Osborne F. X. Almeida

Dynamic DNA methylation programs persistent adverse effects of early-life stress

Adverse early life events can induce long-lasting changes in physiology and behavior. We found that early-life stress (ELS) in mice caused enduring hypersecretion of corticosterone and alterations in passive stress coping and memory. This phenotype was accompanied by a persistent increase in arginine vasopressin (AVP) expression in neurons of the hypothalamic paraventricular nucleus and was reversed by an AVP receptor antagonist. Altered Avp expression was associated with sustained DNA hypomethylation of an important regulatory region that resisted age-related drifts in methylation and centered on those CpG residues that serve as DNA-binding sites for the methyl CpG–binding protein 2 (MeCP2). We found that neuronal activity controlled the ability of MeCP2 to regulate activity-dependent transcription of the Avp gene and induced epigenetic marking. Thus, ELS can dynamically control DNA methylation in postmitotic neurons to generate stable changes in Avp expression that trigger neuroendocrine and behavioral alterations that are frequent features in depression.

The Prefrontal Cortex as a Key Target of the Maladaptive Response to Stress

Research on the detrimental effects of stress in the brain has mainly focused on the hippocampus. Because prefrontal cortex (PFC) dysfunction characterizes many stress-related disorders, we here analyzed the impact of chronic stress in rats on the integrity of the hippocampal–PFC pathway, monitored by behavioral and electrophysiological function and morphological assessment. We show that chronic stress impairs synaptic plasticity by reducing LTP induction in the hippocampal–PFC connection; in addition, it induces selective atrophy within the PFC and severely disrupts working memory and behavioral flexibility, two functions that depend on PFC integrity. We also demonstrate that short periods of stress exposure induce spatial reference memory deficits before affecting PFC-dependent tasks, thus suggesting that the impairment of synaptic plasticity within the hippocampus-to-PFC connection is of relevance to the stress-induced PFC dysfunction. These findings evidence a fundamental role of the PFC in maladaptive responses to stress and identify this area as a target for intervention in stress-related disorders.

The mood-improving actions of antidepressants do not depend on neurogenesis but are associated with neuronal remodeling.

The mechanisms underlying the initiation/onset of, and the recovery from, depression are still largely unknown; views that neurogenesis in the hippocampus may be important for the pathogenesis and amelioration of depressive symptoms have gained currency over the years although the original evidence has been challenged. In this study, an unpredictable chronic mild stress protocol was used to induce a depressive-like phenotype in rats. In the last 2 weeks of stress exposure, animals were treated with the antidepressants fluoxetine, imipramine, CP 156,526 or SSR 1494515, alone or combined with methylazoxymethanol, a cytostatic agent used to arrest neurogenesis. We found that antidepressants retain their therapeutic efficacy in reducing both measured indices of depression-like behavior (learned helplessness and anhedonia), even when neurogenesis is blocked. Instead, our experiments suggest re-establishment of neuronal plasticity (dendritic remodeling and synaptic contacts) in the hippocampus and prefrontal cortex, rather than neurogenesis, as the basis for the restoration of behavioral homeostasis by antidepressants.

The neurosteroid tetrahydroprogesterone attenuates the endocrine response to stress and exerts glucocorticoid-like effects on vasopressin gene transcription in the rat hypothalamus

The neurosteroid tetrahydroprogesterone (5α-pregnan-3α-ol-20-one, allopregnanolone, THP), has been previously shown to counteract the anxiogenic effects of corticotropin-releasing hormone (CRH) and to interfere with noradrenergic and corticosteroid-mediated regulation of CRH release and gene transcription. Those observations indicated that, besides its sedative and analgesic activity, THP may also affect the neuroendocrine response to stress in a mode resembling that of corticosteroids. To examine this possibility, we compared the ability of THP, its precursor progesterone (P4), and the glucocorticoids dexamethasone (DEX) and corticosterone (CORT) to influence the pituitary-adrenal response to acute emotional stress and the adrenalectomy-induced increase in the gene transcription of the stress-related peptide arginine vasopressin (AVP) and of corticosteroid receptors (MR and GR) in the brain. Pretreatment of rats with a single dose of THP or P4 (50 μg/kg) significantly attenuated the elevation of plasma adrenocorticotropin (ACTH) and serum corticosterone after emotional stress; both steroids were, however, less potent than a similar dose of DEX. Administration of 1 mg of THP, CORT, or P4 to adrenalectomized (ADX) rats attenuated the increase in AVP mRNA levels in the ventromedial subdivision of the hypothalamic paraventricular nucleus (PVN), as compared with vehicle-treated ADX rats. However, whereas CORT and P4 influenced the ADX-induced increase in the transcription of both types of corticosteroid receptors in the hippocampus, these were unaffected by THP. In contrast to the glucocorticoids, THP and P4 failed to decrease plasma ACTH levels in rats deprived of endogenous steroids. These results demonstrate that the neurosteroid THP and its precursor P4 resemble glucocorticoids in their suppression of the pituitary-adrenal response to emotional stress; however, THP influences the transcription of glucocorticoid-responsive genes in brain structures involved in the regulation of the hypothalamo-pituitary-adrenal system in a fashion that is quite distinct from that obtained with glucocorticoids.

