Ljubica Gavrilović

Immobilization and cold stress affect sympatho–adrenomedullary system and pituitary–adrenocortical axis of rats exposed to long-term isolation and crowding

Changes in plasma levels of noradrenaline (NA), adrenaline (A), adrenocorticotropic hormone (ACTH) and corticosterone (CORT), as well as in cytosol glucocorticoid receptor (GR) and heat shock protein 70 (Hsp 70) in hippocampus of adult rat males exposed to two long-term types of psychosocial stress, both under basal conditions and in response to immobilization and cold as heterotypic additional stressor were studied. Long-term isolation produced a significant elevation of basal plasma ACTH and CORT levels, but did not affect that of NA and A, while long-term crowding conditions did not elevate the basal plasma levels of these hormones. Long-term isolation of rats exposed to 2 h of immobilization or cold led to a significant elevation of plasma NA, A and CORT in comparison with the controls. Long-term crowding conditions and exposure of animals to immobilization or cold also resulted in an increased plasma NA, A and CORT levels, but to a lesser extent in comparison with the long-term isolation. At the same time, plasma ACTH was significantly more elevated in long-term crowded than in long-term isolated rats. Both kinds of long-term psychosocial stresses (isolation and crowding) had similar but less pronounced effects on cytosol GR and Hsp 70 concentrations in hippocampus comparing to acute immobilization and cold stress. It seems that long-term psychosocial stresses attenuate the effects of an additional stress on hippocampal GR and Hsp 70 concentrations. These data suggest that individual housing of rats appear to act as a stronger stressor than crowding conditions. When the animals suffering a long-term isolation were exposed to either acute immobilization or cold, a stronger activation of the sympatho-adrenomedullary system (SAS) was recorded in comparison with that found in the long-term crowded group subjected to short-term immobilization or cold. No significant differences in the activity of hypotalamo-pituitary-adrenal (HPA) axis were observed between long-term isolated and long-term crowded rats.

Brain glucocorticoid receptor and heat shock protein 70 levels in rats exposed to acute, chronic or combined stress

The pattern and intensity of glucocorticoid receptor (GR) and heat shock 70 protein (Hsp 70) changes in the hippocampus and brain cortex of adult Wistar rat males exposed to acute (immobilization, cold) and chronic (social isolation, crowding, daily swimming) stress or their combinations were followed by Western immunoblotting. Plasma ACTH and CORT were measured by chemiluminescent method and RIA. A significant decrease in cytosol GR and Hsp 70 was observed after acute stress, while chronic stresses led to negligible changes in both these proteins and caused a reduced responsiveness to a novel acute stress. This was valid irrespective of the type of chronic or acute stress combinations for both hippocampal and cortical GR and Hsp 70. The results support the hypothesis that chronic stress-induced deregulation of the LHPA axis may be caused, at least in part, by partial disruption of intracelullar negative feedback control in the higher centers of the brain.

Chronic Individual Housing-Induced Stress Decreased Expression of Catecholamine Biosynthetic Enzyme Genes and Proteins in Spleen of Adult Rats

Objective: Social isolation is regarded as one of the most relevant causes of diseases in mammalian species. The activation of the sympathoneural system represents one of the key components of the stress response. The sympathetic nervous system is one of the major pathways involved in immune-neuroendocrine interactions. The aim of the present study was to determine plasma epinephrine and norepinephrine in individually housed rats, as well as to find out whether splenic gene expression of catecholamine synthesizing enzymes and their protein levels are affected by chronic psychosocial stress. Methods: Tyrosine hydroxylase (TH), dopamine-β-hydroxylase (DBH) and phenylethanolamine N-methyltransferase (PNMT) mRNA levels were quantified by quantitative real-time RT-PCR. The TH, DBH and PNMT immunoproteins were assayed by Western blot. Results: Chronic social isolation of adult male rats produced a significant increase in plasma catecholamine levels and a decrease in splenic TH mRNA, DBH mRNA and PNMT mRNA. Protein levels of TH, DBH and PNMT were also reduced. Conclusion: These results suggest that increased plasma catecholamines and decreased gene expression and protein levels of catecholamine biosynthetic enzymes in the spleen of chronically individually housed animals might reduce catecholamine synthesis, thus leaving the immunocompetent tissues depleted of catecholamines and consequently leading to an impairment of immune response.

