Chronic restraint stress for 6h/21d causes hippocampal CA3 apical dendritic retraction, which parallels spatial memory impairments in male rats. Recent research suggests that chronic immobilization stress for 2h/10d induces CA3 dendritic retraction (Vyas et al., 2002) and questions whether CA3 dendritic retraction and spatial memory deficits can be produced sooner than found following 6h/21d of restraint stress. Therefore, this study investigated the effects of four different durations of chronic restraint stress (varied by hours/day and total number of days) and the subsequent effects on hippocampal CA3 morphology and spatial memory in the same male Sprague-Dawley rats. The results showed that only rats exposed to the 6h/21d restraint paradigm exhibited CA3 apical dendritic retraction, consistent spatial memory deficits, and decreased body weight gain compared to experimental counterparts and controls. While chronically stressing a rat with wire mesh restraint has a physical component, it acts primarily as a psychological stressor, and these findings support the interpretation that chronic psychological stress produces hippocampal-dependent cognitive deficits that are consistent with hippocampal structural changes. Differences in stress effects observed across different studies may be due to rat strain, type of stressor, and housing conditions; however, the current findings support the use of chronic restraint stress, with wire mesh, for 6h/21d as a reliable and efficient method to produce psychological stress and to cause CA3 dendritic retraction and spatial memory deficits in male Sprague-Dawley rats.

Many neurological and psychiatric maladies originate from the deprivation of the human brain from estrogens. However, current hormone therapies cannot be used safely to treat these conditions commonly associated with menopause because of detrimental side-effects in the periphery. The latter also prevents the use of the hormone for neuroprotection. Here we show that a small-molecule bioprecursor prodrug, 10β,17β-dihydroxyestra-1,4-dien-3-one (DHED), converts to 17β-estradiol in the brain after systemic administration, but remains inert in the rest of the body. The localized and rapid formation of estrogen from the prodrug was revealed by a series of in vivo bioanalytical assays and through in vivo imaging in rodents. DHED treatment efficiently alleviated symptoms originated from brain estrogen deficiency in animal models of surgical menopause and provided neuroprotection in a rat stroke model. Concomitantly, we determined that 17β-estradiol formed in the brain from DHED elicited changes in gene expression and neuronal morphology identical to those obtained after direct 17β-estradiol treatment. Altogether, complementary functional and mechanistic data show that our approach is highly relevant therapeutically, because administration of the prodrug selectively produces estrogen in the brain independently from the route of administration and treatment regimen. Therefore, peripheral responses associated with the use of systemic estrogens, such as stimulation of the uterus and estrogen-responsive tumor growth, were absent. Collectively, our brain-selective prodrug approach may safely provide estrogen neuroprotection and medicate neurological and psychiatric symptoms developing from estrogen deficiency, particularly those encountered after surgical menopause, without the adverse side-effects of current hormone therapies.

Emerging data report sex differences in how the brain responds to chronic stress. Here, we investigated the effects of chronic restraint stress (6 h/day/21 days) on hippocampal morphology and function in ovariectomized female rats. Chronic restraint stress caused CA3 apical dendritic retraction in short-and long-shafted neurons, while it reduced basal dendritic arbors in long-shafted neurons only. Chronic restraint did not affect CA1 dendritic arborization, although it increased the proportion of CA1 spine heads compared with controls. Both stressed and control animals performed well on the Y-maze, a spatial memory task. However, chronic stress enhanced Y-maze performance compared with controls, which may reflect facilitated spatial memory or reduced habituation. Ymaze performance correlated with CA1 spine head proportion. This relationship suggests that spatial ability in females may be more tightly coupled with CA1 morphology, which may override the influence of CA3 dendritic retraction. Thus, this research provides additional evidence that CA3 morphology does not always parallel spatial memory.

Two pulses of 17β-estradiol (10µg) are commonly used to increase hippocampal CA1 apical dendritic spine density and alter spatial performance in ovariectomized (OVX) female rats, but rarely are the measures combined. The goal of this study was to use this two-pulse injection protocol repeatedly with intervening wash-out periods in the same rats to: 1) measure spatial ability using different tasks that require hippocampal function and 2) determine whether ovarian hormone depletion for an extended 10-week period reduces 17β-estradiol's effectiveness in elevating CA1 apical dendritic spine density. Results showed that two injections of 10µg 17β-estradiol (72 and 48 hrs prior to testing and timed to maximize CA1 apical spine density at behavioral assessment) corresponded to improved spatial memory performance on object placement. In contrast, two injections of 5µg 17β-estradiol facilitated spatial learning on the water maze compared to rats given two injections of 10µg 17β-estradiol or the sesame oil vehicle. Neither 17β-estradiol dose altered Y-maze performance. As expected, the intermittent two-pulse injection protocol increased CA1 apical spine density, but ten weeks of OVX without estradiol treatment decreased the effectiveness of 10µg 17β-estradiol to increase CA1 apical spine density. Moreover, two pulses of 5µg 17β-estradiol injected intermittently failed to alter CA1 apical spine density and decreased basal spine density. These results demonstrate that extended time without ovarian hormones reduces 17β-estradiol's effectiveness to increase CA1 apical spine density. Collectively, these findings highlight the complex interactions among estradiol, CA1 spine density/morphology, and task requirements, all of which contribute to behavioral outcomes.

