Effects Of Repeated Stress Research Articles

Swim stress alters the behavioural response of mice to GABA-related and some GABA-unrelated convulsants

To elucidate the relationship between stress and seizures, the effect of a single swim stress on the convulsive signs and death produced by several GABA-related and GABA-unrelated convulsants, and the effect of repeated swim stress on picrotoxin-induced convulsions was studied. Mice were subjected to swim stress (10 min swimming at 18–19°C), and the i.v. infusion of convulsants started 15 min thereafter. The latency to the onset of several convulsant signs and death was measured, and the doses of convulsants producing convulsions and death were calculated. Additional experiments included mice swimming at room temperature, and those which were stressed repeatedly (twice a day for four consecutive days, plus one stressful procedure on the fifth day). Swim stress increased the dose needed to produce convulsant signs and death after bicuculline, picrotoxin, pentylenetetrazole, strychnine and 4-aminopyridine, while kainic acid-induced convulsions were not affected. Using picrotoxin infusion, the effect of swimming in room temperature water was less than the effect of swimming in 18–19°C water. In addition, the effect of repeated stress was less than the effect of acute stress on picrotoxin-induced convulsions. The results demonstrate that acute swim stress lowers the convulsive potency of GABA-related and some GABA-unrelated convulsants. Repeatedly stressed animals develop tolerance to anticonvulsive effect of swim stress.

Effect of repeated stress on body weight and body composition of rats fed low- and high-fat diets.

Exposure to the moderate stressor of 3-h restraint for 3 consecutive days causes a temporary drop in food intake but a permanent reduction in body weight in adult rats. Young rats did not show the same response. Food intake of adult rats exposed to repeated restraint was significantly lower than that of controls for 4 days after the end of stress, and there was no rebound hyperphagia. Body weight remained significantly lower for at least 40 days after stress. When the rats were fed a high-fat diet of 80% chow and 20% vegetable shortening (48% kcal fat, 16% protein), lean body mass accounted for all of the weight loss in stressed rats. When the experiment was repeated with a purified high-fat diet containing corn oil and coconut oil as the source of fat (41% kcal fat, 16% protein), weight loss consisted of both lean and fat tissue. There were no sustained changes in single time point measures of corticosterone, insulin, or leptin that could account for the reduced body weight in these rats.

Stress-induced morphohistochemical and functional changes in rat adrenal cortex, testis and major salivary glands.

The effect of repeated stress (1 h of daily immobilization for seven consecutive days) on the adrenal cortex of young adult male albino rats was evaluated by morphohistochemical methods and plasma assays; at the same time, testes and major salivary glands, as steroid-producing and -depending organs, respectively, were examined. Morphological and histochemical changes were found in the adrenal cortex, testis and submaxillary gland, though varying in degree and extent depending on the gland examined. Corticosterone and progesterone plasma levels increased, in agreement with the lipid depletion observed in the zona fasciculata, while testosterone and androstenedione decreased, as confirmed by the less marked enzymatic activity in the Leydig cells. The study thus proves that repeated stress, even of temporary duration, is able to influence directly or indirectly the morphofunctional state of the three examined glands, suggesting a functional linkage.

Regulation of Gene Expression of Catecholamine Biosynthetic Enzymes by Stress

Effects of repeated stress are associated with alterations in gene expression. The effect of immobilization (IMMO) stress on the expression of the genes encoding some catecholamine biosynthetic enzymes, as well as GTPcyclohydrolase I (GTPCH), the rate-limiting enzyme in the tetrahydrobiopterin biosynthetic pathway, has been recognized. This has been studied in the adrenal medulla (AM), a major source of plasma epinephrine; the sympathetic ganglia (SG), a major source of plasma norepinephrine; and the locus ceruleus (LC), the major noradrenergic nucleus in the central nervous system that modulates the response of the hypothalamus and other brain areas to stress. All of these genes have been found to be activated by IMMO. However, there are important differences in the minimal time of IMMO required for the induction of these genes and in the factors mediating the stress response. For example, 5 min of single exposure to stress is found sufficient to elicit a maximal increase in adrenal phenylethanolamine-N-methytransferase (PNMT) mRNA levels. However, to achieve maximal effect on the other genes of interest, longer times were required. The results of these studies indicate that either c-fos is not required for the induction tyrosine hydroxylase (TH) gene expression in AM or that alternative factors are involved. Studies on TH gene expression in cultured cells have identified two major promoter elements, an AP1 site and a cyclic AMP/calcium regulatory element (CRE/CaRE), each of which can independently activate TH transcription.

Regulation of adrenal steroidogenesis during chronic stress.

