Estrogen and Serotonin on Changing Brain Chemistry: Complete Statistical Analysis

1. Trial at a Glance

This page covers the registry results for Estrogen and Serotonin on Changing Brain Chemistry (NCT01208324), a randomized na study. The primary outcome is Percent Change in BOLD Signal.

2. Clinical Question

The study aimed to examine how estrogen and serotonin relate to cognition, emotional processing, and brain activation in healthy menopausal women. Its primary outcome description focuses on verbal memory and asks whether the relation between estrogen therapy and tryptophan depletion is reflected in BOLD signal during brain imaging.

The stated aim also includes cognition and mood, and the study summary describes functional MRI during memory tasks. A secondary outcome concerns changes in verbal working memory mediated through the dorsolateral prefrontal cortex. These objectives provide context for the imaging outcome, but they do not establish that a particular brain region or cognitive function changed in a specific way.

3. Trial Design

The registry identifies this as a randomized, double-masked crossover study. In a crossover design, participants receive more than one condition in sequence, so comparisons may be made within a participant as well as across conditions. That is a general description of the design; the registry record does not report the analysis used to account for within-participant measurements, sequence, or period effects.

The study record lists estrogen patch, amino acids, placebo patch, and placebo pills as interventions. Its arm descriptions specify an estrogen patch regimen of Vivelle Dot® 0.10 - 0.15 mg/day for 8 weeks, and list amino-acid ingredients and quantities. Placebo pills are described as 31.5 g of lactose or microcellulose. The posted descriptions do not establish the full schedule or how each intervention was combined across every study period.

Adverse-event group labels give four sequences involving active or sham patch and active or sham tryptophan. Those labels help clarify that participants could be assigned to different orders of conditions, but they do not provide a complete account of washout periods, sequence allocation, or visit timing. Such details are not reported in the registry record.

The recorded enrollment is 47. The study status is completed, with a start date of 2010-03-04 and primary completion reported as 2018-11. The registry lists phase as NA. No further rationale for the enrollment, allocation procedures, masking implementation, or trial conduct is reported in the registry record.

The crossover structure matters when interpreting the posted group means. Separate means and confidence intervals do not reveal whether the investigators analyzed paired observations, adjusted for period or sequence, or used another model. The registry record reports zero analyses for the posted outcome information and does not give a direct treatment contrast. Therefore, the design label alone is not enough to reconstruct the primary statistical analysis.

4. Participant Flow and Accounting

The registry reports how participants moved through each stage of the study. All counts below are exactly as posted; missing counts are not inferred from other stages.

Before assignment: 2 participants included in the Placebo Patch/ Active then Sham group completed Test Day 1, but did not receive a patch. 1 participant included in the Placebo Patch/ Sham then Active group completed Test Day 1, but did not receive a patch.

Pre-randomization of Patch

Post-randomization of Patch

Reasons for not completing this period:

5. Baseline Characteristics

The table reproduces the baseline characteristics posted in the registry, which describe the participants analysed at baseline and help show whether the randomized groups were comparable.

Population: The participants that were analyzed completed all 4 test days. The study team was not permitted to collect or use the information collected from the 6 participants who did not complete up to test day 4, and so those participants are excluded from the baseline analysis population.

6. Primary Endpoint

The registry lists the primary outcome as Percent Change in BOLD Signal, measured over 8 weeks.

The primary endpoint is percent change in BOLD signal, reported as a mean with a 95% confidence interval and a timeframe of 8 weeks. BOLD, or blood-oxygen-level-dependent, signal is an imaging measure used in functional MRI. The registry description explains that it reflects changes in regional cerebral blood flow and is used to characterize regional brain activity.

According to that description, a positive BOLD signal marks increased regional blood flow and a negative signal marks decreased regional blood flow. A positive percent change means the signal increased between scans; a negative percent change means it decreased between scans. These signs describe direction of change in the measured imaging signal. They do not, on their own, indicate whether memory or another clinical function improved or worsened.

BOLD is used in this study because the investigators aimed to identify differences in brain activation during memory tasks under estrogen and tryptophan-depletion conditions. It provides an imaging outcome related to regional activity, rather than a direct measure of a participant’s memory performance. The study summary also mentions cognition, mood, and emotional processing, but the posted primary endpoint is the imaging measure.

