3.1. Subject characteristics
One hundred and thirteen participants were recruited; of those, 5 (1 PTSD, 4 No-PTSD) were excluded due to incomplete trauma histories, 2 (1 PTSD, 1 No-PTSD) were excluded as outliers as their DST ACTH values were more than 3 standard deviations above the mean, and 6 (all controls) were excluded due to a negative assay for dexamethasone on the DST visit, indicating non-compliance. This resulted in a final sample of 100 participants: 54 who were fear conditioned prior to taking the dexamethasone treatment (No-TMT group) and 46 who were fear conditioned 10 h after taking the dexamethasone treatment (TMT group), see . shows the demographic and clinical information of the subjects across the different groups, as well as their trauma history. The PTSD and No-PTSD groups were matched on sex, age and race. Furthermore, the treatment groups (TMT, No-TMT) did not differ on any demographic or clinical variables (see ).
Table 1 Demographic, trauma history, and PTSD symptom data for the study sample. The participants in the two treatment conditions (TMT, No-TMT) do not differ on any of these variables; the two diagnostic groups (PTSD, No-PTSD control) are matched on demographic (more ...)
We did not exclude subjects who were taking psychotropic medication. Of the 100 participants, 3 (1 PTSD, 2 No-PTSD) used benzodiazepines (χ2 = 0.01, ns), 11 (6 PTSD, 5 No-PTSD) used selective serotonin reuptake inhibitor (SSRI) antidepressants (χ2 = 2.60, ns), and 6 (4 PTSD, 2 No-PTSD) used anti-psychotics (χ2 = 3.72, p = 0.07). None of the medications had an effect on either baseline or fear-potentiated startle.
3.2. Clinical assessments
As shown in , all PTSD subjects had significantly greater levels of childhood (F(1,99) = 24.61, p < 0.0001) and adult (F(1,99) = 17.91, p < 0.0001) trauma compared to the traumatized No-PTSD controls. In addition, PTSD subjects had significantly higher symptoms of depression compared to No-PTSD subjects, F(1,93) = 28.17, p < 0.001. Given the potential confounding effects of these variables on the study results, these variables were used as covariates in analyses between diagnostic PTSD and No-PTSD groups. As expected, the subjects who met current criteria for PTSD had significantly higher total PSS scores, F(1,99) = 145.33, p < 0.0001, than traumatized controls without PTSD. Furthermore, the PTSD subjects had significantly greater symptom severity in each of the three PTSD symptom clusters, i.e., re-experiencing (F(1,99) = 60.31, p < 0.0001), avoidance (F(1,99) = 129.89, p < 0.0001), and hyper-arousal (F(1,99) = 77.37, p < 0.0001). It is important to note, however, that neither trauma history, depression, nor PTSD symptoms differed between dexamethasone treatment groups.
3.3. Dexamethasone suppression
Of the 100 participants with startle and clinical data, baseline cortisol and ACTH levels were available for 90 participants (30 PTSD, 60 No-PTSD), and DST cortisol and ACTH levels were available for 71 participants (27 PTSD, 44 No-PTSD). Missing data were primarily due to: (1) participants not showing up for follow-up appointments, (2) difficulties in drawing blood from participants, despite extensive experience by the nurses, or (3) errors in the hormone assays. shows the baseline and DST cortisol levels in each group. A RM ANOVA of baseline and DST cortisol levels found a significant main effect of visit, F(1,68) = 160.71, p < 0.001, but no effect of diagnosis and no interaction effect of DST by diagnostic group. Because we wanted to confirm significant dexamethasone suppression of cortisol in each group, we repeated the analysis separately in each group, and found significant effects of visit in PTSD subjects, F(1,25) = 98.36, p < 0.001, and No-PTSD subjects, F(1,43) = 91.92, p < 0.001. However, there were no group differences in baseline cortisol, F(1,89) = 0.30, ns, DST cortisol levels, F(1,70) = 1.52, ns, or change scores from baseline to DST cortisol, F(1,70) = 0.04, ns. These data suggest that we did not find hypersuppression of cortisol in the PTSD group in this sample.
Cortisol levels across visits (baseline and DST) and diagnostic groups. Dexamethasone effectively suppressed cortisol levels in both groups.
A RM ANOVA of ACTH levels also showed a significant main effect of visit, F(1,64) = 9.04, p < 0.01, but no significant effect of diagnostic group, and no interaction effect of group by DST visit. A RM ANOVA performed within each diagnostic group separately found a significant reduction in ACTH levels in the No-PTSD subjects, F(1,39) = 6.89, p = 0.01, but only a trend for a reduction in PTSD subjects, F(1,25) = 2.99, p < 0.1. Between-group analyses of baseline and DST ACTH did not show significant group differences, F(1,85) = 0.18, ns, and F(1,66) = 0.05, ns, respectively. The change score from baseline to DST ACTH levels also did not differ between groups, F(1,66) = 0.03, ns. As in the case of cortisol, this study did not find ACTH hypersuppression in PTSD.
