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Post-learning stress enhances long-term memory and differentially influences memory in females depending on menstrual stage
Acta Psychologica 160 (2015) 127–133
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Post-learning stress enhances long-term memory and differentially influences memory in females depending on menstrual stage Phillip R. Zoladz a,⁎, David M. Peters a, Chelsea E. Cadle a, Andrea E. Kalchik a, Rachael L. Aufdenkampe a, Alison M. Dailey a, Callie M. Brown a, Amanda R. Scharf a, McKenna B. Earley a, Courtney L. Knippen a, Boyd R. Rorabaugh b a b
Department of Psychology, Sociology, & Criminal Justice, Ohio Northern University, Ada, OH 45810, USA Department of Pharmaceutical & Biomedical Sciences, Raabe College of Pharmacy, Ohio Northern University, Ada, OH 45810, USA
a r t i c l e
i n f o
Article history: Received 6 June 2015 Received in revised form 17 July 2015 Accepted 20 July 2015 Available online xxxx Keywords: Stress Learning Memory Sex Menstrual Emotion
a b s t r a c t Most work has shown that post-learning stress enhances long-term memory; however, there have been recent inconsistencies in this literature. The purpose of the present study was to examine further the effects of post-learning stress on long-term memory and to explore any sex differences that may exist. Male and female participants learned a list of 42 words that varied in emotional valence and arousal level. Following encoding, participants completed a free recall assessment and then submerged their hand into a bath of ice cold (stress) or lukewarm (no stress) water for 3 min. The next day, participants were given free recall and recognition tests. Stressed participants recalled more words than non-stressed participants 24 h after learning. Stress also enhanced female participants' recall of arousing words when they were in the follicular, but not luteal, phase. These findings replicate previous work examining post-learning stress effects on memory and implicate the involvement of sex-related hormones in such effects. © 2015 Elsevier B.V. All rights reserved.
1. Introduction Stress exerts powerful effects on learning and memory, and understanding these effects may lend insight into cognitive symptoms associated with stress-related psychological disorders, such as post-traumatic stress disorder (PTSD) and major depression. Researchers have frequently dichotomized stress-memory interactions into the effects of stress on consolidation versus the effects of stress on retrieval, as these two phases of memory processing seem to be differentially influenced by stress (Roozendaal, McEwen, & Chattarji, 2009). Indeed, most research has reported that stress enhances consolidation and impairs retrieval, despite similar mechanisms, such as corticosteroid and noradrenergic interactions in the amygdala, underlying both (Beckner, Tucker, Delville, & Mohr, 2006; Cahill, Gorski, & Le, 2003; Felmingham, Tran, Fong, & Bryant, 2012; Nielson, Yee, & Erickson, 2005; Preuss & Wolf, 2009; Roozendaal, 2003). More recently, researchers have discovered that the effects of stress on consolidation depend on whether the stress is administered before or after learning, as well as the temporal proximity of the stress to learning. For instance, pre-learning stress that is experienced in the context of learning (i.e., temporally associated with learning) often enhances ⁎ Corresponding author at: Ohio Northern University, Department of Psychology, Sociology, & Criminal Justice, 525 S. Main St. Hill 013, Ada, OH 45810, USA. E-mail address: [email protected] (P.R. Zoladz).
http://dx.doi.org/10.1016/j.actpsy.2015.07.008 0001-6918/© 2015 Elsevier B.V. All rights reserved.
long-term memory (Diamond, Campbell, Park, Halonen, & Zoladz, 2007; Schwabe, Bohringer, Chatterjee, & Schachinger, 2008; Zoladz et al., 2011; Zoladz et al., 2014), while pre-learning stress that is experienced outside the context of learning (i.e., temporally separated from learning) often impairs long-term memory (Zoladz, Clark, et al., 2011; Zoladz et al., 2013). Post-learning stress, on the other hand, almost unequivocally enhances long-term memory (Cahill et al., 2003; Felmingham, Tran, Fong, & Bryant, 2012; Nielson & Powless, 2007; Nielson et al., 2005; Preuss & Wolf, 2009); however, such enhancement declines as the stress is temporally removed from the learning experience (Nielson & Powless, 2007). Although the effects of stress on learning and memory could be perceived as adaptive when memory is enhanced and maladaptive when memory is impaired, we would contend that all effects of stress on learning and memory serve an adaptive role (Zoladz, Park, & Diamond, 2011). When stress is experienced in the context of learning, long-term memory is enhanced because, hypothetically, the stress experience signals the brain that what is occurring is important to remember, which results in a “Now Print” mechanism (Livingston, 1967), or emotional tag, that enhances memory formation. This enables the individual to remember potentially life-threatening aspects related to the stressor which could aid survival later in life. In support of this reasoning, pre- and post-learning stress often enhances memory for emotional information, while impairing or having no effect on memory for neutral information (Jelicic, Geraerts, Merckelbach, & Guerrieri, 2004;
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Payne et al., 2007; Zoladz, Clark, et al., 2011). When stress is experienced outside the context of learning, long-term memory is impaired because, hypothetically, the brain, and in particular the hippocampus, is focused on storing the memory of the stress experience, thereby rendering new information storage improbable. Though simplistic, this approach to how stress affects memory consolidation emphasizes the adaptive nature of the stress response and is supported by behavioral, molecular and electrophysiological studies in humans and non-human animals. Though many studies have reported that post-learning stress enhances consolidation, some work has presented conflicting data suggesting that such stress impairs long-term memory. For instance, Trammell and Clore (2014) recently reported that post-learning cold pressor stress (1–3 min duration) impaired long-term (48-h) word and picture memory in participants. However, in the studies, all participants exhibited relatively poor long-term memory (recalling b 7% of words studied and only ~ 10% of pictures studied) and, in two of the three experiments, the difference in recall between stressed and nonstressed participants was only approaching significance (p's = 0.06 and 0.07). Additionally, the investigators examined 48-h memory, as opposed to the more commonly used 24-h time point. Thus, the purpose of the present study was to examine further the relationship between post-learning stress and long-term (24-h) memory and to assess how manipulating the emotional valence and arousal level of the learned information could influence such effects. We hypothesized that post-learning stress would facilitate long-term memory, perhaps more strongly for emotionally arousing information. Moreover, because sex differences are prevalent in the stress-memory literature, we also examined whether post-learning stress differentially affected longterm memory in males and females. Some previous work has reported greater memory-enhancing effects of stress in females, relative to males (e.g., Felmingham, Tran, Fong, & Bryant, 2012; Zoladz et al., 2014); thus, we predicted that females may be more susceptible to post-learning stress effects on long-term memory. We also ran exploratory analyses on female data to assess whether menstrual cycle stage interacted with stress effects on memory.
2.2.1. Word presentation Participants were presented with a list of 42 words, which were selected from the Affective Norms for English Words (Bradley & Lang, 1999). Based on standardized valence and arousal ratings, we chose 14 neutral (7 arousing, 7 non-arousing), 14 positive (7 arousing, 7 non-arousing) and 14 negative (7 arousing, 7 non-arousing) words, which, across emotional valence and arousal categories, were balanced for word length and word frequency. As per the methods employed by Schwabe et al. (2008) and previous work from our own laboratory (Zoladz, Clark, et al., 2011; Zoladz et al., 2013, 2014), participants were instructed to read each word aloud and rate its emotional valence on a scale from −4 (very negative) to +4 (very positive) and its arousal level on a scale of 0 (not arousing) to 8 (very highly arousing) on a sheet of paper containing the list of words and the aid of self-assessment manikins. These manipulations were performed to promote encoding of the words, and they allowed us to analyze the final memory data based on participants' own ratings of the words. According to the Affective Norms for English Words (Bradley & Lang, 1999), the mean (± SEM) valence and arousal ratings for the words that made up the list were as follows: positive arousing words (e.g., kiss): valence = 7.79 ± 0.12, arousal = 6.62 ± 0.25; positive non-arousing words (e.g., cozy): valence = 7.50 ± 0.12, arousal = 3.46 ± 0.16; negative arousing words (e.g., poison): valence = 2.21 ± 0.16, arousal = 6.56 ± 0.27; negative non-arousing words (e.g., unhappy): valence = 2.40 ± 0.20, arousal = 3.89 ± 0.19; neutral arousing words (e.g., lightning): valence = 4.93 ± 0.27, arousal = 6.26 ± 0.20; neutral non-arousing words (e.g., poster): valence = 4.90 ± 0.14, arousal = 3.40 ± 0.13. 2.2.2. Immediate memory testing Immediately following word list encoding, participants were given 5 min to write down as many words as they could remember from the list of words they just studied. This immediate free recall test was performed to verify that there were no group differences regarding short-term memory performance and to avoid a potential floor effect during long-term memory assessment (e.g., Zoladz, Clark, et al., 2011).
