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malfunction (n = 2), participant failure to follow directions (n = 2), poor physiology recordings

98

resulting from weak electrode adhesion or other recording artifacts (n = 6), or a lack of SCR response to the US as defined by failure to exhibit positive response magnitudes (greater than 0) to the US during the SIR (n = 7). Significantly more placebo-treated participants were lost in attrition than nicotine-treated participants (4 (4.8%) in the nicotine group and 13 (15.4%) in the placebo group; p < 0.05). Sixty-seven participants were included in the final analysis (nnicotine = 29, 10 female, average age = 19.3 ± 1.46; nplacebo = 38, 12 female, average age = 19.3 ± 1.22).

There were no significant differences between treatment groups in age (t(62) = 0.27, p = 0.79), hours since last nicotine use (Mnicotine = 14.0 ± 10.5, Mplacebo = 11.7 ± 8.28; t(62) = 0.99, p = 0.32), or weekly nicotine consumption (Mnicotine = 30.2 ± 42.0 Mplacebo = 18.7 ± 18.6; t(62) = 1.47, p = 0.15). Additionally, there were no treatment differences on any pre-test questionnaire

measures (Table 13): Fagerstrom (Mnicotine = 2.93 ± 2.70, Mplacebo = 2.97 ± 2.70; t(62) = -0.06, p = 0.95), DASS Stress (Mnicotine = 7.93 ± 7.29, Mplacebo = 6.94 ± 6.15; t(62) = 0.59, p = 0.56), DASS Anxiety (Mnicotine = 8.36 ± 7.14, Mplacebo = 6.83 ± 6.56; t(62) = 0.89, p = 0.38), DASS Depression (Mnicotine = 5.50 ± 5.77, Mplacebo = 6.22 ± 8.79; t(62) = -0.38, p = 0.71), STAI (Mnicotine = 27.5 ± 10.7, Mplacebo = 27.6 ± 10.6; t(62) = -0.06, p = 0.96). However, the nicotine group scored significantly higher than placebo group on the Nausea subscale of the post-test Simulator Sickness Questionnaire (Mnicotine = 4.00 ± 3.04, Mplacebo = 2.65 ± 2.31; t(62) = 2.04, p = 0.05;

Table 13). Therefore, nausea was used as a covariate in exploratory analyses.

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Questionnaire Measure Users (M ± SD)

Non-users

(M ± SD) t df p

Fagerstrom 2.93 ± 2.70 2.97 ± 2.70 -0.06 62 0.95 DASS Stress 7.93 ± 7.29 6.94 ± 6.15 0.59 62 0.56 DASS Anxiety 8.36 ± 7.14 6.83 ± 6.56 0.89 62 0.38 DASS Depression 5.50 ± 5.77 6.22 ± 8.79 -0.38 62 0.71

STAI 27.5 ± 10.7 27.6 ± 10.6 -0.06 62 0.96

Nausea 4.00 ± 3.04 2.65 ± 2.31 2.04 62 0.05

Table 13. No significant differences between nicotine use status on pre-test questionnaire

measures. Nicotine users scored significantly higher than non-users on post-test nausea measure (t(62) = 2.04, p = 0.05).

Acquisition session: To examine the effect of treatment on the acquisition of conditioned fear across both acquisition sessions, SCR data were analyzed using a 2-factor Trial (1-12) X Stimulus Type (CS+, CS-) repeated measures ANOVA with Treatment (nicotine, placebo) as the between-subjects factor. Analysis yielded significant Stimulus Type (F(1, 65) = 12.8, p = 0.001) and Trial main effects (F(5.88, 382.0) = 4.86, p < 0.0001). As expected, there was a larger mean SCR magnitude to the CS+ (M = 0.32 ± 0.34) compared to the CS- (M = 0.20 ± 0.34),

demonstrating conditioned fear responding. Adjusting alpha to 0.01 to account for multiple tests, a series of paired t-tests comparing SCR magnitude for CS+ versus CS- trial pairings revealed that consistent differential fear responding emerged during the last trial of the second acquisition session (t(66) = 6.36, p = 0.0001), but not between earlier presentations of these stimuli (t(66)

=0.95, p = 0.35; Figure 10). None of the effects involving the Treatment factor were significant, indicating that nicotine did not differentially affect the rate of acquisition relative to placebo (F(1, 65) = 0.60, p = 0.44).

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Figure 10. Differential fear responding emerged consistently during the last trial of the second delay cued acquisition session, but not during the first trial (t(66) =0.95, p = 0.35).

Contextual test session: Paired sample t-tests were first conducted to examine our a priori hypothesis that each treatment group, respectively, would demonstrate greater fear to the virtual context previously paired with a shock (CS+) relative to the safe context (CS-). We found that nicotine-treated participants demonstrated significantly greater SCRs to the CS+ relative to the CS- averaged across trials (MCS+= 0.46 ± 0.78, MCS- = 0.13 ± 0.32; t(28) = 2.32, p = 0.03).

Placebo-treated participants did not show conditioned fear to the CS+ relative to the CS- (MCS+ = 0.28 ± 0.60, MCS- = 0.18 ± 0.32; t(38) = 1.43, p = 0.16).

