fraction/g tissue) of both the active form of phosphorylase (phosphorylase a) and total phosphorylase (phosphorylases a and b) were essentially similar in homogenates and 10,000 X g particulate fractions prepared from hearts of control, adrenalectomized, and adrenalectomized dexamethasone-treated rats. Interestingly, however, the specific activities and recoveries of active as well as total phosphorylase were markedly lower (70-80%; P less than 0.001) in cardiac microsomes of adrenalectomized compared to control rats. On the other hand, the specific activities and recoveries of phosphorylase (active and total) were significantly higher (30-60%; P less than 0.01 or 0.001) in the cytosol fraction of heart muscle from adrenalectomized compared to control rats. Treatment of adrenalectomized rats with dexamethasone (a synthetic glucocorticoid) restored the specific activities and recoveries of phosphorylase in heart microsomal and cytosol fractions to control levels. No appreciable differences were observed in the specific activity ratios of phosphorylase a to phosphorylases a and b in any of the subcellular fractions of cardiac muscle from control, adrenalectomized, and adrenalectomized dexamethasone-treated rats. The above findings suggest a potential involvement of glucocorticoids in the maintenance of the membrane-associated (microsomal) pool of phosphorylase and, therefore, in the cellular compartmentalization of this enzyme in heart muscle.

Glucocorticoids facilitate the retention of acquired immobility during forced swimming

The adrenalectomy-induced decrease in the level of immobility during a 5 min retest period in the Porsolt swimming test could be reversed by glucocorticoids administered s.c. 15 min after the initial forced swimming exposure. The synthetic glucocorticoids dexamethasone and RU 28362 were active in the μg dose range while corticosterone was only active at a 500 times higher dose. Aldosterone and progesterone were both ineffective. Treatment of adrenalectomized rats with the synthetic antiglucocorticoid RU 38486 prior to dexamethasone administration dose dependently blocked the effect of the glucocorticoid. Intact rats treated with the antiglucocorticoid RU 38486 prior to the initial forced swimming exposure behaved like adrenalectomized animals during the 5 min retest period. Removal of the adrenal medulla only temporarily impaired swimming behavior. It is concluded that intact adrenocortical secretion of glucocorticoids is sufficient for retention of acquired immobility during forced swimming.

COMPARATIVE EFFECTS OF HYPOPHYSECTOMY AND ADRENALECTOMY UPON PLASMA AND ADRENAL MONOAMINES IN PREGNANT AND NON‐PREGNANT RATS

1 The influence of hypophysectomy and adrenalectomy upon release and storage of adrenaline and noradrenaline during rat oestrous cycle and pregnancy has been studied.2 Hypophysectomy during the pro‐oestrous phase of the oestrous cycle increased plasma noradrenaline concentration which was unchanged, however, by similar treatment in pregnant females. After adrenalectomy during dioestrous and pro‐oestrous phases, as well as on day 15 post‐coitum, plasma noradrenaline showed an increase, although the increase in pregnant females was greater.3 Adrenocorticotrophic hormone (ACTH) administration to non‐pregnant and pregnant females increased plasma adrenaline values significantly, compared with control values. Treatment of adrenalectomized rats with hydrocortisone lowered plasma noradrenaline to control values in both non‐pregnant and pregnant females.4 Hypophysectomy produced a decrease in plasma adrenaline content, whilst ACTH treatment increased it. Plasma adrenaline disappeared completely after adrenalectomy, during the pro‐oestrous phase and pregnancy, but extirpation of adrenals during the dioestrous phase still left appreciable amounts of adrenaline in plasma. Hydrocortisone treatment of both non‐pregnant and pregnant adrenalectomized females resulted in the reappearance of plasma adrenaline5 Adrenal stores of adrenaline and noradrenaline declined after hypophysectomy but the decreases were greater in pregnant females.6 Compensatory administration of ACTH to hypophysectomized females increased adrenaline and noradrenaline content of adrenals to control non‐pregnant female values during the pro‐oestrous phase of the oestrous cycle but similar treatment in pregnant females resulted in an increase in adrenaline.7 Although ACTH administration increased adrenal noradrenaline levels in pregnant hypophysectomized females, the mean value remained below that of non‐operated pregnant females.8 The results suggest that, during oestrous cycle and pregnancy the effects of pituitary and adrenal hormonal deficiency upon storage and release of monoamines differ quantitatively.

The role of glucocorticoids in the regulation of the diurnal rhythm of hepatic beta-hydroxy-beta-methylglutaryl-coenzyme A reductase and cholesterol 7 alpha-hydroxylase.

