Advertisement
Browse Subject Areas
?

Click through the PLOS taxonomy to find articles in your field.

For more information about PLOS Subject Areas, click here.

  • Loading metrics

Open Access

Peer-reviewed

Research Article

Impact of Chronic Stress Protocols in Learning and Memory in Rodents: Systematic Review and Meta-Analysis

  • Pedro Silva Moreira,

    Affiliations Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, Braga, Portugal, ICVS/3B’s, PT Government Associate Laboratory, Braga/Guimarães, Portugal, Clinical Academic Center–Braga, Braga, Portugal

    ⨯
  • Pedro R Almeida,

    Affiliation School of Criminology, Faculty of Law, University of Porto, Porto, Portugal

    ⨯
  • Hugo Leite-Almeida,

    Affiliations Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, Braga, Portugal, ICVS/3B’s, PT Government Associate Laboratory, Braga/Guimarães, Portugal

    ⨯
  • Nuno Sousa ,

    njcsousa@ecsaude.uminho.pt

    Affiliations Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, Braga, Portugal, ICVS/3B’s, PT Government Associate Laboratory, Braga/Guimarães, Portugal, Clinical Academic Center–Braga, Braga, Portugal

    ⨯
  • Patrício Costa

    Affiliations Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, Braga, Portugal, ICVS/3B’s, PT Government Associate Laboratory, Braga/Guimarães, Portugal, Clinical Academic Center–Braga, Braga, Portugal

    ⨯

Abstract

The idea that maladaptive stress impairs cognitive function has been a cornerstone of decades in basic and clinical research. However, disparate findings have reinforced the need to aggregate results from multiple sources in order to confirm the validity of such statement. In this work, a systematic review and meta-analyses were performed to aggregate results from rodent studies investigating the impact of chronic stress on learning and memory. Results obtained from the included studies revealed a significant effect of stress on global cognitive performance. In addition, stressed rodents presented worse consolidation of learned memories, although no significantly differences between groups at the acquisition phase were found. Despite the methodological heterogeneity across studies, these effects were independent of the type of stress, animals’ strains or age. However, our findings suggest that stress yields a more detrimental effect on spatial navigation tests’ performance. Surprisingly, the vast majority of the selected studies in this field did not report appropriate statistics and were excluded from the quantitative analysis. We have therefore purposed a set of guidelines termed PROBE (Preferred Reporting Orientations for Behavioral Experiments) to promote an adequate reporting of behavioral experiments.

Citation: Moreira PS, Almeida PR, Leite-Almeida H, Sousa N, Costa P (2016) Impact of Chronic Stress Protocols in Learning and Memory in Rodents: Systematic Review and Meta-Analysis. PLoS ONE 11(9): e0163245. https://doi.org/10.1371/journal.pone.0163245

Editor: Aditya Bhushan Pant, Indian Institute of Toxicology Research, INDIA

Received: January 14, 2016; Accepted: August 31, 2016; Published: September 23, 2016

Copyright: © 2016 Moreira et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Data Availability: All relevant data are within the paper and its Supporting Information files.

Funding: This work was funded by the European Commission (FP7) “SwitchBox” (Contract HEALTH-F2-2010-259772) project and co-financed by the Portuguese North Regional Operational Program (ON.2 – O Novo Norte) under the National Strategic Reference Framework (QREN), through the European Regional Development Fund (FEDER), and by Fundação Calouste Gulbenkian (Portugal) (Contract grant number: P-139977; project “ Better mental health during ageing based on temporal prediction of individual brain ageing trajectories (TEMPO)”). PSM is supported by an FCT fellowship grant, from the PhD-iHES program, with the reference PDE/BDE/113601/2015.

Competing interests: The authors have declared that no competing interests exist.

1. Introduction

Stress exposure is associated with an activation of the hypothalamic-pituitary-adrenal (HPA) axis[1]. Although this relationship is thought to be bi-directional, here we focus on the causal effect of stress on HPA axis. Repeated stress exposure is known to lead to an excessive HPA axis activation, resulting in an overproduction of glucocorticoids (GCs). As a consequence, neurochemical and neuroanatomical alterations in several brain regions may be observed, including the hippocampus, prefrontal cortex, amygdala[2], dorsal striatum[3], nucleus accumbens[4], bed nucleus of the stria terminalis[5] and brain stem[6]. In the particular case of the hippocampus, a high density of GC receptors has been found[7–10]. Indeed, as a consequence of GCs overproduction, neuronal atrophy as well as decreased neurogenesis have been observed in the dentate gyrus of the hippocampal formation[11].

In experimental settings, several protocols of chronic stress induction have been devised. Among these, the Chronic Mild Stress (CMS) and the Chronic Restraint Stress (CRS) protocols have been the most widely used in behavioral research. In a typical CMS protocol, animals are exposed to unpredictable stressors over a varying period of time (from days to several weeks)[12]. On the contrary, in CRS protocols, the same stressor (restraining) is repeatedly applied[13, 14]. Some authors have demonstrated that the repeated exposition to stress leads to impaired hippocampal-dependent functions[15, 16] (also confront with[17, 18]) in several cognitive paradigms, such as the radial arm maze (RAM)[19], the Morris water maze (MWM)[20], the novel object recognition task (NOR)[21], and the Y Maze (YM)[22] (see also [23],[24] for review). RAM and MWM are widely used experimental apparatus in which navigational and allocentric strategies are required; whereas, NOR and YM evaluate animals’ discrimination ability (novelty and path alternation, respectively).

The impact of chronic stress on cognitive performance is thought depend of biological (such as sex) and chronobiological (age) factors[25, 26]. Other aspects, including stress predictability, may also modulate these effects. For instance, it was reported that the implementation of predictable stressors enhances animals’ cognitive performance[27]. Adding further complexity to this issue, a recent study from our group revealed that the period of the day (diurnal/nocturnal) in which the stress protocol is implemented also modulates cognitive performance[28, 29].

This multi-factorial interplay may explain many of the inconsistencies found in the literature. Nevertheless, the deleterious impact of stress on cognitive functioning has been a cornerstone of decades of research. Many basic and clinical studies have departed from an assumption that has not always been confirmed. Therefore, it is critical to aggregate the data from multiple studies in order to clarify the abovementioned discrepancies. In this context, meta-analysis, though scarcely used in animal studies, is a powerful tool that incorporates the variability across studies, and allows the achievement of an overall estimate. Thus, it constitutes the most suitable means to untangle this issue. For this purpose, in this study we conducted a systematic review and meta-analyses in order to obtain an overall estimate of the impact of chronic stress on learning and memory in rodents. Furthermore, departing from our own observations, we also developed a set of guidelines with the aim of improving the quality of reporting of animal research experiments.

2. Materials and Methods

The systematic-review and meta-analyses adhered to PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses[30]) guidelines, including search strategy, selection criteria, data extraction and data analysis (S1 File).

2.1. Literature search

A comprehensive literature search of electronic databases PubMed (http://www.pubmed.gov) and SCOPUS (http://www.scopus.com) was concluded in March 2014 with the following keywords: [‘learning’ AND ‘memory’] OR ‘morris water maze’ OR ‘novel object recognition’] AND [‘chronic’ AND ‘stress’] AND [‘mice’ OR ‘rats’]. Only experimental studies published in English were included in this analysis. Reviews, commentaries, as well as unpublished studies were not considered. Studies were selected if they met all the following criteria:

  • implementation of a chronic stress protocol in post-weaning rodents, by applying CMS or CRS in one of the experimental groups;
  • at least one control group was required;
  • no other manipulation besides chronic stress was performed (e.g. drug treatments, enriched environment, physical exercise or others);
  • experimental subjects were not genetically altered or had compromised functioning due to lesions or other manipulations;
  • learning and memory were assessed in both control and experimental groups using validated tasks, such as the MWM, NOR, RAM and/or Y-M, after the implementation of the chronic stress protocol. Tasks requiring negative reinforcement, such as fear conditioning and passive avoidance tasks (see [31] and [32] for a review) were excluded. These tasks are characterized by an aversive and stressful nature. As a consequence, they were excluded with the aim of avoiding confounding effects.