2. Materials and method 2.1. Participants Fifty-two healthy men and naturally cycling women (27 men, 25 women; age: M = 20.30, SD = 1.17) from Ohio Northern University volunteered to participate in the experiment. Individuals were excluded from participating if they met any of the following conditions: diagnosis of Raynaud's or peripheral vascular disease; presence of skin diseases, such as psoriasis, eczema or scleroderma; history of syncope or vasovagal response to stress; history of severe head injury; current treatment with psychotropic medications, narcotics, beta-blockers, steroids or any other medication that was deemed to significantly affect central nervous or endocrine system function; mental or substance use disorder; regular nightshift work. Participants were asked to refrain from using recreational drugs (e.g., marijuana) for 3 days prior to the experimental sessions; to refrain from drinking alcohol or exercising extensively for 24 h prior to the experimental sessions; and, to refrain from eating or drinking anything but water for 2 h prior to the experimental sessions. Upon arrival at the laboratory, participants were reminded of the exclusion criteria and study restrictions and verbally affirmed that they had adhered to the requirements. Participants were awarded class credit upon completion of the study. All of the methods for the experiment were approved by the Institutional Review Board at Ohio Northern University. 2.2. Experimental procedures To control for diurnal variations in cortisol levels, all testing was carried out between 1200 and 1800 h.
2.2.3. Cold pressor test (CPT) Following immediate memory testing, participants were asked to submerge their non-dominant hand, up to and including the wrist, in a bath of water for 3 min. Those participants who had been randomly assigned to the stress condition (N = 29; 17 males, 12 females) placed their hand in a bath of ice cold (0–2 °C) water, while participants who had been randomly assigned to the control condition (N = 23; 10 males, 13 females) placed their hand in a bath of warm (35–37 °C) water. The water was maintained at the appropriate temperature by a VWR 1160S circulating water bath. Prior to the actual manipulation, participants were unaware of the possibility of being exposed to the CPT. This was done to reduce the likelihood of a stress response occurring in participants prior to CPT exposure. To maximize the stress response during the CPT, participants were encouraged to keep their hand in the water bath for the entire 3-min period. However, if a participant found the water bath too painful, he or she was allowed to remove his or her hand from the water and continue with the experiment. Only four participants from the stress condition (3 females, 1 male) removed their hand from the water prior to 3 min elapsing (mean water time = 165.24 s), and all participants from the no stress condition kept their hand in the water for the entire 3-min period. Exclusion of the data from the stressed participants who removed their hand from the water early had no significant effects on the observed results. 2.2.4. Subjective pain and stress ratings Participants were asked to rate the painfulness and stressfulness of the water bath manipulation at 1-min intervals on 11-point scales ranging from 0–10, with 0 indicating a complete lack of pain or stress and 10 indicating unbearable pain or stress. If a participant removed his or her
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hand from the water before 3 min had elapsed, the remaining data points were automatically scored as 10s for each measure. 2.2.5. Cardiovascular analysis Heart rate (HR) and blood pressure (BP) measurements were taken before encoding (baseline), halfway through the water bath manipulation and 10 min after cessation of the water bath manipulation. Cardiovascular activity was measured with a vital sign monitor (Mark of Fitness WS-820 Automatic Wrist Blood Pressure Monitor) placed on the wrist of each participant's dominant hand. 2.2.6. Cortisol analysis Saliva samples were collected from participants before learning (baseline) and 22 min following exposure to the water bath manipulation to analyze salivary cortisol concentrations. The samples were collected in a Salivette saliva collection device (Sarstedt, Inc., Newton, NC). Participants were asked to place a synthetic swab in their mouths and chew on it so that it would easily absorb their saliva. Following 1 min of chewing, the synthetic swab was collected and placed in the Salivette conical tube and kept at room temperature until the experimental session was completed. The samples were subsequently stored at − 20 °C until assayed for cortisol. Saliva samples were thawed and extracted by low-speed centrifugation. Salivary cortisol levels were determined by enzyme immunoassay (EIA; Cayman Chemical Co., Ann Arbor, MI) according to the manufacturer's protocol. 2.2.7. Delayed memory testing Twenty-four hours following encoding, participants returned to the laboratory for an unexpected free recall assessment (they had been told to return to the lab the next day to complete paperwork) and were given 5 min to write down as many words as they could remember from the list of words that they studied on the previous day (i.e., delayed free recall). Then, participants sat quietly for 10 min, after which they were given a recognition test. Participants were presented with a list of words containing 42 “old” words (i.e., words that were presented on the previous day) and 42 “new” words (i.e., words that were not presented on the previous day) and were instructed to label each word as “old” or “new.” The “new” words were matched to the “old” words on emotional valence, word length and word frequency, according to the ratings obtained from the Affective Norms for English Words (Bradley & Lang, 1999). To assess participants' ability to discriminate between “old” and “new” words, we calculated a sensitivity index (d' = z[p(hit)] − z[p(false alarm)]) for each category of word (i.e., positive arousing words, positive non-arousing words, negative arousing words, etc.) to be used for statistical analysis. 2.3. Statistical analyses Mixed-model ANOVAs were used to analyze all physiological and behavioral data; the between-subjects factors utilized in these analyses were stress and sex, and the within-subjects factors were word valence and arousal (for recall and recognition analyses) or time (for physiological and subjective ratings analyses). We also performed exploratory analyses on the memory data from female participants to determine whether menstrual cycle stage influence any observed effects. In order to do so, we divided female participants into follicular [0–14 days since last period; N = 12 (8 stress, 4 no stress)] or luteal [≥ 15 days since last period; N = 13 (4 stress, 9 no stress)] phases of the menstrual cycle (Nielsen, Ahmed, & Cahill, 2013). This division was based on selfreport data obtained from female participants regarding how many days it had been since their last period. In these analyses, stress and menstrual stage served as the between-subjects factors. The analyses of participants' valence and arousal ratings and memory for the words (i.e., immediate free recall, delayed free recall, and recognition) were performed based on categorizing the words (i.e., distributing the words to positive arousing, positive non-arousing, negative arousing,
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etc. groups) according to participants' subjective valence and arousal ratings that were obtained during the study. The resulting categories matched almost identically those produced from the standardized ratings obtained from the Affective Norms for English Words. Each valence-arousal category had 7 words, for a total of 42 words. Alpha was set at 0.05 for all analyses, and Bonferroni-corrected post hoc tests were employed when necessary. SPSS (version 18.0; SPSS, Inc.) was used to perform all statistical analyses. 3. Results 3.1. Physiological responses (see Table 1) 3.1.1. Heart rate During the water bath manipulation, stressed participants exhibited an increase in HR, while non-stressed participants exhibited a decrease in HR (significant Condition × Time interaction: F(2,86) = 5.30, p = 0.007, η2 = 0.11, confirmed by post hoc tests with p b 0.05). Overall, participants' HR decreased over time (significant effect of time: F(2,86) = 11.71, p b 0.001, η2 = 0.21, confirmed by post hoc tests with p b 0.05). Males also exhibited greater baseline HR than females (significant Sex × Time interaction: F(2,86) = 3.79, p = 0.026, η2 = 0.08, confirmed by post hoc tests with p b 0.05). 3.1.2. Blood pressure Stressed participants exhibited greater systolic and diastolic BP than non-stressed participants during the water bath manipulation (SYSTOLIC: significant effect of time: F(2,86) = 5.86, p = 0.004, η2 = 0.12; significant Condition × Time interaction: F(2,86) = 3.11, p = 0.049, η2 = 0.07, confirmed by post hoc tests with p b 0.05; DIASTOLIC: significant effect of time: F(2,86) = 10.11, p b 0.001, η2 = 0.19; significant Condition × Time interaction: F(2,86) = 3.40, p = 0.038, η2 = 0.07, confirmed by post hoc tests with p b 0.05). Male participants also exhibited greater systolic and diastolic BP than female participants (SYSTOLIC: significant effect of sex: F(1,43) = 26.25, p b 0.001, η2 = 0.38; DIASTOLIC: significant effect of sex: F(1,43) = 21.99, p b 0.001, η2 = 0.34). 3.1.3. Salivary cortisol (see Fig. 1) Stressed participants exhibited greater salivary cortisol levels than non-stressed participants after the water bath manipulation (significant effect of time: F(1,43) = 6.22, p = 0.017, η2 = 0.13; significant Condition × Time interaction: F(1,43) = 11.24, p = 0.002, η2 = 0.21, confirmed by post hoc tests with p b 0.05). 3.2. Subjective ratings of water bath manipulation Stressed participants reported greater pain (STRESS: M = 5.93, SEM = 0.35; NO STRESS: M = 0.46, SEM = 0.41) and stress (STRESS: M = 5.25, SEM = 0.47; NO STRESS: M = 1.01, SEM = 0.54) ratings than non-stressed participants throughout the water bath manipulation
Table 1 Cardiovascular activity before, during and after the water bath manipulation. DV/condition Heart rate (bpm) Stress No stress
Pre 70.79 (1.98) 75.91 (4.93)
During
Post
72.81 (2.43) 69.15 (3.14)
62.93 (1.56) 68.48 (2.80)
Systolic blood pressure (mm Hg) Stress 119.72 (2.13) No stress 123.59 (2.90)
135.67 (4.73)⁎ 124.75 (4.67)
124.28 (3.41) 122.57 (2.61)
Diastolic blood pressure (mm Hg) Stress 74.00 (2.07) No stress 78.68 (2.50)
88.52 (3.34)⁎ 80.85 (3.43)
77.34 (1.90) 74.81 (2.47)
Data are presented as means ± SEM. ⁎ p b 0.05 relative to the no stress group.
