Topical anti-inflammatory activity of esters of steroid 21-oic acids

Prednisolone derivatives, methyl 20 alpha- and 20 beta-dihydroprednisolonate and methyl 17,20 alpha- and 17,20 beta-acetonidodihydroprednisolonate have been evaluated for their topical anti-inflammatory activity in the croton oil induced ear edema test. The order of anti-inflammatory potency was prednisolone greater than methyl 17,20 alpha-acetonidodihydroprednisolonate greater than methyl 17,20 beta-acetonidodihydroprednisolonate greater than methyl 20 beta-dihydroprednisolonate greater than methyl 20 alpha-dihydroprednisolonate. This order was paralleled by the compounds' octanol-aqueous partition coefficients. Furthermore, after two consecutive days topical administration of an equipotent anti-inflammatory dose, only prednisolone significantly decreased plasma corticosterone levels and relative thymus weight, while the new steroid derivatives had no effect on these parameters, indicating their lack of systemic side effects.     

Comparison of the biopotency of corticosterone and dexamethasone acetate in glucocorticoid receptor down regulation in rat liver

The effects of long treatment with dexamethasone 21-acetate and corticosterone on the glucocorticoid receptor in rat liver cytosol were compared. Dexamethasone acetate (5 micrograms/ml or 10 micrograms/ml water) or corticosterone (100 micrograms/ml water) was given to adrenalectomized animals as drinking solution for 6 days, and glucocorticoid receptor concentration was determined at 0, 12, 24, 48 and 72 h after steroid withdrawal. Dexamethasone acetate caused a dose dependent depletion of cytosol receptor. There was no measurable binding at time 0; the values of Bmax for the glucocorticoid receptor with decreased at 12, 24 and 48 h after the steroid withdrawal. Increased dissociation constant (Kd) were calculated for 12 and 24 h samples. The effect of corticosterone on receptor depletion was less pronounced. Bmax for the receptor was decreased at 0, 12, 24 h after steroid withdrawal with no change in Kd. The extent of steroids-induced receptor depletion showed good correlation with the induction of tyrosine aminotransferase (TAT), however, maximum TAT activity measured immediately after withdrawal of dexamethasone acetate was lower than that found after a single injection of dexamethasone acetate. We conclude that both steroids cause down regulation of the glucocorticoid receptor in rat liver cytosol, with both the extent and the duration of depletion being dependent on the biopotency of the glucocorticoid.     

Hypothalamic-pituitary-adrenal axis following glucocorticoid prophylaxis against acute mountain sickness.

The pituitary-adrenocortical and adrenomedullary response to high altitude (HA) stress was studied following daily single dose administration of prednisolone as a prophylaxis against altitude-induced acute mountain sickness (AMS). Forty healthy men, randomly divided into two groups of twenty, received placebo or prednisolone 20 mg once a day at 08.00 h for two days prior to induction to HA and during an initial three days stay at an altitude of 3450 m. The AMS score and circulatory levels of ACTH, cortisol, epinephrine and norepinephrine were measured at sea level (SL) and during residency at HA. The sensitivity of the hypothalamic-pituitary-adrenal axis in subjects receiving prednisolone therapy was evaluated at SL and on day four of stay at HA. Administration of prednisolone significantly (p < 0.01) decreased the severity of AMS in all the subjects. The steroid dose used did not inhibit endogenous secretion of ACTH, cortisol, epinephrine or norepinephrine, as HA response to adrenocortical and adrenomedullary hormones was identical in placebo and prednisolone treated subjects. The integrity of the hypothalamic-pituitary-adrenal axis was maintained well in subjects receiving low dose prednisolone therapy. These observations suggest that short-term administration of prednisolone is able to curtail AMS without causing suppression of the hypothalamic-pituitary-adrenal axis.     