2.2. Data extraction and management

Abstract selection: Two independent reviewers (PSM and PRA) selected eligible studies based on titles and abstracts’ screening. In the case of disagreements, a third reviewer (PC) decided if the study fulfilled the inclusion criteria.

To rule out subjectivity in the data gathering and entry process, data was independently extracted from eligible studies and recorded in separate databases by three reviewers (PC, PSM and PRA). Data from each study were abstracted using standardized forms in which the following characteristics were recorded: first author, publication year, stress protocol type, stress duration, sample size, animals’ age, gender and strain, and statistical measures for each behavioral parameter (means and standard deviations). Moreover, physiological indices (body weight, sucrose preference or corticosteroids’ levels) and behavioral measures (locomotor activity, anxious-like behavior) were also recorded.

If effect sizes could not be extracted/calculated from the available data, corresponding authors were asked (via e-mail contact) to provide additional statistical information. Afterwards, databases were compared and mismatching entries were identified and corrected upon discussion between the reviewers.

2.3. Data analysis

Heterogeneity was tested with the Cochran Q-test (p<0.10 indicates statistically significant heterogeneity[33]) and I2 statistic. I2 was calculated as I2 = [(Q–degree of freedom)/Q] ×100, where Q is the Cochran’s statistic. I2 values of 25, 50 and 75 represent low, medium and high heterogeneity, respectively. If high and significant heterogeneity (I2>75) was detected, a random-effects model (the Restricted Maximum-Likelihood method) was used to calculate the summary of pooled prevalence estimates. Otherwise, a fixed-effects model (the Mantel-Haenszel method) was preferred.

The presence of potential publication bias was examined through the visual inspection of funnel plot asymmetry, and statistically tested using the rank correlation method from Begg and Mazumdar (p<0.05 represents statistically significant publication bias).

Statistical analysis was conducted using Metafor package[34] in R software.

3. Results

3.1. Study selection

PRISMA diagram (Fig 1) illustrates the process of study selection.

thumbnail
Download:
Fig 1. PRISMA diagram.

Different phases of studies’ selection for conducting qualitative and quantitative analyses.

https://doi.org/10.1371/journal.pone.0163245.g001

The initial search yielded 1042 results, 335 of these were duplicated. Sixty-seven studies were excluded due to (1) not being written in English or (2) consisting of non-original research studies (e.g. reviews). During the initial screening (title and abstract), the main reasons for exclusion were: (1) absence/no implementation of stress protocol; (2) induction of stress (acute or chronic) protocols not considered in this work; (3) stress exposure at prenatal/early life phases; (4) stress implemented as a consequence of drug administration (e.g. dexamethasone); and (5) lack of cognitive assessment. At the end of the initial screening, 148 studies were selected for full-text review. During full-text review, 54 studies were further excluded, mainly due to: (1) absence of learning and memory assessment with validated behavioral tasks; (2) implementation of stress protocols not considered in the current work or absence of implementation of a chronic stress model; (3) use of fear conditioning protocols; (4) non-original research (review articles); (5) absence of control group.

Afterwards, for the selected studies, statistical measures were obtained either from the published paper (n = 15) or through e-mail contact to the corresponding authors (n = 13). Measures for the remaining studies could not be obtained due to lack of response of corresponding authors to email contacts. Thus, experiments in which the necessary measures for conducting the meta-analysis could not be calculated (n = 66), were included only in the systematic review. For the remaining, Cohen’s d (and the associated variance) was calculated as a measure of effect size (and the deviation measure). Cohen’s Kappa revealed a fair/good agreement between raters in the selection of the studies for the systematic review and meta-analyses (Kappa = .413, p < .001[35]).

3.2. Systematic review findings

Among the selected studies, chronic restraint (n = 49) and chronic mild (n = 45) stress protocols were implemented. With respect to learning and memory assessment, MWM was used in 50 experiments, NOR in 16, RAM in eight, and Y-Maze in 14. In five studies, other tasks [Hebb-Williams maze and labyrinth food finding test (n = 2); T-maze (n = 3)] were used. For studies performing more than one task, the results were analyzed individually. The majority of studies included only male animals, with only seven studies reporting findings from female rodents. With respect to animals’ strain, 38 studies used Wistar rats, 32 male Sprague-Dawley and four Long-Evans. In mice studies, seven used C57BL mice, five Kunming mice, four ICR mice, two BALB/C mice, one Laca mice and one study used albino Swiss mice. With respect to physiological changes following stress implementation, the stress group displayed reduction in body weight in 82% of the studies, stressed animals presented decreased sucrose preference in 92% of the studies, and corticosterone levels were augmented in stressed animals in 83% of the studies.

3.2.1. Morris water maze.

Overall, studies revealed that different types of chronic stress produce changes in cognition. Specifically, with respect to the MWM task, the majority of the studies (n = 28) report an absence of significant differences between the latency to reach the platform in the first acquisition day (Table 1). On the other hand, differences seemed to arise in the second, and especially third acquisition day, in which stressed animals displayed significantly more latency to find the hidden platform. Swimming speed was reported in only 12 studies. Of these, no differences were reported in eight studies, lower speed on stressed animals were found in one study, higher swimming speed on stressed animals was reported in three studies.

thumbnail
Download:
Table 1. Description of the studies using the MWM task.

Characteristics: type of stress, duration, sex, age (and/or initial body weight), comparison of performance on acquisition days, probe and reversal parameters and differences in corticosterone (or sucrose preference) and body weight between stressed and control animals.

https://doi.org/10.1371/journal.pone.0163245.t001

Results relative to the probe test were reported in 33 studies. Of these, nine reported no differences between groups in the assessed parameters (time spent and number of crossings over the target quadrant and overall swimming speed). Four studies reported less crossings over the target in the stress group while one study reported the opposite pattern. Seventeen studies reported that stressed animals spent significantly less time in the target quadrant, only one study reported more time spent in the target quadrant by stressed animals, and no differences were found in the remaining seven studies. Swimming speed during the probe trial was reported in four studies with mixed results being reported: stresses animals displayed slower speed in two studies, higher speed in one, and no differences to the control group in stressed animals.

Regarding animals’ performance on the reversal test, stressed animals were found to spent more time in the original quadrant in three studies and fewer crossings in one. With respect to the total time spent in target quadrant, results were less consensual between reports: two studies indicated that stressed rodents spent less time in the target quadrant, and two studies revealed an absence of differences in this parameter.

3.2.2. Novel object recognition.

The performance of control and stressed animals on the NOR task was assessed in 11 studies (Table 2). The majority of studies reported an absence of differences in the total time exploring new objects between stressed and control animals. Only one study revealed a reduced total exploration time in stressed animals. On the other hand, five studies revealed that stressed animals display reduced exploration of new objects (as measured by the difference between novel and familiar objects). Two studies reported no differences between the groups. With respect to the discrimination index (DI, calculated as the difference between time spent exploring novel and familiar objects [36]), five studies reported a significantly reduced DI in stressed animals, while two studies reported no differences between the groups.

thumbnail
Download:
Table 2. Description of the studies using the NOR task.