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effect of valence: F(2,96) = 283.31, p b 0.001, η2 = 0.86). In addition, arousing words were given more positive ratings than non-arousing words (significant effect of arousal: F(1,48) = 4.17, p = 0.047, η2 = 0.08). The latter effect appeared to depend on the valence of the words and the sex of the participant. For instance, positive, but not negative or neutral, words were rated more positively if they were arousing in nature (significant Valence × Arousal interaction: F(2,96) = 13.22, p b 0.001, η2 = 0.22, confirmed by post hoc tests with p b 0.05). In addition, males rated arousing words more positively than non-arousing words, while females did not (significant Sex × Arousal interaction: F(1,48) = 6.76, p = 0.012, η2 = 0.12, confirmed by post hoc tests with p b 0.05).
Fig. 1. Salivary cortisol levels before and after exposure to the water bath manipulation. Stressed participants exhibited significantly higher cortisol levels than non-stressed participants following the water bath manipulation. Data are expressed as means ± SEM. *p b 0.05 relative to no stress.
3.3.2. Arousal ratings Arousing words were rated as more arousing than non-arousing words (significant effect of arousal: F(1,48) = 94.71, p b 0.001, η2 = 0.66). Positive words were rated as more arousing than negative words, which were rated as more arousing than neutral words (significant effect of valence: F(2,96) = 43.92, p b 0.001, η2 = 0.48). 3.4. Memory testing (see Figs. 2 and 3)
(PAIN RATINGS: significant effect of condition: F(1,45) = 103.39, p b 0.001, η2 = 0.70; STRESS RATINGS: significant effect of condition: F(1,45) = 35.25, p b 0.001, η2 = 0.44). 3.3. Word list ratings 3.3.1. Valence ratings Positive words were given more positive ratings than neutral words, which were given more positive ratings than negative words (significant
3.4.1. Immediate free recall For the immediate free recall test, there were no significant differences in overall memory performance between participants assigned to the stress and no stress groups (no significant effect of condition: F(1,45) = 3.21, p = 0.08, η2 = 0.07). However, there was a significant Condition × Valence interaction, F(2,90) = 4.51, p = 0.014, η2 = 0.09, indicating that participants assigned to the stress group recalled more neutral words than participants assigned to the no stress group
Fig. 2. Immediate and delayed free recall performance. Overall, there were no significant group differences in immediate free recall (inset a); however, a significant Condition × Valence interaction was noted, revealing that participants in the stress group recalled more neutral words than participants in the no stress group. As shown in insets b (data expressed as a percentage of immediate free recall performance) and c (raw data expressed as a percentage of total words), stressed participants recalled significantly more words on the delayed free recall assessment than non-stressed participants. This effect was independent of the valence or arousal level of the words. Data are expressed as means ± SEM. *p b 0.05 relative to no stress.
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F(1,21) = 4.77, p = 0.04, η2 = 0.19, confirmed by post hoc tests with p b 0.05; significant Condition × Arousal × Menstrual Stage interaction: F(1,21) = 16.10, p = 0.001, η2 = 0.43, confirmed by post hoc tests with p b 0.05).
Fig. 3. Delayed free recall performance in females as a function of stress, menstrual cycle and arousal levels of the words. Non-stressed females exhibited greater memory for arousing words, relative to non-arousing words, during the luteal phase, and stressed females exhibited greater memory for arousing words, relative to non-arousing words, during the follicular phase. Stressed females also exhibited greater memory for nonarousing words, relative to non-stressed females, during the luteal phase. Data are expressed as means ± SEM, and sample sizes have been provided for each group. *p b 0.05 relative to non-arousing words; β = p b 0.05 relative to no stress.
(confirmed by post hoc tests with p b 0.05). Overall, participants recalled more negative words than positive and neutral words (significant effect of valence: F(2,90) = 5.50, p = 0.006, η2 = 0.11). Participants also recalled more arousing words than non-arousing words, although this effect was only evident in male participants (significant effect of arousal: F(1,45) = 9.52, p = 0.003, η2 = 0.18; significant Sex × Arousal interaction: F(1,45) = 6.57, p = 0.014, η2 = 0.13, confirmed by post hoc tests with p b 0.05). The analysis of immediate free recall performance in females (which included menstrual stage and condition as factors) revealed no significant main effects or interactions.
3.4.2.2. Percent of immediate recall. Because we observed some group differences in immediate free recall performance, we also analyzed the delayed free recall performance as a percent of immediate free recall (i.e., (delayed recall/immediate recall) ∗ 100). This analysis corroborated our findings from the raw data analysis by revealing that stressed participants recalled more words overall than non-stressed participants (significant effect of condition: F(1,48) = 7.90, p = 0.007, η2 = 0.14). Overall, participants recalled more negative words than positive and neutral words (significant effect of valence: F(2,96) = 7.96, p = 0.001, η2 = 0.14). Participants also recalled more arousing words than nonarousing words, particularly when they were negative or neutral in valence (significant effect of arousal: F(1,48) = 6.88, p = 0.012, η2 = 0.13; significant Valence × Arousal interaction: F(2,96) = 9.46, p b 0.001, η2 = 0.17, confirmed by post hoc tests with p b 0.05). The analysis of delayed free recall performance expressed as a percent of immediate free recall in females revealed that the effects observed when analyzing the raw data remained significant. Stressed females recalled more words overall than non-stressed females (significant effect of condition: F(1,21) = 7.80, p = 0.011, η2 = 0.27), and more importantly, the three-way Condition × Arousal × Menstrual Stage interaction described above remained significant, F(1,21) = 5.30, p = 0.032, η2 = 0.20, confirmed by post hoc tests with p b 0.05. 3.4.3. Delayed recognition (see Fig. 4) There were no significant differences between the recognition performance of stressed and non-stressed participants (no significant effect of condition: F(1,48) = 1.38, p = 0.247, η2 = 0.03). Participants did recognize more arousing than non-arousing words when they were negative or neutral in valence (significant Valence × Arousal interaction: F(2,96) = 7.65, p = 0.001, η2 = 0.14, confirmed by post hoc tests with p b 0.05). The analysis of delayed recognition in females revealed no significant main effects or interactions.
3.4.2. Delayed free recall 4. Discussion 3.4.2.1. Raw data. Stressed participants recalled more words overall than non-stressed participants (significant effect of condition: F(1,48) = 6.87, p = 0.012, η2 b 0.13). Overall, participants recalled more negative and neutral words than positive words (significant effect of valence: F(2,96) = 14.30, p b 0.001, η2 = 0.23). Participants also recalled more arousing words than non-arousing words, particularly when they were negative or neutral in valence (significant effect of arousal: F(1,48) = 27.71, p b 0.001, η2 = 0.37; significant Valence × Arousal interaction: F(2,96) = 4.97, p = 0.009, η2 = 0.09, confirmed by post hoc tests with p b 0.05). The analysis of raw delayed free recall performance in females revealed that stressed females recalled more words overall than nonstressed females (significant effect of condition: F(1,21) = 13.84, p = 0.001, η2 = 0.40). Overall, female participants recalled more positive and neutral words than negative words (significant effect of valence: F(2,42) = 8.31, p = 0.001, η2 = 0.28) and more arousing words than non-arousing words (significant effect of arousal: F(1,21) = 6.97, p = 0.015, η2 = 0.25). More interestingly, stress exerted differential effects on emotional memory in females, depending on stage of the menstrual cycle. Specifically, stress enhanced memory for arousing words, relative to non-arousing words, when females were in the follicular stage, while enhancing memory for non-arousing words, relative to non-stressed female memory for non-arousing words, when in the luteal phase. Non-stressed females exhibited greater memory for arousing words, relative to non-arousing words, when in the luteal, but not follicular, phase (significant effect of menstrual stage: F(1,21) = 9.88, p = 0.005, η2 = 0.32; significant Arousal × Menstrual Stage interaction:
Most studies examining the effects of post-learning stress on longterm memory have reported facilitative effects (Beckner et al., 2006; Cahill et al., 2003; Preuss & Wolf, 2009). However, there is some research in this literature reporting no effects or an impairment of long-term memory (e.g., Trammell & Clore, 2014). The purpose of the present study was to examine further the effects of post-learning stress on long-term memory and to explore whether sex and menstrual cycle
Fig. 4. Recognition memory during the 24-h test. No significant group effects were observed for recognition memory. Data are expressed as means ± SEM.
