María Angélica Rivarola

Stress and the Reproductive Axis

There exists a reciprocal relationship between the hypothalamic‐pituitary‐adrenal (HPA) and the hypothalamic‐pituitary‐gonadal (HPG) axes, wherein the activation of one affects the function of the other and vice versa. For example, both testosterone and oestrogen modulate the response of the HPA axis, whereas activation of the stress axis, especially activation that is repeating or chronic, has an inhibitory effect upon oestrogen and testosterone secretion. Alterations in maternal care can produce significant effects on both HPG and HPA physiology, as well as behaviour in the offspring at adulthood. For example, changes in reproductive behaviour induced by altered maternal care may alter the expression of sex hormone receptors such as oestrogen receptor (ER)α that govern sexual behaviour, and may be particularly important in determining the sexual strategies utilised by females. Stress in adulthood continues to mediate HPG activity in females through activation of a sympathetic neural pathway originating in the hypothalamus and releasing norepinephrine into the ovary, which produces a noncyclic anovulatory ovary that develops cysts. In the opposite direction, sex differences and sex steroid hormones regulate the HPA axis. For example, although serotonin (5‐HT) has a stimulatory effect on the HPA axis in humans and rodents that is mediated by the 5‐HT1A receptor, only male rodents respond to 5‐HT1A antagonism to show increased corticosterone responses to stress. Furthermore, oestrogen appears to decrease 5‐HT1A receptor function at presynaptic sites, yet increases 5‐HT1A receptor expression at postsynaptic sites. These mechanisms could explain the heightened stress HPA axis responses in females compared to males. Studies on female rhesus macaques show that chronic stress in socially subordinate female monkeys produces a distinct behavioural phenotype that is largely unaffected by oestrogen, a hyporesponsive HPA axis that is hypersensitive to the modulating effects of oestrogen, and changes in 5‐HT1A receptor binding in the hippocampus and hypothalamus of social subordinate female monkeys that are restored or inverted by oestrogen replacement. This review summarises all of these studies, emphasising the profound effect that the interaction of the reproductive and stress axes may have on human reproductive health and emotional wellbeing.

Neurobiological effects of neonatal maternal separation and post-weaning environmental enrichment

Throughout the lifespan, the brain has a considerable degree of plasticity and can be strongly influenced by sensory input from the outside environment. Given the importance of the environment in the regulation of the brain structure, behavior and physiology, the aim of the present work was to analyze the effects of different environmental qualities during two critical ontogenic periods (early life and peripuberty) on behavior and hippocampal physiology. Male Wistar rats were separated from their mothers for 4.5h daily during the first 3 weeks of life. They were weaned on day 21 and housed under either standard or enriched conditions. At 60 d of age, all animals were then housed in same-treatment groups, two per cage, until testing began on day 74. Emotional and cognitive responses were tested using the open field, novel object recognition test and step-down inhibitory avoidance learning. In the dorsal hippocampus, glucocorticoid receptor expression and neuronal activity were examined by immunoreactivity. Grooming behavior in the open field was found to be significantly lower in maternally separated animals, but post-weaning environmental enrichment completely reversed this tendency. Inhibitory avoidance but not object recognition memory was impaired in maternally separated animals, suggesting that early maternal separation alters learning and memory in a task-specific manner. Again, environmental enrichment reversed the effects of maternal separation on the inhibitory avoidance task. Even though maternal separation did not significantly affect Fos and glucocorticoid receptor (GR) expression, environmental enrichment increased both Fos expression in the total hippocampal area and also the overall number of GR positive cells per hippocampal area, mainly due to the changes in CA1. These findings suggest that differential rearing is a useful procedure to study behavioral and physiological plasticity in response to early experience and that, although the effects of adverse experience early in life such as maternal separation can persist until adulthood, some of them can be compensated by early favorable environments, possibly through nervous system plasticity.

