Octavi Martí

Effects of chronic stress on food intake in rats : influence of stressor intensity and duration of daily exposure

The effect of exposure to chronic intermittent stressors of differing intensities [handling, restraint, and immobilization (IMO)] and daily duration (15, 60, and 240 min of IMO) on changes in food intake was studied in adult male rats. Whereas handling did not caused anorexia, restraint slightly reduced food intake and IMO drastically reduced it. The effects were very similar after the 7th and 27th day of exposure to the stressors. Fourteen days of chronic IMO (2 h daily) resulted in decreased food intake as measured on days 1, 10, and 14 of treatment, the inhibition being slightly greater after the first stress session. The circadian rhythm of food intake, expressed as a percent of the total food eaten in a 24 h period, was found to be almost unaffected by chronic IMO, although IMO rats appeared to satiate sooner than control rats. Exposure of rats to chronic IMO (7 days) for 15, 60, and 240 min daily reduced food intake to the same extent in all IMO groups. Taken together, these results suggest that: a) the magnitude of the changes in food intake after chronic exposure to stressors is closely related to their intensity, and b) a severe stressor such as IMO reduces food intake to a certain level that was independent on its daily duration. After repeated exposure to the same stressor, only a slight tendency to recover normal food intake was observed.

Recovery of the Hypothalamic-Pituitary-Adrenal Response to Stress

Pathological consequences of stress-induced activation of the hypothalamic-pituitary-adrenal (HPA) axis may be related to the duration rather than to the intensity of HPA axis activation after exposure to the stressor. Consequently a fine analysis of post-stress events is of importance. The present experiments were designed to study the importance of three key factors in HPA recovery: intensity of the stressor (experiment 1), duration of exposure to the stressor (experiment 2) and previous experience of the animals with the situation (experiments 3 and 4). In experiment 1, analysis of both the response to the stressor and the poststress period showed that the stronger the stressor, the greater the area under the curve of HPA activation. In experiment 2, different groups of rats were exposed to different periods of immobilization (IMO) (20 min, 1 h and 2 h) and sampled before, during and after exposure to IMO. The speed of recovery of plasma corticotropin (ACTH) levels was not related to the duration of exposure to the stressor. In experiments 3 and 4, the influence of previous experience with the stressor was studied in rats daily exposed to 20 min IMO or daily injected with hypertonic saline (HS) for 8 days and sampled on days 1, 2, 5 and 8. Whereas a significant decline in plasma ACTH levels was not observed immediately after IMO until day 8, a single previous exposure to IMO was enough to enhance recovery 90 min after the end of exposure to IMO. Corticosterone levels were related to the number of previous experiences with the stressor only in the post-IMO period. In response to a novel stressor (forced swimming), chronic IMO rats showed a slightly impaired recovery as compared to stress-naive rats, suggesting that enhanced recovery of the HPA axis was specific for the homotypic stressor. After daily HS injections, a pattern similar to that after IMO was observed, the delayed, but not the early response of the HPA axis being reduced as a function of the number of previous experiences with the situation. Taken together, the present results suggest that the speed of recovery of the HPA axis after its activation by stressors is sensitive to the intensity of the stressors but not to their duration, and that adaptation to a repeated stressor is more apparent during the delayed HPA response.

ANTERIOR PITUITARY RESPONSE TO STRESS : TIME-RELATED CHANGES AND ADAPTATION

A wide array of physical and psychological stressors alter the secretion of anterior pituitary hormones. However, both the qualitative and the quantitative features of the stressors as well as its duration markedly influence the final endocrine response. In addition, among all anterior pituitary hormones, only ACTH and prolactin levels appear to reflect the intensity of the stress experienced by the animals. Although physical stressors show a somewhat specific neuroendocrine profile, the response of the pituitary‐adrenal (PA) and sympathomedulloadrenal axes are common to almost all stressors. After an initial stimulatory effect of stress, an inhibition of all anterior pituitary hormones, except ACTH, can be found provided the stressor is intense enough. The mechanisms responsible for this biphasic response to stress are likely to be located at sites above the pituitary. When the animals are repeatedly exposed to the same stressor, some behavioural and physiological consequences of stress exposure are reduced, suggesting that the animals become adapted to the stimulus. This process has been also termed habituation. Among all the pituitary hormones, only ACTH and prolactin levels are reduced as a consequence of repeated exposure to the same (homotypic) stressor, although some negative results have been reported. However, it has been recently reported that subtle changes in the characteristics of the stressors or in their regularity can greatly influence adaptation, and these factors might explain failure to find adaptation of ACTH and prolactin in some works. Habituation of ACTH and prolactin, when observed, appears to be specific for the chronically applied stressor so that the potentiality of the PA axis and prolactin to respond to a novel (heterotypic) stressor can be preserved. In the case of the PA axis, an intact or potentiated response to a novel stressor is observed in spite of presumably negative feedback exerted by daily stress‐induced glucocorticoid release and the high resting levels of glucocorticoids. This phenomenon has been termed as facilitation and can be unmasked alternating stress. Although with the exception of the PA axis, developmental aspects of anterior pituitary response to stress have been poorly studied, available data suggest that dramatic changes occur in some hormones during weaning, with some, but less profound, changes thereafter. Responsiveness to stressors appears to mature with age, but developmental patterns differ among the various anterior pituitary hormones.

