Function of hypothalamic-pituitary-adrenocortical system in hypertensive rats of ISIAH strain

Functional activity of hypothalamic-pituitary-adrenocortical axis has been studied under control and restraint stress conditions in rats with inherited stress-sensitive arterial hypertension (ISIAH strain) and in normotensive WAG (Wistar Albino Glaxo) strain. The levels of hypothalamic CRH-mRNA (in control and 2 hrs stress), pituitary and plasma ACTH and plasma corticosterone (in control and after 5, 15 or 30 min of restraint stress), were evaluated. Hypothalamic CRH-mRNA level was found to be approximately the same in the control rats of both strains. In control conditions, the pituitary and plasma ACTH content in ISIAH rats was significantly lower whereas the corticosterone level in the plasma differed from each other in both strain. The restraint stress resulted in a statistically significant increase of the CRH-mRNA in ISIAH rats and not in the WAG rats. Moreover, in spite of the lower ACTH level in stressed ISIAH rats, the corticosterone blood plasma concentration in hypetensive rats was significantly higher. The data obtained confirm the idea that the stress-dependent hypertension might be related to an enhanced sensitivity of the main endocrine links involved in the stress response organization.     

Corticotropin-releasing hormone and pituitary-adrenocortical responses in chronically stressed rats

Brain corticotropin-releasing hormone (CRH) concentration and pituitary adreno-cortical responses were examined in chronically stressed rats: body restraint stress (6 h/day) for 4 or 5 weeks. Stressed rats showed a reduction in weight gain. CRH concentration in the median eminence and the rest of the hypothalamus were not different between control and chronically immobilized rats. The anterior pituitary adenocorticotropic hormone (ACTH) concentration was elevated in chronically stressed rats, whereas plasma ACTH and corticosterone levels did not differ from the control values. The median eminence CRH concentration was reduced to the same extent at 5 min after onset of ether exposure (1 min) in chronically immobilized rats and controls. However, plasma ACTH and corticosterone showed greater responses to ether stress in chronically immobilized rats than in control rats. Plasma ACTH and corticosterone responses to exogenous CRH were not different between control and chronically immobilized rats, while the response to arginine vasopressin (AVP) was significantly greater in chronically immobilized rats. These results suggest that chronic stress caused an increase in the ACTH-secreting mechanism and that pituitary hypersensitivity to vasopressin might at least be partly responsible for this.     

Repeated handling, restraint, or chronic crowding impair the hypothalamic-pituitary-adrenocortical response to acute restraint stress.

The purpose of the present study was to assess whether, and to what extent prior handling, restraint or social crowding stress during 3-10 days affects the hypothalamic-pituitary-adrenocortical (HPA) response to an acute short-lasting restraint stress. Also the effect of a feedback inhibitory mechanism of corticosterone in the impairment of HPA axis by these stressors was investigated. Male Wistar rats were pretreated with handling 1 min/day for 3-10 days, restraint 2 times daily for 3-7 days and crowding stress for 7 days before exposure to acute restraint stress in metal tubes for 10 min. Some group of rats received exogenous s.c. corticosterone either once 25 mg/kg or 2 times daily 10 mg/kg for 3-10 days before restraint stress. After the last restraint the rats were decapitated and their trunk blood was collected for the measurement of plasma ACTH and serum corticosterone levels. Handling for 3-7 days, restraint for 3-7 days, and crowding for 7 days and a single pretreatment with corticosterone--all significantly and to a similar extent inhibited the restraint stress-induced increase in ACTH and corticosterone secretion. Chronic pretreatment with corticosterone blunted the restraint stress-induced increase in HPA axis activity. These results indicate that repeated short-lasting stress induced by handling, restraint, or crowding potently attenuates the acute restraint stress-induced stimulatory action of the HPA axis. They also indicate adaptive action of moderate stress on the HPA axis response to acute stress. The results also suggest that a short-lasting hypersecretion of corticosterone during psychological stress may induce a prolonged feedback inhibition of the HPA axis activity. The attenuation of HPA axis response by prior handling has also obvious methodological implications.     

Adenohypophyseal hormone response to chronic stress in dexamethasone-treated rats.