The microsomal activities of the hepatic enzymes hydroxymethylglutaryl-CoA reductase and cholesterol 7 alpha-hydroxylase exhibit a diurnal rhythm with maximum activities observed during the dark period and minimum activities around noon (12:00h). This diurnal rhythm was maintained for both enzymes after adrenalectomy, but the amplitude of variation for the activity of both enzymes was greatly decreased. A single injection of cortisol administered to adrenalectomized rats 3h before the expected maximum in enzyme activity resulted in a twofold increase in the activity of both enzymes 3h later, at values similar to those observed for control rats killed at the same time. This response appeared to require protein synthesis, since it was blocked by actinomycin D. However, the administration of cortisol to adrenalectomized rats 3 h before the expected minimum did not result in significant change in the activity of hydroxymethylglutaryl-CoA reductase and cholesterol 7 alpha-hydroxylase 3 h later. Kinetic studies of cholic acid metabolism in vivo demonstrated that adrenalectomy results in a significant decrease in the rate of synthesis of cholic acid and a considerable decrease in the pool size of cholic acid and its metabolic products. Treatment of adrenalectomized rats with cortisol increased the rate oonsistent with the effects of adrenalectomy and cortisol treatment on the activity of cholesterol 7alpha-hydroxylase.

Activation and inactivation of phosphorylase and glycogen synthetase during perfusion of rat liver as influenced by epinephrine, glucagon and hydrocortisone

1. 1. The changes in phosphorylase activity and glycogen synthetase I (active form) activity during perfusion of rat liver were studied together with their responses to added epinephrine and glucagon. 2. 2. Phosphorylase activity of the rat liver from fed or fasted rats fell rapidly during perfusion, regardless of whether the perfusate was added with glucose or not. The addition of epinephrine or glucagon at the start or at 60 min of perfusion caused a prompt restoration of the initial high activity. Both glycogen breakdown and glucose liberation proceeded in parallel with the changes in phosphorylase activity. 3. 3. The I-form of glycogen synthetase in the liver from fed rats increased rapidly when the concentration of perfusate glucose was raised to near 10 mM. This increase was promptly prevented by the addition of epinephrine or glucagon. In contrast, glycogen synthetase of the liver from fasted rats responded to neither flucose nor epinephrine (or glucagon) during perfusion. 4. 4. Phosphorylase in the liver of fasted, adrenalectomized rats did not respond to a low concentration of epinephrine (3 · 10 −8 M) or glucagon (5 · 10 −9 M), but was increased by higher concentrations of the hormones. The treatment of adrenalectomized rats with hydrocortisone restored the response of liver phosphorylase to the low concentrations of the hormones. Thus, glucocorticoid plays a “permissive” role by increasing the affinity of liver phosphorylase to epinephrine or glucagon.

Suppression of acute and chronic inflammation in adrenalectomized rats by pharmacologic amounts of prostaglandins.

AbstractProstaglandin E1 (PGE1) suppressed acute carrageenan‐induced inflammation and chronic joint inflammation characteristic of adjuvant disease in the rat. Placement of PGE1 into subcutaneous air blebs at the time of carrageenan injection retarded movement of polymorphonuclear leukocytes into the focus of inflammation and produced a concentration‐dependent reduction in exudate β‐glucuronidase and lactate dehydrogenase activities, an effect also observed in adrenalectomized rats. PGE2 also significantly reduced enzyme activities released into inflammatory exudates. Ultrastructural studies indicated that carrageenan was ingested by invading polymorphs and enclosed within phagocytic vacuoles, the membranes of which subsequently ruptured. More lysosomes remained intact after carrageenan uptake in bleb leukocytes from PGE1‐treated than from control rats and there was no loss of phagosomal membrane integrity in cells from treated rats. Treatment of adrenalectomized rats with PGE1 (300 μg twice daily) prevented adjuvant‐induced systemic arthritis, reduced the acute inflammatory reaction at the site of adjuvant injection, and reduced the number of circulating small lymphocytes. Together, these two sets of experiments suggest that pharmacologic doses of prostaglandins are anti‐inflammatory in acute and chronic models, and that the effect is not mediated entirely by stimulation of the pituitary and/or adrenal glands.