Characteristics: type of stress, duration sex, age (and/or body weight) and comparison of scores on object recognition and discrimination index between stressed and control animals.

https://doi.org/10.1371/journal.pone.0163245.t002

3.2.3. Y-Maze.

Among the selected studies, 14 experiments were conducted with the YM (Table 3). Of these, two studies did not report comparisons between groups. In the remaining studies, cognitive performance was assessed based on the number of entries in novel arms. No differences between groups were observed in six studies; in four studies, stressed animals had less entries; and in four studies, stressed animals over-performed control animals in four studies.

thumbnail
Download:
Table 3. Description of the studies using the RAM task.

Characteristics: type of stress, duration, sex, age (and/or body weight) and comparison of learning performance between stressed and control animals.

https://doi.org/10.1371/journal.pone.0163245.t003

3.2.4. Radial arm maze.

In the eight studies where the RAM task was used (Table 4), an obvious effect of stress on cognitive performance was noted: with the exception of two studies, stressed animals displayed more errors and less correct choices.

thumbnail
Download:
Table 4. Description of the studies using the Y-M task.

Characteristics: type of stress, duration, sex, age (and/or body weight) and comparison of performance between stressed and control animals.

https://doi.org/10.1371/journal.pone.0163245.t004

3.2.5. Integration of findings.

The comparison between control and stressed animals for the different tasks is summarized on Table 5. The test parameters that yielded the most significant differences between groups were the latency on the acquisition phase in the MWM test (Χ2(2) = 23.3, p < .001), time spent in the target quadrant (Χ2(2) = 17.2, p < .001) and the object recognition in the NOR test (p = .044). In addition, trends for statistical significance were also observed in the speed on the acquisition phase in the MWM test (p = .054) and in the time spent at the old quadrant (p = .054). Of note, stressed animals had reduced performance in all these parameters.

thumbnail
Download:
Table 5. Summary of groups’ comparisons on different MWM phases.

https://doi.org/10.1371/journal.pone.0163245.t005

3.3. Meta-analytic results

3.3.1. Global analysis.

A general cognitive parameter was calculated based on the scores on learning and memory measures assessed in the different tasks, by averaging the computed effect sizes for each reported measure. Significant heterogeneity among studies was verified (Q(27) = 100.8, p < .001) and therefore a Random-Effects model was selected. An overall effect of stress on general cognition was noted (d = -0.75, SE = 0.18, p < .001), such that stressed animals present overall lower cognitive scores, compared with control animals. The overall effect is graphically represented on the Forest Plot (Fig 2). The test for funnel plot asymmetry revealed a significant result (t(26) = -4.03, p < .001).

thumbnail
Download:
Fig 2. Forest plot.

Overall effect of stress influence on general cognition. Individual lines represent each study included in the meta-analysis. The vertical dashed line represents absence of effects between stressed and control animals. The diamond located at the bottom of the figure represents the overall effect. As can be observed, the diamond does not cross the vertical line, which indicates that overall effect is significant.

https://doi.org/10.1371/journal.pone.0163245.g002

3.3.2. Morris water maze.

Omnibus analysis: The parameters assessed during the acquisition phase (latency, distance and swimming speed) and the probe trial (time spent in the target quadrant, number of crossings and swimming speed) were used to estimate a general cognitive effect size for each study. The total variability in the model was significantly affected by heterogeneity (Q(15) = 40.5, p < .001). The Random-Effects model revealed a significant overall effect of stress on general MWM performance (d = -0.32, SE = 0.11, p < .001). A trend for a significant asymmetry was observed between studies included in this analysis (t(14) = -2.11, p = .053).

Regarding the acquisition days on the MWM task, no significant differences were found in the latency to reach the platform on each of the first two days between groups: Day 1: d = 0.11, SE = 0.13, p = .399; Day 2: d = 0.13, SE = 0.13, p = .353. Regarding the third day, following the assessment of heterogeneity, (Q(7) = 16.4, p = .022, accounting for 56.2% of the variability of the model), it was observed a significant overall effect of stress on the latency to reach the platform(d = 0.58, SE = 0.21, p = .007), with stressed animals presenting worse performance.

With respect to the probe trial, a marginally significant effect of heterogeneity was observed (Q(5) = 10.6, p = .061), contributing for 53.7% (I2) of the variance in this model. Consequently, a Random-Effects model was preferred. A significant overall effect of stress was observed (d = -0.46, SE = 0.03, p = 0.029).

3.3.3. Sensitivity analysis.

Leave-one-out (sensitivity) analyses were conducted for each independent meta-analysis. It was observed that the exclusion of a single study did not yield significant changes in overall effects.

3.3.4. Moderator and mediator effects.

To account for the impact of stress implementation on the overall effect, moderator and mediator meta-analyses were conducted. Using random-effects categorical moderator models, it was observed that both CMS (d = -0.67, SE = 0.20, p = .001) and CRS (d = -0.83, SE = 0.28, p = .003) displayed significantly impact on learning and memory. In addition, restricted maximum-likelihood meta-regression analysis revealed that stress duration did not significantly affected the overall effect (R = -0.003, SE = 0.02, p = .841).

4. Discussion

4.1. General

In this work, we conducted a systematic review and meta-analytic procedures to study the effect of chronic stress on cognitive performance in mice and rats. Despite the observed heterogeneity, it can be generally concluded that the implementation of different protocols of chronic stress leads to alterations on cognitive functioning, particularly on the consolidation of learned memories. For instance, in accordance with a previously hypothesized biphasic effect of chronic stress on the central and peripheral nervous system, it could be expected that shorter exposure to stress would be beneficial to the organism, whereas longer stress exposures would lead to detrimental consequences[18]. Although this effect was apparent in the systematic review findings, the results obtained from the meta-regression failed to confirm a significant association between the duration of stress implementation and the overall estimates. Nevertheless, the absence of significant results may stem from the fact that in the majority of studies (64.3%) stress is implemented during 21 or 28 days, leading to a reduced variability between studies. As previously indicated, it is frequently accepted that the impact of chronic stress on cognitive function is dependent on GCs’ overproduction and concomitant hippocampal atrophy [37]. Yet, some studies included in this analysis report impairments on cognitive performance as a consequence of chronic stress co-occurring with normal levels of circulating corticosterone. It might be hypothesized that GCs exert a differential role on cognition, particularly on memory performance. In fact, it has been proposed that GCs have a dissociative impact on memory consolidation and retrieval [38]. Alternatively, it is also possible to hypothesize that the exposure to stress potentially affects cognitive performance, without affecting corticosterone levels. In line with this, it has been hypothesized that stress impairs cognition through a down-regulation of hippocampal glucocorticoid receptors’ levels and production of CA3 dendritic retraction[39]. It is also relevant to highlight that although in this work, we have focused on the effects of stress on HPA axis, this relationship is thought to be bi-directional. Indeed, HPA axis deregulation is known to contribute to the development of psychosomatic and psychiatric conditions, with its hyper-reactivity being itself associated to an inadequate response to stress[40].

4.2 Strengths and Limitations

Meta-analytic studies are characterized by high level of evidence, as they allow the computation of omnibus results from multiple studies, while accounting for the variability between individual works. Thus, one major contribution of this work relies on the estimation of overall effects. We expect that this work may serve as a rigorous means of estimating sample sizes, which will be critical for detecting true positive effects (i.e. to avoid type II errors). Simultaneously, this approach will also limit the maximum number of animals to use, which is in line with the Russell and Burch[41] recommendations expressed in the principles of 3Rs.