3.4. Fear-potentiated startle (FPS)
shows startle magnitude to the noise alone, CS+, and CS− trials across each block and dexamethasone treatment condition for each diagnostic group. Within each block there was a significant effect of trial type (block 1, F(2,192) = 30.41, p < 0.001; block 2, F(2,192) = 35.16, p < 0.001; block 3, F(2,192) = 28.45, p < 0.001). In the first two blocks there were no other significant main effects or interaction effects. In the third block, there was a significant 3-way interaction, F(2,192) = 4.51, p = 0.01. Follow-up analyses indicated that PTSD subjects had significantly higher startle magnitude than the controls on the CS+ and CS− trials, but not the NA trials. In all blocks, regardless of diagnostic or treatment group, all subjects showed significant fear conditioning to the CS+ compared to NA (see ). In order to reduce the impact of individual differences in baseline (NA) startle magnitude, we calculated a value for fear-potentiated startle; FPS was assessed using the difference score between baseline startle and startle in the presence of the conditioned stimuli. Although there were no significant differences in baseline startle between groups, given that the NA startle was higher on average in the PTSD group than the control group (see ), we performed a bivariate correlation between NA startle and the difference score (i.e., FPS) for the CS+ and CS− in the PTSD group. Neither score was correlated with NA startle (CS+, r = 0.12, ns; CS−, r = −0.08, ns).
Startle magnitude across groups and dexamethasone (DEX) treatment conditions across the three blocks of conditioning.
Fear-potentiated startle data were analyzed across 3 blocks of conditioning using a 4-way mixed ANOVA with block (3 levels) × trial type (CS+, CS−) × group (PTSD, No-PTSD) × dexamethasone treatment (TMT, No-TMT) and childhood and adult trauma history as covariates. This analysis revealed a significant 4-way interaction, F(2,192) = 4.79, p = 0.009. Follow-up analyses within each block of conditioning indicated a significant 3-way interaction of trial type -× group × treatment in the 3rd block of conditioning, F(21,94) = 3.91, p = 0.05. There were no significant interactions or main effects of group or dexamethasone treatment in the earlier conditioning blocks. In order to assess group differences in differential conditioning (i.e., FPS to CS+ vs. CS−) without the dexamethasone treatment, we examined the effect of trial type in the last block of conditioning with a 2-way mixed ANOVA with trial type (CS+, CS−) × group (PTSD, No-PTSD). This analysis revealed significantly higher FPS to the CS+ than the CS− across both groups, F(1,52) = 13.25, p = 0.001. There was no interaction effect of trial type and group; however, there was a significant main effect of group, with PTSD having higher FPS to both trial types, F(1,52) = 8.19, p = 0.006. We repeated the same 2-way ANOVA of trial type (CS+, CS−) × group (PTSD, No-PTSD) with dexamethasone treatment and found that there was still a significant, albeit not as strong, effect of trial type, F(1,44) = 5.32, p = 0.03. Again, there was no interaction of trial type and group, and there was also no longer a significant main effect of group, F(1,44) = 0.11, ns.
3.5. Dexamethasone effect on FPS
In order to examine the effect of dexamethasone treatment on each trial type, we performed a univariate analysis of FPS to the CS+ in the 3rd block of conditioning, with diagnostic group (PTSD, No-PTSD) and dexamethasone treatment (TMT, No-TMT) as between-group variables, with and without trauma history and depression as covariates. The analysis was repeated with the CS− trial type. The analysis of the CS+ trial type revealed a 2-way interaction of diagnosis by treatment, F(1,99) = 6.83, p = 0.01. Adding trauma history as covariates to the model increased the interaction effect, F(1,99) = 7.54, p = 0.007, which was further strengthened by adding depression to the ANCOVA, F(1,93) = 8.07, p = 0.006. This interaction was followed by comparing diagnostic groups separately for each treatment, and by comparing treatment conditions within each diagnostic category. The first analysis found significantly greater fear potentiation to the CS+ in PTSD subjects compared to No-PTSD subjects (ANOVA F(1,53) = 8.08, p = 0.006; ANCOVA with trauma, F(1,53) = 7.52, p = 0.008; ANCOVA with trauma and depression, F(1,50) = 4.33, p = 0.04) when subjects were tested at baseline cortisol levels (within the No-TMT condition, see ). However, in the TMT condition, the group difference was no longer present (ANOVA F(1,45) = 0.77, ns; ANCOVA with trauma, F(1,45) = 0.28, ns; ANCOVA with trauma and depression, F(1,42) = 0.39, ns). When we compared the dexamethasone treatment conditions within each diagnostic group, there was a significant treatment effect in PTSD subjects, with dexamethasone reducing fear-potentiated startle to the CS+ (F(1,32) = 4.00, p = 0.05). However, there was no significant treatment effect in the No-PTSD control group (F(1,66) = 1.94, ns).
