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.     

Dose dependent effects of oxytocin on cognitive defects and anxiety disorders in adult rats following acute infantile maternal deprivation stress

ABSTRACT

Maternal deprivation at an early age is a powerful stressor that causes permanent alterations in cognitive and behavioral functions during the later stages of life. We investigated the effects of oxytocin on cognitive defects and anxiety disorders caused by acute infantile maternal deprivation in adult rats. We used 18-day-old Wistar albino rats of both sexes. The experimental groups included control (C), maternally deprived (MD), maternally deprived and treated with 0.02 μg/kg oxytocin (MD-0.02 µg/kg oxy), maternally deprived and treated with 2 μg/kg oxytocin (MD-2 µg/kg oxy). When the rats were 60 days old, the open field (OF) and elevated plus maze (EPM) behavioral tests, and the Morris water maze (MWM) test for spatial learning and memory were performed. In addition, the number of neurons in the hippocampus, prefrontal cortex (PFC) and amygdala were determined using quantitative histology. We also measured vascular endothelial growth factor (VEGF) and brain-derived neurotrophic factor (BDNF) levels in the PFC. In both sexes, the MD group failed the learning test and the MD-2 μg/kg oxy group failed in the memory test. The MD-0.02 μg/kg oxy group spent more time in the open arm of the EPM device and their locomotor activities were greater in the OF test. The VEGF and BDNF levels in the PFC were higher in the MD-0.02 μg/kg oxy groups than the other maternally deprived groups (oxytocin ±). The number of PFC neurons was low in all male maternally deprived (oxytocin ±) groups, while the number of amygdala neurons was low in both female and male maternally deprived (oxytocin ±) groups. Male rats were more affected by maternal deprivation; administration of oxytocin had dose-dependent biphasic effects on learning, memory and anxiety.     

Medullo adrenal response to lesion of anterodorsal thalamic nuclei in rats

Anterodorsal thalamic nuclei (ADTN) exert an inhibitory influence on hypophyso-adrenal system (HAS) in rats. With the purpose of evaluating if ADTN are also involved in the control of medullo adrenal activity, experiments were conducted on female rats with bilateral lesion of these nuclei. Thirty days after lesion, plasma epinephrine (E) concentration in lesioned rats was higher than that in sham-lesioned control group (P < 0.02). Meanwhile, adrenal E content was significantly lower in lesioned animals than that found in the control group (P < 0.005). Plasma norepinephrine (NE) values in lesioned rats were not significantly different from those in the control ones, however, there was a significant decrease in adrenal NE when compared to the control one (P < 0.02). Basal values of plasma ACTH and plasma and adrenal corticosterone (C) were signicantly higher than those in sham lesioned rats (P < 0.05; P < 0. 001; P < 0.001 respectively). These findings demonstrate that the ADTN in rats are involved in the regulation of both cortico and medullo adrenal activity.     

Deprivation of maternal care has long-lasting consequences for the hypothalamic-pituitary-adrenal axis of zebra finches

Early-life stress caused by the deprivation of maternal care has been shown to have long-lasting effects on the hypothalamic-pituitary-adrenal (HPA) axis in offspring of uniparental mammalian species. We asked if deprivation of maternal care in biparental species alters stress responsiveness of offspring, using a biparental avian species--the zebra finch, Taeniopygia guttata. In our experiment, one group of birds was raised by both male and female parents (control), and another was raised by males alone (maternally deprived). During adulthood, offspring of both groups were subjected to two stressors (restraint and isolation), and corticosterone concentrations were measured. Additionally, we measured baseline levels of the two corticosteroid receptors--glucocorticoid receptor (GR) and mineralocorticoid receptor (MR)--in the hippocampus, hypothalamus and cerebellum. Our results suggest that maternally deprived offspring are hyper-responsive to isolation in comparison with controls. Furthermore, mRNA levels of both GR and MR receptors are altered in maternally deprived offspring in comparison with controls. Thus, absence of maternal care has lasting consequences for HPA function in a biparental species where paternal care is available.     