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stage influenced such effects. We found that post-learning stress enhanced 24-h free recall, even when memory performance was expressed as a percent of immediate free recall performance. This enhancement was independent of sex, as well as the valence and arousal level of the words. We also found that emotional memory in females was differentially affected by post-learning stress, depending on which stage of the menstrual cycle they were experiencing at the time of stress. Specifically, stress enhanced memory for arousing words, relative to non-arousing words, in females who were in the follicular phase and enhanced memory for non-arousing words, relative to nonstressed female memory, in females who were in the luteal phase. Together, our results replicate the majority of post-learning stress research and extend these findings to implicate female hormonal mechanisms in such effects. 4.1. Neurobiological mechanisms of post-learning stress We purposely induced post-learning stress as close in time as possible to word list learning in order for the stressor to be experienced in the context of learning. That we observed an enhancement of long-term memory as a result of this manipulation is consistent with previous research revealing that the timing of stress relative to learning is important in dictating the types of effects observed on long-term memory (Diamond et al., 2007; Quaedflieg, Schwabe, Meyer, & Smeets, 2013; Zoladz, Clark, et al., 2011; Zoladz et al., 2014). Physiologically, stress results in the activation of two major systems, the sympathetic nervous system and the hypothalamus-pituitary-adrenal axis. Activation of these two systems results in increased adrenergic and corticosteroid activity, respectively, which can exert biphasic effects on cognitive processing. When the stress-induced increases in adrenergic and corticosteroid activity converge in time with learning, research suggests that these mechanisms, coupled with amygdala activity, facilitates memory processing in structures such as the hippocampus, which enables phenomena such as flashbulb memories to occur (Diamond et al., 2007; Schwabe, Joels, Roozendaal, Wolf, & Oitzl, 2012). Consistent with this idea, we found that CPT exposure in the present study resulted in significant increases in cardiovascular measures, indicators of autonomic and adrenergic activity, and salivary cortisol concentrations. These effects were produced with a relatively brief (3-min) stressor, suggesting that even brief stress of a particular intensity can promote memory consolidation. The CPT is a physiologically-based stressor, as it produces a stress response via cold-induced pain. However, the effects of the CPT on cortisol levels, learning and memory are comparable to those induced by other stressors, such as psychosocial stress (e.g., Trier Social Stress Test).
can impact the acquisition and extinction of emotional memory. Felmingham, Fong, and Bryant (2012) reported that heightened progesterone, which typically peaks in the mid-late luteal phase, is positively correlated with memory recall of threatening images and may mediate cortisol response. Similarly, a study examining sex differences in acoustic startle response found that females in the late luteal phase exhibited enhanced startle response compared to both males and females in the follicular phase (Armbruster, Strobel, Kirschbaum, & Brocke, 2014). One hypothesis put forth to explain progesterone's enhancing effects for arousing material is that heightened progesterone levels increase sensitivity to sources associated with threat or danger (Conway et al., 2007; Derntl et al., 2008). There appears to be a plausible interaction occurring between progesterone and cortisol, such that in the absence or diminished levels of one hormone, the heightened levels of the opposing chemical acts in such a way as to mimic the effects of the other. This could explain why females in the follicular phase (low progesterone) exhibited greater delayed recall for arousing words only in the stressed condition (high cortisol), while females in the luteal phase (high progesterone) had enhanced delayed recall for arousing material when in the control condition (low cortisol). 4.3. Caveats Although the present results indicate that post-learning stress might differentially influence females depending on menstrual stage, it is important to emphasize that the analyses performed on such measures here were exploratory in nature and consisted of relatively small sample sizes (as indicated in Fig. 3). Furthermore, we did not measure the levels of progesterone or estradiol in females, and our measure of menstrual stage was based on self-report. Therefore, although our findings are suggestive of a link between menstrual cycle activity and stress-memory interactions, further research is clearly warranted to substantiate these claims. 4.4. Conclusions We have provided additional evidence that post-learning stress facilitates long-term memory, an effect that, at least in the present study, is independent of the emotional nature of the learned information. We have also shown that females are differentially affected by post-learning stress depending on stage of the menstrual cycle. These findings further validate post-learning stress-induced enhancements of long-term memory and implicate female sex hormones in such effects. References
4.2. Menstrual cycle interactions with delayed recall An extensive review of the stress-memory literature reveals that females have been largely underrepresented in this research. This is especially surprising and concerning, as females are significantly more likely to develop stress-related psychological disorders, such as PTSD and major depression, that involve pathological stress-memory interactions. Although recent studies have begun to report sex differences more frequently, many researchers still fail to include measures that could help explain sex differences in stress-memory interactions. By collecting data from female participants about menstrual cycle activity, we found that menstrual cycle stage was associated with emotional memory under baseline and stress conditions. Specifically, nonstressed females exhibited greater memory for arousing words, relative to non-arousing words, when they were in the luteal phase. In contrast, stressed females demonstrated greater memory for arousing words, relative to non-arousing words, when they were in the follicular phase. Interestingly, stressed females in the luteal phase exhibited enhanced memory, relative to non-stressed females, for non-arousing words. Recent research has shown that both estradiol and progesterone
Armbruster, D., Strobel, A., Kirschbaum, C., & Brocke, B. (2014). The impact of sex and menstrual cycle on the acoustic startle response. Behavioural Brain Research, 274, 326–333. Beckner, V. E., Tucker, D. M., Delville, Y., & Mohr, D. C. (2006). Stress facilitates consolidation of verbal memory for a film but does not affect retrieval. Behavioral Neuroscience, 120, 518–527. Bradley, M. M., & Lang, P. J. (1999). Affective norms for English words (ANEW): Instruction manual and affective ratings. Technical Report C-1, The Center for Research in Psychophysiology. University of Florida. Cahill, L., Gorski, L., & Le, K. (2003). Enhanced human memory consolidation with postlearning stress: Interaction with the degree of arousal at encoding. Learning and Memory, 10, 270–274. Conway, C. A., Jones, B. C., DeBruine, L. M., Welling, L. L., Law Smith, M. J., Perrett, D. I., et al. (2007). Salience of emotional displays of danger and contagion in faces is enhanced when progesterone levels are raised. Hormones and Behavior, 51, 202–206. Derntl, B., Windischberger, C., Robinson, S., Lamplmayr, E., Kryspin-Exner, I., Gur, R. C., et al. (2008). Facial emotion recognition and amygdala activation are associated with menstrual cycle phase. Psychoneuroendocrinology, 33, 1031–1040. Diamond, D. M., Campbell, A. M., Park, C. R., Halonen, J., & Zoladz, P. R. (2007). The temporal dynamics model of emotional memory processing: A synthesis on the neurobiological basis of stress-induced amnesia, flashbulb and traumatic memories, and the Yerkes–Dodson law. Neural Plasticity, 2007, 60803. Felmingham, K. L., Fong, W. C., & Bryant, R. A. (2012). The impact of progesterone on memory consolidation of threatening images in women. Psychoneuroendocrinology, 37, 1896–1900.
P.R. Zoladz et al. / Acta Psychologica 160 (2015) 127–133 Felmingham, K. L., Tran, T. P., Fong, W. C., & Bryant, R. A. (2012). Sex differences in emotional memory consolidation: The effect of stress-induced salivary alpha-amylase and cortisol. Biological Psychology, 89, 539–544. Jelicic, M., Geraerts, E., Merckelbach, H., & Guerrieri, R. (2004). Acute stress enhances memory for emotional words, but impairs memory for neutral words. The International Journal of Neuroscience, 114, 1343–1351. Livingston, R. B. (1967). Reinforcement. In G. Quarton, T. Melnechuk, & F. Schmitt (Eds.), The neurosciences: A study program (pp. 514–576). New York: Rockefeller Press. Nielsen, S. E., Ahmed, I., & Cahill, L. (2013). Sex and menstrual cycle phase at encoding influence emotional memory for gist and detail. Neurobiology of Learning and Memory, 106, 56–65. Nielson, K. A., & Powless, M. (2007). Positive and negative sources of emotional arousal enhance long-term word-list retention when induced as long as 30 min after learning. Neurobiology of Learning and Memory, 88, 40–47. Nielson, K. A., Yee, D., & Erickson, K. I. (2005). Memory enhancement by a semantically unrelated emotional arousal source induced after learning. Neurobiology of Learning and Memory, 84, 49–56. Payne, J. D., Jackson, E. D., Hoscheidt, S., Ryan, L., Jacobs, W. J., & Nadel, L. (2007). Stress administered prior to encoding impairs neutral but enhances emotional long-term episodic memories. Learning and Memory, 14, 861–868. Preuss, D., & Wolf, O. T. (2009). Post-learning psychosocial stress enhances consolidation of neutral stimuli. Neurobiology of Learning and Memory, 92, 318–326. Quaedflieg, C. W., Schwabe, L., Meyer, T., & Smeets, T. (2013). Time dependent effects of stress prior to encoding on event-related potentials and 24 h delayed retrieval. Psychoneuroendocrinology, 38, 3057–3069. Roozendaal, B. (2003). Systems mediating acute glucocorticoid effects on memory consolidation and retrieval. Progress in Neuropsychopharmacology and Biological Psychiatry, 27, 1213–1223.