The mechanism of the altered adrenal responsiveness during chronic stress was studied by analysis of ACTH and Ang II responses and the expression and activity of steroidogenic enzymes in the adrenal cortex of rats subjected to repeated immobilization (2 hr/day for 14 days), or repeated i.p. injection of 1.5 M NaCl. Concomitant with increased pregnenolone production and reduced aldosterone secretion by isolated adrenal glomerulosa cells of chronically stressed rats, P-450scc mRNA were increased and P-450aldo mRNA levels were decreased in adrenal zona glomerulosa. Consistent with elevated plasma corticosterone levels, isolated adrenal fasciculata cells from stressed rats showed higher cAMP, pregnenolone and corticosterone responses to ACTH. Adrenal fasciculata area and levels of P-450scc, but not those of P-450(11s) hydroxylase were significantly increased. The effects of repeated stress on adrenal steroidogenesis were mimicked by repeated ACTH injections. The half life of corticosterone in plasma measured with [3H]corticosterone was increased in stressed rats but not in ACTH injected rats. This study shows that chronic stress leads to a) inhibition of mineralocorticoid secretion due to inhibition of the late biosynthetic pathway, and b) increased circulating glucocorticoids due to increased ACTH receptor activity, expression and activity of the early pathway, and decreased glucocorticoid clearance. Altered adrenal glomerulosa and fasciculata function, but not changes in glucocorticoid clearance, are probably mediated by increased ACTH secretion during chronic stress.

Effect of repeated stress on a number of plasma amino acids and related compounds in young and old rats

A number of amino compounds were studied in the plasma of young and old rats exposed to an immobilization stress on 3 different days. Baseline values were similar for all compounds between age groups except for slightly higher phenylalanine levels in old animals. Most differences were seen in the stress responses. Stress response intensities for alanine, aspartic acid, methionine, phenylalanine, and tyrosine were generally higher in old rats and recovery times from stress were usually longer. Although the stress responses decreased in young rats for some compounds over the three exposures, old rats showed no or less adaptation during this time. All the other amino acids showed no marked age effects. This study shows that certain plasma amino acids can serve as selective indicators of biological age and that their stress responses might be better markers of the aging process than their resting levels.

Effect of development of habituation to restraint stress on hypothalamic noradrenaline release and adrenocorticotropin secretion.

The effect of repeated stress has been studied on noradrenaline release in the hypothalamic paraventricular nucleus and on adrenocorticotropin levels. Rats were stressed by 20-min immobilization once a day for 5 days. On day 6 they were exposed to the same stress or to a different one (ether vapors for 2 min). Immobilization and ether stress increased noradrenaline release in naive rats (271 +/- 43 and 197 +/- 9%, respectively) and raised adrenocorticotropin levels, showing activation of the hypothalamus-pituitary axis. Repeated daily restraint did not modify basal noradrenaline or adrenocorticotropin levels. The further immobilization session on day 6 did not change noradrenaline levels at any observation time (20-120 min). The adrenocorticotropin response was still present, although significantly reduced. In repeatedly restrained rats, exposure to ether vapors induced a maximal increase in noradrenaline level similar to that observed in naive rats, although prolonged. In these rats the adrenocorticotropin response did not differ from that in acutely stressed rats. These results suggest that habituation may develop to a stressful stimulus leading to suppression of the hypothalamic noradrenergic response and that this phenomenon is stress specific. Moreover, modifications of noradrenaline release in the paraventricular nucleus are not solely responsible for the adrenocorticotropin response during stress, suggesting that other pathways and/or neurotransmitters are involved too.

Effect of acute and chronic epinephrine administration on clearance and metabolism of [3H]-epinephrine in trout (Oncorhynchus mykiss)

Several studies have measured the rate of catecholamine clearance, metabolism, and tissue accumulation in fish. However, no information is available on the effect of repeated stress or high circulating catecholamine levels on catecholamine clearance and metabolism. We measured the clearance and metabolism of [3H]-epinephrine (approximately 0.1 μg·kg-1) in SW-acclimated rainbow trout subjected to acute (five injections in 1 day) and chronic (4 days; five injections per day) administration of 4.0 μg·kg-1 epinephrine or saline. In addition, a saturation experiment, where 4.0 μg·kg-1 of unlabelled epinephrine was injected concurrently with [3H]-epinephrine, investigated whether catecholamine clearance and metabolism are affected by high circulatin levels. Neither the rate constants for catecholamine clearance, nor the post-injection proportions of unmetabolised [3H]-epinephrine, deaminated [3H]-epinephrine and O-methylated [3H]-epinephrine were affected by the acute or chronic injection protocols. The concurrent injection of [3H]-epinephrine and 4.0 μg·kg-1 of unlabelled epinephrine resulted in an elevated postinjection 3H:14C ratio, but increased proportions of O-methylated [3H]-epinephrine and reduced proportions of unmetabolised [3H]-epinephrine. We conclude that in fish (1) catecholamine clearance and metabolism are unlikely to be compromised by repeated exposure to acute stressors; (2) catecholamine extraction and/or metabolism is enhanced when circulating levels are high; and (3) there is a marked capacity to rapidly (minutes) clear and inactivate catecholamines that are released in response to stressful stimuli.

