Sarit Avishai-Eliner

Down-Regulation of Hypothalamic Corticotropin-Releasing Hormone Messenger Ribonucleic Acid (mRNA) Precedes Early-Life Experience-Induced Changes in Hippocampal Glucocorticoid Receptor mRNA1

Early-life experiences, including maternal interaction, profoundly influence hormonal stress responses during adulthood. In rats, daily handling during a critical neonatal period leads to a significant and permanent modulation of key molecules that govern hormonal secretion in response to stress. Thus, hippocampal glucocorticoid receptor (GR) expression is increased, whereas hypothalamic CRH-messenger RNA (mRNA) levels and stress-induced glucocorticoid release are reduced in adult rats handled early in life. Recent studies have highlighted the role of augmented maternal sensory input to handled rats as a key determinant of these changes. However, the molecular mechanisms, and particularly the critical, early events leading from enhanced sensory experience to long-lasting modulation of GR and CRH gene expression, remain largely unresolved. To elucidate the critical primary genes governing this molecular cascade, we determined the sequence of changes in GR-mRNA levels and in hypothalamic and amygdala CRH-mRNA expression at three developmental ages, and the temporal relationship between each of these changes and the emergence of reduced hormonal stress-responses. Down-regulation of hypothalamic CRH-mRNA levels in daily-handled rats was evident already by postnatal day 9, and was sustained through postnatal days 23 and 45, i.e. beyond puberty. In contrast, handling-related up-regulation of hippocampal GR-mRNA expression emerged subsequent to the 23rd postnatal day, i.e. much later than changes in hypothalamic CRH expression. The hormonal stress response of handled rats was reduced starting before postnatal day 23. These findings indicate that early, rapid, and persistent changes of hypothalamic CRH gene expression may play a critical role in the mechanism(s) by which early-life experience influences the hormonal stress-response long-term.

Altered regulation of gene and protein expression of hypothalamic-pituitary-adrenal axis components in an immature rat model of chronic stress.

Chronic stress early in postnatal life influences hormonal and behavioural responses to stress persistently, but the mechanisms and molecular cascades that are involved in this process have not been clarified. To approach these issues, a chronic stress paradigm for the neonatal rat, using limited bedding material to alter the cage environment, was devised. In 9‐day‐old rats subjected to this chronic stress for 1 week, significant and striking changes in the expression and release patterns of key molecules that govern the neuroendocrine stress responses were observed. The presence of sustained stress was evident from enhanced activation of peripheral elements of the neuroendocrine stress response, i.e. increased basal plasma corticosterone concentrations, high adrenal weight and decreased body weight. Central regulatory elements of the neuroendocrine stress response were perturbed, including reduced expression of hypothalamic corticotropin‐releasing hormone that, surprisingly, was accompanied by reduced glucocorticoid receptor expression. Thus, the effects of chronic sustained stress in the neonatal rat on the hypothalamic‐pituitary‐adrenal axis included substantial changes in the expression and activity of major regulators of this axis. Importantly, the changes induced by this chronic stress differed substantially from those related to acute or recurrent stress, providing a novel model for studying the long‐term effects of chronic, early life stress on neuroendocrine functions throughout life.

Naturalistic rodent models of chronic early-life stress

A close association between early-life experience and cognitive and emotional outcomes is found in humans. In experimental models, early-life experience can directly influence a number of brain functions long-term. Specifically, and often in concert with genetic background, experience regulates structural and functional maturation of brain circuits and alters individual neuronal function via large-scale changes in gene expression. Because adverse experience during sensitive developmental periods is often associated with neuropsychiatric disease, there is an impetus to create realistic models of distinct early-life experiences. These can then be used to study causality between early-life experiential factors and cognitive and emotional outcomes, and to probe the underlying mechanisms. Although chronic early-life stress has been linked to the emergence of emotional and cognitive disorders later in life, most commonly used rodent models of involve daily maternal separation and hence intermittent early-life stress. We describe here a naturalistic and robust chronic early-life stress model that potently influences cognitive and emotional outcomes. Mice and rats undergoing this stress develop structural and functional deficits in a number of limbic-cortical circuits. Whereas overt pathological memory impairments appear during adulthood, emotional and cognitive vulnerabilities emerge already during adolescence. This naturalistic paradigm, widely adopted around the world, significantly enriches the repertoire of experimental tools available for the study of normal brain maturation and of cognitive and stress-related disorders including depression, autism, post-traumatic stress disorder, and dementia.

Stress and the developing hippocampus: a double-edged sword?