Sex differences in rats: Effects of chronic stress on sympathetic system and anxiety

In this study we tested whether periodic maternal deprivation (MD) (4.5 h daily during the first 3 weeks of life) caused chronic changes in anxiety and medullo-adrenal responses to chronic stress in either male or female adult (2.5 months of age) rats, or both. Repeated maternal deprivation had a sex-specific effect on epinephrine (E) and norepinephrine (NE) levels: an increase in both measures was observed only in females. Unpredictable stress did not produce changes on plasma catecholamine levels either in males or females. However, when the females were maternally deprived as well as stressed they showed an increase in plasma NE p < 0.05. On the other hand, non-maternally deprived (NMD), maternally-deprived and stressed males showed high levels of catecholamines compared to females p < 0.001. In the elevated plus maze test, MD-treated males displayed a slight increase in anxiety-related behavior compared with NMD rats. This was indicated by a reduction in the time spent on the open arms, whereas females showed less anxiety, indicated by an increase in the number of entries, and in the time spent on the open arms. After exposure to chronic stress only the females displayed decreased anxiety-related behavior. These results suggest that there are sex-induced effects in emotional reactivity, perception of the stressor and in the evaluation of novel situations. Thus, maternal deprivation and chronic variable stress caused both long-term alterations in sympathetic response and gender-dependent changes in the anxiety index of adult rats.

Early maternal separation: neurobehavioral consequences in mother rats.

Repeated separation of dams from their pups during the postpartum period may evoke emotional stress in the dam. In the present study we investigated whether prolonged maternal separation is stressful for rat dams by studying different behavioral and central responses known to be affected by stress. After delivery, female Wistar rats were subjected to either animal facility rearing (AFR) conditions or daily 4.5 h of mother-litter separation from postpartum day (PPD) 1-21. Maternal care (pup retrieval) was evaluated at PPD 3. After weaning on PPD 21, anxiety (elevated plus maze) and depression-like behaviors (forced swimming test) were assessed in the dams. Memory abilities (one-trial step down inhibitory avoidance) were tested either 1 h (short-term memory) or 24 h (long-term memory) after training session. Finally, c-Fos expression was examined in the central nucleus of the amygdala. The results revealed that pup retrieval efficiency at PPD 3 was significantly impaired by maternal separation. AFR dams retrieved their pups sooner and engaged in more pup-directed activities (nest building and carrying pups). Separation from pups increased the number of entries in open arms of the plus maze and decreased latency times in the inhibitory avoidance test for both short and long-term memory in the dams. There were no differences in depression-related behavior as assessed using the forced swimming test. Furthermore, maternal separation yielded high c-Fos expression in the central nucleus of the amygdala. Together, these data indicate that repeated maternal separation in the early postpartum period reduces maternal care and impairs the retention memory, providing further evidence for the detrimental neurobehavioral effects of maternal separation in dams.

Periodic maternal deprivation and lesion of anterodorsal thalami nuclei induce alteration on hypophyso adrenal system activity in adult rats.

The hypothalamic-pituitary-adrenal (HPA) axis is normally regulated by extrahypothalamic limbic structures, among these, the anterodorsal thalami nuclei (ADTN), which exert an inhibitory influence on HPA, in basal and acute stress conditions in rats. In the present work we have investigated whether neonatal maternal deprivation (MD) produces long-term changes in the ADTN regulation of HPA activity. Maternal deprivation, in female rats, for 4.5 hs daily, during the first 3 weeks of life, produced at 3 months old, a significant decrease in plasma ACTH concentration (p<0.001) and an increase in plasma corticosterone (C) (p<0.001), compared to control non-deprived rats (NMD). Also MD showed higher plasma epinephrine (E) and norepinephrine (NE) levels than NMD rats. The increase of NE (66.6% p<0.001) was higher than that observed in E (19%). After 30 days of ADTN lesion, plasma ACTH values were higher than in sham lesioned rats, in both NMD and MD animals. ACTH response was greater in MD rats. Plasma C, in NMD, was higher, whereas in MD lesioned animals, it was significantly lower than in sham lesioned. In MD rats, lesion produced a significant increase in plasma E and NE (p<0.001), and again, NE increase was higher than E increase. The more accentuated increase of NE than E, suggests sympathetic nervous system hyperactivity. In summary, neonatal maternal deprivation induces long-term alterations on HPA axis sensitivity and medullo adrenal secretion; enhanced sympathetic nervous system activity and, therefore affected the ADTN inhibitory influence on ACTH and adrenal glands secretion.

