Use of an Eight-arm Radial Water Maze to Assess Working and Reference Memory Following Neonatal Brain Injury

Working and reference memory are commonly assessed using the land based radial arm maze. However, this paradigm requires pretraining, food deprivation, and may introduce scent cue confounds. The eight-arm radial water maze is designed to evaluate reference and working memory performance simultaneously by requiring subjects to use extra-maze cues to locate escape platforms and remedies the limitations observed in land based radial arm maze designs. Specifically, subjects are required to avoid the arms previously used for escape during each testing day (working memory) as well as avoid the fixed arms, which never contain escape platforms (reference memory). Re-entries into arms that have already been used for escape during a testing session (and thus the escape platform has been removed) and re-entries into reference memory arms are indicative of working memory deficits. Alternatively, first entries into reference memory arms are indicative of reference memory deficits. We used this maze to compare performance of rats with neonatal brain injury and sham controls following induction of hypoxia-ischemia and show significant deficits in both working and reference memory after eleven days of testing. This protocol could be easily modified to examine many other models of learning impairment.     

Social isolation improves working memory at reversal but not primary radial-arm maze learning in rats

Social isolation starting from the 21st day of birth affected neither a short-term nor a long-term memory in male rats at primary acquisition learning in an 8-arm radial maze. A number of the short-term and long-term memory errors were substantially decreased during primary learning but the difference between groups was not significant. Isolates were faster to start a search in an individual trial and took less time to finish offa trial. During the reversal learning, when baited and non-baited arms were reversed, the isolates outperformed of socially reared rats on working but not reference memory task. In overall they made twice less working memory errors than socially reared animals. During the reversal learning the isolates were also faster than non-isolates in initiation and completion of a trial. Maternal separation of rat's pups on the postnatal days 1-21 for 4 hr per day did not affect either working or reference memory on both primary and reversal learning. The data obtained are discussed on basis of influence of stress in early postnatal life on hypothalamo-pituitary axis and its effects on behavior of adult animals.     

Estrogen Enhances Performance of Female Rats during Acquisition of a Radial Arm Maze

Estrogen can influence the expression of behaviors not associated directly with reproduction, including learning and memory. However, the effects of estrogen on learning and memory in mammals are complex, dependent on a variety of factors. The radial arm maze is a traditional experimental task that takes advantage of the natural foraging strategy of rats and provides an appropriate measure for studying the effects of estrogen on working memory in this species. In the experiments reported here, ovariectomized rats were implanted subcutaneously with 5-mm Silastic capsules containing 25% estradiol diluted with cholesterol. Control females received 5-mm Silastic capsules containing 100% cholesterol. Results of three separate experiments demonstrated that estradiol administered by Silastic implants for 30 days prior to eight-arm radial maze training, during the 24 days of maze training, or both significantly improved working memory performance compared to females treated with cholesterol alone, as indicated by improved arm choice accuracy over trials. The positive effect of estradiol exposure prior to training suggests that estrogen may induce neuronal changes that persist beyond the period of exposure with functional consequences for behavior.     

The effects of chronic estradiol treatment on working memory deficits induced by combined infusion of beta-amyloid (1-42) and ibotenic acid

Estrogen limits in vitro neuron death induced by application of beta-amyloid, the cytotoxic peptide linked to Alzheimer's disease. However, the ability of estrogen to protect neurons and preserve cognitive function in vivo following exposure to beta-amyloid has not been demonstrated. Our objective was to evaluate the potential of estrogen to reduce spatial working memory deficits in female rats induced by administration of a neurotoxic form of beta-amyloid in combination with the excitotoxin, ibotenic acid. The interaction of beta-amyloid with excitotoxic factors may underlie cognitive deficits associated with Alzheimer's disease. Therefore, to create an experimental model typical of early Alzheimer's disease a low dose of ibotenic acid was administered with beta-amyloid into the dorsal hippocampus. Ovariectomized rats were implanted subcutaneously with Silastic capsules that produce physiological levels of 17beta-estradiol 10 days before bilateral intrahippocampal injections of aggregated beta-amyloid (1-42) and ibotenic acid. Capsules remained in situ throughout behavioral testing. When tested 3-10 weeks after neurotoxin treatment, females without estrogen capsules exhibited delay-dependent impairments in working memory performance on a water maze and a radial arm maze. Females treated with estrogen and combined neurotoxins displayed working memory performance comparable to unlesioned females on both tasks. Neurotoxin treatment increased immunoreactivity for glial fibrillary acidic protein but this measure was unaffected by estradiol treatment indicating that estrogen did not limit glial proliferation. Results indicate that estrogen prevented deficits in spatial working memory induced by neurotoxin treatments intended to mimic the pathology of early Alzheimer's disease.     