Adaptive response of hepatic carbohydrate metabolism to oral administration of p, p′-1,1,1 -trichloro-2,2- bis ( p-chlorophenyl)ethane in rats

The effects of an acute dose of p, p′-1,l,1-trichloro-2,2- bis ( p-chlorophenyl)ethane ( p, p′-DDT) have been investigated on the activities of pyruvate carboxylase, phosphoenolpyruvate carboxykinase, fructose 1,6-diphosphatase and glucose 6-phosphatase in rat liver. Administration of p, p′-DDT (600 mg/kg, p.o.) produced 2- to 2.5-fold stimulation of these key gluconeogenic enzymes in 5 hr, although statistically significant increases could be detected at 1 hr. Blood glucose increased as early as 0.5 hr, attained peak values at 1 hr, and then declined to reach control levels m 3–5 hr. In contrast, hepatic glycogen decreased gradually to 47 per cent of the control values in 1 hr, and was restored by 3 hr following treatment with this insecticide. Administration of p, p′-DDT to adrenalectomized rats produced similar changes in blood glucose and hepatic glycogen, suggesting that the insecticide-induced hyperglycemia and glycogenolysis may not be mediated by a release of catecholamines from the adrenals. Doseresponse studies revealed that 100 mg/kg of p, p′-DDT was the minimal amount necessary to induce statistically significant increases in the activities of hepatic pyruvate carboxylase, phosphoenolpyruvate carboxykinase, fructose 1,6-diphosphatase and glucose 6-phosphatase. Maximal stimulation of the four key gluconeogenic enzymes was seen generally at 5 hr when rats were treated with a 400–600 mg/kg dose of p, p′-DDT. Daily administration of small doses of p, p′-DDT (5 or 25 mg/kg) for 45 days also resulted in significant enhancement in the activities of various hepatic gluconeogenic enzymes. Actinomycin D, cycloheximide or ethionine failed to affect the basal levels of either of these hepatic enzymes, but effectively reduced the insecticide-induced increases in various enzyme activities. Treatment of adrenalectomized rats with p, p′-DDT enhanced various hepatic enzymes to a degree similar to that observed in intact animals. Furthermore, administration of triamcinolone to p, p′-DDT-treated adrenalectomized rats did not potentiate the action of the insecticide on any of the gluconeogenic enzymes examined. Our results suggest that treatment with p, p′-DDT produces marked increases in the quartet of key, rate-limiting gluconeogenic enzymes in hepatic tissue which are not mediated through a release of corticosteroids from adrenal glands.

Effect of irradiation on the biosynthesis of liver ribonucleic acids in intact and adrenalectomized rats.

Intact and adrenalectomized rats were irradiated with 900 R or treated with 10 mg per 100 g body weight of hydrocortisone. The incorporation of orotic acid-6-14C for 2 hours into liver free uridine nucleotides, nuclear and cytoplasmic RNA and RNA fractions obtained by agar gel electrophoresis, were studied. The obtained results show that in intact animals both irradiation and hydrocortisone induce a higher labelling of cytoplasmic and nuclear liver RNA. The higher labelling of RNA is not correlated with a higher labelling of the free uridine nucleotides. The labelling of all electrophoretic RNA fractions is increased to approximately the same extent under the action of either hydrocortisone or irradiation. Irradiation or hydrocortisone treatment of adrenalectomized rats causes also a higher labelling of liver nuclear and cytoplasmic RNA. The higher labelling of RNA is not correlated with that of free uridine nucleotides and affects all electrophoretic RNA fractions.. The combined action of irradiation and hydrocortisone shows an additive effect on the labelling of nuclear and cytoplasmic liver RNA. The obtained results indicate that whole body irradiation causes an increased synthesis of both ribosomal and non-ribosomal RNA’s of rat liver. Since the same effect is observed in intact and adrenalectomized animals it may be concluded that the stimulation of liver RNA synthesis by irradiation is not mediated by the adrenals.

In vivo studies on the mechanism of action of 6-dehydro-16-methylene-hydrocortisone (STC 407) on the hypothalamo-pituitary-adrenal axis in rats.

The mechanism of action of a new synthetic steroid, 6-dehydro-16-methylene-hydrocortisone (STC 407), which has been found useful in the treatment of Cushing’s syndrome due to adrenal hyperplasia, was studied in the rat. After treatment of normal rats with STC 407 for 11–14 days, the CRF content of the median eminence of the hypothalamus (SME), the concentration of ACTH in the anterior pituitary, the wet weight of the adrenal glands/ 100 g b.w., the adrenal corticosterone concentration/100 mg, and the plasma corticosterone level were significantly decreased. After treatment of adrenalectomized rats with STC 407 for 4 days, the concentration of ACTH in the pituitary was increased, but there was no appreciable change in CRF activity in the SME. Plasma levels of ACTH, which were elevated after adrenalectomy, decreased to undetectable levels after 4 days’ treatment with 400 <i>µ</i>g/100 g b.w. daily. Treatment of adrenalectomized rats with this steroid for 13 days considerably reduced CRF activity in the SME, and significantly raised the pituitary ACTH content. These results indicate that STC 407 suppresses both the release and the synthesis of ACTH by the pituitary in normal rats, this effect being mediated at least in part by a reduction in hypothalamic CRF activity. In adrenalectomized rats, the predominant effect of this steroid seems to be suppression of release of ACTH by the pituitary.