Figure 3 (A) Fear-potentiated startle to the CS+ and CS− in the last block of conditioning across treatment conditions and diagnostic groups. The significant group difference in the no dexamethasone condition was eliminated in the dexamethasone condition. (more ...)
In an effort to standardize the startle data, we converted the raw startle magnitude data to T
scores according to the methods used in the NIMH Center for Emotion and Attention (e.g. McTeague et al., 2010
). shows the above data presented as T
scores, with the individual’s average NA startle magnitude set as the mean, i.e., T
= 50. Using the T
scores in the same analyses as above, we replicated the effect of dexamethasone treatment on the group differences in fear potentiation to CS+ trials. There was significantly greater fear potentiation to the CS+ in PTSD subjects compared to No-PTSD subjects, F
(1,53) = 4.98, p
= 0.03 (covariates did not change the results), when No-TMT subjects were tested (no dexamethasone condition, see ). However, in the TMT condition, the group difference was no longer present, F
(1,45) = 0.40, ns. Furthermore, when we compared the dexamethasone treatment conditions within each diagnostic group, the analyses of T
scores did not reveal a significant effect of dexamethasone in either group.
The analysis of the CS− trial type showed a significant main effect of diagnostic group, with higher fear-potentiated startle in the PTSD subjects compared to No-PTSD subjects (ANOVA F(1,99) = 3.99, p = 0.05; ANCOVA with trauma, F(1,99) = 4.35, p = 0.04). However, this was not the case when depression was added to the model (ANCOVA with trauma and depression, F(1,93) = 2.23, ns). There was no effect of treatment and no interaction effect of group by treatment. The analysis was repeated with T scores of the startle magnitude to the CS− trials; there were no significant effects of group or treatment.
3.6. Contingency awareness
Response pad data were available for 59 participants (19 PTSD, 40 controls, the reduction in sample size was due to computer error); these data were analyzed with a RM ANOVA comparing US expectancy responses to CS+ and CS− across blocks of conditioning, with diagnosis and treatment as between-group factors. This analysis revealed a significant main effect of trial type, F(1,53) = 130.27, p < 0.0001, but no main effects of dexamethasone treatment or diagnostic group. There was also a significant interaction effect of block and trial type, F(2,106) = 15.23, p < 0.001, showing that awareness increased over conditioning blocks. In order to better compare awareness with the startle data, we analyzed US expectancy specifically in the 3rd block of conditioning, with and without trauma history and depression as covariates. The results of the response keypad data showed that, across both groups, subjects understood the experimental contingencies demonstrating significant differential conditioning between the CS+ and CS− () (RM ANOVA F(1,55) = 117.88, p < 0.0001; RM ANOVA with trauma covariates F(1,53) = 9.39, p = 0.003; RM ANOVA with trauma and depression covariates F(1,49) = 4.88, p = 0.03). Furthermore, contingency awareness was not affected by dexamethasone. Analysis of US expectancy did not show main effects of diagnostic group, treatment, or interaction effects of the two factors. Although most participants demonstrated awareness, there were a few who did not (2 PTSD, 7 No-PTSD). Therefore, we analyzed the main findings in only the aware participants (defined as having a greater US expectancy on the CS+ than CS− on the final conditioning block). The results did not change, i.e., without dexamethasone PTSD subjects had exaggerated FPS to the CS+ compared to No-PTSD subjects, F(1,28) = 8.39, p = 0.007, while with dexamethasone there was no difference between groups, F(1,20) = 2.37, ns. These data suggest that PTSD symptoms did not affect cognitive awareness of the CS contingency and that awareness was also unaffected by dexamethasone.
Figure 4 Response keypad data for the CS+ and CS− trials in the last block of conditioning across dexamethasone treatment conditions and diagnostic groups. Expectancy of the US was significantly higher to the CS+ than CS− in every condition, but (more ...)
3.7. Cortisol levels and FPS
Given the significant suppression of cortisol with dexamethasone, we wanted to see whether cortisol levels were associated with FPS to the CS+. We performed correlations on baseline and DST cortisol levels in the total sample as well as separately for each diagnostic group. In the total sample, we did not find a correlation between baseline cortisol and FPS (r = 0.09, ns), or DST cortisol and FPS (r = −0.16, ns). However, when we examined each diagnostic group separately, we found a significant positive correlation in the PTSD group between baseline cortisol and FPS to the CS+ (r = 0.46, p = 0.01), see ; but not between DST cortisol and FPS (r = −0.01, ns). Neither association was significant in the No-PTSD group. These data suggest that HPA axis activity, represented by baseline cortisol, is associated with heightened fear in PTSD, and that the suppression of HPA function in the above studies may be correlated with normalization of fear responses.
Figure 5 Scatter plot of fear-potentiated startle responses to the CS+ and baseline cortisol levels in the two diagnostic groups. A positive correlation was found only in the PTSD group. Abbreviation: CS+ = conditioned stimulus reinforced with the airblast (reinforced). (more ...)