Exploring the Effects of Exercise on Memory and Cognition in Parkinson´s Disease
Phase: NA
Conditions: Parkinson Disease
Sponsor: Heidelberg University

 
Exploring the Effects of Exercise on Memory and Cognition in 
Parkinson´s Disease (EMCo) 
- Study Protocol & Statistical Analysis Plan - 
 
Principal Investigators: 
Prof. Dr. Simon Steib 
Institute for Sports und Sports Sciences 
Heidelberg University 
E-Mail: simon.steib@issw.uni-
heidelberg.de  
Dr. Philipp Wanner 
Institute for Sports und Sports Sciences 
Heidelberg University 
E-Mail: philipp.wanner@issw.uni-
heidelberg.de  
 
Sub-investigators: 
Charlotte Hohloch 
Institute for Sports und Sports Sciences 
Heidelberg University 
E-Mail: charlotte.hohloch@issw.uni-
heidelberg.de  
  
 
Collaborators (alphabetical order): 
Dr. med. Andreas Becker 
Abteilung für Neurologie 
SRH Kurpfalzkrankenhaus Heidelberg 
PD Dr. med. Julian Conrad 
Abteilung Neurodegeneration 
Universitätsklinikum Mannheim 
Prof. Dr. med. Brigit Friedmann-Bette 
Sportmedizin 
Universitätsklinikum Heidelberg 
PD Dr. med. Heike Jacobi 
Neurologie und Poliklinik 
Universitätsklinikum Heidelberg 
Prof. Dr. med. Rebecca Schüle-Freyer 
Sektion Neurodegeneration 
Universitätsklinikum Heidelberg 
Prof. Dr. med. Jochen Weishaupt 
Abteilung Neurodegeneration 
Universitätsklinikum Mannheim 
Supporting Biometrician: 
Dr. Marietta Kirchner 
Institut für medizinische Biometrie 
Universitätsklinikum Heidelberg 
 
 
 
Date: 21.04.2024 
Version 1 
 
 
 
 

 
Statistical analysis plan V1; 21.04.2024 
 
2 
Table of contents 
1 
Aims and hypotheses ................................................................................................3 
1.1 
Primary aims ............................................................................................................3 
1.2 
Secondary aims .......................................................................................................4 
2 
Study design ..............................................................................................................6 
2.1 
General procedure ...................................................................................................6 
2.2 
Intervention ..............................................................................................................6 
2.3 
Study sample ...........................................................................................................7 
2.4 
Sample size justification ...........................................................................................7 
3 
Outcome Measures ....................................................................................................8 
3.1 
Primary outcomes ....................................................................................................8 
3.2 
Secondary outcomes ................................................................................................9 
3.3 
Other measures ..................................................................................................... 10 
4 
Data analysis ............................................................................................................ 11 
5 
Appendix .................................................................................................................. 13 
6 
References ............................................................................................................... 14 
 

 
Statistical analysis plan V1; 21.04.2024 
 
3 
1 
Aims and hypotheses 
The study aims to assess the effects of cardiovascular (aerobic) training on memory formation 
and cognitive function in people with Parkinson’s disease (pwPD). Participants will be 
randomly allocated to one of two groups either performing cardiovascular training 
(experimental group) or stretching (control group) for twelve weeks, three times a week. We 
examine whether moderate-intense cardiovascular training (MICT) improves procedural 
memory formation (primary outcome) compared to stretching and mobility training. Secondary 
outcomes include episodic memory formation, cognitive function, cardiorespiratory fitness, 
sleep quality, and brain-derived neurotrophic factor (BDNF) blood concentration levels. 
1.1 
Primary aims 
Our primary research question is to investigate the effects of cardiovascular training on non-
declarative (procedural) memory formation. A first study in pwPD demonstrated an improved 
encoding of motor sequences after a twelve-week cardiovascular training in a pre-post design 
(Duchesne et al., 2015). Further randomized controlled trials in populations with different 
neurological conditions reported enhanced encoding (Quaney et al., 2009) and consolidation 
(Lo et al., 2023) of motor sequences. We, therefore, expect that cardiovascular training 
improves all aspects of procedural memory formation. To assess the effects of cardiovascular 
training on non-declarative (procedural) memory formation, we will calculate a global motor 
learning score (i.e., change from start of encoding to 24h recall test) for explicit and implicit 
components of motor sequence learning and analyze the change (delta) in the global motor 
learning scores from pre- to post-intervention assessments. This procedure results in two 
primary outcome variables: (i) explicit global motor learning score and (ii) implicit global motor 
learning score. The primary questions (Q) are: 
Q1: Does twelve weeks of cardiovascular training improve the explicit components of non-
declarative (procedural) memory formation in pwPD? 
We expect that a twelve-week cardiovascular training program improves the global motor learning 
score (i.e., performance changes from start of encoding to a 24 h recall test) for explicit components 
of motor sequence learning. 
Q2: Does twelve weeks of cardiovascular training improve the implicit components of non-
declarative (procedural) memory formation in pwPD? 
We expect that a twelve-week cardiovascular training program improves the global motor learning 
score (i.e., performance changes from start of encoding to a 24 h recall test) for implicit components 
of motor sequence learning. 