Maternal Deprivation Induces Depressive-like Behaviour and Alters Neurotrophin Levels in the Rat Brain

The present study was aimed to evaluate the behavioral and molecular effects of maternal deprivation in adult rats. To this aim, male rats deprived and non-deprived were assessed in the forced swimming and open-field tests in adult phase. In addition adrenocorticotrophin hormone (ACTH) levels was assessed in serum and brain-derived-neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and nerve growth factor (NGF) protein levels were assessed in prefrontal cortex, hippocampus and amygdala. We observed that maternal deprivation increased immobility time, and decreased climbing time, without affecting locomotor activity. ACTH circulating levels were increased in maternal deprived rats. Additionally, BDNF protein levels were reduced in the amygdala and NT-3 and NGF were reduced in both hippocampus and amygdala in maternal deprived rats, compared to control group. In conclusion, our results support the idea that behavioral, ACTH circulating levels and neurotrophins levels altered in maternal deprivation model could contribute to stress-related diseases, such as depression.     

Social support attenuates the adverse consequences of social deprivation stress in domestic piglets

Social deprivation is a severe stressor affecting a number of behavioral and physiological functions of gregarious species. It is assumed that, dependent upon the level of familiarity, social support given by a conspecific may attenuate the adverse consequences of stress. We investigated the effects of a 4 h maternal and littermate deprivation on behavioral reactions, stress hormone responses and brain corticosteroid receptor expression in 7-, 21- and 35-day-old domestic piglets (Sus scrofa) that were left alone or in the presence either of a familiar or unfamiliar age-matched piglet. Compared to control animals, all of the socially deprived piglets showed significant stress responses, such as impaired habituation in repeated open-field/novel-object tests, enhanced ACTH and cortisol release, and altered corticosteroid receptor expression in the hypothalamus. In addition, our results demonstrated that younger piglets had more difficulty coping with stress. The presence of an age-matched conspecific had a direct calming effect on the deprived piglet during the deprivation procedure, which was revealed by diminished stress-induced HPA activity and altered reactions in the behavioral test situations (e.g., activity, escape, and vocalization). Furthermore, because the presence of a familiar piglet causes a more pronounced buffering effect, we have shown for the first time that the degree of familiarity between the piglets may influence the effectiveness of social support. Our study emphasizes the benefits of social partners on positive welfare and the ability for pigs to cope with stress; therefore, our results should be taken into account during handling practices such as weaning and mixing.     

Delayed maturation and altered proliferation within the rat rostral migratory stream following maternal deprivation

The objective of this study was to investigate whether stressful experience during early postnatal period may influence morphological characteristics of the rat neurogenic pathway--the rostral migratory stream (RMS) and proliferation of neuronal precursors in three successive areas of the RMS: in the vertical arm, the elbow and the horizontal arm. To induce stress, the pups were subjected to repeated maternal deprivation during the first postnatal week after birth. Brains were analyzed at the seventh postnatal day. The controls matched the age of maternally deprived animals. Observation of hematoxylin-eosin stained sections showed that maternal deprivation did not affect the general morphological appearance of the RMS. The shape of the RMS of maternally deprived rats resembles the RMS of control animals. Maternal deprivation caused slight, not significant increase in the RMS thickness in comparison with control rats. Significant difference between the control and maternally deprived rats concerns the olfactory ventricle. While in seven days old control rats the olfactory ventricle is completely closed, in maternally deprived rats of the same age the olfactory ventricle was regularly visible as a narrow lumen at the axis of the RMS horizontal arm. This finding indicates delayed maturation of the migratory pathway as a consequence of stress. Proliferation activity has been assessed by immunoreactivity of the endogenous cell cycle protein Ki-67. The results of Ki-67 immunohistochemistry showed that seven days' maternal separation for 3 h daily induces significant quantitative changes in the number of proliferating cells within the RMS. The response of Ki-67-positive cells to stress differed in individual part of the RMS, with a marked decrease in the vertical arm and a significant increase in the elbow, suggesting heterogeneity of neural stem cells along the RMS; while in the RMS vertical arm the number of dividing cells significantly decreased, there was a marked increase of Ki-67-positive cells in the RMS elbow. This suggests heterogeneity of neural stem cells along the RMS.     