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Roozendaal, B., McEwen, B. S., & Chattarji, S. (2009). Stress, memory and the amygdala. Nature Review Neuroscience, 10, 423–433. Schwabe, L., Bohringer, A., Chatterjee, M., & Schachinger, H. (2008). Effects of pre-learning stress on memory for neutral, positive and negative words: Different roles of cortisol and autonomic arousal. Neurobiology of Learning and Memory, 90, 44–53. Schwabe, L., Joels, M., Roozendaal, B., Wolf, O. T., & Oitzl, M. S. (2012). Stress effects on memory: An update and integration. Neuroscience and Biobehavioral Reviews, 36, 1740–1749. Trammell, J. P., & Clore, G. L. (2014). Does stress enhance or impair memory consolidation? Cogn Emot, 28, 361–374. Zoladz, P. R., Clark, B., Warnecke, A., Smith, L., Tabar, J., & Talbot, J. N. (2011). Pre-learning stress differentially affects long-term memory for emotional words, depending on temporal proximity to the learning experience. Physiology and Behavior, 103, 467–476. Zoladz, P. R., Kalchik, A. E., Hoffman, M. M., Aufdenkampe, R. L., Lyle, S. M., Peters, D. M., et al. (2014). ADRA2B deletion variant selectively predicts stress-induced enhancement of long-term memory in females. Psychoneuroendocrinology, 48, 111–122. Zoladz, P. R., Park, C. R., & Diamond, D. M. (2011). Neurobiological basis of the complex effects of stress on memory and synaptic plasticity. In C. D. Conrad (Ed.), The Handbook of Stress: Neuropsychological Effects on the Brain (pp. 157–178). Oxford, UK: Wiley-Blackwell. Zoladz, P. R., Warnecke, A. J., Woelke, S. A., Burke, H. M., Frigo, R. M., Pisansky, J. M., et al. (2013). Pre-learning stress that is temporally removed from acquisition exerts sexspecific effects on long-term memory. Neurobiology of Learning and Memory, 100, 77–87.
Contents lists available at ScienceDirect
Acta Psychologica journal homepage: www.elsevier.com/ locate/actpsy
Post-learning stress enhances long-term memory and differentially influences memory in females depending on menstrual stage Phillip R. Zoladz a,⁎, David M. Peters a, Chelsea E. Cadle a, Andrea E. Kalchik a, Rachael L. Aufdenkampe a, Alison M. Dailey a, Callie M. Brown a, Amanda R. Scharf a, McKenna B. Earley a, Courtney L. Knippen a, Boyd R. Rorabaugh b a b
Department of Psychology, Sociology, & Criminal Justice, Ohio Northern University, Ada, OH 45810, USA Department of Pharmaceutical & Biomedical Sciences, Raabe College of Pharmacy, Ohio Northern University, Ada, OH 45810, USA
a r t i c l e
i n f o
Article history: Received 6 June 2015 Received in revised form 17 July 2015 Accepted 20 July 2015 Available online xxxx Keywords: Stress Learning Memory Sex Menstrual Emotion
a b s t r a c t Most work has shown that post-learning stress enhances long-term memory; however, there have been recent inconsistencies in this literature. The purpose of the present study was to examine further the effects of post-learning stress on long-term memory and to explore any sex differences that may exist. Male and female participants learned a list of 42 words that varied in emotional valence and arousal level. Following encoding, participants completed a free recall assessment and then submerged their hand into a bath of ice cold (stress) or lukewarm (no stress) water for 3 min. The next day, participants were given free recall and recognition tests. Stressed participants recalled more words than non-stressed participants 24 h after learning. Stress also enhanced female participants' recall of arousing words when they were in the follicular, but not luteal, phase. These findings replicate previous work examining post-learning stress effects on memory and implicate the involvement of sex-related hormones in such effects. © 2015 Elsevier B.V. All rights reserved.
1. Introduction Stress exerts powerful effects on learning and memory, and understanding these effects may lend insight into cognitive symptoms associated with stress-related psychological disorders, such as post-traumatic stress disorder (PTSD) and major depression. Researchers have frequently dichotomized stress-memory interactions into the effects of stress on consolidation versus the effects of stress on retrieval, as these two phases of memory processing seem to be differentially influenced by stress (Roozendaal, McEwen, & Chattarji, 2009). Indeed, most research has reported that stress enhances consolidation and impairs retrieval, despite similar mechanisms, such as corticosteroid and noradrenergic interactions in the amygdala, underlying both (Beckner, Tucker, Delville, & Mohr, 2006; Cahill, Gorski, & Le, 2003; Felmingham, Tran, Fong, & Bryant, 2012; Nielson, Yee, & Erickson, 2005; Preuss & Wolf, 2009; Roozendaal, 2003). More recently, researchers have discovered that the effects of stress on consolidation depend on whether the stress is administered before or after learning, as well as the temporal proximity of the stress to learning. For instance, pre-learning stress that is experienced in the context of learning (i.e., temporally associated with learning) often enhances ⁎ Corresponding author at: Ohio Northern University, Department of Psychology, Sociology, & Criminal Justice, 525 S. Main St. Hill 013, Ada, OH 45810, USA. E-mail address: [email protected] (P.R. Zoladz).
http://dx.doi.org/10.1016/j.actpsy.2015.07.008 0001-6918/© 2015 Elsevier B.V. All rights reserved.
long-term memory (Diamond, Campbell, Park, Halonen, & Zoladz, 2007; Schwabe, Bohringer, Chatterjee, & Schachinger, 2008; Zoladz et al., 2011; Zoladz et al., 2014), while pre-learning stress that is experienced outside the context of learning (i.e., temporally separated from learning) often impairs long-term memory (Zoladz, Clark, et al., 2011; Zoladz et al., 2013). Post-learning stress, on the other hand, almost unequivocally enhances long-term memory (Cahill et al., 2003; Felmingham, Tran, Fong, & Bryant, 2012; Nielson & Powless, 2007; Nielson et al., 2005; Preuss & Wolf, 2009); however, such enhancement declines as the stress is temporally removed from the learning experience (Nielson & Powless, 2007). Although the effects of stress on learning and memory could be perceived as adaptive when memory is enhanced and maladaptive when memory is impaired, we would contend that all effects of stress on learning and memory serve an adaptive role (Zoladz, Park, & Diamond, 2011). When stress is experienced in the context of learning, long-term memory is enhanced because, hypothetically, the stress experience signals the brain that what is occurring is important to remember, which results in a “Now Print” mechanism (Livingston, 1967), or emotional tag, that enhances memory formation. This enables the individual to remember potentially life-threatening aspects related to the stressor which could aid survival later in life. In support of this reasoning, pre- and post-learning stress often enhances memory for emotional information, while impairing or having no effect on memory for neutral information (Jelicic, Geraerts, Merckelbach, & Guerrieri, 2004;
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Payne et al., 2007; Zoladz, Clark, et al., 2011). When stress is experienced outside the context of learning, long-term memory is impaired because, hypothetically, the brain, and in particular the hippocampus, is focused on storing the memory of the stress experience, thereby rendering new information storage improbable. Though simplistic, this approach to how stress affects memory consolidation emphasizes the adaptive nature of the stress response and is supported by behavioral, molecular and electrophysiological studies in humans and non-human animals. Though many studies have reported that post-learning stress enhances consolidation, some work has presented conflicting data suggesting that such stress impairs long-term memory. For instance, Trammell and Clore (2014) recently reported that post-learning cold pressor stress (1–3 min duration) impaired long-term (48-h) word and picture memory in participants. However, in the studies, all participants exhibited relatively poor long-term memory (recalling b 7% of words studied and only ~ 10% of pictures studied) and, in two of the three experiments, the difference in recall between stressed and nonstressed participants was only approaching significance (p's = 0.06 and 0.07). Additionally, the investigators examined 48-h memory, as opposed to the more commonly used 24-h time point. Thus, the purpose of the present study was to examine further the relationship between post-learning stress and long-term (24-h) memory and to assess how manipulating the emotional valence and arousal level of the learned information could influence such effects. We hypothesized that post-learning stress would facilitate long-term memory, perhaps more strongly for emotionally arousing information. Moreover, because sex differences are prevalent in the stress-memory literature, we also examined whether post-learning stress differentially affected longterm memory in males and females. Some previous work has reported greater memory-enhancing effects of stress in females, relative to males (e.g., Felmingham, Tran, Fong, & Bryant, 2012; Zoladz et al., 2014); thus, we predicted that females may be more susceptible to post-learning stress effects on long-term memory. We also ran exploratory analyses on female data to assess whether menstrual cycle stage interacted with stress effects on memory.