Effects of repeated stress on plasma arginine levels in young and old rats

The effect of age and recurring stress responses on endogenous plasma arginine (ARG) concentrations were determined. Blood from catheterized young (3 months) and old (24 months) were used to ascertain the plasma concentrations of ARG under baseline conditions, during a 30-min immobilization experience and during a 3.5 h poststress period. Rats were again immobilized and blood drawn following a 1 day rest period (day 3) and then after an additional 3 day rest period (day 7). For young animals, ARG levels decrease significantly after 30 min of stress and remain suppressed during the entire poststress period (3.5 h). After a 1 day rest period, these young rats may have exhibited adaptation to stress on day 3. Old animals showed little or no reduction in plasma ARG concentrations during and after immobilization stress exposure on all three occasions. Baseline levels of old animals were lower in comparison to young animals and their stress responses were smaller. Thus, it is possible that significant changes in plasma ARG levels (as the nitric oxide precursor) during or after stress could serve as an indicator of impending pathology such as cardiovascular disease or immunocompetency.

Effect of repeated stress on plasma catecholamines and taurine in young and old rats

The effect of age and multiple stress responses on plasma norepinephrine (NE), epinephrine (EPI), and taurine (TAU) levels were determined in F344 rats. Blood samples obtained from catheterized young (3 months) and old (24 months) animals were used to examine plasma levels of NE, EPI, and TAU under baseline conditions and in the same animals after a 30-min immobilization stress. Rats were again immobilized and blood drawn (Day 3) following a 1-day rest period and, after an additional 4-day rest period (Day 7). Age differences seen between young and old rats were not the same for the three neurochemicals measured but were relatively unique for the specific biochemicals. In old animals baseline values of NE but not EPI and TAU were higher then young animals and all three values did not change for the baseline during the two additional stress exposures. Initial stress responses were similar for all three biochemicals in both age groups. Although no signs of adaptation were evident in the old animals, adaptation to immobilization was seen for EPI and TAU but not NE on the third occasion in the young animals. Correlations seen between NE and EPI in young and old rats on the first day disappeared during the second stress period but were again seen during the third stress exposure suggesting subtle indicators of repeated stress subject to adaptation. TAU values for young but not old rats correlated positively with EPI concentrations during the first stress exposure and negatively after the immobilization was terminated indicating a regulatory interaction between EPI and TAU present in young but lost in old animals. Thus, changes in levels and interrelationships of specific biochemicals during repeated stress experiences in rats may provide a good model of the aging process.

A chronic stress state in rats: Effects of repeated stress on basal corticosterone and behavior

The chronic stress state has previously been defined as persistent visceral arousal coupled with behavioral abnormalities. To determine the number of stressor exposures necessary to induce a chronic stress state, male rats were given 2 hours of inescapable shock on 10, 7, 4, or 3 consecutive days. The 3-day stress group had the most pervasive changes in the variables measured: persistently elevated basal plasma corticosterone (CORT), continued weight loss in the post-stressor period, and abnormal behavior. More exposures to the stress regimen did not produce higher CORT levels or greater behavioral changes. Acutely stressed rats, exposed to 1 day of inescapable shock, had persistent CORT elevations without the other changes seen in the 3-day stress group. The data suggest that 3 days of our stress regimen are sufficient to produce a state of chronic stress and that some signs of this state begin to appear as early as the first exposure to our inescapable stress regimen.

Repeated stress enhances vasopressin synthesis in corticotropin releasing factor neurons in the paraventricular nucleus

The effect of repeated stress on corticotropin-releasing factor (CRF) and vasopressin (AVP) synthesis in parvocellular neurons of the paraventricular nucleus (PVN) was studied by means of double label immunocytochemistry. Once daily immobilization of male rats for 16 days leads to a 75% increase in the number of CRF immunoreactive neurons, but a 5-fold elevation in the number of AVP containing CRF cell bodies. These results demonstrate that repeated stress activates AVP synthesis in CRF neurons.