Early stress affects neurogenesis in the rat rostral migratory stream

Stressful experience during the early postnatal period may influence processes associated with neurogenesis (i.e. proliferation, cell death, appearance of astrocytes or cell differentiation) in the neonatal rat rostral migratory stream (RMS). To induce stress, pups were subjected to maternal deprivation daily for three hours, starting from the first postnatal day till the seventh postnatal day. Immunohistochemical methods were used to visualize proliferating cells and astrocytes; dying cells and nitrergic cells were visualized using histochemical staining. Quantitative analysis showed that maternal deprivation decreased the number of proliferating cells and significantly increased the number of dying cells in the RMS. Maternal deprivation did not influence the appearance of astrocytes in the RMS, but caused premature differentiation of nitrergic cells. In control rats, nitrergic cells can be observed in the RMS as early as the tenth postnatal day. In maternally deprived pups, these cells were detected as early as the seventh postnatal day. The observed earlier appearance of nitrergic cells in the RMS was associated with altered proliferation and increased cell dying and this observation supports the hypothesis that nitric oxide has an anti-proliferative role in the RMS. Our study demonstrates that maternal deprivation represents a stressful condition with a profound impact on early postnatal neurogenesis.     

Effect of prenatal stress on the hypothalamo-hypophyseal-adrenal system activity and fat metabolism in the male water vole Arvicola terrestris

The HPA axis function and the fat metabolism were studied in adult male water voles born to intact mothers and mothers deprived of food on the 15th and 18th days of their pregnancy. The HPA response to emotional stress was relatively low in experimental males in winter. The hormonal response to a 24-hrs food deprivation diminished in experimental group only in February. The blood level of free fatty acids was increased in experimental group as compared to the control one. The short maternal food deprivation modified adaptive abilities of their male progeny.     

Long Term Hippocampal and Cortical Changes Induced by Maternal Deprivation and Neonatal Leptin Treatment in Male and Female Rats

Maternal deprivation (MD) during neonatal life has diverse long-term behavioral effects and alters the development of the hippocampus and frontal cortex, with several of these effects being sexually dimorphic. MD animals show a marked reduction in their circulating leptin levels, not only during the MD period, but also several days later (PND 13). A neonatal leptin surge occurs in rodents (beginning around PND 5 and peaking between PND 9 and 10) that has an important neurotrophic role. We hypothesized that the deficient neonatal leptin signaling of MD rats could be involved in the altered development of their hippocampus and frontal cortex. Accordingly, a neonatal leptin treatment in MD rats would at least in part counteract their neurobehavioural alterations. MD was carried out in Wistar rats for 24 h on PND 9. Male and female MD and control rats were treated from PND 9 to 13 with rat leptin (3 mg/kg/day sc) or vehicle. In adulthood, the animals were submitted to the open field, novel object memory test and the elevated plus maze test of anxiety. Neuronal and glial population markers, components of the glutamatergic and cannabinoid systems and diverse synaptic plasticity markers were evaluated by PCR and/or western blotting. Main results include: 1) In some of the parameters analyzed, neonatal leptin treatment reversed the effects of MD (eg., mRNA expression of hippocampal IGF1 and protein expression of GFAP and vimentin) partially confirming our hypothesis; 2) The neonatal leptin treatment, per se, exerted a number of behavioral (increased anxiety) and neural effects (eg., expression of the following proteins: NG2, NeuN, PSD95, NCAM, synaptophysin). Most of these effects were sex dependent. An adequate neonatal leptin level (avoiding excess and deficiency) appears to be necessary for its correct neuro-programing effect.     