2.2.1. Word presentation Participants were presented with a list of 42 words, which were selected from the Affective Norms for English Words (Bradley & Lang, 1999). Based on standardized valence and arousal ratings, we chose 14 neutral (7 arousing, 7 non-arousing), 14 positive (7 arousing, 7 non-arousing) and 14 negative (7 arousing, 7 non-arousing) words, which, across emotional valence and arousal categories, were balanced for word length and word frequency. As per the methods employed by Schwabe et al. (2008) and previous work from our own laboratory (Zoladz, Clark, et al., 2011; Zoladz et al., 2013, 2014), participants were instructed to read each word aloud and rate its emotional valence on a scale from −4 (very negative) to +4 (very positive) and its arousal level on a scale of 0 (not arousing) to 8 (very highly arousing) on a sheet of paper containing the list of words and the aid of self-assessment manikins. These manipulations were performed to promote encoding of the words, and they allowed us to analyze the final memory data based on participants' own ratings of the words. According to the Affective Norms for English Words (Bradley & Lang, 1999), the mean (± SEM) valence and arousal ratings for the words that made up the list were as follows: positive arousing words (e.g., kiss): valence = 7.79 ± 0.12, arousal = 6.62 ± 0.25; positive non-arousing words (e.g., cozy): valence = 7.50 ± 0.12, arousal = 3.46 ± 0.16; negative arousing words (e.g., poison): valence = 2.21 ± 0.16, arousal = 6.56 ± 0.27; negative non-arousing words (e.g., unhappy): valence = 2.40 ± 0.20, arousal = 3.89 ± 0.19; neutral arousing words (e.g., lightning): valence = 4.93 ± 0.27, arousal = 6.26 ± 0.20; neutral non-arousing words (e.g., poster): valence = 4.90 ± 0.14, arousal = 3.40 ± 0.13. 2.2.2. Immediate memory testing Immediately following word list encoding, participants were given 5 min to write down as many words as they could remember from the list of words they just studied. This immediate free recall test was performed to verify that there were no group differences regarding short-term memory performance and to avoid a potential floor effect during long-term memory assessment (e.g., Zoladz, Clark, et al., 2011).
2. Materials and method 2.1. Participants Fifty-two healthy men and naturally cycling women (27 men, 25 women; age: M = 20.30, SD = 1.17) from Ohio Northern University volunteered to participate in the experiment. Individuals were excluded from participating if they met any of the following conditions: diagnosis of Raynaud's or peripheral vascular disease; presence of skin diseases, such as psoriasis, eczema or scleroderma; history of syncope or vasovagal response to stress; history of severe head injury; current treatment with psychotropic medications, narcotics, beta-blockers, steroids or any other medication that was deemed to significantly affect central nervous or endocrine system function; mental or substance use disorder; regular nightshift work. Participants were asked to refrain from using recreational drugs (e.g., marijuana) for 3 days prior to the experimental sessions; to refrain from drinking alcohol or exercising extensively for 24 h prior to the experimental sessions; and, to refrain from eating or drinking anything but water for 2 h prior to the experimental sessions. Upon arrival at the laboratory, participants were reminded of the exclusion criteria and study restrictions and verbally affirmed that they had adhered to the requirements. Participants were awarded class credit upon completion of the study. All of the methods for the experiment were approved by the Institutional Review Board at Ohio Northern University. 2.2. Experimental procedures To control for diurnal variations in cortisol levels, all testing was carried out between 1200 and 1800 h.
2.2.3. Cold pressor test (CPT) Following immediate memory testing, participants were asked to submerge their non-dominant hand, up to and including the wrist, in a bath of water for 3 min. Those participants who had been randomly assigned to the stress condition (N = 29; 17 males, 12 females) placed their hand in a bath of ice cold (0–2 °C) water, while participants who had been randomly assigned to the control condition (N = 23; 10 males, 13 females) placed their hand in a bath of warm (35–37 °C) water. The water was maintained at the appropriate temperature by a VWR 1160S circulating water bath. Prior to the actual manipulation, participants were unaware of the possibility of being exposed to the CPT. This was done to reduce the likelihood of a stress response occurring in participants prior to CPT exposure. To maximize the stress response during the CPT, participants were encouraged to keep their hand in the water bath for the entire 3-min period. However, if a participant found the water bath too painful, he or she was allowed to remove his or her hand from the water and continue with the experiment. Only four participants from the stress condition (3 females, 1 male) removed their hand from the water prior to 3 min elapsing (mean water time = 165.24 s), and all participants from the no stress condition kept their hand in the water for the entire 3-min period. Exclusion of the data from the stressed participants who removed their hand from the water early had no significant effects on the observed results. 2.2.4. Subjective pain and stress ratings Participants were asked to rate the painfulness and stressfulness of the water bath manipulation at 1-min intervals on 11-point scales ranging from 0–10, with 0 indicating a complete lack of pain or stress and 10 indicating unbearable pain or stress. If a participant removed his or her
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hand from the water before 3 min had elapsed, the remaining data points were automatically scored as 10s for each measure. 2.2.5. Cardiovascular analysis Heart rate (HR) and blood pressure (BP) measurements were taken before encoding (baseline), halfway through the water bath manipulation and 10 min after cessation of the water bath manipulation. Cardiovascular activity was measured with a vital sign monitor (Mark of Fitness WS-820 Automatic Wrist Blood Pressure Monitor) placed on the wrist of each participant's dominant hand. 2.2.6. Cortisol analysis Saliva samples were collected from participants before learning (baseline) and 22 min following exposure to the water bath manipulation to analyze salivary cortisol concentrations. The samples were collected in a Salivette saliva collection device (Sarstedt, Inc., Newton, NC). Participants were asked to place a synthetic swab in their mouths and chew on it so that it would easily absorb their saliva. Following 1 min of chewing, the synthetic swab was collected and placed in the Salivette conical tube and kept at room temperature until the experimental session was completed. The samples were subsequently stored at − 20 °C until assayed for cortisol. Saliva samples were thawed and extracted by low-speed centrifugation. Salivary cortisol levels were determined by enzyme immunoassay (EIA; Cayman Chemical Co., Ann Arbor, MI) according to the manufacturer's protocol. 2.2.7. Delayed memory testing Twenty-four hours following encoding, participants returned to the laboratory for an unexpected free recall assessment (they had been told to return to the lab the next day to complete paperwork) and were given 5 min to write down as many words as they could remember from the list of words that they studied on the previous day (i.e., delayed free recall). Then, participants sat quietly for 10 min, after which they were given a recognition test. Participants were presented with a list of words containing 42 “old” words (i.e., words that were presented on the previous day) and 42 “new” words (i.e., words that were not presented on the previous day) and were instructed to label each word as “old” or “new.” The “new” words were matched to the “old” words on emotional valence, word length and word frequency, according to the ratings obtained from the Affective Norms for English Words (Bradley & Lang, 1999). To assess participants' ability to discriminate between “old” and “new” words, we calculated a sensitivity index (d' = z[p(hit)] − z[p(false alarm)]) for each category of word (i.e., positive arousing words, positive non-arousing words, negative arousing words, etc.) to be used for statistical analysis. 2.3. Statistical analyses Mixed-model ANOVAs were used to analyze all physiological and behavioral data; the between-subjects factors utilized in these analyses were stress and sex, and the within-subjects factors were word valence and arousal (for recall and recognition analyses) or time (for physiological and subjective ratings analyses). We also performed exploratory analyses on the memory data from female participants to determine whether menstrual cycle stage influence any observed effects. In order to do so, we divided female participants into follicular [0–14 days since last period; N = 12 (8 stress, 4 no stress)] or luteal [≥ 15 days since last period; N = 13 (4 stress, 9 no stress)] phases of the menstrual cycle (Nielsen, Ahmed, & Cahill, 2013). This division was based on selfreport data obtained from female participants regarding how many days it had been since their last period. In these analyses, stress and menstrual stage served as the between-subjects factors. The analyses of participants' valence and arousal ratings and memory for the words (i.e., immediate free recall, delayed free recall, and recognition) were performed based on categorizing the words (i.e., distributing the words to positive arousing, positive non-arousing, negative arousing,