 
Statistical analysis plan V1; 21.04.2024 
 
4 
1.2 
Secondary aims 
To disentangle the effects of cardiovascular training on the different processes of non-
declarative (procedural) memory formation, we will investigate the encoding and consolidation 
of the motor sequence learning task in secondary analyses. We will calculate the encoding 
and consolidation performance for the explicit and implicit components resulting in four 
outcome variables. We expect a global effect of cardiovascular exercise on procedural memory 
formation, and thus an improvement in all variables.  
Q3: Does twelve weeks of cardiovascular training improve the encoding and consolidation of 
non-declarative (procedural) memory in pwPD? 
We expect that a twelve-week cardiovascular training program improves the encoding of explicit 
components of motor sequences. 
We expect that a twelve-week cardiovascular training program improves the encoding of implicit 
components of motor sequences. 
We expect that a twelve-week cardiovascular training program improves the consolidation of explicit 
components of motor sequences. 
We expect that a twelve-week cardiovascular training program improves the consolidation of implicit 
components of motor sequences. 
In a next step, we will investigate the effects of cardiovascular training on (i) declarative 
(episodic) memory and (ii) cognitive function (Kim et al., 2023). We expect that if the exercise-
induced effects on brain function are global, declarative memory and cognitive function should 
also be enhanced. For declarative (episodic) memory, similar to non-declarative (procedural) 
memory we will first analyze the global episodic learning score (i.e., performance change from 
start of encoding to recall test) followed by a separate analysis of encoding and consolidation. 
For global cognitive function we will first analyze a composite score including the domains (i) 
short-term memory, (ii) working memory, (iii) inhibition, and (vi) cognitive flexibility followed by 
a separate analysis of each domain.   
Q4: Does twelve weeks of cardiovascular training improve declarative (episodic) memory in 
pwPD? 
We expect that a twelve-week cardiovascular training program improves the global episodic learning 
score (i.e., performance changes from start of encoding to a 24 h recall test). 
Q5: Does twelve weeks of cardiovascular training improve the encoding and consolidation 
declarative (episodic) memory in pwPD? 
We expect that a twelve-week cardiovascular training program improves the encoding of episodic 
memory. 
We expect that a twelve-week cardiovascular training program improves the consolidation of episodic 
memory. 
 
 