Nonetheless, results herein presented should be interpreted with some caution. The systematic review process is prone to criticism. On this, one can argue that the process of selecting studies may be itself biased, due to different factors such as the initial exclusion based only on abstracts’ reading or to the inclusion of studies from the same group of researchers. However, this was based on the widely recommended and most accepted practices for conducting systematic reviews. Another criticism may be related with the exclusion of tasks encompassing aversive learning. Several studies demonstrate that the implementation of stress conducts to an impaired performance in these tasks. Nevertheless, we decided to exclude these tasks with the goal of avoiding the influence of potential confounders.

Furthermore, a major concern raised in this work is related with the reduced number of studies included in the meta-analysis. This was particularly disappointing since a considerable number of studies met the inclusion criteria. However, most of the studies did not report appropriate statistics required for the computation of effect size measures. As a consequence, the meta-analytic calculations were estimated based on a reduced number of studies. This also precluded the appropriate control of covariates of interest, such as animals’ strain and age. As a good practice and following other research areas, research with animal models would benefit from a better data reporting. In particular, a comprehensive description of the appropriate statistics is of critical relevance, as it will allow an aggregation of results from different studies employing similar experimental manipulations. This aspect was also referred in a recent review that focused on the quality of experimental design in the field[42]. Another relevant issue highlighted in our work is related with the presence of publication bias. Although we were not able to test for publication bias in individual parameters due to a reduced number of studies reporting the same outcome, significant asymmetry was found on the global analyses.

In addition, aspects pertaining to the experiment organization, including lack of appropriate randomization or experimenter blinding, raise additional concerns. In particular, randomization was not reported in a considerable part of the studies and blindness was rarely referred. These factors highlight the relevance of improving experimental designs and the current guidelines in the reporting of the experiments with animals as a means to ensure an appropriate level of research evidence.

A further limitation is related with the reduced number of studies with female animals, which precluded the analysis of the moderating effect of sex. This would be of upmost importance, since it has been acknowledged that the effects of stress on anatomical, neuroendocrine and neurochemical variables and on cognitive performance measures varies between sexes[18, 43]. For instance, there is evidence showing that male rats, but not female, show impaired performance in the NOR task after 21 days of chronic stress. These results are also reflected at the neural and endocrine levels, where male rats show significant atrophy of apical dendritic branches of the CA3c pyramidal neurons. In contrast, female rats showed a decreased number of branch points in the basal dendritic area and revealed higher levels of plasma corticosterone both at baseline and during stress implementation[43]. These differences highlight the importance of characterizing the effects of stress, taking into account the sex of the animals. Moreover, based on our findings, it seems evident that more research with females should be undertaken, with the goal of better understanding the neurophysiological mechanisms, and protective factors, of cognitive decline following stress.

Finally, it was noted that structured procedures for the implementation of stress are still missing. As an example, CRS protocols varied between studies concerning duration of the restraint sessions (one to six hours) and extension of the protocol (from 14 to 28 days). Additionally, a recent study from our group demonstrated that methodological differences such as the implementation of stress protocol in the resting (light phase) or activity (dark phase) of the animal can differentially impact the performance on probe test[29]. Also, there is considerable heterogeneity with respect to behavioral assessments. For instance, there is high variability in the number of acquisition days to assess the animals’ performance on the MWM. Consequently, animals will have different training levels from experiment to experiment, which will likely induce alterations in the animals’ performance during the probe trial. Reported parameters are also exceedingly heterogeneous, with different measures being reported across studies, such as swimming speed, latency or distance to the hidden platform. Curiously, some authors reported an average of the assessed parameter during the acquisition days, while others presented these parameters during individual days/blocks/sessions. This also limits the assessment of animals’ learning curve throughout days.

4.3. PROBE–Preferred Reporting Orientations for Behavioral Experiments

In order to overcome the abovementioned limitations, herein we propose a guidance for reporting results in animal research, henceforth termed PROBE (Preferred Reporting Orientations for Behavioral Experiments) (Table 6). In this set of guidelines, we focus on distinct classes of factors that were experienced as crucial in the development of this work. Specifically, this guidance focus on several parameters, including: experimental conditions, biological factors, experimental organization (emphasizing both general aspects and those related to the implementation of stress protocols), experimental design and statistical analysis. As previously mentioned, the rationale behind the selection of these factors relies on our experience in the selection of studies to conduct this work. Overall, these guidelines are aimed to constitute a checklist to be progressively established in the animal research field in order to enhance the quality and accuracy of data reporting. We consider that this will allow an easier communication between different researchers and laboratories, by enabling the understanding of possible methodological differences that may lead to contrasting (and even contradicting) outcomes.

thumbnail
Download:
Table 6. PROBE–Preferred Reporting Orientations for Behavioral Experiments.

https://doi.org/10.1371/journal.pone.0163245.t006

5. Conclusions

Cognitive dysfunction is a hallmark of chronic stress in humans. However, in rodents, divergent findings regarding the effects of chronic stress on cognitive performance have been reported. This raises serious concerns to the translation value of rodent models of chronic stress. Despite this heterogeneity, our meta-analytic work provides solid evidence that indeed rodents mimic this feature of human pathology. As a corollary of this work, we suggest a set of guidelines for adequate reporting of animal results. We expect this to be helpful in facilitating the aggregation of results in the field and potentiating an increased level of research evidence. Taken together, the present work may be a relevant reference for future studies, by potentiating a better research planning and reporting in work involving animal experimentation. This will also potentiate the validity (face, predictive and construct validities) of animal models and their translation value.

Acknowledgments

The authors are thankful to the authors of the included studies in the meta-analysis for the collaboration in providing the asked measures and to Nadine Santos for carefully reading the manuscript.

Author Contributions

  1. Conceptualization: PSM HLA PC NS.
  2. Data curation: PSM PRA PC.
  3. Formal analysis: PSM PC.
  4. Funding acquisition: NS.
  5. Investigation: PSM PRA PC.
  6. Methodology: PSM PRA PC.
  7. Project administration: NS.
  8. Software: PSM PC.
  9. Supervision: NS PC.
  10. Validation: PSM PRA PC NS.
  11. Visualization: HLA NS.
  12. Writing – original draft: PSM PRA PC.
  13. Writing – review & editing: PSM HLA NS PC.