The registry record does not report a direct estimate of the contrast between treatment conditions for this endpoint. The posted entries are labeled High Ace and Low Ace; their definitions and relation to the intervention sequences are not explained in the outcome information. Their means should therefore be read as the entries’ reported group summaries, not as a fully specified treatment effect.

7. Why No Re-analysis Is Shown

8. Statistical Interpretation

9. Statistical Methods Explained

The primary outcome is continuous: it is a mean percent change in BOLD signal. The registry identifies the reported summary as a mean and gives a 95% confidence interval. This is a descriptive presentation of the outcome, but the record does not report an inferential analysis comparing the intervention conditions.

What does a confidence interval communicate?

In general, a confidence interval expresses uncertainty around an estimated quantity under the method used to construct it. A narrower interval indicates greater precision for that estimate than a wider interval, all else being equal. A 95% confidence interval is not a range containing 95% of individual participants’ values, nor does it by itself give the probability that a particular treatment effect is true. Its interpretation depends on the statistical method and assumptions.

The posted intervals are attached to the two reported means, not to a contrast between them. A treatment comparison would ordinarily require an estimate of the difference or another prespecified contrast, together with its uncertainty and an appropriate test or interval. The registry record does not report such a contrast. It is therefore not possible to use the listed means and intervals alone to recover the sponsor’s comparative analysis or to conclude whether the primary question was statistically supported.

Why does the crossover design matter?

In general, crossover studies collect measurements under different conditions from the same participants. This can make within-participant comparisons informative because each participant can serve as their own reference. It also creates analytical considerations: measurements from one participant are related, and outcomes can depend on period, order, or carryover from an earlier condition. A valid analysis needs to account for the actual design and data structure.

The registry identifies randomization, double masking, and a crossover model. It does not report the statistical model, how sequence or period was handled, whether a washout was used, or how any carryover concerns were assessed. It would therefore be unjustified to assume that the analysis used a paired test, a mixed model, or any other particular method. These are general methodological possibilities, not claims about the investigators’ analysis.

What can the posted summaries establish?

The two means give the direction and magnitude of the recorded group summaries in the endpoint’s units. Their confidence intervals show uncertainty in those means as reported. Neither item directly tests the stated interaction between estrogen therapy and tryptophan depletion. The record also lists zero analyses for the outcome information and includes a note that values are missing or not estimable for a two-arm comparison of the continuous endpoint. That note is consistent with the absence of a direct comparison in the posted results.

The record does not report a p-value, model coefficient, standard error, treatment-effect estimate, or adjustment variables for the primary outcome. It does not state whether the interval construction used a particular distributional assumption or whether results were adjusted for baseline measurements. Without these details, readers should not try to reconstruct an inferential result from the group summaries.

How should planning calculations be understood?

The registry’s planning description characterizes sample-size illustrations as based on the observed effect, with a two-sided significance level of 0.05, equal allocation, and a normal approximation. It describes the effect to detect as a stated fraction of the observed effect. Such calculations are planning illustrations, not evidence about the probability that this completed study detected an effect. They are not observed power and do not replace a reported analysis. The registry record does not provide the actual illustration values in the information available for this interpretation.

In general, planning estimates depend on assumptions about the effect to detect, variability, allocation, significance level, and analytical approach. Illustrations based on an observed effect can be especially uncertain because the observed effect itself is an estimate. The record does not report the original design calculation or its assumptions, so the planning description should not be mistaken for the rationale used to choose enrollment.

Because this is a continuous endpoint, the meaning of an estimate depends on its scale, variability, and uncertainty. The BOLD percentage-change convention gives a sign interpretation, but not a clinical threshold for importance. The registry record does not report a threshold for a meaningful difference in BOLD signal or link the numeric changes to an established change in memory performance.

10. Multiplicity, Missing Data and Analysis Populations

The registry lists 2 outcome measures and 1 posted secondary outcome. The secondary outcome is described as evaluating changes in verbal working memory mediated through the dorsolateral prefrontal cortex. The record reports zero analyses for the posted outcome information. It does not report a multiplicity strategy, such as a hierarchy of outcomes or an adjustment for testing several outcomes.