 
Statistical analysis plan V1; 21.04.2024 
 
5 
Q6: Does twelve weeks of cardiovascular training improve cognitive function in pwPD? 
We expect that a twelve-week cardiovascular training program improves global cognitive function in 
pwPD. 
We expect that a twelve-week cardiovascular training program improves short-term memory, working 
memory, inhibition, and cognitive flexibility in pwPD. 
We will then examine the effects on (i) cardiorespiratory fitness (Schootemeijer et al., 2020), 
(ii) sleep (Cristini et al., 2021), and (iii) BDNF blood concentration (Kaagman et al., 2024) to 
explore potential mechanisms of the exercise-induced effects on memory and cognition (see 
exploratory analyses) (Loprinzi et al., 2021). 
Q7: Does twelve weeks of cardiovascular training improve cardiovascular fitness in pwPD? 
We expect that a twelve-week cardiovascular training program improves peak oxygen consumption 
(VO2peak). 
Q8: Does twelve weeks of cardiovascular training improve sleep in pwPD? 
We expect that a twelve-week cardiovascular training program improves objectively measured sleep 
efficiency. 
We expect that a twelve-week cardiovascular training program improves subjective disease-related 
sleep disturbances. 
Q9: Does twelve weeks of cardiovascular training improve brain-derived neurotrophic factor 
(BDNF) blood concentration level in pwPD? 
We expect that a twelve-week cardiovascular training program increases resting serum BDNF blood 
concentration levels. 
In exploratory analyses, we will lastly investigate potential determinants of the exercise-
induced effects on memory formation and cognition (Loprinzi et al., 2021). We will explore the 
associations between changes in memory formation and (i) cognitive function (especially 
working memory), (ii) cardiorespiratory fitness, (iii) sleep, and (iv) BDNF levels, respectively. 
We will further test whether changes in cognitive function are associated with changes in (i) 
cardiorespiratory fitness, (ii) sleep, and (iii) BDNF levels, respectively. We expect that changes 
in these variables will be associated with changes in memory formation and cognition. The 
following questions are analyzed in exploratory analyses: 
Q10: Are exercise-induced changes in non-declarative (procedural) memory formation 
associated with changes in (i) cognitive function (especially working memory), (ii) 
cardiorespiratory fitness, (iii) sleep, and (iv) BDNF blood concentration in pwPD? 
Q11: Are exercise-induced changes in declarative (episodic) memory formation associated 
with changes in (i) cognitive function (especially working memory), (ii) cardiorespiratory 
fitness, (iii) sleep, and (iv) BDNF blood concentration in pwPD? 
Q12: Are exercise-induced changes in cognitive function associated with changes in (i) 
cardiorespiratory fitness, (ii) sleep, and (iii) BDNF blood concentration in pwPD? 

 
Statistical analysis plan V1; 21.04.2024 
 
6 
2 
Study design 
2.1 
General procedure  
In a randomized controlled trial, pwPD with mild to moderate disease stage (i.e., Hoehn & Yahr 
≤3) will either perform moderate-intensity cardiovascular training (experimental group) or 
stretching training (control group) for twelve weeks (three times per week, totaling 36 training 
sessions; duration per training session 30 to 55 min). For group allocation, we will use block 
randomization stratified by biological sex (male / female) and age (<62 / ≥ 62), since these two 
factors may influence procedural memory (Smith et al., 2005) and the effects of exercise on 
cognition (Kramer & Colcombe, 2018). The outcome variables will be assessed at baseline 
(pre-assessment, within two weeks before the intervention) and after the intervention (post-
assessment, within two weeks after the intervention has terminated). 
2.2 
Intervention 
Both, cardiovascular training (experimental group) as well as stretching training (control 
group), will be performed in supervised sessions and in small groups of 2-5 persons at the 
study site. 
Experimental group (Duchesne et al., 2015; Sacheli et al., 2019): Cardiovascular training is 
performed on a cycle ergometer at 60% of peak power output (Wmax) for 30 min (i.e., 5 min 
warm-up, 20 min at 60% Wmax, 5 min cool-down) three times a week for twelve weeks. The 
exercise program will be increased by 5 min (max. 55 min) and 5% of intensity (in Watt) every 
two weeks. If participants report a too low (i.e., Borg scale ≤ 11) or too high (i.e., Borg scale ≥ 
16) subjective rate of perceived exertion (RPE; Borg scale 6-20) intensity will be increased or 
decreased until the target levels are met (i.e., Borg scale 12-15). During training, heart rate, 
intensity (in Watt) and RPE will be monitored. Participants missing training sessions will be 
offered an additional week to perform up to three training sessions. 
Control group (Duchesne et al., 2015; Sacheli et al., 2019): Stretching training consists of lying, 
seated, and standing flexibility exercises (according to the recommendations of the American 
Parkinson´s Disease Foundation and the Parkinson Society Canada) for 30 min (i.e., 5 min 
warm-up, 20 min main part, 5 min cool-down) three times a week for twelve weeks. The 
exercise program will be increased by 5 min (i.e., adding additional repetitions and exercises; 
max. 55 min). Intensity of stretching exercises will be assessed by subjective rate of perceived 
muscle tension on a visual analog scale (VAS) from 0 to 10. We aim for a moderate muscle 
tension defined as 5 to 7 on the VAS. During training VAS, heart rate, and RPE will be 