Effect of clorgyline and of alpha-methyl-p-tyrosine on the plasma levels of corticosterone in the rat

The effect of a type A MAO inhibitor, clorgyline, injected alone or with alpha-methyl-p-tyrosine (aMpT) on the plasmatic corticosterone levels estimated at 9 a.m. and 5 p.m. has been studied in male Wistar rats. The clorgyline injected alone produced significant decreases in corticosterone values, especially at 5 p.m., determining a variation lack between the morning and the afternoon levels. When alpha-MpT is associated to IMAO, increases at both points of the day considered in this experiment take place. The amount of NA and 5-HT in the brain was also estimated; clorgyline high increases in Na and 5-HT contents, 5-HT; aMpT reduces the effect of clorgyline, especially as regards content. The results are discussed in relation to this highly specific MAO inhibitor and with the role of these amines as modulators of ACTH secretion. In view of the changes introduced by the aMpT injection, the modifications produced by clorgyline alone are related to NA, but to 5-HT, when the NA synthesis has been interrupted by the tyrosine hydroxylase inhibitor.     

Role of brain noradrenergic mechanisms in disturbing the circadian rhythm of the functioning hypothalamo-hypophyseo-adrenal cortex system in white rats in early ontogeny

The effects of hormone action and disturbance in catecholamine synthesis in the early postnatal ontogenesis on the circadian rhythm in the hypothalamic-hypophysial-adrenocortical system function were compared in the adult albino rat males. Injection of prednisolone on the 17-19th days of life blocked completely the diurnal rhythm of the corticosterone basal level in blood, the rhythm of adrenocortical response to an emotional stressor and to injection of noradrenaline into the brain lateral ventricle in 3-4 month old animals. Injection of an inhibitor of tyrosine hydroxylase, alpha-methyl-p-tyrosine, at the same period resulted in disappearance of the diurnal rhythm of the corticosterone basal level in adult animals, although the rhythm of response to an emotional stressor or injection of noradrenaline into the brain remained unchanged. A conclusion has been reached that disturbances in catecholamine synthesis in the early postnatal period induces long-term changes of predominantly tonic corticosterone secretion, while the hormone action on the circadian rhythm of the corticosterone basal level and stress response is only partly due to changes in noradrenergic regulation of the hypothalamic-hypophysial-adrenocortical system.     

Facilitation of Feedback Inhibition Through Blockade of Glucocorticoid Receptors in the Hippocampus

In the present study the effects of intracerebroventricular (icv) and intrahippocampal administration of corticosteroid antagonists on basal hypothalamic-pituitary-adrenal (HPA) activity around the diurnal peak were compared in male Wistar rats. In two separate experiments the glucocorticoid receptor (GR) antagonist RU 38486 and the mineralocorticoid receptor (MR) antagonist RU 28318 were tested. One hour after GR antagonist injection, significant increases in plasma ACTH and corticosterone levels were observed in the icv treated rats, when compared to vehicle. In contrast, a significant decrease in ACTH levels, and a slight, but non-significant decrease in corticosterone concentrations were attained one hour after intrahippocampal injection of the GR antagonist. Injection of the MR antagonist, on the other hand, resulted in enhanced ACTH and corticosterone levels irrespective of the site of injection. These findings suggest that negative feedback inhibition at the circadian peak involves hippocampal MRs and extrahippocampal (hypothalamic) GRs. The latter feedback inhibition overrides a positive feedback influence exerted by endogenous corticosteroids through hippocampal GRs.     

Effect of ACTH, dexamethasone, and adrenalectomy on the secretion of testosterone in the rat

The effect of ACTH (100 micrograms/animal/day, i.p.), dexamethasone (75 micrograms/animal/day, s.c.), both for three consecutive days, and adrenalectomy, with or without dexamethasone, maintained according to the group, one, two or three days, on the plasmatic testosterone and corticosterone levels, has been studied in adult male Wistar rats. ACTH and adrenalectomy produced a high decrease in testosterone levels (p less than 0.001 for the three days studied). Dexamethasone produced lower testosterone levels in the first day followed by partial recuperation between the second and the third days of its administration. Dexamethasone produced the effects mentioned for intact animals. The changes in corticosterone levels were according to an adequate response of the hypothalamus-pituitary-adrenal system under these experimental circumstances. ACTH exerts an inhibitory effect on testosterone secretion in the rat, so that such an effect from the data obtained after adrenalectomy and simultaneous dexamethasone injections, does not seems to be mediated either by the presence of adrenals or high corticosterone levels.     