Lesions of the anteroventral third ventricle region exaggerate neuroendocrine and thermogenic but not behavioral responses to a novel environment

Mild psychological stressors provoke an acute rise in core temperature (T(C)), stimulate the hypothalamo-pituitary-adrenocortical (HPA) axis, and induce various stress-related behaviors. In the present study, we examined the effect of ablation of the anteroventral third ventricle region (AV3V) on both physiological and behavioral responses to a novel environment. T(C) was monitored in male Sprague-Dawley rats, with either sham or AV3V lesions, during a 5-h exposure to a novel environment. Trunk blood was collected, in a second group of rats, for the assessment of plasma levels of ACTH and corticosterone. Novelty-induced grooming and rearing behaviors were assessed in a third group of animals. T(C) was elevated in all animals after 30 min in the novel environment, but the rise was exaggerated in rats with AV3V lesions ( approximately 0.5 degrees C). AV3V-lesion rats maintained a higher core temperature for 2 h before it returned to the same level as the control group. Plasma levels of ACTH and corticosterone were also exaggerated in the AV3V lesion group after 30 min in a novel environment. In contrast to the physiological responses, the behavioral measures of grooming and rearing revealed no differences between the groups. The results from the current study suggest that neurons within the AV3V region exert an inhibitory influence on the HPA axis and fever developed in response to stressful psychological stimuli. They also confirm that the physiological and hormonal components of the stress response are independent of certain behavioral measures of stress.     

Maternal Deprivation Exacerbates the Response to a High Fat Diet in a Sexually Dimorphic Manner

Maternal deprivation (MD) during neonatal life has diverse long-term effects, including affectation of metabolism. Indeed, MD for 24 hours during the neonatal period reduces body weight throughout life when the animals are maintained on a normal diet. However, little information is available regarding how this early stress affects the response to increased metabolic challenges during postnatal life. We hypothesized that MD modifies the response to a high fat diet (HFD) and that this response differs between males and females. To address this question, both male and female Wistar rats were maternally deprived for 24 hours starting on the morning of postnatal day (PND) 9. Upon weaning on PND22 half of each group received a control diet (CD) and the other half HFD. MD rats of both sexes had significantly reduced accumulated food intake and weight gain compared to controls when raised on the CD. In contrast, when maintained on a HFD energy intake and weight gain did not differ between control and MD rats of either sex. However, high fat intake induced hyperleptinemia in MD rats as early as PND35, but not until PND85 in control males and control females did not become hyperleptinemic on the HFD even at PND102. High fat intake stimulated hypothalamic inflammatory markers in both male and female rats that had been exposed to MD, but not in controls. Reduced insulin sensitivity was observed only in MD males on the HFD. These results indicate that MD modifies the metabolic response to HFD intake, with this response being different between males and females. Thus, the development of obesity and secondary complications in response to high fat intake depends on numerous factors.     

What can we know from pituitary-adrenal hormones about the nature and consequences of exposure to emotional stressors?

Exposure to stress induces profound physiological and behavioral changes in the organisms and some of these changes may be important regarding stress-induced pathologies and animal models of psychiatric diseases. Consequences of stress are dependent on the duration of exposure to stressors (acute, chronic), but also of certain characteristics such as intensity, controllability, and predictability. If some biological variables were able to reflect these characteristics, they could be used to predict negative consequences of stress. Among the myriad of physiological changes caused by stress, only a restricted number of variables appears to reflect the intensity of the situation, mainly plasma levels of ACTH and adrenaline. Peripheral hypothalamic-pituitary-adrenal (HPA) hormones (ACTH and corticosterone) are also able to reflect fear conditioning. In contrast, the activation of the HPA axis is not consistently related to anxiety as evaluated by classical tests such as the elevated plus-maze. Similarly, there is no consistent evidence about the sensitivity of the HPA axis to psychological variables such as controllability and predictability, despite the fact that: (a) lack of control over aversive stimuli can induce behavioral alterations not seen in animals which exert control, and (b) animals showed clear preference for predictable versus unpredictable stressful situations. New studies are needed to re-evaluate the relationship between the HPA axis and psychological stress characteristics using ACTH instead of corticosterone and taking advantages of our current knowledge about the regulation of this important stress system.     