The influence of dexamethasone treatment on the basal values of corticosterone, GH, prolactin (PRL), LH and FSH, as well as on the adenohypophyseal hormone response to chronic stress was studied in female rats. Dexamethasone acetate (25 micrograms/100 b.w.), given by gavage twice daily for 10 days, decreased the resting plasma levels of corticosterone, GH, LH and PRL, whereas the FSH titers remained normal. The secretion of ACTH (evaluated indirectly through corticosterone concentrations) and of GH appeared to be most sensitive to the suppressive effect of dexamethasone. The same hormonal response pattern was induced by 8 h of daily immobilization for 10 days, except that ACTH release was enhanced and the plasma LH titers dropped more drastically. Dexamethasone administration in combination with restraint did not alter the characteristic hormonal profile of chronic stress, despite the fact that ACTH secretion was completely blocked. These data suggest that the inhibition of PRL, LH and GH secretion following severe, chronic stress is not causally related to the sustained elevation of plasma ACTH.     

Effect of synthetic atrial natriuretic polypeptide on hemorrhage-induced adrenocorticotropin secretion of the rat

The effect of synthetic alpha-human atrial natriuretic polypeptide (alpha-hANP) on the in vivo and in vitro release of ACTH and corticosterone was examined. In the in vivo study ACTH and corticosterone responses to rapid 2-ml/rat hemorrhage were measured in sixteen conscious rats after alpha-hANP administration. The hemorrhage increased plasma ACTH and corticosterone concentrations in the control group of rats (p greater than 0.01). ANP inhibited hemorrhage-induced ACTH secretion (p less than 0.05), but the plasma corticosterone response was not affected. In the in vitro study a high concentration of ANP (1 microM) reduced basal corticosterone secretion from the isolated rat adrenal gland (p less than 0.05), but the response to ACTH (10 ng/ml) and dibutyryl cyclic AMP (0.5 mM, 5.0 mM) was not affected. Our data suggest that ANP inhibits hemorrhage-induced ACTH secretion from the anterior pituitary but inhibits corticosterone secretion from the adrenal gland very weakly.     

In situ detection of cyclic AMP-phosphodiesterase activity in the heart of Lewis and Sprague-Dawley rats: the effect of restraint stress or amphetamine application

Cyclic AMP plays an important role in heart functions under normal as well as pathological conditions. Since phosphodiesterase (PDE), responsible for the hydrolysis of cAMP, is equally important as synthesizing adenylyl cyclase, we decided to determine its activity by cytochemical procedure after exposure of rats to restraint stress or an acute dose of amphetamine. Sprague-Dawley (S-D) and Lewis (LE) rats, the latter known to have a deficient hypothalamo-pituitary-adrenal axis activity, were used in order to disclose the possible significance of rat strain on PDE activity. Animals were divided into 3 groups: controls, rats treated with an acute dose of amphetamine (8 mg/kg, i.p., for 60 min) and rats under restraint stress for 60 min. Control hearts of both strains revealed PDE activity on sarcolemma of cardiomyocytes and plasmalemma of endothelial cells of microvessels. In LE rats we observed an additional enzyme reaction in junctional sarcoplasmic reticulum. In addition, cardiomyocytes of LE rats revealed a higher PDE activity when compared to S-D rats. Restraint stress decreased PDE activity in cardiomyocytes of LE rats while amphetamine markedly inhibited enzyme activity in cardiomyocytes of S-D rats. Endothelial PDE was more resistant to stress. Our results indicate differences in PDE localization and variations in sensitivity of myocardial cAMP-PDE of LE and S-D rat strains to restraint stress and amphetamine application.     

Dissociation between adrenocorticotropin and corticosterone responses to restraint after previous chronic exposure to stress

The effect on emotional reactivity produced by a model for chronic stress in which different types of acute stresses were randomly combined for 29 days was studied in adult male rats. Chronically stressed rats showed a slight decrease in body weight gain and an increase in relative adrenal weight. Chronic stress did not modify defecation rate but reduced exploratory activity in the holeboard. Neither basal nor acute-stress induced levels of adrenocorticotropin (ACTH) were modified by previous chronic stress. Likewise, basal corticosterone levels were similar in both groups. However, corticosterone response to acute restraint stress was higher in chronically stressed than in control rats. The mechanisms underlying the dissociation between ACTH and corticosterone as well as its possible implications are discussed.     