Gender-Dependent Effects of Early Maternal Separation and Variable Chronic Stress on Vasopressinergic Activity and Glucocorticoid Receptor Expression in Adult Rats

The aim of the present study was to investigate the influence of early maternal separation on Fos, arginine vasopressin (AVP) and glucocorticoid receptor (GR) expression in the medial parvocellular portion of the paraventricular hypothalamic nucleus (PaMP), and GR expression in the hippocampus of adult male and female rats subjected to variable chronic stress (VCS). Male and female Wistar rats were isolated 4.5 h daily, during the first 3 weeks of life. At 48 days of age, the rats were exposed to VCS. Nonmaternally separated (NMS) females had a higher number of activated AVP neurons than NMS male rats. Maternally separated (MS) females subjected to VCS also showed a higher number of Fos/AVP double-labeled neurons than males with the same treatment. Males and females subjected to early maternal separation and VCS, compared with the MS animals, showed a decrease in the expression of GR in the PaMP. As regards GR expression in the hippocampus, MS animals subjected to VCS as adults, both males and females, showed an increase in GR expression in the subfields CA1, CA2 and CA3. The increase in AVP-immunoreactive neurons coexpressing Fos in response to stress in females exposed to early maternal separation suggests that perhaps early life stress results in a more reactive neuroendocrine stress response in females. Furthermore, our results demonstrate that the different anatomical levels of the hypothalamic-pituitary-adrenal axis have different roles related to its stress response and support the evidence of regional specificity in GR regulation.

Early maternal separation and chronic variable stress in adulthood changes the neural activity and the expression of glucocorticoid receptor in limbic structures.

There is increasing evidence that early adverse experience contributes to the development of stress susceptibility, and increases the onset of stress‐related psychiatric disorders in stressful environments in adulthood. This study addressed whether or not prolonged maternal separation, a well‐established model of early stress, affects adult limbic areas related to the regulation of the hypothalamic‐pituitary‐adrenal axis in exposure to chronic variable stress in adulthood. Rats were subjected to daily maternal separation for 4.5 h during postnatal days 1–21. As adults, the animals were exposed to a variable chronic stress paradigm of 24 days. Persistent changes were assessed in glucocorticoid receptor density and Fos activity in the anterodorsal thalamic nuclei, mammillary nuclei and retrosplenial cortex. Immunohistochemical analysis revealed that adult maternally separated animals had increased levels of c‐Fos immunoreactivity in the anterodorsal thalamic nuclei as well as in the mammillary nuclei compared to normal non‐maternally separated animals. Chronic variable stress in maternally separated and non‐maternally separated animals diminished glucocorticoid receptor density in the anterodorsal thalamic nuclei but not in the rest of the nuclei analyzed. These results indicate that c‐Fos immunoreactivity as well as glucocorticoid receptor expression in the anterodorsal thalamic nuclei and mammillary nuclei exhibit long‐term alterations in adult rats following repeated maternal separation and subsequent stress exposure. Recognition of these adaptations helps to define the brain regions and neural circuitry associated with persistent alterations induced by early life environment and the development of stress‐associated disorders.

The Role of the Anterodorsal Thalami Nuclei in the Regulation of Adrenal Medullary Function, Beta-adrenergic Cardiac Receptors and Anxiety Responses in Maternally Deprived Rats under Stressful Conditions