Morphological Correlates of Corticosteroid-Induced Changes in Prefrontal Cortex-Dependent Behaviors

Imbalances in the corticosteroid milieu have been implicated in several neuropsychiatric disorders, including depression and schizophrenia. Prefrontal cortex (PFC) dysfunction is also a hallmark of these conditions, causing impairments in executive functions such as behavioral flexibility and working memory. Recent studies have suggested that the PFC might be influenced by corticosteroids released during stress. To test this possibility, we assessed spatial working memory and behavioral flexibility in rats submitted to chronic adrenalectomy or treatment with corticosterone (25 mg/kg) or the synthetic glucocorticoid dexamethasone (300 μg/kg); the behavioral analysis was complemented by stereological evaluation of the PFC (prelimbic, infralimbic, and anterior cingulate regions), the adjacent retrosplenial and motor cortices, and the hippocampal formation. Dexamethasone treatment resulted in a pronounced impairment in working memory and behavioral flexibility, effects that correlated with neuronal loss and atrophy of layer II of the infralimbic, prelimbic, and cingulate cortices. Exposure to corticosterone produced milder impairments in behavioral flexibility, but not in working memory, and reduced the volume of layer II of all prefrontal areas. Interestingly, adrenalectomy-induced deleterious effects only became apparent on the reverse learning task and were not associated with structural alterations in the PFC. None of the experimental procedures influenced the morphology of retrosplenial or motor cortices, but stereological measurements confirmed previously observed effects of corticosteroids on hippocampal structure. Our results describe, for the first time, that imbalances in the corticosteroid environment can induce degeneration of specific layers of the PFC; these changes appear to be the morphological correlate of corticosteroid-induced impairment of PFC-dependent behavior(s).

Implications of estrogen-dependent brain organization for gender differences in hypothalamo-pituitary-adrenal regulation.

Estrogens, derived from the aromatization of testosterone in the brain, account for sex‐specific organization of neural circuits controlling gonadotropin release and sexual behavior. This study examines the possible organizing role of perinatal gonadal steroids in the manifestation of known, albeit unexplained, male‐female differences in basal and stress‐related adrenocortical secretion. We document here the existence of gender‐specific differences in the gene expression of hypothalamic corticotropin‐releasing hormone (CRH), and hippocampal and hypothalamic glucocorticoid receptors (GR), diurnal corticosterone secretion, as well as in the responsiveness of CRH and GR mRNA levels to exogenous estradiol. In addition, we report that neonatal estrogenization of female rats profoundly affects several regulatory substrates of the hypothalamo‐pituitary‐adrenal (HPA) axis, namely, the gene expression of CRH, arginine‐vasopressin (AVP) and GR in the brain, and the responsiveness of these parameters to estrogen. The neonatal treatment appeared to “defeminize” a number of neuroendocrine mechanisms related to HPA function; these changes were reminiscent of those observed in earlier studies on sexual differentiation of reproductive behavior and hormonal secretion. The results indicate a pivotal role for estrogens during early development for the determination of gender‐specific differences in HPA function in the mature animal and demonstrate for the first time that the brain‐organizing actions of gonadal steroids may extend to nonreproductive neuroendocrine axes.—Patchev, V. K., Hayashi, S., Orikasa, C., Almeida, O. F. X. Implications of estrogen‐dependent brain organization for gender differences in hypothalamo‐pituitary‐adrenal regulation. FASEB J. 9, 419–423 (1995)

Exacerbation of apoptosis in the dentate gyrus of the aged rat by dexamethasone and the protective role of corticosterone.