The Effect of Repeated Physical Exercise on Hippocampus and Brain Cortex in Stressed Rats

Abstract:  Sensitivity of target cells to glucocorticoids is regulated by the expression of intracellular glucocorticoid receptor (GR), which mediates the effects of glucocorticoids. The level of GR and of its nuclear transporter protein 70 (Hsp70) were followed in hippocampus and brain cortex of adult Wistar rat males exposed to acute (immobilization, cold) and chronic (social isolation, isolation, and 15 min daily swimming) stress or their combinations. Changes in plasma levels of adenocorticotropic hormone and corticosterone were also studied. A significant decrease in cytosol GR and Hsp70 was observed after acute stress. Opposite to that, chronic stress led to negligible changes in both cytosol GR and Hsp70 levels. Isolation, as chronic psychosocial stressor, caused reduced responsiveness to novel acute stressors, judged by the cytosol GR and Hsp70 levels. This was not observed if chronic isolation was combined with 15 min daily swimming prior to acute exposure to immobilization. The data suggest that repeated physical exercise may, at least in some cases, diminish detrimental effects of chronic social isolation on limbic‐hypothalamic‐pituitary‐adrenocortical axis, as judged by the levels of GR and Hsp70 in the Wistar rat brain.

Effects of repeated maprotiline and fluoxetine treatment on gene expression of catecholamine synthesizing enzymes in adrenal medulla of unstressed and stressed rats.

1 Repeated maprotiline (a noradrenaline reuptake inhibitor) and fluoxetine (a serotonin reuptake inhibitor) treatment on gene expression of catecholamine biosynthetic enzymes were examined in adrenal medulla of unstressed control and chronic unpredictable mild stressed rats. 2 Maprotiline did not change gene expression of catecholamine biosynthetic enzymes in control and stressed rats. 3 Fluoxetine increased gene expression of tyrosine hydroxylase (TH) and dopamine-β-hydroxylase (DBH), but did not phenylethanolamine N-methyltransferase in both unstressed and chronic unpredictable mild stressed animals. 4 In conclusion, we have demonstrated that repeated administration of fluoxetine enhanced gene transcription of TH and DBH and subsequently stimulates noradrenaline synthesis in adrenal medulla of control and stressed rats.

Psychosocial stress-related changes in gene expression of norepinephrine biosynthetic enzymes in stellate ganglia of adult rats

In this study we investigated the changes in norepinephrine biosynthetic enzymes tyrosine hydroxylase (TH), dopamine beta-hydroxylase (DBH) and phenylethanolamine N-methyltransferase (PNMT) gene expression in the stellate ganglia of naive controls and long-term socially isolated (12 weeks) adult rats and the response of these animals to additional immobilization stress. Psychosocial stress produced a significant increase of both TH mRNA and DBH mRNA levels in stellate ganglia. Additional immobilization of long-term psychosocially stressed rats expressed no effect on gene expression of these enzymes. The results presented here suggest that psychosocial stress-induced increase in gene expression of norepinephrine biosynthetic enzymes in stellate ganglia may be connected to the increased risk of cardiovascular disease.

Novel stressors affected catecholamine stores in socially isolated normotensive and spontaneously hypertensive rats

Catecholamines in some central (hypothalamus and hippocampus) and peripheral tissues (adrenal glands and heart auricles) of long-term socially isolated normotensive and spontaneously hypertensive rats exposed to novel immobilization stress were determined by a simultaneous single isotope radioenzymatic assay. Long-term isolation (21 days) produced depletion of hypothalamic norepinephrine (NE) stores and hippocampal dopamine (DA) stores in both normotensive and spontaneously hypertensive rats. Acute immobilization stress (2 h) significantly decreased NE and DA stores in hypothalamus and hippocampus of naive normotensive and spontaneously hypertensive rats controls. However, novel immobilization stress applied to normotensive rats previously subjected to long-term isolation produced no changes in catecholamine levels in hypothalamus, while resulting in somewhat higher depletion of NE stores in hypothalamus of spontaneously hypertensive rats treated in the same way. Novel immobilization stress decreased NE and DA stores in hippocampus of normotensive but was without effect on NE and DA stores of spontaneously hypertensive rats. Social isolation did not affect catecholamine stores in peripheral tissues but novel immobilization stress produced a significant decrease in catecholamine content. The results suggest that some central and peripherals tissues of spontaneously hypertensive rats and normotensive rats differ with regard to catecholamine content and that there are certain differences in their responsiveness to stress.