References

  1. 1. Lupien SJ, Maheu F, Tu M, Fiocco A, Schramek TE. The effects of stress and stress hormones on human cognition: Implications for the field of brain and cognition. Brain and cognition. 2007;65(3):209–37. Epub 2007/05/01. pmid:17466428.
  2. 2. McEwen BS. Protective and damaging effects of stress mediators: central role of the brain. Dialogues in clinical neuroscience. 2006;8(4):367. pmid:17290796
  3. 3. Dias-Ferreira E, Sousa JC, Melo I, Morgado P, Mesquita AR, Cerqueira JJ, et al. Chronic stress causes frontostriatal reorganization and affects decision-making. Science (New York, NY). 2009;325(5940):621–5. Epub 2009/08/01. pmid:19644122.
  4. 4. Kalivas PW, Duffy P. Selective activation of dopamine transmission in the shell of the nucleus accumbens by stress. Brain research. 1995;675(1):325–8.
  5. 5. Erb S, Stewart J. A role for the bed nucleus of the stria terminalis, but not the amygdala, in the effects of corticotropin-releasing factor on stress-induced reinstatement of cocaine seeking. Journal of Neuroscience. 1999;19:RC35 (1–6).
  6. 6. Chappell P, Smith M, Kilts C, Bissette G, Ritchie J, Anderson C, et al. Alterations in corticotropin-releasing factor-like immunoreactivity in discrete rat brain regions after acute and chronic stress. The Journal of neuroscience. 1986;6(10):2908–14. pmid:3020187
  7. 7. Kim JJ, Foy MR, Thompson RF. Behavioral stress modifies hippocampal plasticity through N-methyl-D-aspartate receptor activation. Proceedings of the National Academy of Sciences of the United States of America. 1996;93(10):4750–3. Epub 1996/05/14. pmid:8643474; PubMed Central PMCID: PMCPmc39350.
  8. 8. McEwen BS. The neurobiology of stress: from serendipity to clinical relevance. Brain Res. 2000;886(1–2):172–89. Epub 2000/12/20. 11119695. pmid:11119695
  9. 9. Belanoff JK, Kalehzan M, Sund B, Fleming Ficek SK, Schatzberg AF. Cortisol activity and cognitive changes in psychotic major depression. The American journal of psychiatry. 2001;158(10):1612–6. Epub 2001/10/02. pmid:11578992.
  10. 10. Baker KB, Kim JJ. Effects of stress and hippocampal NMDA receptor antagonism on recognition memory in rats. Learning & memory (Cold Spring Harbor, NY). 2002;9(2):58–65. Epub 2002/05/07. pmid:11992016; PubMed Central PMCID: PMCPmc155932.
  11. 11. Sousa N, Almeida OF. Disconnection and reconnection: the morphological basis of (mal)adaptation to stress. Trends in neurosciences. 2012;35(12):742–51. Epub 2012/09/25. pmid:23000140.
  12. 12. Hill MN, Hellemans KG, Verma P, Gorzalka BB, Weinberg J. Neurobiology of chronic mild stress: parallels to major depression. Neuroscience & Biobehavioral Reviews. 2012;36(9):2085–117.
  13. 13. Green MR, McCormick CM. Effects of social instability stress in adolescence on long-term, not short-term, spatial memory performance. Behavioural brain research. 2013;256:165–71. Epub 2013/08/21. pmid:23948213.
  14. 14. Nagata K, Nakashima-Kamimura N, Mikami T, Ohsawa I, Ohta S. Consumption of molecular hydrogen prevents the stress-induced impairments in hippocampus-dependent learning tasks during chronic physical restraint in mice. Neuropsychopharmacology: official publication of the American College of Neuropsychopharmacology. 2009;34(2):501–8. Epub 2008/06/20. pmid:18563058.
  15. 15. Rinwa P, Kumar A. Modulation of nitrergic signalling pathway by American ginseng attenuates chronic unpredictable stress-induced cognitive impairment, neuroinflammation, and biochemical alterations. Naunyn-Schmiedeberg's archives of pharmacology. 2014;387(2):129–41. pmid:24132508
  16. 16. Ghadrdoost B, Vafaei AA, Rashidy-Pour A, Hajisoltani R, Bandegi AR, Motamedi F, et al. Protective effects of saffron extract and its active constituent crocin against oxidative stress and spatial learning and memory deficits induced by chronic stress in rats. European journal of pharmacology. 2011;667(1–3):222–9. Epub 2011/05/28. pmid:21616066.
  17. 17. Walesiuk A, Braszko JJ. Preventive action of Ginkgo biloba in stress- and corticosterone-induced impairment of spatial memory in rats. Phytomedicine: international journal of phytotherapy and phytopharmacology. 2009;16(1):40–6. Epub 2007/05/08. pmid:17482446.
  18. 18. Bowman RE, Beck KD, Luine VN. Chronic stress effects on memory: sex differences in performance and monoaminergic activity. Hormones and behavior. 2003;43(1):48–59. Epub 2003/03/05. pmid:12614634.
  19. 19. Olton DS, Samuelson RJ. Remembrance of places passed: Spatial memory in rats. Journal of Experimental Psychology: Animal Behavior Processes. 1976;2(2): 97–116.
  20. 20. Morris R, Garrud P, Rawlins J, O'Keefe J. Place navigation impaired in rats with hippocampal lesions. Nature. 1982;297(5868):681–3. pmid:7088155
  21. 21. Ennaceur A, Delacour J. A new one-trial test for neurobiological studies of memory in rats. 1: Behavioral data. Behavioural brain research. 1988;31(1):47–59. Epub 1988/11/01. pmid:3228475.
  22. 22. Conrad CD, Galea LA, Kuroda Y, McEwen BS. Chronic stress impairs rat spatial memory on the Y maze, and this effect is blocked by tianeptine treatment. Behavioral neuroscience. 1996;110(6):1321. pmid:8986335
  23. 23. Sousa N, Almeida O, Wotjak C. A hitchhiker's guide to behavioral analysis in laboratory rodents. Genes, Brain and Behavior. 2006;5(s2):5–24.
  24. 24. Paul C- M, Magda G, Abel S. Spatial memory: Theoretical basis and comparative review on experimental methods in rodents. Behavioural brain research. 2009;203(2):151–64. pmid:19467271
  25. 25. Ricon T, Toth E, Leshem M, Braun K, Richter-Levin G. Unpredictable chronic stress in juvenile or adult rats has opposite effects, respectively, promoting and impairing resilience. Stress. 2012;15(1):11–20. pmid:21682654
  26. 26. Ter Horst J, De Kloet E, Schächinger H, Oitzl M. Relevance of stress and female sex hormones for emotion and cognition. Cellular and molecular neurobiology. 2012;32(5):725–35. pmid:22113371
  27. 27. Parihar VK, Hattiangady B, Kuruba R, Shuai B, Shetty AK. Predictable chronic mild stress improves mood, hippocampal neurogenesis and memory. Molecular psychiatry. 2011;16(2):171–83. Epub 2009/12/17. pmid:20010892; PubMed Central PMCID: PMCPmc2891880.
  28. 28. Perez-Cruz C, Simon M, Flügge G, Fuchs E, Czéh B. Diurnal rhythm and stress regulate dendritic architecture and spine density of pyramidal neurons in the rat infralimbic cortex. Behavioural brain research. 2009;205(2):406–13. pmid:19643147
  29. 29. Aslani S, Harb MR, Costa PS, Almeida OF, Sousa N, Palha JA. Day and night: diurnal phase influences the response to chronic mild stress. Frontiers in behavioral neuroscience. 2014;8:82. Epub 2014/03/29. pmid:24672446; PubMed Central PMCID: PMCPmc3954061.