fMRI Hippocampal Activity During a VirtualRadial Arm Maze

Numerous studies have shown that the hippocampus is critical for spatial memory. Within nonhuman research, a task often used to assess spatial memory is the radial arm maze. Because of the spatial nature of this task, this maze is often used to assess the function of the hippocampus. Our goal was to extrapolate this task to humans and examine whether healthy undergraduates utilize their hippocampus while performing a virtual reality version of the radial arm maze task. Thirteen undergraduates performed a virtual radial arm maze during functional magnetic resonance imaging. The brain maps of activity reveal bilateral hippocampal BOLD signal changes during the performance of this task. However, paradoxically, this BOLD signal change decreases during the spatial memory component of the task. Additionally, we note frontal cortex activity reflective of working memory circuits. These data reveal that, as predicted by the rodent literature, the hippocampus is involved in performing the virtual radial arm maze in humans. Hence, this virtual reality version may be used to assess the integrity of hippocampus so as to predict risk or severity in a variety of psychiatric disorders.     

The effects of exogenous testosterone on spatial memory in rats

Testosterone (T) is known to affect spatial abilities in men and women. Studies focusing on this relationship showed that both endogenous variability of T and administration of exogenous T, altered mental rotation and spatial visualization. Organizational and activational effects of T can be separately identified. The aim of our study was to evaluate the activational effects of exogenous T on spatial memory in male and female rats. T was administered 3 times a week over a two week period in either 1 mg/kg for low testosterone group or 10 mg/kg for high testosterone group. The Morris water maze was performed to assess the rat’s working and reference spatial memory. T and estradiol levels were measured in plasma. Increase in plasma T levels was confirmed in the experimental groups in comparison to the control groups (receiving sterile oil, 3 times a week over a two week period). Low dose T impaired working, but improved reference memory in female rats. In male rats the negative effects of T (both doses) on reference memory were shown. This experiment showed that the activational effects of exogenous testosterone on spatial memory of rats were gender and dose-dependent.     

Testosterone modulates performance on a spatial working memory task in male rats

Gonadal hormones have been shown to modulate memory retention in female rats. The current experiments examine the role of testicular hormones in modulating the performance of male rats on two spatial water maze tasks. In the first study, castrated and intact rats were trained on the visible platform and hidden platform versions of the Morris water maze task. Castration did not affect performance on either version of this reference memory task with castrated and intact rats demonstrating similar performance both during acquisition and on post-training probe trials. In the second experiment, castrated and intact rats were tested on a delayed-matching-to-place version of the water maze. Rats received a series of trial pairs in the maze with a hidden platform located in the same pool location on the exposure and retention trials of each pair; between pairs of trials, however, the platform was repositioned to a novel pool location. The interval between trials was either 10- or 60-min and memory retention, taken as the difference between the pathlengths on the exposure and retention trials, declined as the interval increased. Relative to intact males, castrated males demonstrated impaired working memory retention at 60-min but not at 10-min retention intervals. This interval-dependent impairment in working memory retention was reversed by physiologic levels of testosterone replacement. These findings indicate that castration does not significantly affect acquisition or probe trial performance on a classic reference memory task but does impair spatial working memory retention, an effect that is reversed by exogenous testosterone.     