Effects of repeated stress on turgor pressure and cell elasticity changes in black spruce seedlings

One-year-old black spruce (Piceamariana (Mill.) B.S.P.) seedlings were preconditioned by exposing them to either one or two dehydration–rehydration cycles by using the osmoticum polyethylene glycol 3350. Preconditioned and unconditioned seedlings were then subjected to a more severe osmotic (water) stress by exposing them to a higher concentration of polyethylene glycol. Effects of repeated dehydration–rehydration cycles on cell-water relations were studied after 3, 7, and 13 days of stress relief using pressure–volume curve analysis. Repeated dehydration–rehydration cycles caused a cumulative increase in turgor potentials at full saturation. In these preconditioned plants there was also a progressive lowering of osmotic potentials and relative water contents at zero turgor, which increased over time with stress relief. The decline in osmotic potentials at zero turgor in osmotically stressed black spruce was associated with increased cell wall relaxation, followed by increased turgor potentials, in preconditioned but not in unconditioned seedlings. Saturated osmotic potentials were not altered by repeated, short-term conditioning stresses, suggesting that tissue elasticity was more important for turgor regulation than osmotic adjustment.

Effects of repeated stress on T cell numbers and function in rats

Although stress has been reported to affect various functions of the immune system, the mechanism that mediate these effects remain unclear. Thus we examined the effects that 1, 7, and 14 days of stress could have on various aspects of immune and endocrine function in rats. Rats subjected to repeated stress (7 and 14 days) showed significant decreases in the total number of mononuclear cells, particularly suppressor/cytotoxic (CD8) T cells, in the spleen and blood. The mitogenic responses of T cells to phytohemagglutinin (PHA) and concanavalin-A (Con A) were also significantly diminished at these times, as well as after acute (1 day) stress in the case of PHA stimulation. The mechanisms of this impaired T cell mitogenesis were explored by assessing the effects of stress on T cell interleukin 2 (IL-2) production and T cell responsiveness to IL-2. T cells from repeatedly stressed rat showed a decreased production of IL-2 in response to PHA, although their proliferative response to exogenous IL-2 was normal. Repeated stress also decreased body weight and spleen weight, increased adrenal weight, and decreased plasma levels of triiodothyronine and testosterone. These results suggest that lower levels of IL-2 production during stress could be one reason for the decreased mitogen responsiveness of T cells, often seen with stress. This is important because defective IL-2 production could also lead to significant impairment of immunoregulatory T cell generation and thus a predisposition to malignancy or autoimmune disease that some have associated with stress.

The anterior midline cortex and adaptation to stress ulcers in rats

Rats with bilateral radio-frequency lesions of the anterior midline cortex were subjected to either single or repeated immobilization stress treatments. As previously demonstrated, the lesions greatly reduced gastric pathology in the acute stress treatment. Conversely, the lesions were found to aggravate gastric pathology in animals stressed repeatedly, thus eliminating the adaptive (i.e., pathology-reducing) effects of repeated stress seen in controls. The role of the anterior midline cortex in stress and stress adaptation is discussed with reference to noradrenergic mechanisms. Anterior midline cortex Rat Immobilization stress Stress adaptation Gastric pathology

Repeated membrane plasma separation with on-line sorbent treatment of plasma in the conscious rat.

Repeated membrane plasma separation with on-line sorbent treatment of plasma was performed in unrestrained and conscious rats, thus avoiding the possible effects of repeated stress and exposure to anesthetic agents. The procedure was well tolerated, even after four consecutive perfusions. Blood flow was 0.7 ml/min, with a transmembrane pressure of 18.6 mm Hg and plasma filtration rate of 0.21 ml/min. This allowed 12 ml of plasma (greater than 1 plasma volume) to be treated within 1 h. Levels of albumin, immunoglobulin G, and C3 remained constant during each perfusion and were not significantly different from those of animals subjected to control procedures. Results obtained from filtrate were comparable at 15 and 60 min with values obtained from whole blood, with sieving coefficients of approximately 1. Leukocyte, erythrocyte, and thrombocyte counts remained unchanged during each plasma perfusion. A progressive rise in leukocyte counts occurred following successive perfusions, but this was true also of animals subjected to control procedures.

Effects of repeated stress on pituitary cyclic AMP, and plasma prolactin, corticosterone and growth hormone in male rats

The effects of five putative stressors (saline injection, cold exposure, forced running, immobilization, and footshock) on levels of pituitary cyclic AMP, plasma prolactin, corticosterone and growth hormone were examined. In naive rats exposed to 15 min of these stressors for the first time, running, immobilization and footshock increased levels of pituitary cyclic AMP, plasma corticosterone and prolactin and decreased growth hormone, typical of stress response in the rat. Cold exposure only increased corticosterone and saline injection did not affect any measured parameter. In rats chronically exposed to the same stressor (once a day for 15 min) for 10 days immediately prior to the experiment, an attenuated pituitary cyclic AMP and plasma prolactin response was seen upon application of 15 min of that stressor on the day of the experiment, compared to the responses observed in the naive rats.