 
Statistical analysis plan V1; 21.04.2024 
 
7 
monitored. Participants missing training sessions will be offered an additional week to perform 
up to three training sessions. 
2.3 
Study sample 
Inclusion criteria: 
- 
Diagnosed Parkinson’s disease (Postuma et al., 2015) 
- 
Disease stage ≤3 on the Hoehn & Yahr scale 
- 
Age ranging from 50 – 80 years 
- 
Naive to the memory tasks (primary outcome) 
- 
Ability to stand and walk at least 10 meters independently 
Exclusion Criteria: 
- 
Atypical Parkinsonism 
- 
Significant level of cognitive impairment (i.e., Montreal Cognitive Assessment 
<21) (Dalrymple-Alford et al., 2010) 
- 
Deep brain stimulation or brain pacemaker 
- 
Diagnosed psychiatric illness 
- 
Known clinically relevant neurological, internal or orthopedic conditions besides 
Parkinsonism that would interfere with the exercise paradigm 
- 
Exceeding the recommended level of cardiovascular exercise for older adults 
(i.e., cardiovascular exercise done ≥150 min per week of moderate-intensity or 
≥75 min per week of vigorous-intensity) 
2.4 
Sample size justification 
Following the recommendations by Lakens (2022), the sample size is based on the available 
resources and the potential gain in information. Due to resource constraints the maximum 
sample that can be collected is n = 60 participants (n = 30 per group). We used this sample 
size to perform a sensitivity power analysis (G*Power, version 3.1.9.7) for the primary analysis 
(see 4). The analysis of covariance (ANCOVA) includes the respective primary outcome (i.e., 
explicit global motor learning score and implicit global motor learning score) as dependent 
variable, group (i.e., experimental vs. control group) as between-subject factor, and the 
baseline value of the primary outcome variable and the stratification criteria (i.e., age and 
biological sex) as covariates. With an alpha level of 2.5% (adjusted for two primary outcomes) 
the study has 80% power to detect an effect of at least f = 0.41, which is considered a large 
effect according to Cohen (1988). With an anticipated drop-out rate of ~15-20% resulting in a 
total sample size of n = 50 the study has a power of 80% to detect an effect of at least f = 0.45 
(due to imputation of missing values, a power increase can be expected). Therefore, the study 