Glucocorticoid-induced cell loss in the dentate gyrus of rats of various ages was studied using the TUNEL procedure to detect apoptotic cells. A highly significant increase in the incidence of apoptosis was observed within the dentate hilus and granule cell layer within 24 h of a single injection of dexamethasone (DEX) in rats aged between 1 and 36 months; DEX-induced apoptosis was more pronounced with increasing age. Corticosterone (CORT) did not cause an increase in the rate of apoptosis above that found in age-matched controls. However, CORT pretreatment (3 h) resulted in a significantly attenuated DEX-induced apoptosis in both areas of the dentate gyrus. Serum CORT levels in saline-treated rats peaked at 6 months of age and reached a nadir at 36 months of age. The results indicate that (i) aged subjects are more susceptible to DEX in terms of dentate gyrus cell loss by apoptosis, (ii) CORT, which binds to Type I corticosteroid receptors with a high affinity, might serve to protect against the damaging effects of DEX which is a ligand of the Type II glucocorticoid receptor, and (iii) declining endogenous levels of CORT may increase the vulnerability of the dentate gyrus of aged rats to insult by DEX.

Neuropathology of stress

Environmental challenges are part of daily life for any individual. In fact, stress appears to be increasingly present in our modern, and demanding, industrialized society. Virtually every aspect of our body and brain can be influenced by stress and although its effects are partly mediated by powerful corticosteroid hormones that target the nervous system, relatively little is known about when, and how, the effects of stress shift from being beneficial and protective to becoming deleterious. Decades of stress research have provided valuable insights into whether stress can directly induce dysfunction and/or pathological alterations, which elements of stress exposure are responsible, and which structural substrates are involved. Using a broad definition of pathology, we here review the “neuropathology of stress” and focus on structural consequences of stress exposure for different regions of the rodent, primate and human brain. We discuss cytoarchitectural, neuropathological and structural plasticity measures as well as more recent neuroimaging techniques that allow direct monitoring of the spatiotemporal effects of stress and the role of different CNS structures in the regulation of the hypothalamic–pituitary–adrenal axis in human brain. We focus on the hypothalamus, hippocampus, amygdala, nucleus accumbens, prefrontal and orbitofrontal cortex, key brain regions that not only modulate emotions and cognition but also the response to stress itself, and discuss disorders like depression, post-traumatic stress disorder, Cushing syndrome and dementia.

Stimulation of hypothalamic β-endorphin and dynorphin release by corticotropin-releasing factor (in vitro)

Corticotropin-releasing factor (CRF) at doses of 10(-12)-10(-8) M significantly stimulated the release of beta-endorphin and dynorphin from superfused rat hypothalamic slices. These effects were shown to be mediated by the CRF receptor since they were antagonized by the CRF receptor antagonist alpha-helical CRF9-41 (10(-6) M). The two opioid peptides showed different time courses of response and in the case of beta-endorphin, an attenuation of the response upon continued exposure to CRF was observed.

Stress Acts Cumulatively To Precipitate Alzheimer's Disease-Like Tau Pathology and Cognitive Deficits

Stressful life experiences are likely etiological factors in sporadic forms of Alzheimers disease (AD). Many AD patients hypersecrete glucocorticoids (GCs), and their GC levels correlate with the rate of cognitive impairment and extent of neuronal atrophy. Severity of cognitive deficits in AD correlates strongly with levels of hyperphosphorylated forms of the cytoskeletal protein TAU, an essential mediator of the actions of amyloid β (Aβ), another molecule with a key pathogenic role in AD. Our objective was to investigate the sequential interrelationships between these various pathogenic elements, in particular with respect to the mechanisms through which stress might precipitate cognitive decline. We thus examined whether stress, through the mediation of GCs, influences TAU hyperphosphorylation, a critical and early event in the cascade of processes leading to AD pathology. Results from healthy, wild-type, middle-aged rats show that chronic stress and GC induce abnormal hyperphosphorylation of TAU in the hippocampus and prefrontal cortex (PFC), with contemporaneous impairments of hippocampus- and PFC-dependent behaviors. Exogenous GC potentiated the ability of centrally infused Aβ to induce hyperphosphorylation of TAU epitopes associated with AD and cytoplasmic accumulation of TAU, while previous exposure to stress aggravated the biochemical and behavioral effects of GC in Aβ-infused animals. Thus, lifetime stress/GC exposure may have a cumulative impact on the onset and progress of AD pathology, with TAU hyperphosphorylation serving to transduce the negative effects of stress and GC on cognition.