Maternal glucocorticoid deficit affects hypothalamic-pituitary-adrenal function and behavior of rat offspring

Detrimental consequences of prenatal stress include increased hypothalamic-pituitary-adrenal (HPA) function, anxiety and depression-like behavior in adult offspring. To identify the role of maternal corticosterone milieu in the fetal programming of adult function, we measured these same behavioral and hormonal endpoints after maternal adrenalectomy (ADX) and replacement with normal or moderately high levels of corticosterone (CORT). Adult male and female offspring exhibited differing HPA responses to maternal ADX. In female offspring of ADX mothers, exaggerated plasma ACTH stress responses were reversed by the higher, but not the lower, dose of maternal CORT. In contrast, male offspring of both ADX and ADX dams with higher CORT replacement showed exaggerated ACTH stress responses. Hypothalamic glucocorticoid receptor (GR) expression was decreased in these latter groups, while hippocampal GR increased only in the ADX offspring. Activity of young offspring of ADX dams replaced with the higher dose of CORT decreased in the open field test of exploration/anxiety, while immobility behavior of adult offspring in the forced swim test of depression increased following maternal ADX or higher levels of CORT replacement. Interestingly, for some measures, none or moderately high CORT replacement resulted in similar deficits in this study. These findings are in accord with consequences of prenatal stress or prenatal dexamethasone exposure, suggesting that a common mechanism may underlie the effects of too low or too high maternal glucocorticoids on adult HPA function and behavior.     

Interaction of some limbic structures which exert inhibitory effect on corticosterone secretion.

The interaction between limbic structures which exert inhibitory influence on corticosterone secretion was investigated in the rat. The following experiments were performed: 1) electrical stimulation at mammillary medial nucleus (MMN) in rats with lesioned anterodrosal thalami nucleus (ADTN) or intermediate tegmental area; 2) electrical stimulation at ADTN in rats with lesioned retrosplenial cortex (RC). Bilateral stimulation at MMN in ADTN or RC-lesioned rats produces an increase in plasma corticosterone concentration. In animals with lesioned RC, values of plasma corticosterone after stimulation at ADTN were higher than before stimulation. Taking into consideration that electrical stimulation of MMN or ADTN in intact rats produces a decrease in plasma corticosterone concentration, these studies demonstrate that MMN and ADTN exert inhibitory influence on corticoadrenal activity only when their projection areas remain intact.     

A role for retinal brain-derived neurotrophic factor in ocular dominance plasticity

Visual deprivation is a classical tool to study the plasticity of visual cortical connections. After eyelid closure in young animals (monocular deprivation, MD), visual cortical neurons become dominated by the open eye, a phenomenon known as ocular dominance (OD) plasticity . It is commonly held that the molecular mediators of OD plasticity are cortically derived and that the retina is immune to the effects of MD . Recently, it has been reported that visual deprivation induces neurochemical, structural, and functional changes in the retina , but whether these retinal changes contribute to the effects of MD in the cortex is unknown. Here, we provide evidence that brain-derived neurotrophic factor (BDNF) produced in the retina influences OD plasticity. We found a reduction of BDNF expression in the deprived retina of young rats. We compensated this BDNF imbalance between the two eyes by either injecting exogenous BDNF in the deprived eye or reducing endogenous BDNF expression in the nondeprived eye. Both treatments were effective in counteracting the OD shift induced by MD. Retinal BDNF could also influence OD distribution in normal animals. These results show for the first time that OD plasticity is modulated by BDNF produced in the retina.     

Gender-dependent cellular and biochemical effects of maternal deprivation on the hippocampus of neonatal rats: a possible role for the endocannabinoid system

Adult animals submitted to a single prolonged episode of maternal deprivation (MD) [24 h, postnatal days (PND) 9-10] show behavioral alterations that resemble specific symptoms of schizophrenia. These behavioral impairments may be related to neuronal loss in the hippocampus triggered by elevated glucocorticoids. Furthermore, our previous data suggested functional relationships between MD stress and the endocannabinoid system. In this study, we addressed the effects of MD on hippocampal glial cells and the possible relationship with changes in plasma corticosterone (CORT) levels. In addition, we investigated the putative involvement of the endocannabinoid system by evaluating (a) the effects of MD on hippocampal levels of endocannabinoids (b) The modulation of MD effects by two inhibitors of endocannabinoids inactivation, the fatty acid amide hydrolase inhibitor N-arachidonoyl-serotonin (AA-5-HT), and the endocannabinoid reuptake inhibitor, OMDM-2. Drug treatments were administered once daily from PND 7 to PND 12 at a dose of 5 mg/kg, and the animals were sacrificed at PND 13. MD induced increased CORT levels in both genders. MD males also showed an increased number of astrocytes in CA1 and CA3 areas and a significant increase in hippocampal 2-arachidonoylglycerol. The cannabinoid compounds reversed the endocrine and cellular effects of maternal deprivation. We provide direct evidence for gender-dependent cellular and biochemical effects of MD on developmental hippocampus, including changes in the endocannabinoid system.     