 
Statistical analysis plan V1; 21.04.2024 
 
8 
has sufficient power to detect an effect size reported in comparable studies investigating the 
effects of cardiovascular training on procedural memory formation in people with neurological 
conditions (Lo et al., 2023; Quaney et al., 2009). Furthermore, this will be the study with the 
largest sample of people with neurological conditions to date assessing the effects of 
cardiovascular training on non-declarative (procedural) memory.   
3 
Outcome Measures 
An overview of all primary and secondary outcomes as well as screening and descriptive 
variables is listed in the table in the appendix. The outcome variables will be assessed at 
baseline (pre-assessment, within two weeks before the intervention) and after the intervention 
(post-assessment, within two weeks after the intervention has terminated). For primary and 
secondary outcomes measures, we will calculate the change (delta) from pre- to post-
intervention assessments to evaluate the effects of cardiovascular training.  
3.1 
Primary outcomes 
 Domain 
Measures 
Outcome measure 
Non-declarative 
(procedural) memory: 
global motor learning 
score 
To evaluate non-declarative (procedural) memory 
formation, we will use the Visuomotor Serial 
Targeting Task (VSTT). The VSTT has been widely 
used to study motor sequence learning in pwPD and 
other neurological condition (Ghilardi et al., 2003; 
Ghilardi et al., 2007; Marinelli et al., 2017). The task 
consists of eight radially arranged circles (targets) 
around a central circle (center) on a computer 
screen (distance from center to targets: 4 cm; radius 
of targets: 1 cm). One of the targets turn red in a 
fixed interval of 1.5 sec in synchrony with a tone. 
The participants are instructed to move a cursor 
using a digitizing tablet (Wacom Intuos Pro Large, 
Wacom; Wacom; Toyonodai, Kazo-shi, Saitama, 
Japan) with the dominant hand from the center to 
the displayed target with a single, uncorrected out 
movement, reverse sharply inside the target and do 
a similar back movement to the center. To assess 
motor sequence learning, the eight targets appear in 
a repeating order. The participants are instructed to 
learn the sequence while moving the cursor to the 
targets and reach the target in synchrony with its 
appearance (i.e., movement start before 
appearance) when the location is known (i.e., 
learned). Encoding consists of six blocks with each 
block including eight repetitions of the eight-target 
sequence (i.e., 64 movements per block), whereas 
the recall test contains three blocks of similar size. 
For each movement, we will calculate (i) onset time 
(i.e., time between target display and out movement 
start), (ii) movement time (i.e., time from out 
movement start to end), (iii) peak velocity, (iv) 
directional error at peak velocity (i.e., angular 
deviation between real and ideal movement 
trajectory at peak velocity), (v) spatial error to target 
(i.e., shortest distance of the out movement end to 
1) Change in explicit global 
motor learning score from 
pre- to post-assessment  
2) Change in implicit global 
motor learning score from 
pre- to post- assessment 

 
Statistical analysis plan V1; 21.04.2024 
 
9 
the target center), (vi) spatial error to ideal 
movement (i.e., root mean squared error between 
real and ideal movement trajectory), and (vii) 
normalized hand-path area (i.e., area enclosed by 
the out and in movement divided by the squared 
path length) to analyze practice-related changes in 
temporal and kinematic movement characteristics. 
We will use the number of correct anticipatory 
movements defined as movements with onset times 
lower than in random trials and directed to the 
correct target (directional error at peak velocity 
<22°) to analyze the learning of explicit components 
of the motor sequence (Ghilardi et al., 2009). To 
assess the learning of implicit components of the 
motor sequence, we will use spatial error to target 
(shortest distance of the movement end point from 
the center of the target) (Ghilardi et al., 2009). We 
will calculate a global motor learning score defined 
as relative change from first block of encoding to 
first block of 24h recall test for the explicit and 
implicit component of the task, respectively. To 
assess the effects on non-declarative (procedural) 
memory formation, we will analyze the change 
(delta) in the explicit and implicit global motor 
learning score from pre- to post-intervention 
assessment. Hence, two primary outcome 
measures from the VSTT are considered to answer 
the primary research questions. 
 
3.2 
Secondary outcomes 
An overview of all secondary outcome measures is listed in the table below. 
 Domain 
Measures 
Outcome measure 
Non-declarative 
(procedural) memory: 
encoding and 
consolidation 
To evaluate the effects on the different processes of 
non-declarative (procedural) memory formation, we 
will investigate the encoding and consolidation of 
the motor sequence for the implicit and explicit 
components, respectively. Encoding will be defined 
as relative performance change (delta) from 
beginning (first block of encoding) to end (last block 
of encoding) of practice, whereas consolidation will 
be calculated as relative performance change 
(delta) from end of practice (last block of encoding) 
to a 24h recall test (first block of recall). This results 
in four secondary outcome variables for the VSTT. 
1) Change in encoding of the 
explicit component from pre- 
to post assessment 
2) Change in encoding of the 
implicit component from pre- 
to post assessment 
3) Change in consolidation of 
the explicit component from 
pre- to post assessment 
4) Change in consolidation of 
the implicit component from 
pre- to post assessment 
Declarative (episodic) 
memory 
To evaluate the effects on declarative (episodic) 
memory, we will use the German version of the Rey 
Auditory Verbal Learning Test (Helmstaedter et al., 
2001). In addition to the standard retrieval after 20 
min, we conduct a recall test after 24h to investigate 
the consolidation following nocturnal sleep and to be 
comparable to the procedural memory task. Similar 
to non-declarative (procedural) memory we will first 
analyze the (i) global episodic learning score 
(correctly recalled words at 24 h recall test) followed 
by a separate analysis of (ii) encoding (correctly 
recalled words after listening five times to the 
wordlist) and (iii) consolidation (change in correctly 
recalled words from end of encoding to recall test). 
1) Change in global episodic 
learning from pre- to post 
assessment 
2) Chang in encoding from pre- 
to post assessment 
3) Change in consolidation 
from pre- to post 
assessment  