  30. 30. Moher D, Liberati A, Tetzlaff J, Altman DG. Preferred reporting items for systematic reviews and meta-analyses: the PRISMA statement. Annals of internal medicine. 2009;151(4):264–9, w64. Epub 2009/07/23. pmid:19622511.
  31. 31. Kim JJ, Jung MW. Neural circuits and mechanisms involved in Pavlovian fear conditioning: a critical review. Neuroscience & Biobehavioral Reviews. 2006;30(2):188–202.
  32. 32. Nagel JA, Kemble ED. Effects of amygdaloid lesions on the performance of rats in four passive avoidance tasks. Physiology & behavior. 1976;17(2):245–50.
  33. 33. Higgins JPT, Green S, Collaboration C. Cochrane Handbook for Systematic Reviews of Interventions: Wiley-Blackwell; 2008.
    • 34. Viechtbauer W. Conducting meta-analyses in R with the metafor package. Journal of Statistical Software. 2010;36(3):1–48.
    • 35. Fleiss JL, Levin B, Paik MC. Statistical Methods for Rates and Proportions: Wiley; 2013.
      • 36. Antunes M, Biala G. The novel object recognition memory: neurobiology, test procedure, and its modifications. Cognitive processing. 2012;13(2):93–110. pmid:22160349
      • 37. Kim JJ, Diamond DM. The stressed hippocampus, synaptic plasticity and lost memories. Nature Reviews Neuroscience. 2002;3(6):453–62. pmid:12042880
      • 38. Roozendaal B. Stress and memory: opposing effects of glucocorticoids on memory consolidation and memory retrieval. Neurobiology of learning and memory. 2002;78(3):578–95. pmid:12559837
      • 39. Wright RL, Lightner EN, Harman JS, Meijer OC, Conrad CD. Attenuating corticosterone levels on the day of memory assessment prevents chronic stress-induced impairments in spatial memory. The European journal of neuroscience. 2006;24(2):595–605. Epub 2006/08/15. pmid:16903861; PubMed Central PMCID: PMCPmc1550977.
      • 40. Kirschbaum C, Kudielka BM, Gaab J, Schommer NC, Hellhammer DH. Impact of gender, menstrual cycle phase, and oral contraceptives on the activity of the hypothalamus-pituitary-adrenal axis. Psychosomatic medicine. 1999;61(2):154–62. Epub 1999/04/16. pmid:10204967.
      • 41. Russell WMS, Burch RL. The principles of humane experimental technique: Methuen; 1959.
        • 42. Kilkenny C, Parsons N, Kadyszewski E, Festing MFW, Cuthill IC, Fry D, et al. Survey of the Quality of Experimental Design, Statistical Analysis and Reporting of Research Using Animals. PLOS ONE. 2009;4(11):e7824. pmid:19956596
        • 43. Galea LA, McEwen BS, Tanapat P, Deak T, Spencer RL, Dhabhar FS. Sex differences in dendritic atrophy of CA3 pyramidal neurons in response to chronic restraint stress. Neuroscience. 1997;81(3):689–97. Epub 1997/10/08. pmid:9316021.
        • 44. Bessa JM, Mesquita AR, Oliveira M, Pego JM, Cerqueira JJ, Palha JA, et al. A trans-dimensional approach to the behavioral aspects of depression. Frontiers in behavioral neuroscience. 2009;3:1. Epub 2009/02/06. pmid:19194528; PubMed Central PMCID: PMCPmc2634526.
        • 45. de Vasconcellos AP, Zugno AI, Dos Santos AH, Nietto FB, Crema LM, Goncalves M, et al. Na+,K(+)-ATPase activity is reduced in hippocampus of rats submitted to an experimental model of depression: effect of chronic lithium treatment and possible involvement in learning deficits. Neurobiol Learn Mem. 2005;84(2):102–10. Epub 2005/06/18. pmid:15961330.
        • 46. First M, Gil-Ad I, Taler M, Tarasenko I, Novak N, Weizman A. The effects of fluoxetine treatment in a chronic mild stress rat model on depression-related behavior, brain neurotrophins and ERK expression. Journal of molecular neuroscience: MN. 2011;45(2):246–55. Epub 2011/04/12. pmid:21479508.
        • 47. Kasar M, Mengi M, Yildirim EA, Yurdakos E. Different effects of tianeptine pretreatment in rats exposed to acute stress and repeated severe stress. Methods and findings in experimental and clinical pharmacology. 2009;31(3):157–63. Epub 2009/06/19. pmid:19536358.
        • 48. Quan M, Zheng C, Zhang N, Han D, Tian Y, Zhang T, et al. Impairments of behavior, information flow between thalamus and cortex, and prefrontal cortical synaptic plasticity in an animal model of depression. Brain research bulletin. 2011;85(3–4):109–16. Epub 2011/03/15. pmid:21396989.
        • 49. Sandi C, Touyarot K. Mid-life stress and cognitive deficits during early aging in rats: individual differences and hippocampal correlates. Neurobiology of aging. 2006;27(1):128–40. Epub 2005/11/22. pmid:16298248.
        • 50. Sun CY, Qi SS, Lou XF, Sun SH, Wang X, Dai KY, et al. Changes of learning, memory and levels of CaMKII, CaM mRNA, CREB mRNA in the hippocampus of chronic multiple-stressed rats. Chinese medical journal. 2006;119(2):140–7. Epub 2006/02/04. pmid:16454996.
        • 51. Tagliari B, Scherer EB, Machado FR, Ferreira AG, Dalmaz C, Wyse AT. Antioxidants prevent memory deficits provoked by chronic variable stress in rats. Neurochemical research. 2011;36(12):2373–80. Epub 2011/08/09. pmid:21822921.
        • 52. Touyarot K, Venero C, Sandi C. Spatial learning impairment induced by chronic stress is related to individual differences in novelty reactivity: search for neurobiological correlates. Psychoneuroendocrinology. 2004;29(2):290–305. Epub 2003/11/08. pmid:14604607.
        • 53. Cunningham JI, Raudensky J, Tonkiss J, Yamamoto BK. MDMA pretreatment leads to mild chronic unpredictable stress-induced impairments in spatial learning. Behav Neurosci. 2009;123(5):1076–84. Epub 2009/10/15. 10.1037/a0016716. pmid:19824774; PubMed Central PMCID: PMCPmc2786777.
        • 54. First M, Gil-Ad I, Taler M, Tarasenko I, Novak N, Weizman A. The effects of reboxetine treatment on depression-like behavior, brain neurotrophins, and ERK expression in rats exposed to chronic mild stress. Journal of molecular neuroscience: MN. 2013;50(1):88–97. Epub 2012/09/13. pmid:22968760.
        • 55. Gouirand AM, Matuszewich L. The effects of chronic unpredictable stress on male rats in the water maze. Physiology & behavior. 2005;86(1):21–31.
        • 56. Isgor C, Kabbaj M, Akil H, Watson SJ. Delayed effects of chronic variable stress during peripubertal-juvenile period on hippocampal morphology and on cognitive and stress axis functions in rats. Hippocampus. 2004;14(5):636–48. Epub 2004/08/11. pmid:15301440.
        • 57. Kallarackal AJ, Kvarta MD, Cammarata E, Jaberi L, Cai X, Bailey AM, et al. Chronic Stress Induces a Selective Decrease in AMPA Receptor-Mediated Synaptic Excitation at Hippocampal Temporoammonic-CA1 Synapses. The Journal of Neuroscience. 2013;33(40):15669–74. pmid:24089474
        • 58. Li H, Zhang L, Huang Q. Differential expression of mitogen-activated protein kinase signaling pathway in the hippocampus of rats exposed to chronic unpredictable stress. Behavioural brain research. 2009;205(1):32–7. Epub 2009/07/07. pmid:19576250.