We previously found that chronic stress conditions producing CA3 dendritic retraction and spatial memory deficits make the hippocampus vulnerable to the neurotoxin ibotenic acid (IBO). The purpose of this study was to determine whether exposure to chronic corticosterone (CORT) under conditions that produce CA3 dendritic retraction would enhance CA3 susceptibility to IBO. Male Sprague Dawley rats were chronically treated for 21 d with CORT in drinking water (400 g/ml), and half were given daily injections of phenytoin (40 mg/kg), an antiepileptic drug that prevents CA3 dendritic retraction. Three days after treatments stopped, IBO was infused into the CA3 region. Conditions producing CA3 dendritic retraction (CORT and vehicle) exacerbated IBO-induced CA3 damage compared with conditions in which CA3 dendritic retraction was not observed (vehicle and vehicle, vehicle and phenytoin, CORT and phenytoin). Additionally, spatial recognition memory was assessed using the Y-maze, revealing that conditions producing CA3 dendritic retraction failed to impair spatial recognition memory. Furthermore, CORT levels in response to a potentially mild stressor (injection and Y-maze exposure) stayed at basal levels and failed to differ among key groups (vehicle and vehicle, CORT and vehicle, CORT and phenytoin), supporting the interpretations that CORT levels were unlikely to have been elevated during IBO infusion and that the neuroprotective actions of phenytoin were not through CORT alterations. These data are the first to show that conditions with prolonged glucocorticoid elevations leading to structural changes in hippocampal dendritic arbors can make the hippocampus vulnerable to neurotoxic challenges. These findings have significance for many disorders with elevated glucocorticoids that include depression, schizophrenia, Alzheimer's disease, and Cushing's disease.

Chronic stress produces sex-specific neuromorphological changes in a variety of brain regions, which likely contribute to the gender differences observed in stress-related illnesses and cognitive ability. Here, we review the literature investigating the relationship between chronic stress and sex differences on brain plasticity and function, with an emphasis on morphological changes in dendritic arborization and spines in the hippocampus, prefrontal cortex, and amygdala. These brain structures are highly interconnected and sensitive to stress and gonadal hormones, and influence a variety of cognitive abilities. Although much less work has been published using female subjects than with male subjects, the findings suggest that the relationship between brain morphology and function is very different between the sexes. After reviewing the literature, we present a model showing how chronic stress influences the morphology of these brain regions and changes the dynamic of how these limbic structures interact with each other to produce altered behavioral outcomes in spatial ability, behavioral flexibility/executive function, and emotional arousal.

Chronic stress may have different effects on hippocampal CA3 and CA1 neuronal morphology and function depending upon hormonal status, but rarely are manipulations of stress and gonadal steroids combined. Experiment 1 investigated the effects of chronic restraint and 17β-estradiol replacement on CA3 and CA1 dendritic morphology and spatial learning in ovariectomized female Sprague-Dawley rats. Ovariectomized rats were implanted with 25% 17β-estradiol, 100% cholesterol or blank silastic capsules, and then chronically restrained (6h/d/21d) or kept in home cages. 17β-estradiol or cholesterol prevented stress-induced CA3 dendritic retraction, increased CA1 apical spine density, and altered CA1 spine shape. The combination of chronic stress and 17β-estradiol facilitated water maze acquisition compared to chronic stress + blank implants and nonstressed controls + 17β-estradiol. To further investigate the interaction between 17β-estradiol and stress on hippocampal morphology, Experiment 2 was conducted on gonadally intact, cycling female rats that were chronically restrained (6h/d/21d) and then euthanized at proestrus (high ovarian hormones) or estrus (low ovarian hormones). Cycling female rats failed to show chronic stress-induced CA3 dendritic retraction at either estrous phase. Chronic stress enhanced the ratio of CA1 basal spine heads to headless spines as found in Experiment 1. In addition, proestrous rats displayed increased CA1 spine density regardless of stress history. These results show that 17β-estradiol or cholesterol protect against chronic stress-induced CA3 dendritic retraction in females. These stress-and 17β-estradiol-induced morphological changes may provide insight into how dendritic complexity and spine properties contribute to spatial ability.

Chronic stress has detrimental effects on hippocampal integrity, while environmental enrichment (EE) has beneficial effects when initiated early in development. In this study, we investigated whether EE initiated in adulthood would mitigate chronic stress effects on cognitive function and hippocampal neuronal architecture, when EE started one week before chronic stress began, or two weeks after chronic stress onset. Adult male Sprague Dawley rats were chronically restrained (6h/d) or assigned as non-stressed controls and subdivided into EE or non-EE housing. After restraint ended, rats were tested on a radial arm water maze (RAWM) for 2-d to assess spatial learning and memory. The first study showed that when EE began prior to 3-weeks of chronic stress, EE attenuated chronic stress-induced impairments in acquisition, which corresponded with the prevention of chronic stress-induced reductions in CA3 apical dendritic length. A second study showed that when EE began 2-weeks after the onset of a 5-week stress regimen, EE blocked chronic stress-induced impairments in acquisition and retention at 1-h and 24-h delays. RAWM performance corresponded with CA3 apical dendritic complexity. Moreover, rats in EE housing (control or stress) exhibited similar corticosterone profiles across weeks, which differed from the muted corticosterone response to restraint by the chronically stressed pair-housed rats. These data support the interpretation that chronic stress and EE may act on similar mechanisms within the hippocampus, and that manipulation of these factors may yield new directions for optimizing brain integrity and resilience under chronic stress or stress related neuropsychological disorders in the adult.