Multiplicity matters in general when several outcomes, time points, subgroups, or contrasts are examined: the chance of at least one apparently unusual result can increase as the number of tests grows. This general concern does not establish how many tests the investigators conducted. The registry record does not report the complete analysis set, all comparisons considered, or whether any multiplicity adjustment was used. No conclusion about a correction—or its absence in the actual analysis—can be drawn from the registry entries alone.

The outcome information does not report missing-data counts, reasons for missingness, imputation, or sensitivity analyses. It does not identify an intention-to-treat, per-protocol, or other analysis population for the primary endpoint. Although the registry records enrollment of 47 and endpoint group sizes of 16 and 24, it does not explain how participants moved from enrollment to the reported outcome summaries. Those figures should not be treated as a participant-flow account.

The registry record also does not state whether all randomized participants completed each crossover condition or contributed measurements at each scan. In general, missing observations can affect precision and can bias estimates if the reasons for missingness are related to the outcome. Without reporting on completeness and the analysis population, the extent or direction of any such effect cannot be assessed.

11. Safety Results

The registry reports serious adverse events as affected participants divided by participants at risk.

Deaths

Other (non-serious) adverse events

Participants with at least one non-serious event at or above the registry's 0% reporting threshold.

The serious adverse-event entries are presented by sequence group. For Active Patch: Active Tryptophan, Then Sham Tryptophan, the registry reports 14 at risk and 0 affected for serious events, 14 at risk and 0 affected for other events, and 0 deaths. For Active Patch: Sham Tryptophan, Then Active Tryptophan, the corresponding figures are 15, 0, 15, 0, and 0.

For Sham Patch: Active Tryptophan, Then Sham Tryptophan, the figures are 10 at risk and 0 affected for serious events, 10 at risk and 0 affected for other events, and 0 deaths. For Sham Patch: Sham Tryptophan, Then Active Tryptophan, the figures are 8 at risk and 0 affected for serious events, 8 at risk and 0 affected for other events, and 0 deaths. No serious-event terms are listed.

These entries describe what the registry reports for the listed groups; they do not establish that no adverse events occurred outside the recorded categories or reporting period. The record does not provide additional details about event definitions, collection procedures, follow-up duration, or whether all participants were assessed in the same way. Zero reported events should be read as a registry result, not as a guarantee of safety.

12. What the Result Does, and Does Not, Mean

The posted data establish that the registry contains mean percent changes in BOLD signal for entries labeled High Ace and Low Ace, with 95% confidence intervals. They show a negative reported mean for High Ace and a positive reported mean for Low Ace, using the endpoint’s stated sign convention. The data also give group sizes for these entries and sequence-specific adverse-event counts.

The posted data do not establish that estrogen therapy changed BOLD signal, that tryptophan depletion changed it, or that the two factors interacted. The registry does not report a direct treatment contrast, test statistic, p-value, or adjusted estimate for the primary question. The group labels are not defined in the endpoint information, so they cannot be reliably mapped to the specific intervention conditions from these outcome entries alone.

The BOLD outcome is an imaging measure that reflects changes in regional cerebral blood flow, as described in the registry. A change in that signal is not itself a demonstration of better or worse memory, mood, or emotional processing. The study’s aims include those functions, but the posted primary results do not provide a direct functional interpretation or a threshold for a meaningful imaging change.

The confidence intervals give uncertainty around the reported means under the methods used to calculate them, but the record does not describe those methods. They are not intervals for a difference between groups. Consequently, the intervals do not answer whether the groups differ or whether the treatment interaction is supported.

The safety entries report zero serious events, zero other events, and zero deaths in each listed group. This is a description of the registry’s entries, not evidence that an intervention is free of risk. The study’s results should be read with the limitations of the registry outcome reporting in mind, especially the absent comparative analysis, the undefined outcome-group labels, and the unreported analysis population and missing-data handling.