Time course of vasopressin and oxytocin secretion after stress in adrenalectomized rats.

To characterize the participation of vasopressin (AVP) and oxytocin (OT) in hypothalamus-pituitary-adrenal regulation after adrenalectomy (ADX), we evaluated corticosterone, ACTH, AVP and OT plasma concentrations and AVP and OT content of the paraventricular nucleus (PVN) at different periods (3 h, 1, 3, 7 and 14 days) in sham or ADX rats under basal conditions and after immobilization stress. ADX animals showed undetectable corticosterone levels, while sham animals showed a marked increase in corticosterone and ACTH 3 h after surgery, then lowering to basal control levels. ADX rats showed high basal ACTH levels with a triphasic response without changes after immobilization. After three hours, the ADX group showed higher OT levels than the sham group. OT was increased after immobilization stress in sham and ADX groups. AVP plasma levels did not change throughout the basal or stress studies in either group. There was a decrease in hypothalamic AVP content 1 and 3 days after ADX under basal and stress conditions. Plasma osmolality showed a significant decrease in the ADX group at 3, 7, and 14 days. In conclusion, there are different pituitary-adrenal axis set points after removal of the glucocorticoid negative feedback. The role of vasopressinergic and oxytocinergic neurons in the ACTH secretion after ADX or immobilization stress appears to differ. Magnocellular AVP is unlikely to contribute to ACTH secretion in response to ADX or immobilization stress. On the other hand, OT is elicited by immobilization stress and might contribute to the ACTH secretion during short-term ADX.     

Involvement of the suprachiasmatic nucleus in diurnal ACTH and corticosterone responsiveness to stress

We explored the contribution of the suprachiasmatic nucleus (SCN) in ACTH and corticosterone (CORT) diurnal responsiveness of the rat to restraint stress applied either in the morning (AM) or in the evening (PM). Ablation of the SCN caused the diurnal rhythmicity of the CORT response to disappear but had no effects on AM vs. PM differences in the ACTH response. Stress-response curves in SCN-lesioned rats that had prestress levels of CORT either in the AM range or in the PM range, when compared with those obtained for AM and PM controls, showed that the SCN differentially regulates the stress response depending on the underlying secretory activity of the adrenal cortex. When basal CORT secretion is at its lowest, the SCN inhibits CORT responsiveness to stress by controlling pituitary corticotrophs; but when it is at its highest, it has a permissive action that will bypass the hypophysis and reach the adrenals to adjust the response of the gland to ACTH.     

Increased hypothalamic-pituitary-adrenal axis activity and hepatic insulin resistance in low-birth-weight rats

Individuals born with a low birth weight (LBW) have an increased prevalence of type 2 diabetes, but the mechanisms responsible for this association are unknown. Given the important role of insulin resistance in the pathogenesis of type 2 diabetes, we examined insulin sensitivity in a rat model of LBW due to intrauterine fetal stress. During the last 7 days of gestation, rat dams were treated with dexamethasone and insulin sensitivity was assessed in the LBW offspring by a hyperinsulinemic euglycemic clamp. The LBW group had liver-specific insulin resistance associated with increased levels of PEPCK expression. These changes were associated with pituitary hyperplasia of the ACTH-secreting cells, increased morning plasma ACTH concentrations, elevated corticosterone secretion during restraint stress, and an approximately 70% increase in 24-h urine corticosterone excretion. These data support the hypothesis that prenatal stress can result in chronic hyperactivity of the hypothalamic-pituitary-adrenal axis, resulting in increased plasma corticosterone concentrations, upregulation of hepatic gluconeogenesis, and hepatic insulin resistance.     

Psychosocial stress affects the involvement of prostaglandins and nitric oxide in the lipopolysaccharide-induced hypothalamic-pituitary-adrenal response.