 
Statistical analysis plan V1; 21.04.2024 
 
10 
Cognitive function 
To evaluate the effects on cognitive function, we will 
assess the domains (i) short-term memory (Digit 
and Spatial Span Task forward: length of correctly 
reproduced sequence of digits/locations) 
(Helmstaedter et al., 2001), (ii) working memory 
(Digit and Spatial Span Task backward: length of 
correctly reproduced sequence of digits/locations) 
(Helmstaedter et al., 2001), (iii) inhibition (Stroop 
Test interference score: number of correctly named 
colors in the color-word condition compared to the 
predicted color-word score) (MacLeod, 1991), and 
(vi) cognitive flexibility (Trail Making Test A & B trail 
flexibility score: time to complete test B compared to 
test A) (Salthouse et al., 2000). We will first 
calculate a composite score including all domains to 
analyze global cognitive function followed by a 
separate analysis of the domains. 
1) Change in global cognitive 
function from pre- to post 
assessment 
2) Change in short-term 
memory from pre- to post 
assessment  
3) Change in working memory 
from pre- to post 
assessment 
4) Change in inhibition from 
pre- to post assessment 
5) Change in cognitive 
flexibility from pre- to post 
assessment 
Cardiorespiratory 
fitness 
To evacuate the effects on cardiorespiratory fitness, 
we will perform a graded exercise test (GXT) on a 
cycle ergometer with spiroergometry (peak oxygen 
consumption). Depending on anticipated peak 
power output participants will start with 25 W or 50 
W. The load will be increased every 3 min by 25 W 
until exhaustion. 
1) Change in cardiorespiratory 
fitness from pre- to post 
assessment 
Sleep 
To evaluate the effects on sleep, we will assess (i) 
sleep efficiency using wrist-worn actigraphy 
(ActiGraph GT9X; ActiGraph; Pensacola; USA) for 
seven nights, and (ii) subjective disease-related 
sleep disturbance using the Parkinson’s Disease 
Sleep Scale 2 (PDDS-2) total score (Trenkwalder et 
al., 2011). 
1) Change in objective sleep 
efficiency from pre- to post 
assessment 
2) Change in subjective 
disease-related sleep 
disturbance from pre- to 
post assessment 
Brain-derived 
neurotrophic factor 
(BDNF)   
To evaluate the effects on brain-derived 
neurotrophic factor (BDNF), we will collect serum 
blood samples at rest. Samples will be analyzed 
using ELISA kits. 
1) Change in brain-derived 
neurotrophic factor (BDNF) 
from pre- to post 
assessment 
3.3 
Other measures 
An overview of all screening and descriptive variables is listed in the table below. 
Domain 
Measures 
Sociodemographic & physical 
characteristics 
Age, biological sex 
Years of education 
Height, weight, body mass index 
Handedness: Edinburgh Handedness Inventory (Oldfield, 1971) 
Medical information & health status 
Time since diagnosis 
Severity of disease (Hoehn & Yahr, 1967) 
Motor & non-motor symptoms: Movement Disorder Society Unified 
Parkinson´s Disease Rating Scale part I & III (Goetz et al., 2008) 
Drugs & levodopa equivalent daily dose (LEDD) (Schade et al., 
2020) 
Quality of life: Parkinson´s Disease Questionnaire-39 (Jenkinson 
et al., 1997) 
Depression. Beck-Depression-Inventory (Beck & Beamesderfer, 
1974) 
Anxiety: Beck-Anxiety-Index (Beck et al., 1988) 
Self-reported falls in the past 12 months (Schwenk et al., 2012) 
Other disease besides Parkinsonism or acute injuries 
Cognitive status 
Montreal Cognitive Assessment (Dalrymple-Alford et al., 2010) 
Sleep 
Sleep quality: Pittsburgh Sleep Quality Index (Monk et al., 1994) 
Daytime Sleepiness. Epworth Sleepiness Scale (Johns, 1991) 
Physical activity 
Sensor-based physical activity (ActiGraph GT9X) for 7 days/nights 