Treadmill exercise does not change gene expression of adrenal catecholamine biosynthetic enzymes in chronically stressed rats

Chronic isolation of adult animals represents a form of psychological stress that produces sympatho-adrenomedullar activation. Exercise training acts as an important modulator of sympatho-adrenomedullary system. This study aimed to investigate physical exercise-related changes in gene expression of catecholamine biosynthetic enzymes (tyrosine hydroxylase, dopamine-ß-hydroxylase and phenylethanolamine N-methyltransferase) and cyclic adenosine monophosphate response element-binding (CREB) in the adrenal medulla, concentrations of catecholamines and corticosterone (CORT) in the plasma and the weight of adrenal glands of chronically psychosocially stressed adult rats exposed daily to 20 min treadmill running for 12 weeks. Also, we examined how additional acute immobilization stress changes the mentioned parameters. Treadmill running did not result in modulation of gene expression of catecholamine synthesizing enzymes and it decreased the level of CREB mRNA in the adrenal medulla of chronically psychosocially stressed adult rats. The potentially negative physiological adaptations after treadmill running were recorded as increased concentrations of catecholamines and decreased morning CORT concentration in the plasma, as well as the adrenal gland hypertrophy of chronically psychosocially stressed rats. The additional acute immobilization stress increases gene expression of catecholamine biosynthetic enzymes in the adrenal medulla, as well as catecholamines and CORT levels in the plasma. Treadmill exercise does not change the activity of sympatho-adrenomedullary system of chronically psychosocially stressed rats.

Behavioral and endocrine responses of socially isolated rats to long-term diazepam treatment

The effects of diazepam (0.2 mg/kg/ during 21 days, i.p.) on behavior, pituitary-adrenocortical and sympatho-adrenomedullary system of socially isolated and group-housed adult male rats additionally exposed to immobilization, were studied. Social isolation led to a shorter duration of grooming and longer latency to start grooming. Diazepam in social isolated rats reduced incorrect transitions percentage, but the number of grooming bouts, duration and latency to start grooming remained unchanged. Long-term isolation significantly elevated plasma ACTH and corticosterone, while not affecting noradrenaline and adrenaline. Diazepam decreased only plasma ACTH. Social isolation and immobilization significantly elevated all examined hormones. Immobilization of diazepam-treated isolated rats enhanced plasma ACTH , the increase being significantly lower, comparing to isolated vehicle-treated rats. Immobilization significantly increased plasma adrenaline, noradrenaline and corticosterone of diazepam- or vehicle-treated socially isolated rats. No differences in adrenaline, noradrenaline and corticosterone level between these two groups were observed. This indicates that chronic diazepam treatment of socially isolated rats changes some grooming behavior parameters, but insignificantly affects stress-related adrenomedullary and adrenocortical alterations.

Antioxidant enzymes in women with endometrial polyps: relation with sex hormones

OBJECTIVE To investigate whether antioxidant enzyme activities (superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase) and lipid hydroperoxide levels in patients with endometrial polyps are influenced by the changes in sex hormones (estradiol, progesterone, FSH, and LH) during the menstrual cycle and in postmenopause. STUDY DESIGN The material consisted of blood and endometrial tissue specimens from women diagnosed with endometrial polyps. Patients were divided into groups depending on the phase of the menstrual cycle--follicular or luteal--and the postmenopause. The activities of antioxidant enzymes and the lipid hydroperoxide levels were compared among the phases and a linear regression model was used to evaluate the associations between hormones and antioxidant/oxidant variables. RESULTS In the blood of examined women, a significant difference in superoxide dismutase activity and lipid hydroperoxide levels was recorded among the phases. There was also a positive correlation between the estradiol concentration and superoxide dismutase. In polyp tissue, we recorded a phase-related difference in superoxide dismutase and glutathione peroxidase activities as well as in the lipid hydroperoxide levels. A negative correlation was observed between FSH/LH and glutathione peroxidase, and between LH and superoxide dismutase. CONCLUSION Antioxidant enzymes and lipid hydroperoxide levels in patients with endometrial polyps are influenced by the changes in sex hormones during the menstrual cycle and after the menopause, pointing to a role of the observed relationship in polyp etiology.