        • 59. Xi G, Hui J, Zhang Z, Liu S, Zhang X, Teng G, et al. Learning and Memory Alterations Are Associated with Hippocampal N-acetylaspartate in a Rat Model of Depression as Measured by 1H-MRS. PLOS ONE. 2011;6(12):e28686. pmid:22194886
        • 60. Zheng H, Liu Y, Li W, Yang B, Chen D, Wang X, et al. Beneficial effects of exercise and its molecular mechanisms on depression in rats. Behavioural brain research. 2006;168(1):47–55. pmid:16290283
        • 61. Hill MN, Patel S, Carrier EJ, Rademacher DJ, Ormerod BK, Hillard CJ, et al. Downregulation of endocannabinoid signaling in the hippocampus following chronic unpredictable stress. Neuropsychopharmacology: official publication of the American College of Neuropsychopharmacology. 2005;30(3):508–15. Epub 2004/11/05. pmid:15525997.
        • 62. Bian Y, Pan Z, Hou Z, Huang C, Li W, Zhao B. Learning, memory, and glial cell changes following recovery from chronic unpredictable stress. Brain research bulletin. 2012;88(5):471–6. pmid:22537595
        • 63. Liao MJ, Lin LF, Zhou X, Zhou XW, Xu X, Cheng X, et al. Daphnetin prevents chronic unpredictable stress-induced cognitive deficits. Fundamental & clinical pharmacology. 2013;27(5):510–6. Epub 2012/06/22. pmid:22715971.
        • 64. Zhang X, Dong YL, Yang N, Liu YY, Gao RF, Zuo PP. Effects of ning shen ling granule and dehydroepiandrosterone on cognitive function in mice undergoing chronic mild stress. Chinese journal of integrative medicine. 2007;13(1):46–9. Epub 2007/06/21. pmid:17578318.
        • 65. Song L, Che W, Min-Wei W, Murakami Y, Matsumoto K. Impairment of the spatial learning and memory induced by learned helplessness and chronic mild stress. Pharmacology, biochemistry, and behavior. 2006;83(2):186–93. Epub 2006/03/08. pmid:16519925.
        • 66. Bisaz R, Schachner M, Sandi C. Causal evidence for the involvement of the neural cell adhesion molecule, NCAM, in chronic stress-induced cognitive impairments. Hippocampus. 2011;21(1):56–71. Epub 2009/11/19. pmid:19921700.
        • 67. Cuadrado-Tejedor M, Ricobaraza A, Del Rio J, Frechilla D, Franco R, Perez-Mediavilla A, et al. Chronic mild stress in mice promotes cognitive impairment and CDK5-dependent tau hyperphosphorylation. Behavioural brain research. 2011;220(2):338–43. Epub 2011/01/18. pmid:21238494.
        • 68. Liu Y, Yang N, Zuo P. cDNA microarray analysis of gene expression in the cerebral cortex and hippocampus of BALB/c mice subjected to chronic mild stress. Cell Mol Neurobiol. 2010;30(7):1035–47. Epub 2010/06/10. pmid:20532976.
        • 69. Abidin I, Yargicoglu P, Agar A, Gumuslu S, Aydin S, Ozturk O, et al. The effect of chronic restraint stress on spatial learning and memory: relation to oxidant stress. The International journal of neuroscience. 2004;114(5):683–99. Epub 2004/06/19. pmid:15204074.
        • 70. Kitraki E, Kremmyda O, Youlatos D, Alexis M, Kittas C. Spatial performance and corticosteroid receptor status in the 21-day restraint stress paradigm. Annals of the New York Academy of Sciences. 2004;1018:323–7. Epub 2004/07/09. pmid:15240385.
        • 71. Kumar RS, Narayanan SN, Nayak S. Ascorbic acid protects against restraint stress-induced memory deficits in Wistar rats. Clinics (Sao Paulo, Brazil). 2009;64(12):1211–7. Epub 2009/12/29. pmid:20037710; PubMed Central PMCID: PMCPmc2797591.
        • 72. Sandi C, Davies HA, Cordero MI, Rodriguez JJ, Popov VI, Stewart MG. Rapid reversal of stress induced loss of synapses in CA3 of rat hippocampus following water maze training. The European journal of neuroscience. 2003;17(11):2447–56. Epub 2003/06/20. pmid:12814376.
        • 73. Trofimiuk E, Braszko JJ. Concomitant docosahexaenoic acid administration ameliorates stress-induced cognitive impairment in rats. Physiol Behav. 2013;118:171–7. Epub 2013/05/16. pmid:23672853.
        • 74. Walesiuk A, Trofimiuk E, Braszko JJ. Gingko biloba extract diminishes stress-induced memory deficits in rats. Pharmacological reports: PR. 2005;57(2):176–87. Epub 2005/05/12. pmid:15886416.
        • 75. Wattanathorn J, Tong-un T, Muchimapura S, Wannanon P, Sripanidkulchai B, Phachonpai W. Anti-stress effects of kaempferia parviflora in immobilization subjected rats. American Journal of Pharmacology and Toxicology. 2013;8(1):31–8.
        • 76. Meng Z-Z, Chen J-X, Jiang Y-M, Zhang H-T. Effect of Xiaoyaosan Decoction on Learning and Memory Deficit in Rats Induced by Chronic Immobilization Stress. Evidence-Based Complementary and Alternative Medicine. 2013;2013:8.
        • 77. Wang YT, Tan QR, Sun LL, Cao J, Dou KF, Xia B, et al. Possible therapeutic effect of a Traditional Chinese Medicine, Sinisan, on chronic restraint stress related disorders. Neuroscience letters. 2009;449(3):215–9. Epub 2008/11/15. pmid:19007859.
        • 78. Wright RL, Conrad CD. Enriched environment prevents chronic stress-induced spatial learning and memory deficits. Behavioural brain research. 2008;187(1):41–7. Epub 2007/10/02. pmid:17904657; PubMed Central PMCID: PMCPmc2629380.
        • 79. Xu Y, Lin D, Li S, Li G, Shyamala SG, Barish PA, et al. Curcumin reverses impaired cognition and neuronal plasticity induced by chronic stress. Neuropharmacology. 2009;57(4):463–71. Epub 2009/06/23. pmid:19540859.
        • 80. Radecki DT, Brown LM, Martinez J, Teyler TJ. BDNF protects against stress-induced impairments in spatial learning and memory and LTP. Hippocampus. 2005;15(2):246–53. Epub 2004/10/12. pmid:15476265.
        • 81. Liu Y, Zhuang X, Gou L, Ling X, Tian X, Liu L, et al. Protective effects of nizofenone administration on the cognitive impairments induced by chronic restraint stress in mice. Pharmacology, biochemistry, and behavior. 2013;103(3):474–80. Epub 2012/10/03. pmid:23026061.
        • 82. Tian X, Sun L, Gou L, Ling X, Feng Y, Wang L, et al. Protective effect of l-theanine on chronic restraint stress-induced cognitive impairments in mice. Brain Res. 2013;1503:24–32. Epub 2013/02/12. pmid:23395732.
        • 83. Muto J, Hosung L, Uwaya A, Isami F, Ohno M, Mikami T. Morinda citrifolia fruit reduces stress-induced impairment of cognitive function accompanied by vasculature improvement in mice. Physiol Behav. 2010;101(2):211–7. Epub 2010/04/27. pmid:20416332.
        • 84. Delgado-Morales R, del Rio E, Gomez-Roman A, Bisagno V, Nadal R, de Felipe C, et al. Adrenocortical and behavioural response to chronic restraint stress in neurokinin-1 receptor knockout mice. Physiol Behav. 2012;105(3):669–75. Epub 2011/10/25. pmid:22019828.
        • 85. Pawlak R, Rao BS, Melchor JP, Chattarji S, McEwen B, Strickland S. Tissue plasminogen activator and plasminogen mediate stress-induced decline of neuronal and cognitive functions in the mouse hippocampus. Proceedings of the National Academy of Sciences of the United States of America. 2005;102(50):18201–6. Epub 2005/12/07. pmid:16330749; PubMed Central PMCID: PMCPmc1312427.