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etc. groups) according to participants' subjective valence and arousal ratings that were obtained during the study. The resulting categories matched almost identically those produced from the standardized ratings obtained from the Affective Norms for English Words. Each valence-arousal category had 7 words, for a total of 42 words. Alpha was set at 0.05 for all analyses, and Bonferroni-corrected post hoc tests were employed when necessary. SPSS (version 18.0; SPSS, Inc.) was used to perform all statistical analyses. 3. Results 3.1. Physiological responses (see Table 1) 3.1.1. Heart rate During the water bath manipulation, stressed participants exhibited an increase in HR, while non-stressed participants exhibited a decrease in HR (significant Condition × Time interaction: F(2,86) = 5.30, p = 0.007, η2 = 0.11, confirmed by post hoc tests with p b 0.05). Overall, participants' HR decreased over time (significant effect of time: F(2,86) = 11.71, p b 0.001, η2 = 0.21, confirmed by post hoc tests with p b 0.05). Males also exhibited greater baseline HR than females (significant Sex × Time interaction: F(2,86) = 3.79, p = 0.026, η2 = 0.08, confirmed by post hoc tests with p b 0.05). 3.1.2. Blood pressure Stressed participants exhibited greater systolic and diastolic BP than non-stressed participants during the water bath manipulation (SYSTOLIC: significant effect of time: F(2,86) = 5.86, p = 0.004, η2 = 0.12; significant Condition × Time interaction: F(2,86) = 3.11, p = 0.049, η2 = 0.07, confirmed by post hoc tests with p b 0.05; DIASTOLIC: significant effect of time: F(2,86) = 10.11, p b 0.001, η2 = 0.19; significant Condition × Time interaction: F(2,86) = 3.40, p = 0.038, η2 = 0.07, confirmed by post hoc tests with p b 0.05). Male participants also exhibited greater systolic and diastolic BP than female participants (SYSTOLIC: significant effect of sex: F(1,43) = 26.25, p b 0.001, η2 = 0.38; DIASTOLIC: significant effect of sex: F(1,43) = 21.99, p b 0.001, η2 = 0.34). 3.1.3. Salivary cortisol (see Fig. 1) Stressed participants exhibited greater salivary cortisol levels than non-stressed participants after the water bath manipulation (significant effect of time: F(1,43) = 6.22, p = 0.017, η2 = 0.13; significant Condition × Time interaction: F(1,43) = 11.24, p = 0.002, η2 = 0.21, confirmed by post hoc tests with p b 0.05). 3.2. Subjective ratings of water bath manipulation Stressed participants reported greater pain (STRESS: M = 5.93, SEM = 0.35; NO STRESS: M = 0.46, SEM = 0.41) and stress (STRESS: M = 5.25, SEM = 0.47; NO STRESS: M = 1.01, SEM = 0.54) ratings than non-stressed participants throughout the water bath manipulation
Table 1 Cardiovascular activity before, during and after the water bath manipulation. DV/condition Heart rate (bpm) Stress No stress
Pre 70.79 (1.98) 75.91 (4.93)
During
Post
72.81 (2.43) 69.15 (3.14)
62.93 (1.56) 68.48 (2.80)
Systolic blood pressure (mm Hg) Stress 119.72 (2.13) No stress 123.59 (2.90)
135.67 (4.73)⁎ 124.75 (4.67)
124.28 (3.41) 122.57 (2.61)
Diastolic blood pressure (mm Hg) Stress 74.00 (2.07) No stress 78.68 (2.50)
88.52 (3.34)⁎ 80.85 (3.43)
77.34 (1.90) 74.81 (2.47)
Data are presented as means ± SEM. ⁎ p b 0.05 relative to the no stress group.
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effect of valence: F(2,96) = 283.31, p b 0.001, η2 = 0.86). In addition, arousing words were given more positive ratings than non-arousing words (significant effect of arousal: F(1,48) = 4.17, p = 0.047, η2 = 0.08). The latter effect appeared to depend on the valence of the words and the sex of the participant. For instance, positive, but not negative or neutral, words were rated more positively if they were arousing in nature (significant Valence × Arousal interaction: F(2,96) = 13.22, p b 0.001, η2 = 0.22, confirmed by post hoc tests with p b 0.05). In addition, males rated arousing words more positively than non-arousing words, while females did not (significant Sex × Arousal interaction: F(1,48) = 6.76, p = 0.012, η2 = 0.12, confirmed by post hoc tests with p b 0.05).
Fig. 1. Salivary cortisol levels before and after exposure to the water bath manipulation. Stressed participants exhibited significantly higher cortisol levels than non-stressed participants following the water bath manipulation. Data are expressed as means ± SEM. *p b 0.05 relative to no stress.
3.3.2. Arousal ratings Arousing words were rated as more arousing than non-arousing words (significant effect of arousal: F(1,48) = 94.71, p b 0.001, η2 = 0.66). Positive words were rated as more arousing than negative words, which were rated as more arousing than neutral words (significant effect of valence: F(2,96) = 43.92, p b 0.001, η2 = 0.48). 3.4. Memory testing (see Figs. 2 and 3)
(PAIN RATINGS: significant effect of condition: F(1,45) = 103.39, p b 0.001, η2 = 0.70; STRESS RATINGS: significant effect of condition: F(1,45) = 35.25, p b 0.001, η2 = 0.44). 3.3. Word list ratings 3.3.1. Valence ratings Positive words were given more positive ratings than neutral words, which were given more positive ratings than negative words (significant
3.4.1. Immediate free recall For the immediate free recall test, there were no significant differences in overall memory performance between participants assigned to the stress and no stress groups (no significant effect of condition: F(1,45) = 3.21, p = 0.08, η2 = 0.07). However, there was a significant Condition × Valence interaction, F(2,90) = 4.51, p = 0.014, η2 = 0.09, indicating that participants assigned to the stress group recalled more neutral words than participants assigned to the no stress group
Fig. 2. Immediate and delayed free recall performance. Overall, there were no significant group differences in immediate free recall (inset a); however, a significant Condition × Valence interaction was noted, revealing that participants in the stress group recalled more neutral words than participants in the no stress group. As shown in insets b (data expressed as a percentage of immediate free recall performance) and c (raw data expressed as a percentage of total words), stressed participants recalled significantly more words on the delayed free recall assessment than non-stressed participants. This effect was independent of the valence or arousal level of the words. Data are expressed as means ± SEM. *p b 0.05 relative to no stress.
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F(1,21) = 4.77, p = 0.04, η2 = 0.19, confirmed by post hoc tests with p b 0.05; significant Condition × Arousal × Menstrual Stage interaction: F(1,21) = 16.10, p = 0.001, η2 = 0.43, confirmed by post hoc tests with p b 0.05).
Fig. 3. Delayed free recall performance in females as a function of stress, menstrual cycle and arousal levels of the words. Non-stressed females exhibited greater memory for arousing words, relative to non-arousing words, during the luteal phase, and stressed females exhibited greater memory for arousing words, relative to non-arousing words, during the follicular phase. Stressed females also exhibited greater memory for nonarousing words, relative to non-stressed females, during the luteal phase. Data are expressed as means ± SEM, and sample sizes have been provided for each group. *p b 0.05 relative to non-arousing words; β = p b 0.05 relative to no stress.
(confirmed by post hoc tests with p b 0.05). Overall, participants recalled more negative words than positive and neutral words (significant effect of valence: F(2,90) = 5.50, p = 0.006, η2 = 0.11). Participants also recalled more arousing words than non-arousing words, although this effect was only evident in male participants (significant effect of arousal: F(1,45) = 9.52, p = 0.003, η2 = 0.18; significant Sex × Arousal interaction: F(1,45) = 6.57, p = 0.014, η2 = 0.13, confirmed by post hoc tests with p b 0.05). The analysis of immediate free recall performance in females (which included menstrual stage and condition as factors) revealed no significant main effects or interactions.