The effect of anastrozol on spatial memory in adult rats

Androgens are known to affect cognitive functions via organizational and activational effects. It is unknown whether the effects are mediated via the androgen receptor or after conversion to estradiol with aromatase via estrogen receptors. The aim of our study was to find out whether testosterone affects spatial memory directly or through its metabolite estradiol. Rats were treated with testosterone; with testosterone and the aromatase blocker anastrozole or saline. An 8 radial arm maze was used for testing spatial memory twice daily for 4 days. Each arm was baited with food, and the ability of animals to learn the location of food was assessed. Testosterone treated rats and control rats achieved comparable coefficients of spatial memory, although the plasma levels of testosterone differed markedly. Anastrozole treatment resulted in the worst performance in the maze. The differences between groups did not reach the level of significance. It can be concluded that aromatase and, thus, the conversion of testosterone to estradiol may play a role in spatial memory, as pharmacological blockade of aromatase led to a decrease in maze performace of adult male rats. Detailed molecular mechanisms should be the focus of further studies.     

Exposure to social defeat stress in adolescence improves the working memory and anxiety-like behavior of adult female rats with intrauterine growth restriction,independently of hippocampal neurogenesis

Intrauterine growth restriction (IUGR) is a risk factor for memory impairment and emotional disturbance during growth and adulthood. However, this risk might be modulated by environmental factors during development. Here we examined whether exposing adolescent male and female rats with thromboxane A2-induced IUGR to social defeat stress (SDS) affected their working memory and anxiety-like behavior in adulthood. We also used BrdU staining to investigate hippocampal cellular proliferation and BrdU and NeuN double staining to investigate neural differentiation in female IUGR rats. In the absence of adolescent stress, IUGR female rats, but not male rats, scored significantly lower in the T-maze test of working memory and exhibited higher anxiety-like behavior in the elevated-plus maze test compared with controls. Adolescent exposure to SDS abolished these behavioral impairments in IUGR females. In the absence of adolescent stress, hippocampal cellular proliferation was significantly higher in IUGR females than in non-IUGR female controls and was not influenced by adolescent exposure to SDS. Hippocampal neural differentiation was equivalent in non-stressed control and IUGR females. Neural differentiation was significantly increased by adolescent exposure to SDS in controls but not in IUGR females. There was no significant difference in the serum corticosterone concentrations between non-stressed control and IUGR females; however, adolescent exposure to SDS significantly increased serum corticosterone concentration in control females but not in IUGR females. These results demonstrate that adolescent exposure to SDS improves behavioral impairment independent of hippocampal neurogenesis in adult rats with IUGR.     

Effect of losartan and enalapril on cognitive deficit caused by Goldblatt induced hypertension

The present study was designed to evaluate the learning and memory, in an altered physiological state associated with increased blood pressure and activated renin angiotensin system in Wistar rats. The role of angiotensin in cognitive function was assessed by treatment with angiotensin converting enzyme (ACE) inhibitor enalapril (2 mg/kg), angiotensin 1 receptor (AT(1)) antagonist losartan (5 mg/kg) and their combination. The experimental renal hypertension was induced by the method of Goldblatt. Learning and memory was assessed using the radial arm maze test. Acetylcholine esterase (AChE) levels in the pons medulla, hippocampus, striatum and frontal cortex were measured as a cholinergic marker of learning and memory. Results indicate that in comparison to normotensive rats, renal hypertensive rats committed significantly higher number of errors and took more trials and days to learn the radial arm maze learning and exhibited memory deficit in the radial arm maze retrieval after two weeks of retention interval, indicating impaired acquisition and memory. Treatment with enalapril, losartan and their combination attenuated the observed memory deficits indicating a possible role of renin angiotensin system in cognitive function. AChE level was reduced in hippocampus and frontal cortex of renal hypertensive rats which could be attributed to the observed memory deficit in hypertensive rats. It can be concluded that, renal hypertensive rats had a poor acquisition, retrieval of the learned behavior, perhaps a possible disturbance in memory consolidation process and that this state was reversed with ACE inhibitor enalapril and AT 1 receptor antagonist losartan.     