Forced swimming test in rats: effect of desipramine administration and the period of exposure to the test on struggling behavior, swimming, immobility and defecation rate

The effect of desipramine administration and the duration of the daily exposure to forced swimming on some variables has been studied in adult male rats. Desipramine administration (15 mg/kg) significantly increased struggling behavior in the first and second 5-min periods of a single exposure to forced swimming. Swimming was reduced in the first 5 min and remained unchanged thereafter. Immobility was decreased in the second and the third 5-min periods. After a pre-exposure to forced swimming for 15 min the day before, the drug was effective in increasing struggling behavior and reducing immobility during a subsequent 5-min test. Swimming was not modified. Daily exposure to forced swimming for 3 days caused a decline in struggling behavior and swimming, while increasing immobility and the defecation rate. The duration of daily exposure to forced swimming did not alter the changes in the variables measured. The present results indicate that a one-day test can be used to discriminate between saline- and desipramine-treated rats, and that struggling behavior could be a reliable measure of the positive action of antidepressants. The finding that behavioral changes over the 3 days were independent of the duration of exposure to swimming argues against the interpretation of the results which suggest that the responses are caused by the appearance of a behavioral despair state, and suggests that these behaviors might be trait-markers in the rat. In addition, the changes in struggling behavior and immobility over the 3 days cannot be attributed to a behavioral adaptation to the test because the defecation rate increased rather than decreased during successive forced swimming tests.

Direct evidence for chronic stress-induced facilitation of the adrenocorticotropin response to a novel acute stressor.

The ACTH response to CRF and the role of glucocorticoids on the pituitary-adrenal responsiveness to acute stressors after a period of chronic stress were assessed in Sprague-Dawley rats. After chronic immobilization (IMO) an enhanced ACTH response to CRF administration was observed. In another experiment, control and chronic IMO rats were adrenalectomized (ADX) or sham-adrenalectomized (SHAM) and 2 days later killed in resting conditions or after having been subjected to acute IMO or tail-shock for 30 min. Chronic IMO increased basal corticosterone but not adrenocorticotropin (ACTH) levels in SHAM rats. As expected, ADX increased ACTH levels in all conditions. In response to the novel acute stressor (shock), ACTH levels were drastically dependent on the presence of corticosterone: thus, whereas in SHAM rats chronic IMO reduced the ACTH response to shock, in ADX rats a clear enhancement of the ACTH response to shock was observed in chronic IMO rats. In order to demonstrate that, in our experimental conditions, chronic stress also induces down-regulation of glucocorticoid receptors in the hippocampus, an additional experiment was done: rats subjected chronically to IMO or administered 5 mg corticosterone (B) were adrenalectomized and killed 20 h later under basal conditions. Both chronic IMO and chronic B administration decreased glucocorticoid type II binding in the hippocampus. From these results, it is concluded that chronic IMO induces facilitation of the ACTH response to novel acute stressors which is uncovered after corticosterone removal.

Evidence that a single exposure to aversive stimuli triggers long-lasting effects in the hypothalamus-pituitary-adrenal axis that consolidate with time.

Because of its use as a negative reinforcer in animal studies and its potential pathological impact (e.g. post‐traumatic stress disorder and depression), exposure to aversive stimuli is a relevant model for studying CNS plasticity. We present evidence that a single exposure to two predominantly emotional stressors [restraint in tubes and immobilization on wooden boards (IMO)] can modify the response of the hypothalamo–pituitary–adrenal (HPA) axis to a subsequent exposure to the same stressor days later in that a more rapid return to the baseline was observed in the poststress period. In addition, the effect was greater with IMO, the more severe stressor. Using IMO, we have further demonstrated that the effect of a previous single exposure to the stressor (i) increased with days elapsed between the two exposures; (ii) was specific for the previously experienced stressor; and (iii) was mediated via central‐mediated effects [corticotropin‐releasing factor (CRF) mRNA in the paraventricular nucleus of the hypothalamus]. These data suggest that animals retain memory about a single experience with stressors, resulting in an acceleration of the poststress recovery of the HPA axis that enhances progressively over a period of weeks. The extent to which the present data are relevant regarding post‐traumatic stress disorders is unclear, but the study of the HPA response to severe stressors may be suitable for the study of the neurobiological basis of the progressive consolidation of learning over a long period of time (days to weeks).