3.4.2.2. Percent of immediate recall. Because we observed some group differences in immediate free recall performance, we also analyzed the delayed free recall performance as a percent of immediate free recall (i.e., (delayed recall/immediate recall) ∗ 100). This analysis corroborated our findings from the raw data analysis by revealing that stressed participants recalled more words overall than non-stressed participants (significant effect of condition: F(1,48) = 7.90, p = 0.007, η2 = 0.14). Overall, participants recalled more negative words than positive and neutral words (significant effect of valence: F(2,96) = 7.96, p = 0.001, η2 = 0.14). Participants also recalled more arousing words than nonarousing words, particularly when they were negative or neutral in valence (significant effect of arousal: F(1,48) = 6.88, p = 0.012, η2 = 0.13; significant Valence × Arousal interaction: F(2,96) = 9.46, p b 0.001, η2 = 0.17, confirmed by post hoc tests with p b 0.05). The analysis of delayed free recall performance expressed as a percent of immediate free recall in females revealed that the effects observed when analyzing the raw data remained significant. Stressed females recalled more words overall than non-stressed females (significant effect of condition: F(1,21) = 7.80, p = 0.011, η2 = 0.27), and more importantly, the three-way Condition × Arousal × Menstrual Stage interaction described above remained significant, F(1,21) = 5.30, p = 0.032, η2 = 0.20, confirmed by post hoc tests with p b 0.05. 3.4.3. Delayed recognition (see Fig. 4) There were no significant differences between the recognition performance of stressed and non-stressed participants (no significant effect of condition: F(1,48) = 1.38, p = 0.247, η2 = 0.03). Participants did recognize more arousing than non-arousing words when they were negative or neutral in valence (significant Valence × Arousal interaction: F(2,96) = 7.65, p = 0.001, η2 = 0.14, confirmed by post hoc tests with p b 0.05). The analysis of delayed recognition in females revealed no significant main effects or interactions.
3.4.2. Delayed free recall 4. Discussion 3.4.2.1. Raw data. Stressed participants recalled more words overall than non-stressed participants (significant effect of condition: F(1,48) = 6.87, p = 0.012, η2 b 0.13). Overall, participants recalled more negative and neutral words than positive words (significant effect of valence: F(2,96) = 14.30, p b 0.001, η2 = 0.23). Participants also recalled more arousing words than non-arousing words, particularly when they were negative or neutral in valence (significant effect of arousal: F(1,48) = 27.71, p b 0.001, η2 = 0.37; significant Valence × Arousal interaction: F(2,96) = 4.97, p = 0.009, η2 = 0.09, confirmed by post hoc tests with p b 0.05). The analysis of raw delayed free recall performance in females revealed that stressed females recalled more words overall than nonstressed females (significant effect of condition: F(1,21) = 13.84, p = 0.001, η2 = 0.40). Overall, female participants recalled more positive and neutral words than negative words (significant effect of valence: F(2,42) = 8.31, p = 0.001, η2 = 0.28) and more arousing words than non-arousing words (significant effect of arousal: F(1,21) = 6.97, p = 0.015, η2 = 0.25). More interestingly, stress exerted differential effects on emotional memory in females, depending on stage of the menstrual cycle. Specifically, stress enhanced memory for arousing words, relative to non-arousing words, when females were in the follicular stage, while enhancing memory for non-arousing words, relative to non-stressed female memory for non-arousing words, when in the luteal phase. Non-stressed females exhibited greater memory for arousing words, relative to non-arousing words, when in the luteal, but not follicular, phase (significant effect of menstrual stage: F(1,21) = 9.88, p = 0.005, η2 = 0.32; significant Arousal × Menstrual Stage interaction:
Most studies examining the effects of post-learning stress on longterm memory have reported facilitative effects (Beckner et al., 2006; Cahill et al., 2003; Preuss & Wolf, 2009). However, there is some research in this literature reporting no effects or an impairment of long-term memory (e.g., Trammell & Clore, 2014). The purpose of the present study was to examine further the effects of post-learning stress on long-term memory and to explore whether sex and menstrual cycle
Fig. 4. Recognition memory during the 24-h test. No significant group effects were observed for recognition memory. Data are expressed as means ± SEM.
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stage influenced such effects. We found that post-learning stress enhanced 24-h free recall, even when memory performance was expressed as a percent of immediate free recall performance. This enhancement was independent of sex, as well as the valence and arousal level of the words. We also found that emotional memory in females was differentially affected by post-learning stress, depending on which stage of the menstrual cycle they were experiencing at the time of stress. Specifically, stress enhanced memory for arousing words, relative to non-arousing words, in females who were in the follicular phase and enhanced memory for non-arousing words, relative to nonstressed female memory, in females who were in the luteal phase. Together, our results replicate the majority of post-learning stress research and extend these findings to implicate female hormonal mechanisms in such effects. 4.1. Neurobiological mechanisms of post-learning stress We purposely induced post-learning stress as close in time as possible to word list learning in order for the stressor to be experienced in the context of learning. That we observed an enhancement of long-term memory as a result of this manipulation is consistent with previous research revealing that the timing of stress relative to learning is important in dictating the types of effects observed on long-term memory (Diamond et al., 2007; Quaedflieg, Schwabe, Meyer, & Smeets, 2013; Zoladz, Clark, et al., 2011; Zoladz et al., 2014). Physiologically, stress results in the activation of two major systems, the sympathetic nervous system and the hypothalamus-pituitary-adrenal axis. Activation of these two systems results in increased adrenergic and corticosteroid activity, respectively, which can exert biphasic effects on cognitive processing. When the stress-induced increases in adrenergic and corticosteroid activity converge in time with learning, research suggests that these mechanisms, coupled with amygdala activity, facilitates memory processing in structures such as the hippocampus, which enables phenomena such as flashbulb memories to occur (Diamond et al., 2007; Schwabe, Joels, Roozendaal, Wolf, & Oitzl, 2012). Consistent with this idea, we found that CPT exposure in the present study resulted in significant increases in cardiovascular measures, indicators of autonomic and adrenergic activity, and salivary cortisol concentrations. These effects were produced with a relatively brief (3-min) stressor, suggesting that even brief stress of a particular intensity can promote memory consolidation. The CPT is a physiologically-based stressor, as it produces a stress response via cold-induced pain. However, the effects of the CPT on cortisol levels, learning and memory are comparable to those induced by other stressors, such as psychosocial stress (e.g., Trier Social Stress Test).
can impact the acquisition and extinction of emotional memory. Felmingham, Fong, and Bryant (2012) reported that heightened progesterone, which typically peaks in the mid-late luteal phase, is positively correlated with memory recall of threatening images and may mediate cortisol response. Similarly, a study examining sex differences in acoustic startle response found that females in the late luteal phase exhibited enhanced startle response compared to both males and females in the follicular phase (Armbruster, Strobel, Kirschbaum, & Brocke, 2014). One hypothesis put forth to explain progesterone's enhancing effects for arousing material is that heightened progesterone levels increase sensitivity to sources associated with threat or danger (Conway et al., 2007; Derntl et al., 2008). There appears to be a plausible interaction occurring between progesterone and cortisol, such that in the absence or diminished levels of one hormone, the heightened levels of the opposing chemical acts in such a way as to mimic the effects of the other. This could explain why females in the follicular phase (low progesterone) exhibited greater delayed recall for arousing words only in the stressed condition (high cortisol), while females in the luteal phase (high progesterone) had enhanced delayed recall for arousing material when in the control condition (low cortisol). 4.3. Caveats Although the present results indicate that post-learning stress might differentially influence females depending on menstrual stage, it is important to emphasize that the analyses performed on such measures here were exploratory in nature and consisted of relatively small sample sizes (as indicated in Fig. 3). Furthermore, we did not measure the levels of progesterone or estradiol in females, and our measure of menstrual stage was based on self-report. Therefore, although our findings are suggestive of a link between menstrual cycle activity and stress-memory interactions, further research is clearly warranted to substantiate these claims. 4.4. Conclusions We have provided additional evidence that post-learning stress facilitates long-term memory, an effect that, at least in the present study, is independent of the emotional nature of the learned information. We have also shown that females are differentially affected by post-learning stress depending on stage of the menstrual cycle. These findings further validate post-learning stress-induced enhancements of long-term memory and implicate female sex hormones in such effects. References
4.2. Menstrual cycle interactions with delayed recall An extensive review of the stress-memory literature reveals that females have been largely underrepresented in this research. This is especially surprising and concerning, as females are significantly more likely to develop stress-related psychological disorders, such as PTSD and major depression, that involve pathological stress-memory interactions. Although recent studies have begun to report sex differences more frequently, many researchers still fail to include measures that could help explain sex differences in stress-memory interactions. By collecting data from female participants about menstrual cycle activity, we found that menstrual cycle stage was associated with emotional memory under baseline and stress conditions. Specifically, nonstressed females exhibited greater memory for arousing words, relative to non-arousing words, when they were in the luteal phase. In contrast, stressed females demonstrated greater memory for arousing words, relative to non-arousing words, when they were in the follicular phase. Interestingly, stressed females in the luteal phase exhibited enhanced memory, relative to non-stressed females, for non-arousing words. Recent research has shown that both estradiol and progesterone
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