The role of prostaglandins and nitric oxide (NO), generated after peripheral lipopolysaccharide (LPS) administration, in the adaptation of hypothalamic-pituitary-adrenal (HPA) axis under stressful circumstances remains to be elucidated. The aim of the present study was to assess the effect of chronic repetitive restraint or social crowding stress on the involvement of nitric oxide and prostaglandins in the LPS-induced pituitary-adrenocortical response. Male Wistar rats were restrained in metal tubes 2 x 10 min/day or crowded in cages for 7 days prior to treatment. All compounds were injected i.p., cyclooxygenase (COX) and nitric oxide synthase (NOS) inhibitors 15 min before LPS. Two hrs after injection LPS induced a significant increase in ACTH and corticosterone secretion. Repeated restraint impaired more potently than crowding stress the LPS-induced HPA-response. Indomethacin, a non-selective COX inhibitor, considerably reduced the LPS-induced HPA response in non-stressed rats and to a lesser extent diminished this response in repeatedly restrained or crowded rats. Neuronal NOS inhibitor, Nomega-nitro-L-arginine decreased the LPS-induced HPA response, more potently in control than crowded rats. Aminoguanidine, an iNOS inhibitor, diminished the LPS-elicited ACTH response in crowded rats. These results indicate that prostaglandins and NO generated by neuronal and inducible NOS are involved in the LPS-induced HPA axis response under basal conditions and during its adaptation to chronic social stress circumstances.     

Nitric oxide mediates the interleukin-1beta- and nicotine-induced hypothalamic-pituitary-adrenocortical response during social stress.

We investigated the role of nitric oxide (NO) in the interleukin 1beta (IL-1beta) and nicotine induced hypothalamic-pituitary-adrenal axis (HPA) responses, and a possible significance of CRH and vasopressin in these responses under basal and social stress conditions. Male Wistar rats were crowded in cages for 7 days prior to treatment. All compounds were injected i.p., nitric oxide synthase (NOS) inhibitors, alpha-helical CRH antagonist and vasopressin receptor antagonist 15 min before IL-1beta or nicotine. Identical treatment received control non-stressed rats. Plasma ACTH and serum corticosterone levels were measured 1 h after IL-1beta or nicotine injection. L-NAME (2 mg/kg), a general nitric oxide synthase (NOS) inhibitor, considerably reduced the ACTH and corticosterone response to IL-1beta (0.5 microg/rat) the same extent in control and crowded rats. CRH antagonist almost abolished the nicotine-induced hormone responses and vasopressin antagonist reduced ACTH secretion. Constitutive endothelial eNOS and neuronal nNOS inhibitors substantially enhanced the nicotine-elicited ACTH and corticosterone response and inducible iNOS inhibitor, aminoguanidine, did not affect these responses in non-stressed rats. Social stress significantly attenuated the nicotine-induced ACTH and corticosterone response. In crowded rats L-NAME significantly deepened the stress-induced decrease in the nicotine-evoked ACTH and corticosterone response. In stressed rats neuronal NOS antagonist did not alter the nicotine-evoked hormone responses and inducible NOS inhibitor partly reversed the stress-induced decrease in ACTH response to nicotine. These results indicate that NO plays crucial role in the IL-1beta-induced HPA axis stimulation under basal and social stress conditions. CRH and vasopressin of the hypothalamic paraventricular nucleus may be involved in the nicotine induced alterations of HPA axis activity. NO generated by eNOS, but not nNOS, is involved in the stress-induced alterations of HPA axis activity by nicotine.     

Atrial natriuretic factor is unlikely to be involved in the reduced aldosterone production in the Brattleboro rat

We have previously reported that basal and stimulated aldosterone production in Brattleboro rat (DI) lacking hypothalamic arginine vasopressin is lower than that observed in control Long-Evans rat (LE). In the present study, we investigated the secretion under various experimental conditions, adrenal binding sites, and the aldosterone-inhibiting effect of atrial natriuretic factor (ANF). In the conscious resting state, the plasma ANF concentration was similar between LE and DI rats. Pentobarbital anaesthesia (5 mg/100 g body wt.) reduced the plasma ANF concentration equally in both groups, with or without captopril pretreatment. Morphine (10 mg/100 g body wt.) increased ANF secretion dramatically and equally in the two groups of pentobarbital anaesthetized (2 mg/100 g body wt.) rats. In dexamethasone pretreated-pentobarbital anaesthetized rats, a concurrent i.v. ANF infusion (50 ng/min) did not change significantly the corticosterone response to ACTH (1-24) (1 mI.U./100 g body wt.) but steeply depressed ACTH-induced aldosterone production to a similar extent between DI and LE rats. A single class of adrenal ANF receptor sites was found with a similarity in high affinity and maximum binding capacity between the two groups of rats. Taken together, these results suggest that the reduced aldosterone production by Brattleboro rat adrenals is unlikely to be related to the inhibitory effect of ANF.     