Effects of bilateral olfactory bulbectomy on the anterior pituitary corticotropic cell activity in male rats.

Bilateral olfactory bulbectomy (OB) has drastic biochemical and behavioral effects and is often associated with an increase in plasma corticosterone concentrations. This experiment examined the effects of OB on adrenocorticotropin (ACTH) and corticosterone release under basal and stress conditions and on proopiomelanocortin (POMC) gene expression. Bulbectomy potentiated hypophysal ACTH and adrenal corticosterone release induced by ether stress but had no effect on ACTH release under basal conditions, despite a significant increase of circulating corticosterone. POMC gene expression was stronger (+60%) in OB rats than in sham-operated rats. These results suggest that olfactory bulbectomy substantially altered the negative feed-back exerted by glucocorticoids on anterior pituitary corticotropic cells in the male rat.     

Involvement of corticosteroids in the processing of stressful life-events. A possible implication for the development of depression

In a sub-population of endogenously depressed patients, disturbances of the hypothalamic-pituitary-adrenal axis can be observed. Increased cortisol and CRH levels combined with normal ACTH concentrations have often been reported. Corticosteroids appear to play a role in the mood changes, in depressed subjects. However, their mechanism of action is unknown. In animal experiments, the involvement of corticosteroids in stressor-induced learning was investigated. Three paradigms were used. In the Porsolt swimtest an animal had to learn to adapt to an inescapable situation. In the lithium chloride conditioned taste aversion an animal learned to avoid sugar water. In the amphetamine sensitization a second injection of amphetamine caused a potentiated response, because of conditioning. All three conditions appeared to be stressful because they induced a corticosterone release. When adrenalectomized (ADX) mice were compared to control animals it appeared that, in all three paradigms, their memory function was disturbed. The data indicated that this was a specific glucocorticoid-mediated effect since corticosterone and dexamethasone injections were able to reverse the ADX-induced deficit. The ADX-induced disturbances were only observable at moderate stress levels. More severe stressor (lower water temperature in the Porsolt swimtest, higher lithium chloride and amphetamine doses) also made ADX mice remember their previous experiences. The results suggest that corticosteroids are involved in the consolidation of stressful events and the corresponding coping responses. They play, however, only a role in the case of moderate stressors. In ADX animals no stressor-induced corticosterone increase can occur and therefore these animals only remember severe stressors. In a depressed patient basal steroid levels are increased and consequently very mild stressors, which induce only a small extra steroid release, will be remembered. The remembering of all these negative experiences might be of importance for the development and maintenance of the depression.     

Effects of U-50,488H and U-50,488H withdrawal on catecholaminergic neurons of the rat hypothalamus

Previous report from our laboratory showed that morphine produces a stimulatory effect of hypothalamic noradrenaline (NA) turnover concurrently with enhanced pituitary-adrenal response after its acute injection and during withdrawal. In the present work we have studied the effects of acute and chronic administration of the kappa agonist U-50,488H as well as the influence of U-50,488H withdrawal on the activity of hypothalamic NA and dopamine (DA) neurons and on the activity of hypothalamic-pituitary-adrenal (HPA) axis. A single dose of U-50,488H (15 mg/kg i.p.) significantly increased hypothalamic NA and decreased DA turnover at the time of an enhanced corticosterone release. Rats rendered tolerant to the kappa agonist by administration of U-50,488H twice a day for 4 days showed no changes in corticosterone secretion. Additionally, a decrease in both hypothalamic MHPG (the cerebral NA metabolite) production and NA turnover was observed, whereas DOPAC concentration and DA turnover were enhanced, which indicate the development of tolerance towards the neuronal and endocrine actions of U-50,488H. After naloxone (3 mg/kg s.c.) administration to U-50,488H-tolerant rats, we found neither behavioural signs of physical dependence nor changes in hypothalamic catecholaminergic neurotransmission. In addition, corticosterone secretion was not altered in U-50,488H withdrawn rats. Present data clearly indicate that tolerance develops towards the NA turnover accelerating and DA turnover decreasing effect of U-50,488H. Importantly and by contrast to mu agonists, present results demonstrate that U-50,488H withdrawal produce no changes in hypothalamic catecholamines turnover or in corticosterone release (an index of the hypothalamus-pituitary-adrenal activity), which indicate the absence of neuroendocrine dependence on the kappa agonist. As has been proposed, this would suggest that the mu and the kappa receptor be regulated through different cellular mechanisms, as kappa agonists have a lower proclivity to induce dependence.     