Maternal separation can interfere with growth and development of the brain and represents a significant risk factor for adult psychopathology. In rodents, prolonged separation from the mother affects the behavioral and endocrine responses to stress for the lifetime of the animal. Limbic structures such as the anterodorsal thalamic nuclei (ADTN) play an important role in the control of neuroendocrine and sympathetic-adrenal function. In view of these findings we hypothesized that the function of the ADTN may be affected in an animal model of maternal deprivation. To test this hypothesis female rats were isolated 4.5 h daily, during the first 3 weeks of life and tested as adults. We evaluated plasma epinephrine (E) and norepinephrine (NE), cardiac adrenoreceptors and anxiety responses after maternal deprivation and variable chronic stress (VCS) in ADTN-lesioned rats. Thirty days after ADTN lesion, in non-maternally deprived rats basal plasma NE concentration was greater and cardiac β-adrenoreceptor density was lower than that in the sham-lesioned group. Maternal deprivation induced a significant increase in basal plasma NE concentration, which was greater in lesioned rats, and cardiac β-adrenoreceptor density was decreased in lesioned rats. After VCS plasma catecholamine concentration was much greater in non-maternally deprived rats than in maternally-deprived rats; cardiac β-adrenoreceptor density was decreased by VCS in both maternally-deprived and non-deprived rats, but more so in non-deprived rats, and further decreased by the ADTN lesion. In the plus maze test, the number of open arm entries was greater in the maternally deprived and in the stressed rats. Thus, sympathetic-adrenal medullary activation produced by VCS was much greater in non-deprived rats, and was linked to a down regulation of myocardial β-adrenoceptors. The ADTN are not responsible for the reduced catecholamine responses to stress in maternally-deprived rats. Maternal deprivation or chronic stress also induced a long term anxiolytic effect, which was also not affected by ADTN lesion.

Effects of maternal deprivation on adrenal and behavioural responses in rats with anterodorsal thalami nuclei lesions

There is evidence that repeated maternal isolation of neonatal rats may influence both emotional behavior and Hypothalamic-Pituitary Adrenal (HPA) activity. On the other hand the Anterodorsal Thalami Nuclei (ADTN) exerts an inhibitory influence on the hypophyso-adrenal system under basal and stressful conditions. In the present work we investigated whether neonatal maternal deprivation produces long term effects on the ADTN regulation of behavioral patterns (open field test) and on HPA axis activity. Specifically, we sought to determine whether adult female rats with ADTN lesions, previously isolated for 4.5 hours daily during the first 3 weeks of life, react in endocrinologically and behaviourally distinct manner as compared to controls. The examined groups were: non maternally deprived (NMD)/sham lesioned, NMD/lesioned, maternally deprived (MD)/sham lesioned, MD/lesioned with and without the open field test. At 3 months MD/sham lesioned animals showed a marked decrease in ambulation (P < 0.01), and with ADTN lesion, the rearing values were lower (P < 0.01) and grooming higher (P < 0.05) than NMD. This last data would indicate a high emotional index. Regarding the activity of the HPA axis, maternal deprivation induced a significant decrease in plasma ACTH concentration both in sham and lesioned animals (P < 0.001), and plasma Corticosterone (C) increased in sham animals (P < 0.001). This data would indicate a higher sensitivity of the adrenal glands. After the open field test ACTH and C were different between deprived and non-deprived animals depending on the ADTN lesion. Taking into consideration the increase of ACTH levels in sham lesioned MD animals exposed to the test, we could conclude that this new situation was a stressful situation. Finally in the present work, it was very difficult to relate the behavioral parameters with the endocrine data. It is known that depending on the context, corticosteroids may produce opposite effects on emotional behavior via different receptors in the brain.In summary, neonatal maternal deprivation induced alterations of behavioral patterns and affected the ADTN inhibitory influence on ACTH and C secretion.

Maternal Separation and Lesion of Adtn Alters Anxiety And Adrenal Activity in Male Rats

The present study investigated the effect of early maternal separation on anxiety and hypophyso-adrenal system activity to anterodorsal thalamic nuclei (ADTN) lesion in male rats as adults in order to compare this with previous results with female rats. During the first 3 weeks of life, male rats were isolated 4.5 hr daily and tested as adults. Thirty days after ADTN lesion we found that adrenocorticotropic hormone (ACTH) plasma levels were affected neither by maternal separation nor by ADTN lesion. Plasma corticosterone (CORT) concentration was increased with lesion of the ADTN in maternally separated rats. A significant increase in plasma catecholamine concentration was induced by early maternal separation. In ADTN-lesioned rats, plasma norepinephrine (NE) concentration was significantly lower than in the respective sham-lesioned groups. In terms of anxiety, there were no significant effects of early experience. However, the ADTN lesion tended to decrease anxiety-related behavior.