To test whether nicotine enhances contextual fear conditioning, SCR data were analyzed using a Trial (2) X Stimulus Type (CS+, CS-) repeated measures ANOVA with Treatment (nicotine, placebo) as the between-subjects factor. The ANOVA produced significant main effects of Stimulus Type (F(1, 65) = 8.45, p = 0.005) and Trial (F(1, 65) = 4.52, p = 0.04), as well as a Trial x Treatment (F(1, 65) = 4.08, p = 0.05; Figure 11). As expected, mean SCR magnitude was greater for CS+ (M = 0.36 ± 0.69) relative to CS- trials (M = 0.16 ± 0.31), indicating that participants exhibited greater conditioned fear to the context previously paired

-0.20 -0.10 0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70

Trial 1 Trial 12

SCR

Skin Conductance Responses between the First and Last Acquisition sessions

CS+

CS-101

with the aversive shock relative to the neutral, shock-free context. Raw SCR scores were converted to difference scores for post-hoc analysis of the Trial x Treatment interaction by subtracting SCRs to the CS- from those to the CS+. Independent sample t-tests between SCR difference scores revealed that compared to placebo, SCR magnitude was significantly enhanced by nicotine during the first trial of the context test (Mnicotine = 0.33 ±0.79,Mplacebo = 0.04 ± 0.57;

t(65) = 1.73, p = 0.04, one-tailed; Figure 11). There were no significant differences in contextual fear between treatments for the second trial (Mnicotine = 0.32 ±0.93,Mplacebo = 0.17 ± 0.79; t(65) = 0.76, p = 0.45; Figure 11). To ensure that nausea did not disrupt fear conditioning, a multivariate ANOVA was conducted with nausea as a covariate, SCR difference score as the dependent variable and treatment as the between subjects factor. When controlling for nausea, nicotine significantly enhanced conditioned fear on trial 1 (F(1, 62) = 3.76, p = .05), but had no effect on trial 2 (F(1, 62) = 0.086, p = 0.77), or the average across trials (F(1, 62) = 1.66, p = 0.20).

Figure 11. Relative to placebo, nicotine significantly enhanced fear to the context during the first trial of the context test session (t(65) = 1.73, p = 0.04, one-tailed). There were no

significant differences between treatments in contextual fear for the second trial (t(65) = 0.76, p

= 0.45).

-0.10 0.00 0.10 0.20 0.30 0.40 0.50 0.60

Trial 1 Trial 2

SCR Difference Score

Skin Conductance Difference Scores between Treatments for Each Trial of the Context Test

Nicotine Placebo

102

Delay cued test session: Paired sample t-tests were first conducted to examine our a priori hypothesis that each treatment group, respectively, would demonstrate greater fear to the cue previously paired with a shock (CS+) relative to the safe cue (CS-). Nicotine-treated participants demonstrated significantly greater SCRs to the CS+ cue relative to the CS- cue averaged across trials (MCS+ = 0.46 ± 0.67, MCS- = 0.20 ± 0.31; t(28) = 2.20, p = 0.04; Figure 12).

Placebo-treated participants, however, did not exhibit conditioned fear to the CS+ cue (MCS+ = 0.51 ± 0.53, MCS- = 0.35 ± 0.80; t(37) = 1.15, p = 0.26; Figure 12). Examining the effect of nicotine on cued fear conditioning relative to placebo, an ANOVA with repeated measures revealed a significant main effect of Stimulus Type (F(1, 65) = 4.70, p = 0.03; Figure 12) where responses to the CS+ (M= 0.49 ± 0.60) were significantly greater than to the CS- (M= 0.29 ± 0.63). However, none of the effects involving the Treatment factor were significant, indicating that there were no differences between nicotine and placebo on conditioned fear to the cue (F(1, 65) = 0.60, p = 0.44; Figure 12).

Figure 12. No significant differences in delay cued fear response between nicotine or placebo treatments (F(1, 65) = 0.60, p = 0.44).

0.0 0.1 0.2 0.3 0.4 0.5 0.6

Nicotine Placebo

SCR

Skin Conductance Responses Averaged Across Trials during the Delay Cued Test Session

CS+

CS-103

Correlations between Questionnaire Metrics and Fear Conditioning: Correlations were conducted to examine potential relationships between delay cued or contextual fear and any of the questionnaire metrics. We hypothesized that individuals with greater levels of nicotine dependence would be more likely to demonstrate nicotine-enhanced contextual FC. Looking across all participants, no significant correlations were found between the Fagerstrom Test for Nicotine Dependence (FTND) and context difference scores during the first (r = -0.62, p = 0.62) nor second trials (r = -0.34, p = 0.79). When specifically examining individuals who received nicotine, no significant relationships were found between FTND and context FC during the first (r = -0.21, p = 0.29) nor second trials (r = -0.15, p = 0.45).

We also hypothesized that individuals who scored highly on measures of stress and anxiety would be more likely to demonstrate learned fear. Examining all participants or those who were administered nicotine, no relationships were found between any subscale (anxiety, depression, or stress) of the Depression Anxiety Stress Scales and contextual FC during any trial (p > 0.05). However, in examining all participants, there was a significant positive correlation between the Trait Anxiety subscale of the State Trait Anxiety Inventory and delay cued fear when averaged across trials (r = -0.25, p = 0.04). The Trait Anxiety subscale defines a general tendency to perceive situations as threatening, which differs from State Anxiety which assesses temporary anxiety experienced in specific situations or in the present moment. Furthermore, when specifically examining individuals who received nicotine, delay cued FC significantly correlated with both Trait Anxiety when averaged across trials (r = -0.51, p = 0.01) and State Anxiety during the first CS presentations (r = -0.38, p = 0.05).

4.3.3. Trace Cued and Contextual Fear Conditioning: Of the eighty-three undergraduates