Antagonistic effects of aldosterone on corticosterone-mediated changes in exploratory behavior of adrenalectomized rats

The effect of aldosterone administration on exploratory activity of chronic adrenalectomized (10 days) male rats was investigated. Aldosterone (30 μg/100 g body wt sc) administered 1 hr or 30 min prior to the behavioral test failed to normalize disturbed exploratory activity of adrenalectomized rats, in contrast to the restoration observed after corticosterone, the naturally occurring glucocorticoid of the rat. Administration of the mineralocorticoid 30 min prior to corticosterone prevented the normalization of the behavioral response by the latter steroid. Administration of the same dose of aldosterone 30 min prior to a tracer amount of [³H]corticosterone effectively blocked cell nuclear uptake of radioactive-labeled hormone in the hippocampus. The specific action of corticosterone on exploratory behavior corresponds with the stringent specificity of the neuronal hippocampal corticosterone receptor system. Mineralocorticoid receptors do not seem to be involved in effects on this behavior. The antagonistic action of aldosterone is probably exerted by competitive binding to the corticosterone receptor.     

Stress triggered changes in expression of genes for neurosecretory granules in adrenal medulla

With acute stress, the release of adrenomedullary catecholamines is important for handling the emergency situation. However, when chronic or repeated, stress alters the allostatic load and leads to a hyperadrenergic state, resulting in the development or worsening of a wide range of diseases. To help elucidate the mechanism, we examined the effects of single and repeated immobilization stress on gene expression of components of neurosecretory vesicles in the adrenal medulla. Male Sprague-Dawley rats were exposed to immobilization stress once for 2?h (1× IMO) or daily for six consecutive days (6× IMO). Compared to unstressed controls, 1× IMO elevated gene expression of vesicular monoamine transporter 2 (VMAT2). In response to 6× IMO, not only was VMAT2 mRNA still elevated, but chromogranin A (CgA) and chromogranin B (CgB) mRNAs were also increased two to three-fold above basal levels. To investigate the possible role of the hypothalamic-pituitary-adrenal axis in the induction of VMAT2, PC12 cells were treated with the synthetic glucocorticoid dexamethasone, which was found to elevate VMAT2 mRNA expression. The findings suggest that following repeated stress, elevations of various components of neurosecretory vesicles in the adrenal can facilitate more efficient utilization of the well-characterized heightened catecholamine levels.     

Corticosteroid actions from neuronal membrane to behavior: neurophysiological mechanisms underlying rapid behavioral effects of corticosterone.

Investigation of the rapid suppression of male courtship clasping behavior by corticosterone in roughskin newts (Taricha granulosa) has led to the identification of a specific neuronal membrane receptor for this stress steroid. This paper describes studies of the neurophysiological effects of the rapid, membrane receptor mediated action of corticosterone on neurons that are involved in the control of clasping. In freely behaving newts, medullary neurons, including reticulospinal neurons, process clasp-triggering sensory signals and participate in control of clasping movements. Corticosterone injection causes these brainstem neurons to show selective depression of clasping-related sensorimotor function. These corticosterone effects appear in 3-10 min and are closely associated with the simultaneous depression of clasping. In addition to these functionally specific effects, corticosterone simultaneously causes widespread, primarily depressive effects on neuronal activity and excitability in the medulla and elsewhere in the brain. Thus, the membrane actions of corticosterone lead to diverse neural effects, including changes in membrane excitability as well as specific, network-level actions that are apparent only during behavior. These rapid corticosterone effects strongly interact with actions of the neuropeptides vasotocin and corticotropin-releasing factor, such that the form and magnitude of the steroid's effects depend on the prevailing neuroendocrine state of the brain.     

The effect of intrastriatal injection of liposome-entrapped tyrosinase on the dopamine levels in the rat brain.