Effect of regularity of exposure to chronic immobilization stress on the circadian pattern of pituitary adrenal hormones, growth hormone, and thyroid stimulating hormone in the adult male rat

Circadian variation of serum levels of adrenocorticotropin hormone (ACTH), corticosterone, growth hormone (GH), and thyroid-stimulating hormone (TSH) were studied in three groups of adult male rats exposed to chronic intermittent immobilization stress (IMO) for 2 hr daily under different schedules. IMO resulted in reduced food intake, body weight loss, and increased adrenal weight. ACTH levels were not affected but corticosterone levels were increased in all IMO rats as compared to control ones during the diurnal phase of the circadian cycle. IMO decreased serum GH and TSH levels but the circadian pattern of secretion was influenced in a complex way depending on the specific pattern of daily exposure to IMO. Differences observed between the IMO groups were not caused by differences in food intake because its circadian rhythm was very similar in all IMO groups. These results suggest that regularity of exposure to immobilization alters in a complex fashion circadian GH and TSH rhythms.

Acute stress markers in humans: Response of plasma glucose, cortisol and prolactin to two examinations differing in the anxiety they provoke

Although the endocrine response to psychological stressors has been extensively studied both in animals and humans, the sensitivity of these variables to the intensity of stress experienced by exposure to psychologically stressful situations has not been studied in humans. In the present work this was assessed by measuring plasma levels of glucose, cortisol and prolactin in female medical students just before taking two examinations clearly differing in the anxiety they provoke. It was found that both examinations increased anxiety just before taking them, but the physiology examination (EPh) caused higher anxiety than the psychology (EPs). Prolactin increased in response to both examinations as compared to the non-stress condition, but its levels were greater in the EPh than in the EPs. Cortisol followed the same pattern as prolactin, but increased only marginally in the EPs. Finally, glycemia rose to the same extent in response to both examinations. A significant positive correlation was found between anxiety and glucose, and between cortisol and prolactin when data from all situations were included. On the basis of these results, it appears that the three variables might be useful as putative markers of stress in humans, although glucose might reflect different underlying psychological processes than cortisol and prolactin. In addition, it was found for the first time that prolactin is able to discriminate between stressful situations of different intensity. The response of these physiological variables to other stressful situations differing both in quantitative and in qualitative terms merits to be studied in further work.

Influence of single or repeated experience of rats with forced swimming on behavioural and physiological responses to the stressor

In the present work behavioural (struggling and immobility), physiological (hypothermia, glycaemia) and endocrine (hypothalamo-pituitary-adrenal (HPA) hormones) response to repeated forced swimming (FS) for 15 days was studied in adult male rats and compared with the response of rats having only one single experience with FS either 1 or 14 days before the last exposure to the stressor. Repeated experiences with FS reduced struggling and increased immobility as compared with stress-naïve rats, whereas a single previous exposure to FS, regardless of the time elapsed, had the same, but less marked, effect. Hypothermia followed the same trend. FS-induced hyperglycaemia was not sensitive to a previous single experience, but rather it was totally abolished in chronically stressed rats. Neither a single nor chronic exposure to FS modified the secretion of ACTH in response to the last FS session. However, repeated FS enhanced the speed of recovery of plasma corticosterone as compared to control rats, suggesting a dissociation between the two hormones. The present results revealed great differences in the sensitivity of various behavioural and physiological responses to repeated FS stress and suggest that reduced response to repeated FS, when found, is not a consequence of the time elapsed between exposures but to the repetition of the stressful situation.

Stress-induced activation of the immediate early gene Arc (activity-regulated cytoskeleton-associated protein) is restricted to telencephalic areas in the rat brain: relationship to c-fos mRNA

Arc is an effector immediate early gene whose expression is induced in situations of increased neuronal activity. However, there is no report on the influence of stress on Arc expression. Here, we compared the induction of both c‐fos and Arc mRNAs in the brain of rats exposed to one of three different stressful situations: novel environment, forced swimming and immobilization. An absent or weak c‐fos mRNA signal was observed in control rats, whereas those exposed to one of three stressors showed enhanced c‐fos expression in a wide range of brain areas. Constitutive Arc expression was observed in some areas such as cortex, striatum, hippocampus, reticular thalamic nucleus and cerebellar cortex. In response to stressors, a strong induction of Arc was observed, but the pattern was different from that of c‐fos. For instance, activation of Arc but not c‐fos was observed in the nucleus accumbens after immobilization and in the hippocampus after novel environment. No Arc induction was observed in diencephalic and brainstem areas. The present data show that Arc has a neuroanatomically restricted pattern of induction in the brain after emotional stress. Telencephalic activation suggests that a more intense induction of synaptic plasticity is occurring in this area after exposure to emotional stressors.