The mechanisms that regulate neuronal function are a sum of genetically determined programs and experience. The effect of experience on neuronal function is particularly important during development, because early-life positive and adverse experience (stress) may influence the still “plastic” nervous system long-term. Specifically, for hippocampal-mediated learning and memory processes, acute stress may enhance synaptic efficacy and overall learning ability, and conversely, chronic or severe stress has been shown to be detrimental. The mechanisms that enable stress to act as this “double-edged sword” are unclear. Here, we discuss the molecular mediators of the stress response in the hippocampus with an emphasis on novel findings regarding the role of the neuropeptide known as corticotropin-releasing hormone (CRH). We highlight the physiological and pathological roles of this peptide in the developing hippocampus, and their relevance to the long-term effects of early-life experience on cognitive function during adulthood.

Differential regulation of glucocorticoid receptor messenger RNA (GR-mRNA) by maternal deprivation in immature rat hypothalamus and limbic regions

Maternal deprivation (MDep) of neonatal rats significantly influences the hypothalamic-pituitary-adrenal (HPA) axis. This study hypothesized that GR-mRNA modulation constituted an early, critical mechanism for the acute effects of MDep on neuroendocrine stress-responses. GR-mRNA hybridization signal in hippocampal CA1, hypothalamic paraventricular nucleus (PVN) and frontal cortex was significantly reduced immediately following 24 h MDep. In amygdala, cingulate cortex, PVN and CA1, apparent gender-dependent MDep effects on GR-mRNA expression were observed, without significant differences in absolute levels. Thus, rapid, region-specific MDep effects on GR-mRNA expression in HPA-regulating areas are shown, consistent with involvement of GR-expression in mechanisms of MDep influence on HPA tone.

The developmental profile of the corticotropin releasing factor receptor (CRF2) in rat brain predicts distinct age-specific functions

Corticotropin releasing factor (CRF) activates two known receptor types, CRF1, and CRF2. In the adult rat brain, CRF2 has a distinct distribution pattern, suggesting that it may mediate functions exclusive of CRF1. The goal of this study was to determine the age-dependent distribution of CRF2-messenger RNA (CRF2-mRNA) in the rat brain. Brains from rats sacrificed under stress-free conditions on fetal days (F) 15, 16, 17 and 19, and postnatal days 1, 3, 5, 7, 9, 12, 15, 25, 49, and 90 (adult) were analyzed using semiquantitative in situ hybridization histochemistry. The onset and distribution of CRF2-mRNA in the developing rat brain revealed important differences from the adult expression pattern: earliest expression of CRF2-mRNA was observed in the ventromedial hypothalamus (VMH) on F16. High levels of CRF2-mRNA were present in the fronto-parietal cortex in the fetal and early postnatal brain but not later. Conversely, no CRF2-mRNA was detectable in the ventroposterior (lateral and medial) thalamic nuclei prior to postnatal day 7. Distinct developmental profiles of CRF2-mRNA were also observed in the lateral septum, medial, basal and cortical amygdala nuclei, and in several hippocampal fields. In conclusion, CRF2 is expressed in the hypothalamus on F16, prior to the detection of CRF itself in the paraventricular nucleus. The differential levels and distributions of CRF2-mRNA in hypothalamic and limbic brain regions indicate a precise regulation of this receptors expression during development, as shown for CRF1. Regulation of the levels of CRF2 may modulate the effects of CRF (and related ligands) on target neurons, consistent with differential maturation of the functions mediated by this receptor.

Corticotropin Releasing Hormone Antagonist Does Not Prevent Adrenalectomy-Induced Apoptosis in the Dentate Gyrus of the Rat Hippocampus

Adrenalectomy in the mature rat leads to death of granule cells in the dentate gyrus of the hippocampal formation. The mechanisms underlying this cell death have not been fully clarified: It has been considered that the granule cells require adrenal steroids for their survival, since corticosterone replacement prevents their death. However, adrenalectomy-induced loss of negative feedback also increases levels of corticotropin releasing hormone (CRH) in several limbic brain regions. CRH is known to induce neuronal death in hippocampal regions rich in CRH receptors. This study tested the hypothesis that adrenalectomy-induced granule cell death is mediated via the enhanced activation of CRH receptors. The extent of granule cell degeneration was compared among 4 groups of young adult male rats: Sham-adrenalectomy controls, adrenalectomized rats, adrenalectomized rats infused with a CRH antagonist from the onset of steroid deprivation to the time of sacrifice, and adrenalectomized rats infused with vehicle only. (9-41)-alpha-helical CRH was administered using an osmotic pump into the cerebral ventricles. Adrenalectomy led to robust granule cell degeneration, which was maximal in the suprapyramidal blade of the dentate gyrus. Infusion of the CRH antagonist in doses shown to block CRH actions on limbic neurons did not decrease the number of degenerating granule cells compared with the untreated or vehicle-infused adrenalectomized groups. Therefore, blocking the actions of CRH does not prevent adrenalectomy-induced granule cell death, consistent with a direct effect of corticoids on the survival of these neurons.