Gonadotropin inhibition during chronic stress: role of the adrenal gland

The effect of adrenalectomy, metyrapone and dexamethasone treatments on gonadotropin response to chronic stress were studied. Adult male rats were submitted to chronic restraint (6 h daily over 4 days). At the end of the last stress period animals were decapitated and trunk blood was collected. Chronic restraint evoked a decrease in plasma LH and to a lesser degree in plasma FSH in the intact rat. Adrenalectomy did not prevent the LH reduction induced by stress and magnified the inhibitory effect of restraint on FSH secretion. Administration of the corticosterone synthesis blocker metyrapone increased the inhibitory effect of restraint on plasma LH and to a lesser degree on plasma FSH. Dexamethasone treatment did not significantly modify plasma gonadotropin levels in adrenalectomized unstressed rats, but this treatment totally blocked plasma LH and FSH reduction after chronic restraint. These results indicate that plasma LH and FSH reduction during chronic restraint is not due to the increase in glucocorticoid secretion, but seems to be mediated by the increase of the hypothalamic-pituitary components of the adrenal axis.     

Dissociation of oxytocin, vasopressin and corticotropin secretion during different types of stress

Oxytocin (OT), vasopressin (AVP), and corticotropin (ACTH) levels were measured in peripheral plasma of male rats subjected to one of three models of stress: restraint, cold, or ether. ACTH secretion was increased in all three groups compared to unstressed controls. OT secretion was increased in rats subjected to restraint or ether but not cold. AVP secretion was increased only by ether stress. The data suggest that the hypothalamic and neurohypophysial contribution to the control of ACTH secretion may vary in response to different types of stress.     

Inhibitory effect of ouabain on epinephrine-induced stimulation of adrenal corticosterone secretion in rats.

Chronic administration of ouabain (3 mg/Kg body weight, subcutaneously, once daily for consecutive 15 days) definitely inhibited epinephrine-induced increase of adrenal corticosterone secretion. The inhibition rate increased along with frequency of ouabain administration. Increase in adrenal corticosterone synthesis and secretion by ACTH (20-50 mU/rat) administration was partially suppressed by pretreatment with chronic ouabain administration. A slight but significant increase of adrenal corticosterone secretion caused by epinephrine administration in hypophysectomized rats was also inhibited by pretreatment with ouabain administration. Chronic administration of neither phentolamine (1 mg/rat, intraperitoneally, once daily for consecutive 15 days) nor propranolol (3 mg/Kg body weight, subcutaneously, once daily for consecutive 15 days) caused significant changes in adrenal corticosterone secretion in response to ACTH as well as to epinephrine. Chronic administration of ouabain in rats causes not only elevated secretion of ACTH from anterior pituitary but also functional change in adrenals leading to suppression of corticosterone secretion in response to ACTH or epinephrine administration.     

Suppressive effect of pentagastrin on pituitary-adrenocortical secretion.

The effect of pentagastrin on the pituitary-adrenocortical secretion was examined in male rats. In the morning the intraperitoneal injection of this peptide produced a slight, but significant, decrease in the plasma corticosterone levels, but it had no effect on the evening rise due to circadian periodicity and the stress-induced elevation of the plasma corticosterone level. Following intracerebroventricular administration of pentagastrin, plasma corticosterone tended to decrease and in in vitro incubation of rat pituitary tissue the addition of pentagastrin elicited a suppressive effect on the ACTH release from the tissue into the medium. It was suggested that gastrin-like peptide might control the secretion of ACTH.     

Vasopressin, corticosterone levels, and gastric ulcers during food-restriction stress.