Spatial memory deficits in maternal iron deficiency paradigms are associated with altered glucocorticoid levels

"The goal of this study was to examine the effect of maternal iron deficiency on the developing hippocampus in order to define a developmental window for this effect, and to see whether iron deficiency causes changes in glucocorticoid levels. The study was carried out using pre-natal, post-natal, and pre + post-natal iron deficiency paradigm. Iron deficient pregnant dams and their pups displayed elevated corticosterone which, in turn, differentially affected glucocorticoid receptor (GR) expression in the CA1 and the dentate gyrus. Brain Derived Neurotrophic Factor (BDNF) was reduced in the hippocampi of pups following elevated corticosterone levels. Reduced neurogenesis at P7 was seen in pups born to iron deficient mothers, and these pups had reduced numbers of hippocampal pyramidal and granule cells as adults. Hippocampal subdivision volumes also were altered. The structural and molecular defects in the pups were correlated with radial arm maze performance; reference memory function was especially affected. Pups from dams that were iron deficient throughout pregnancy and lactation displayed the complete spectrum of defects, while pups from dams that were iron deficient only during pregnancy or during lactation displayed subsets of defects. These findings show that maternal iron deficiency is associated with altered levels of corticosterone and GR expression, and with spatial memory deficits in their pups."     

Where Have I Been? Where Should I Go? Spatial Working Memory on a Radial Arm Maze in a Rat Model of Depression

Disturbances in cognitive functioning are among the most debilitating problems experienced by patients with major depression. Investigations of these deficits in animals help to extend and refine our understanding of human emotional disorder, while at the same time providing valid tools to study higher executive functions in animals. We employ the “learned helplessness” genetic rat model of depression in studying working memory using an eight arm radial maze procedure with temporal delay. This so-called delayed spatial win-shift task consists of three phases, training, delay and test, requiring rats to hold information on-line across a retention interval and making choices based on this information in the test phase. According to a 2×2 factorial design, working memory performance of thirty-one congenitally helpless (cLH) and non-helpless (cNLH) rats was tested on eighteen trials, additionally imposing two different delay durations, 30 s and 15 min, respectively. While not observing a general cognitive deficit in cLH rats, the delay length greatly influenced maze performance. Notably, performance was most impaired in cLH rats tested with the shorter 30 s delay, suggesting a stress-related disruption of attentional processes in rats that are more sensitive to stress. Our study provides direct animal homologues of clinically important measures in human research, and contributes to the non-invasive assessment of cognitive deficits associated with depression.     

Spatial working memory in rats: the effect of the dipeptide gamma-L-glutamyl-taurine and haloperidol

To assess the possible involvement of the dipeptide gamma-L-glutamyl-taurine (Litoralon) and some of its analogues in the maintenance of spatial working memory, rats were treated with the dipeptides immediately or 2 hours after completing the first four choices in an 8 arm radial maze, or 3 hours before the test to exclude proactive effects of the compounds. Treatment with Litoralon, SZJ 3381 and 3361 at doses of 100 and 500 micrograms/kg (i.p.) did not impair spatial memory in rats, regardless of when these substances were injected during the session. By contrast, haloperidol (500 micrograms/kg, i.p.) treatment resulted in a dramatic decrease of performance.     

Effects of testosterone on spatial learning and memory in adult male rats

A male advantage over females for spatial tasks has been well documented in both humans and rodents, but it remains unclear how the activational effects of testosterone influence spatial ability in males. In a series of experiments, we tested how injections of testosterone influenced the spatial working and reference memory of castrated male rats. In the eight-arm radial maze, testosterone injections (0.500 mg/rat) reduced the number of working memory errors during the early blocks of testing but had no effect on the number of reference memory errors relative to the castrated control group. In a reference memory version of the Morris water maze, injections of a wide range of testosterone doses (0.0625-1.000 mg/rat) reduced path lengths to the hidden platform, indicative of improved spatial learning. This improved learning was independent of testosterone dose, with all treatment groups showing better performance than the castrated control males. Furthermore, this effect was only observed when rats were given testosterone injections starting 7 days prior to water maze testing and not when injections were given only on the testing days. We also observed that certain doses of testosterone (0.250 and 1.000 mg/rat) increased perseverative behavior in a reversal-learning task. Finally, testosterone did not have a clear effect on spatial working memory in the Morris water maze, although intermediate doses seemed to optimize performance. Overall, the results indicate that testosterone can have positive activational effects on spatial learning and memory, but the duration of testosterone replacement and the nature of the spatial task modify these effects.     