Parkinson's disease is a neurodegenerative disorder which is mainly characterized by degeneration of the dopaminergic cells in the nigro-striatal system. Due to a lowered L-tyrosine 3-monooxygenase activity, L-tyrosine is not sufficiently transformed to L-DOPA. To date the most common therapy is the administration of the dopamine precursor L-DOPA, with severe collateral effects. Therefore, the substitution of the lacking tyrosine hydroxylase with tyrosinase might be a novel therapeutical approach that would generate specifically L-DOPA from L-tyrosine. We present here evidence that stereotaxic injection of liposome-entrapped tyrosinase is able to significatively increase the levels of dopamine in the rat brain. The catecholamines L-DOPA, dopamine, L-epinephrine, L-norepinephrine were extracted by acid treatment from the brains and detected by HPLC.     

Systemic activities of a new anti-inflammatory steroid: Methyl 20-dihydroprednisolonate

The systemic activities of methyl 20-dihydroprednisolonate (1), a new local anti-inflammatory steroid synthesized by modifying the 17β-ketol side-chain of prednisolone, on pituitary-adrenal function and liver glycogen content were investigated in rats. The parent compound, prednisolone, administered intramuscularly, caused a significant doserelated decrease in plasma levels of corticosterone, adrenocorticotropic hormone (ACTH), and liver glycogen content in rats. In contrast, methyl 20-dihydroprednisolonate caused mild PA suppression and liver glycogen depletion only at high doses. The steroid acid ester did not exert glycogenic activity, unlike prednisolone, in the adrenalectomized rats.     

Repeated stress alters the ability of nicotine to activate the hypothalamic-pituitary-adrenal axis

Acute nicotine administration has been shown to activate the hypothalamic-pituitary-adrenal (HPA) axis and stimulate secretion of adrenocorticotrophic hormone (ACTH), corticosterone/cortisol and beta-endorphin (beta-END) in both rodents and humans, raising the possibility that activation of the HPA axis by nicotine may mediate some of the effects of nicotine. Since stress can increase the risk of drug use and abuse, we hypothesized that repeated stress would increase the ability of nicotine to stimulate the secretion of HPA hormones. To test our hypothesis, mice were exposed to repeated stress (swimming in 15 degrees C water for 3 min/day for 5 days) and killed 15 min after injection of saline or nicotine (0.1 mg/kg, s.c.). Repeated exposure to stress increased the ability of nicotine to stimulate plasma ACTH (p<0.05) and beta-END (p<0.05), but not corticosterone secretion. In contrast, repeated exposure to stress increased the post-saline injection levels of corticosterone (p<0.05), but not ACTH and beta-END. The present results suggest that chronic stress leads to an enhanced sensitivity of some components of the HPA axis to a subsequent nicotine challenge.     

Effects of corticosterone on the negative feedback action of testosterone, 5 alpha-dihydrotestosterone and estradiol in the adult male rat

Corticosterone acetate (10 mg/day) was administered to gonadectomized and adrenalectomized male rats bearing 5, 10 or 15 mm long testosterone filled silicone elastomer capsules. It was found that the serum testosterone levels induced by these capsules were not influenced by corticosterone treatment and that the weights of the prostates in the corticosterone treated rats were not different from their controls. In contrast, corticosterone acetate increased markedly the LH and FSH inhibitory effects of testosterone. Since several brain structures are able to convert testosterone into 17-beta-hydroxy-5-alpha-androstan-3-one (5-alpha-dihydrotestosterone) and/or estradiol, and these metabolites are probably involved in mechanisms controlling gonadotropin secretion, we studied also the effects of corticosterone on the feedback action of dihydrotestosterone and estradiol. 5 alpha-Dihydrotestosterone was administered by 5, 10 or 20 mm long elastomere capsules whereas estradiol was given by daily s.c. injections of 0.125, 0.25 or 0.50 micrograms estradiol benzoate. In the presence of corticosterone acetate the gonadotropin inhibitory action of testosterone, 5 alpha-dihydrotestosterone and estradiol increased more than 2 times.     

Adrenalectomy does not change CRF secretion induced by interleukin-1 from rat perifused hypothalami.