Paternal deprivation induces dendritic and synaptic changes and hemispheric asymmetry of pyramidal neurons in the somatosensory cortex

Similar to maternal care, paternal care is a source of neonatal sensory stimulation, which in primates and rodents has been shown to be essential for developing structure and function of sensory cortices. The aim of our study in the biparental rodent Octodon degus was to assess the impact of paternal deprivation on dendritic and synaptic development in the somatosensory cortex. We (i) quantified the amount of paternal care in relation to total parental investment and (ii) compared dendritic and synaptic development of pyramidal neurons in the somatosensory cortex of animals raised by a single mother or by both parents. On the behavioral level we show that paternal care comprises 37% of total parent‐offspring interactions, and that the somatosensory stimulation provided by the fathers primarily consists of huddling, licking/grooming, and playing. On the morphological level we found that, compared with offspring raised by both parents (mother and father), the father‐deprived animals displayed significantly reduced spine numbers on the basal dendrites of pyramidal neurons. Furthermore, paternal deprivation induces hemispheric asymmetry of the dendritic morphology of somatosensory pyramidal neurons. Father‐deprived animals show shorter and less complex basal dendrites in the left somatosensory cortex compared with the right hemisphere. These findings indicate that paternal deprivation results in delayed or retarded dendritic and synaptic development of somatosensory circuits. © 2009 Wiley Periodicals, Inc. Develop Neurobiol, 2009     

Developmental plasticity of HPA and fear responses in rats: a critical review of the maternal mediation hypothesis

Developmental plasticity of HPA and fear responses in rats has been proposed to be mediated by environment-dependent variation in active maternal care. Here, we review this maternal mediation hypothesis based on the postnatal manipulation literature and on our own recent research in rats. We show that developmental plasticity of HPA and fear responses in rats cannot be explained by a linear single-factor model based on environment-dependent variation in active maternal care. However, by adding environmental stress as a second factor to the model, we were able to explain the variation in HPA and fear responses induced by postnatal manipulations. In this two-factor model, active maternal care and environmental stress (as induced, e.g., by long maternal separations or maternal food restriction) exert independent, yet opposing, effects on HPA reactivity and fearfulness in the offspring. This accounts well for the finding that completely safe and stable, as well as, highly stressful maternal environments result in high HPA reactivity and fearfulness compared to moderately challenging maternal environments. Furthermore, it suggests that the downregulation of the HPA system in response to stressful maternal environments could reflect adaptive developmental plasticity based on the increasing costs of high stress reactivity with increasingly stressful conditions. By contrast, high levels of environmental stress induced by environmental adversity might constrain such adaptive plasticity, resulting in non-adaptive or even pathological outcomes. Alternatively, however, developmental plasticity of HPA and fear responses in rats might be a function of maternal HPA activation (e.g., levels of circulating maternal glucocorticoid hormones). Thus, implying a U-shaped relationship between maternal HPA activation and HPA reactivity and fearfulness in the offspring, increasing maternal HPA activation with increasing environmental adversity would explain the effects of postnatal manipulations equally well. This raises the possibility that variation in active maternal care is an epiphenomenon, rather than a causal factor in developmental plasticity of HPA and fear responses in rats. Developmental plasticity of HPA and fear responses in rats and other animals has important implications for the design of animal experiments and for the well-being of experimental animals, both of which depend on the exact underlying mechanism(s). Importantly, however, more naturalistic approaches are needed to elucidate the adaptive significance of environment-dependent variation of HPA reactivity and fearfulness in view of discriminating between effects reflecting adaptive plasticity, phenotypic mismatch and pathological outcomes, respectively.