Tetrachlorodibenzo-p-dioxin exposure alters radial arm maze performance and hippocampal morphology in female AhR mice

Perinatal exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) has been reported to alter spatial learning in rats tested on a radial arm maze (RAM). TCDD is believed to exert most of its effects through binding to the aryl hydrocarbon receptor (AhR). To determine whether the AhR mediates TCDD-induced alterations in spatial learning, we tested male and female AhR-knockout (AhR-/-), heterozygous (AhR+/-) and wild-type (AhR+/+) mice on the RAM. AhR+/- male and female mice were time mated, and treated dams were dosed with 5 microg TCDD/kg body weight on day 13 of gestation. When offspring reached adulthood, male and female AhR+/+, AhR+/- and AhR-/- mice from TCDD-exposed and unexposed litters were tested on the eight-arm RAM. After testing, we examined hippocampal morphology as visualized by the Timm's silver sulfide stain. TCDD-exposed female AhR+/- mice made more errors than their respective controls on the RAM and exhibited a decrease in the size of the intra- and infrapyramidal mossy fiber (IIP-MF) field of the hippocampus. None of the other TCDD-exposed groups differed from their respective control groups with regard to maze performance or hippocampal morphology. The reduction of IIP-MF field indicates a possible morphological basis for the learning deficit that was observed in the female AhR+/- mice. It is hypothesized that the effect of TCDD exposure is AhR dependent and that TCDD may alter GABAergic activity in the hippocampus of female mice during development.     

Does Chronic Unpredictable Stress during Adolescence Affect Spatial Cognition in Adulthood?

Spatial abilities allow animals to retain and cognitively manipulate information about their spatial environment and are dependent upon neural structures that mature during adolescence. Exposure to stress in adolescence is thought to disrupt neural maturation, possibly compromising cognitive processes later in life. We examined whether exposure to chronic unpredictable stress in adolescence affects spatial ability in late adulthood. We evaluated spatial learning, reference and working memory, as well as long-term retention of visuospatial cues using a radial arm water maze. We found that stress in adolescence decreased the rate of improvement in spatial learning in adulthood. However, we found no overall performance impairments in adult reference memory, working memory, or retention caused by adolescent-stress. Together, these findings suggest that adolescent-stress may alter the strategy used to solve spatial challenges, resulting in performance that is more consistent but is not refined by incorporating available spatial information. Interestingly, we also found that adolescent-stressed rats showed a shorter latency to begin the water maze task when re-exposed to the maze after an overnight delay compared with control rats. This suggests that adolescent exposure to reoccurring stressors may prepare animals for subsequent reoccurring challenges. Overall, our results show that stress in adolescence does not affect all cognitive processes, but may affect cognition in a context-dependent manner.

Highlights

• -Rats were reared with or without chronic unpredictable stress in adolescence.
• -In adulthood, spatial cognitive abilities were tested in a radial arm water maze.
• -Prior-stressed rats began searching faster in the maze after an overnight delay.
• -Prior stress may facilitate faster action in challenging situations.
• -Prior stress did not affect learning, reference or working memory, or retention.

     

Protective effects of methanol extract of Acori graminei rhizoma and Uncariae Ramulus et Uncus on ischemia-induced neuronal death and cognitive impairments in the rat

Acori graminei rhizoma (AGR) and Uncariae Ramulus et Uncus (URE) have been widely used as herbal medicine against ischemia. In order to investigate whether AGR and URE influenced cerebral ischemia-induced neuronal and cognitive impairments, we examined the effect of AGR and URE on ischemia-induced cell death in the striatum, cortex and hippocampus, and on the impaired learning and memory in the Morris water maze and radial eight-arm maze in rats. After middle cerebral artery occlusion (MCAO) for 2 h, rats were administered saline, AGR or URE (100 mg/kg, p.o.) daily for three weeks, followed by their training to the tasks. In the water maze test, the animals were trained to find a platform in a fixed position during 6 days and then received a 60-s probe trial in which the platform was removed from the pool on the 7th day. In the radial eight-arm maze, animals were tested six times per week for 1 week. Rats with ischemic insults showed impaired learning and memory on the tasks. Pretreatment with AGR and URE produced a significant improvement in escape latency to find the platform in the Morris water maze and in the number of choice errors in the radial arm maze test. Consistent with behavioral data, pretreatments with AGR and URE significantly reduced ischemia-induced cell death in the hippocampal CA1 area. These results demonstrated that AGR and URE have a protective effect against ischemia-induced neuronal loss and learning and memory damage. Our studies suggest that AGR and URE may be useful in the treatment of vascular dementia.     