Interleukin-1 (IL-1) is a potent hypothalamic-pituitary-adrenal (H-P-A) axis activator. The hypothalamus is considered one of the main sites of action of IL-1 on the H-P-A axis, inducing CRF secretion, which is modulated by glucocorticoids. Glucocorticoids, which modulate CRF release by a negative feedback inhibition, have been postulated to exert a permissive action on the IL-1 effect on CRF secretion. Using a continuous perifusion system of rat hypothalami, the results of the present study indicate that at the same concentrations, IL-1 beta exerted a more potent effect than IL-1 alpha stimulating CRF secretion. The increase in hypothalamic CRF release induced by IL-1 was rapidly inhibited by both dexamethasone and corticosterone. However, adrenalectomy 2 or 8 days before did not modify CRF secretion induced by IL-1 from the in vitro perifused hypothalami. These data indicate that IL-1 does not seem to induce CRF secretion by interfering with an impeding action of glucocorticoids, although the cytokine effect is negatively modulated by corticosteroids.     

Cerebral serotonin and the hypothalamo-hypophyseal-adrenal system of the rat during aging]

The role of brain serotonin (5HT) on the hypothalamus-pituitary-adrenal system (HPAs) under basal condition and after injections of p-chlorophenylalanine (pCPA) and L-5-hydroxytryptophan (L-5HTP) has been studied in 6, 12 and 28 month old male Wistar rats. Four experimental groups were made for each age: control, saline, injected with pCPA (250 mg/kg i.p.) and L-5HTP (200 mg/kg i.p.), the effects being valued 2 hours after L-5HTP administration and 24 hours after pCPA injection. In all groups the plasmatic ACTH, the corticosterone levels as well as the simultaneous changes of the 5TH content tryptophan hydroxylase activity in whole brain were estimated two hours after the L-5HTP injection and 24 hours after that of pCPA. Significant changes are not found in the plasmatic ACTH and corticosterone values with respect to age under basal condition. Nevertheless, the response of HPAs differs with the age after pCPA or L-5HTP injection. The ACTH and corticosterone levels augment by L-5HTP and decrease by pCPA in all age groups, but this corresponding increase or decrease was less marked in the older rats. The 5HT content as tryptophan hydroxylase activity in brain decreased in old animals. pCPA and L-5HTP determine, respectively, high falls and rise of 5TH values, these changes being more intense for pCPA in old rats and for L-5HTP in young and mature animals. The tryptophan hydroxylase activity is decreased by pCPA as L-5HTP injections.(ABSTRACT TRUNCATED AT 250 WORDS)     

The influence of sex and gonadectomy on the growth hormone and corticosterone response to hexarelin in the rat

The present study is designed to investigate the role of sex and gonadal status in the growth hormone (GH) and corticosterone response to hexarelin (HEXA), a GH-releasing peptide, which also causes a stimulatory action on the hypothalamic-pituitary-adrenal (HPA) axis. HEXA (80 microg/Kg) was administered intracarotid to anesthetized intact or gonadectomized male (ORC) and female (OVX) middle-aged rats. The GH stimulatory response to HEXA was gender-related since the GH increase was significantly (p < 0.001) higher in intact males (area under the curve, AUC= 12560 +/- 1784 ng/ml.45 min) than in females (AUC= 4628 +/- 257 ng/ml.45 min). This sex difference does not depend on circulating gonadal steroids since it persists in ORC (AUC = 11980 +/- 1136 ng/ml.45 min) and OVX (AUC = 5539 +/- 614 ng/ml.45 min) rats. The different effects of HEXA on corticosterone secretion detected in male and female rats are probably dependent on the prevailing activity of the HPA axis. In fact, in male rats that have low basal corticosterone levels, HEXA caused an increase in corticosterone secretion, which was significantly (p< 0.05) higher in ORC than in intact rats. The increase in corticosterone secretion by HEXA both in intact and OVX females was delayed, probably due to the elevated initial corticosterone levels, which could have activated the glucocorticoid negative feedback. We suggest that gender-specific patterns in the regulation of the hypothalamus-pituitary function could be responsible for the GH and corticosterone sexually differentiated responses to HEXA.