Corticosterone levels and ulcers were compared in vasopressin-containing (LE) and vasopressin-deficient (DI) rats under ad lib and food-restricted conditions. In the ad lib situation, DI and LE rats had similar corticosterone levels and no ulcers. After 1 day of food restriction, the corticosterone levels were elevated in DI and LE rats, with a significantly higher level in LE rats. No ulcers were present in either strain. After 2 days of food restriction, the corticosterone levels were similar in DI and LE rats. The level in DI rats was comparable to that of the preceding day, but the level in LE animals dropped significantly from the previous day. Significant ulceration was evident in DI rats, but absent in LE rats. Following 3 days of food restriction, the corticosterone level in LE rats had returned to the ad lib level, whereas, for DI rats, an elevated level was maintained. There were no ulcers in LE rats, but they were present in DI rats. Thus LE and DI rats responded differently to the stress of food restriction. The mechanism underlying the response is most likely related to changes in the hypothalamic-pituitary-adrenocortical axis and its reaction to stress.     

Rat Strain Differences in Responses of Plasma Prolactin and PRL mRNA Expression After Acute Amphetamine Treatment or Restraint Stress

1. The aim of this study was to compare the effects of acute amphetamine (AMPH) treatment and restraint stress on plasma level of prolactin (PRL) and PRL mRNA expression in the adenohypophysis in Sprague–Dawley and Lewis male rats, the latter known to have a deficient hypothalamo–pituitary-adrenal (HPA) axis.2. Both restraint stress and AMPH treatment (i.p. in a dose of 8 mg/kg of b.w.) were applied 15 or 30 min before termination of the experiment. Plasma PRL and corticosterone (CORT) were determined by radioimmunoassay. PRL mRNA expression was estimated by a dot-blot hybridization.3. Restraint stress and AMPH treatment induced a significant increase in theCORT plasma level, as an indicator of stress response. Compared to Sprague–Dawley rats, the magnitude of CORT increase after both stimuli was significantly lower in Lewis rats.4. Although restraint stress significantly increased the PRL plasma levels in both rat strains, AMPH treatment reduced the PRL levels in both rat strains. However, the changes of PRL plasma levels had another pattern in Lewis rats than in Sprague–Dawley rats. Control plasma PRL levels were significantly higher in Lewis rats, and in this rat strain AMPH treatment for 30 min increased the PRL levels as compared to the values obtained after AMPH treatment for 15 min.5. Expression of PRL mRNA in adenohypophysis by restraint stress and AMPH treatment had a similar pattern. After a 15-min lasting restraint stress, the expression of PRL mRNA was decreased insignificantly in both rat strains. AMPH treatment induced in Sprague–Dawley rats a significant decrease of PRL mRNA after a 15-min interval while after 30 min there was a significant increase. However, in Lewis rats AMPH failed to significantly change PRL mRNA.6. The results from the present study indicate that the mechanisms mediatingthe effects of acute restraint stress and acute AMPH treatment differ in PRL response in Sprague–Dawley and Lewis male rat strains. Differences in the observed responses in Lewis rats could be related to the deficient activity of HPA axis in this rat strain.     

Role of hypothalamic inputs in maintaining pituitary-adrenal responsiveness in repeated restraint

The role of hypothalamic structures in the regulation of chronic stress responses was studied by lesioning the mediobasal hypothalamus or the paraventricular nucleus of hypothalamus (PVH). Rats were acutely (60 min) and/or repeatedly (for 7 days) restrained. In controls, a single restraint elevated the plasma adrenocorticotropin (ACTH), corticosterone, and prolactin levels. Repeated restraint produced all signs of chronic stress, including decreased body and thymus weights, increased adrenal weight, basal corticosterone levels, and proopiomelanocortin (POMC) mRNA expression in the anterior pituitary. Some adaptation to repeated restraint of the ACTH response, but not of other hormonal responses, was seen. Lesioning of the mediobasal hypothalamus abolished the hormonal response and POMC mRNA activation to acute and/or repeated restraint, suggesting that the hypothalamo-pituitary-adrenal axis activation during repeated restraint is centrally driven. PVH lesion inhibited the ACTH and corticosterone rise to the first restraint by approximately 50%. In repeatedly restrained rats with PVH lesion, the ACTH response to the last restraint was reduced almost to basal control levels, and the elevation of POMC mRNA level was prevented. PVH seems to be important for the repeated restraint-induced ACTH and POMC mRNA stimulation, but it appears to partially mediate other restraint-induced hormonal changes.