        • 86. Briones A, Gagno S, Martisova E, Dobarro M, Aisa B, Solas M, et al. Stress-induced anhedonia is associated with an increase in Alzheimer's disease-related markers. British journal of pharmacology. 2012;165(4):897–907. Epub 2011/07/30. pmid:21797840; PubMed Central PMCID: PMCPmc3312487.
        • 87. Llorente R, Miguel-Blanco C, Aisa B, Lachize S, Borcel E, Meijer OC, et al. Long term sex-dependent psychoneuroendocrine effects of maternal deprivation and juvenile unpredictable stress in rats. Journal of neuroendocrinology. 2011;23(4):329–44. Epub 2011/01/12. pmid:21219484.
        • 88. Elizalde N, Gil-Bea FJ, Ramirez MJ, Aisa B, Lasheras B, Del Rio J, et al. Long-lasting behavioral effects and recognition memory deficit induced by chronic mild stress in mice: effect of antidepressant treatment. Psychopharmacology. 2008;199(1):1–14. Epub 2008/05/13. pmid:18470507.
        • 89. Solas M, Aisa B, Tordera RM, Mugueta MC, Ramirez MJ. Stress contributes to the development of central insulin resistance during aging: implications for Alzheimer's disease. Biochimica et biophysica acta. 2013;1832(12):2332–9. Epub 2013/10/05. pmid:24090692.
        • 90. Balk Rde S, Silva MH, Bridi JC, Carvalho NR, Portella Rde L, Dobrachinski F, et al. Effect of repeated restraint stress and clomipramine on Na+/K+-ATPase activity and behavior in rats. International journal of developmental neuroscience: the official journal of the International Society for Developmental Neuroscience. 2011;29(8):909–16. Epub 2011/07/19. pmid:21762772.
        • 91. Braszko JJ, Wincewicz D, Jakubow P. Candesartan prevents impairment of recall caused by repeated stress in rats. Psychopharmacology. 2013;225(2):421–8. Epub 2012/08/15. pmid:22890474; PubMed Central PMCID: PMCPmc3537078.
        • 92. Trofimiuk E, Braszko JJ. Single dose of H3 receptor antagonist—ciproxifan—abolishes negative effects of chronic stress on cognitive processes in rats. Psychopharmacology. 2014;231(1):209–19. Epub 2013/08/27. pmid:23975035.
        • 93. Abush H, Akirav I. Cannabinoids ameliorate impairments induced by chronic stress to synaptic plasticity and short-term memory. Neuropsychopharmacology: official publication of the American College of Neuropsychopharmacology. 2013;38(8):1521–34. Epub 2013/02/22. pmid:23426383; PubMed Central PMCID: PMCPmc3682147.
        • 94. Bowman RE, Kelly R. Chronically stressed female rats show increased anxiety but no behavioral alterations in object recognition or placement memory: a preliminary examination. Stress. 2012;15(5):524–32. Epub 2011/12/16. pmid:22168672.
        • 95. Gomez JL, Lewis MJ, Sebastian V, Serrano P, Luine VN. Alcohol administration blocks stress-induced impairments in memory and anxiety, and alters hippocampal neurotransmitter receptor expression in male rats. Hormones and behavior. 2013;63(4):659–66. Epub 2013/02/05. pmid:23376488; PubMed Central PMCID: PMCPmc3646638.
        • 96. Noorafshan A, Abdollahifar MA, Karbalay-Doust S, Asadi-Golshan R, Rashidian-Rashidabadi A. Protective effects of curcumin and sertraline on the behavioral changes in chronic variable stress-induced rats. Experimental neurobiology. 2013;22(2):96–106. Epub 2013/07/09. pmid:23833558; PubMed Central PMCID: PMCPmc3699679.
        • 97. Srikumar BN, Raju TR, Shankaranarayana Rao BS. The involvement of cholinergic and noradrenergic systems in behavioral recovery following oxotremorine treatment to chronically stressed rats. Neuroscience. 2006;143(3):679–88. Epub 2006/09/30. pmid:17008021.
        • 98. Veena J, Srikumar BN, Mahati K, Bhagya V, Raju TR, Shankaranarayana Rao BS. Enriched environment restores hippocampal cell proliferation and ameliorates cognitive deficits in chronically stressed rats. Journal of neuroscience research. 2009;87(4):831–43. Epub 2008/11/14. pmid:19006089.
        • 99. Hutchinson KM, McLaughlin KJ, Wright RL, Bryce Ortiz J, Anouti DP, Mika A, et al. Environmental enrichment protects against the effects of chronic stress on cognitive and morphological measures of hippocampal integrity. Neurobiol Learn Mem. 2012;97(2):250–60. Epub 2012/01/24. pmid:22266288.
        • 100. Mika A, Mazur GJ, Hoffman AN, Talboom JS, Bimonte-Nelson HA, Sanabria F, et al. Chronic stress impairs prefrontal cortex-dependent response inhibition and spatial working memory. Behav Neurosci. 2012;126(5):605–19. Epub 2012/08/22. pmid:22905921; PubMed Central PMCID: PMCPmc3463780.
        • 101. Henningsen K, Andreasen JT, Bouzinova EV, Jayatissa MN, Jensen MS, Redrobe JP, et al. Cognitive deficits in the rat chronic mild stress model for depression: relation to anhedonic-like responses. Behavioural brain research. 2009;198(1):136–41. Epub 2008/11/29. pmid:19038290.
        • 102. Palumbo ML, Canzobre MC, Pascuan CG, Rios H, Wald M, Genaro AM. Stress induced cognitive deficit is differentially modulated in BALB/c and C57Bl/6 mice: correlation with Th1/Th2 balance after stress exposure. Journal of neuroimmunology. 2010;218(1–2):12–20. Epub 2009/11/28. pmid:19942299.
        • 103. Bellani R, Luecken LJ, Conrad CD. Peripubertal anxiety profile can predict predisposition to spatial memory impairments following chronic stress. Behavioural brain research. 2006;166(2):263–70. Epub 2005/10/11. pmid:16214234.
        • 104. Conrad CD, Grote KA, Hobbs RJ, Ferayorni A. Sex differences in spatial and non-spatial Y-maze performance after chronic stress. Neurobiol Learn Mem. 2003;79(1):32–40. Epub 2002/12/17. pmid:12482677.
        • 105. McLaughlin KJ, Gomez JL, Baran SE, Conrad CD. The effects of chronic stress on hippocampal morphology and function: an evaluation of chronic restraint paradigms. Brain Res. 2007;1161:56–64. Epub 2007/07/03. pmid:17603026; PubMed Central PMCID: PMCPmc2667378.
        • 106. Wright RL, Conrad CD. Chronic stress leaves novelty-seeking behavior intact while impairing spatial recognition memory in the Y-maze. Stress. 2005;8(2):151–4. Epub 2005/07/16. pmid:16019606; PubMed Central PMCID: PMCPmc1380302.
        • 107. Kleen JK, Sitomer MT, Killeen PR, Conrad CD. Chronic stress impairs spatial memory and motivation for reward without disrupting motor ability and motivation to explore. Behav Neurosci. 2006;120(4):842–51. Epub 2006/08/09. pmid:16893290; PubMed Central PMCID: PMCPmc1578508.
        • 108. Chen Y, Mao Y, Zhou D, Hu X, Wang J, Ma Y. Environmental enrichment and chronic restraint stress in ICR mice: effects on prepulse inhibition of startle and Y-maze spatial recognition memory. Behavioural brain research. 2010;212(1):49–55. Epub 2010/04/03. pmid:20359501.