Adult rats exposed to early-life social isolation exhibit increased anxiety and conditioned fear behavior, and altered hormonal stress responses

Social isolation of rodents during development is thought to be a relevant model of early-life chronic stress. We investigated the effects of early-life social isolation on later adult fear and anxiety behavior, and on corticosterone stress responses, in male rats. On postnatal day 21, male rats were either housed in isolation or in groups of 3 for a 3 week period, after which, all rats were group-reared for an additional 2 weeks. After the 5-week treatment, adult rats were examined for conditioned fear, open field anxiety-like behavior, social interaction behavior and corticosterone responses to restraint stress. Isolates exhibited increased anxiety-like behaviors in a brightly-lit open field during the first 10 min of the test period compared to group-reared rats. Isolation-reared rats also showed increased fear behavior and reduced social contact in a social interaction test, and a transient increase in fear behavior to a conditioned stimulus that predicted foot-shock. Isolation-reared rats showed similar restraint-induced increases in plasma corticosterone as group-reared controls, but plasma corticosterone levels 2 h after restraint were significantly lower than pre-stress levels in isolates. Overall, this study shows that isolation restricted to an early part of development increases anxiety-like and fear behaviors in adulthood, and also results in depressed levels of plasma corticosterone following restraint stress.     

Prenatal testosterone improves the spatial learning and memory by protein synthesis in different lobes of the brain in the male and female rat

The high density of the steroid hormone receptors in the structures of temporal lobe involved in learning and memory, such as the hippocampus, perirhinal cortex, entorhinal cortex and amigdaloid complex, shows that there must be a direct relationship between gonadal hormones and organizational effects of steroid hormones in those structures during development of the nervous system. The present study was undertaken in order to investigate the effect of testosterone administration during the third week of gestation on the spatial memory formation of the offspring rats and the level of soluble proteins in the temporal lobe and frontal lobe of brain, as evidence of important organizational effects of androgens during prenatal development in brain sexual dimorphism. Animals have received testosterone undecanoate on days 14, 15, 16 and 19, 20, 21 of gestation. Learning and memory tests were started 100 days after the testosterone treatment. At the end of the experiments, the temporal and frontal lobes of brain were removed for assessing the level of soluble proteins. Testosterone treatment significantly improved spontaneous alternations percentage of male offspring in Y-maze task comparative with female offspring and reference memory in radial 8 arm-maze task (decreasing in number of reference memory errors in both male and female offspring groups), suggesting effects of both short and long-term memory. Also, testosterone significantly increased the brain soluble protein level of treated female rats in 14–16 prenatal days compared with the control group as well as the brain soluble protein level of treated male rats. These results suggest that steroid hormones play an important role in the spatial learning and memory formation by means of protein synthesis in different lobes of the brain.     

Effects of the NMDA antagonist MK-801 on radial maze performance in histidine-deficient rats

We examined the effects of a histidine-deficient diet on brain histamine contents as well as on learning and memory using the eight-arm radial maze in rats. A significant decrease in histamine content in the hippocampus was observed after long-term feeding of rats with a histidine-deficient diet. At the same time, significant enhancement of the acquisition process in radial maze performance was also observed. Pyrilamine did not show a significant effect on radial maze performance in histidine-deficient rats. On the other hand, pyrilamine caused a significant spatial memory deficit in control rats. Scopolamine was effective in inhibiting spatial memory in both histidine-deficient and control rats. MK-801 caused spatial memory deficits more potently in histidine-deficient rats than in controls. Brain glycine contents showed a significant increase in the hippocampus in histidine-deficient rats. These results indicated that the spatial memory deficits induced by MK-801 in histidine-deficient rats are closely related to increased glycine levels and activation of NMDA receptors.