13. Why This Trial Matters Statistically

This trial is statistically notable because it combines an imaging endpoint with a randomized crossover design. The endpoint is a continuous measure of percent change in BOLD signal, while the study question concerns the relationship between estrogen therapy and tryptophan depletion. Answering that question requires an analysis that reflects both the multiple conditions and the repeated measurements expected in a crossover study.

The registry identifies randomization, double masking, and crossover structure, but the posted outcome information does not give the comparative model or a treatment contrast. This distinction illustrates why a design description and group-level summaries are not substitutes for an analysis matched to the trial question. The reader needs to know what was compared, how measurements from the same participant were handled, and how uncertainty in the contrast was quantified.

The endpoint also illustrates a broader interpretive issue in clinical research: a biological or imaging measure can be informative about a mechanism while remaining distinct from a direct measure of a clinical function. The registry explains the direction of BOLD signal change, but does not provide a threshold for meaningful change or a direct link between the posted values and memory performance.

Finally, the record distinguishes enrollment from endpoint summaries, but does not explain participant flow or missing outcome data. Clear reporting of analysis populations, missingness, outcome definitions, and model estimates is important for assessing precision and potential bias. The posted record is useful for identifying the study’s purpose and the reported descriptive results, while leaving the central comparative inference unreported.

14. Limitations

• The posted summary statistics are the only data these results rest on; patient-level data, full protocols and statistical analysis plans are not available.
• Recalculated results are simple, unadjusted calculations from posted summaries. They do not replace the sponsor's prespecified analysis.

The registry outcome entries label the reported groups High Ace and Low Ace, but do not define those labels or explain their relation to the randomized intervention sequences. This prevents a direct clinical interpretation of the two means as treatment-specific effects. The record also does not report an estimate or test of the primary treatment comparison or interaction.

The confidence intervals are identified as 95% intervals, but the calculation method is not described. The record does not report the model, assumptions, adjustment variables, or handling of repeated observations. As a result, the precision and inferential meaning of the group summaries cannot be fully evaluated.

Although enrollment is recorded as 47, the endpoint entries report group sizes of 16 and 24. The registry does not explain the relationship between these figures, the number randomized to each sequence, or the number contributing usable scans. No participant-flow information, missing-data account, or analysis population is reported for this endpoint.

The study description gives the general aims and lists interventions, but does not provide a complete treatment schedule, period structure, or details of any washout. The record does not state whether sequence, period, or carryover effects were considered analytically. These omissions matter for interpreting a crossover study, but do not establish that the design or analysis was flawed.

The registry also does not report a multiplicity plan or a threshold for a meaningful BOLD change. The adverse-event entries are limited to the listed groups and counts; further definitions and collection details are not reported. These limitations restrict what can be concluded from the registry record, without indicating what additional information may exist outside it.

15. Related Tutorials

16. Related Statistical Calculators

17. Sources

• ClinicalTrials.gov: Estrogen and Serotonin on Changing Brain Chemistry — NCT01208324.
• Registry record last updated 2021-04-02; status last verified 2021-04.
• Results first posted 2021-04-02.
• Data retrieved from ClinicalTrials.gov on 2026-11-09.
• Group sizes, means, event counts and sponsor-reported p-values are reproduced as posted. Effect estimates, confidence intervals and p-values not attributed to the sponsor are independently calculated from the posted summary statistics.

18. Record Summary

The registry identifies NCT01208324 as Estrogen and Serotonin on Changing Brain Chemistry, with the official title Interaction of Estrogen and Serotonin in Modulating Brain Activation in Menopause. The sponsor is University of Colorado, Denver. The study is recorded as randomized, double-masked, and crossover, with status completed.

The study summary describes examining estrogen and acute tryptophan depletion in relation to cognition, mood, emotional processing, and brain activation in healthy menopausal women. The primary endpoint is mean percent change in BOLD signal, with a 95% confidence interval and an 8 weeks timeframe. The posted entries give means for High Ace and Low Ace but do not report a direct comparison or define those labels in the outcome information.

The registry lists enrollment of 47 and a secondary outcome concerning verbal working memory mediated through the dorsolateral prefrontal cortex. It reports zero analyses for the posted outcome information. The record does not report the primary comparative analysis, analysis population, missing-data methods, or multiplicity approach.