 
Statistical analysis plan V1; 21.04.2024 
 
11 
German Physical Activity Questionnaire 50+ (Huy & Schneider, 
2008) 
International Physical Activity Questionnaire (Craig et al., 2003) 
4 
Data analysis 
We will perform all statistical analyses with IBM SPSS Statistics or RStudio. Boxplot, scatter 
plots or bar charts will be used for an initial inspection of the data. Data will be checked 
graphically for normality (Rochon et al., 2012) and error outliers will be excluded from the 
respective analysis (e.g., measurement errors) but included in the other analyses (Leys et al., 
2019). The alpha level for statistical significance will be set at p ≤ .05 unless otherwise stated. 
We will additionally report effect sizes.  
In a first step, we will use summary statistics (mean and standard deviation or median and 
interquartile range or frequencies depending on the scale of the variable) to provide a 
comprehensive description of the baseline sociodemographic, physical, and health 
characteristics. 
To investigate the effects of the cardiovascular training program on procedural memory 
formation including (i.e., primary aim), we will conduct separate analyses of covariance 
(ANCOVA) for the two primary outcomes (i.e., Q1 & Q2: explicit global motor learning score 
and implicit global motor learning score) (O'Connell et al., 2017; Wan, 2021). The ANCOVA 
will contain the respective primary outcome as dependent variable, the group (i.e., 
experimental vs. control group) as between-subject factor, and the baseline value of the 
primary outcome variable and the stratification criteria (i.e., age and biological sex) as 
covariates. The analysis will be performed according to an intention-to-treat principle including 
all participants randomized after pre-assessment. As sensitivity analysis, a complete-case 
analysis is used to check the robustness of the results. For missing data, we will perform 
multiple imputations using multivariate imputation by chained equations (MICE) with predictive 
mean matching while assuming that data is missing at random. The alpha level for statistical 
significance in the primary analysis will be corrected using the Bonferroni-Holm method to 
adjust for multiple primary outcome measures.  
The effects of the intervention on the secondary outcome measures (i.e., secondary analyses) 
will be tested in separate ANCOVA models using the same procedure as the primary analysis. 
We will first analyze the effects on (i) the encoding and consolidation of the motor sequence 
learning task to disentangle the effects of cardiovascular training on the different processes of 
procedural memory formation (Q3), followed by (ii) episodic memory (Q4 & Q5), and (iii) 
cognitive function (Q6). For cognitive function, we will analyze global cognitive function 

 
Statistical analysis plan V1; 21.04.2024 
 
12 
(composite score of cognitive tests) in a first step and in a second step the individual test 
scores. We will then investigate the effects on (iv) cardiorespiratory fitness (Q7), (v) sleep (Q8), 
and (vi) BDNF blood concentration (Q9). We will familywise adjust the alpha level using 
Bonferroni-Holm correction for secondary analyses with multiple outcome measures.  
To explore the potential influence of subject characteristics on the exercise effects, we will 
separately include cardiorespiratory fitness, sleep parameters, BDNF blood concentration, 
cognitive function, disease stage, and symptoms at baseline (pre-assessment) as covariates 
into the ANCOVA models testing for changes in procedural memory formation (primary 
analysis), episodic memory formation (secondary analysis), and cognitive function (secondary 
analysis). Additionally, potential associations between changes in memory formation or 
cognitive function and subject characteristics at baseline will be explored using Pearson 
product-moment correlations or Spearman’s rank correlation (cardiorespiratory fitness, sleep 
parameters, BDNF blood concentration, cognitive function, disease stage, and symptoms) and 
point-biserial correlation (biological sex). Correlations are performed at group level. To further 
explore if these baseline characteristics moderate exercise-induced changes in memory and 
cognitive function, we plan to run separate multiple linear regression models including changes 
in memory formation or cognitive function (Y) and the baseline subject characteristics listed 
above (i.e., moderation analysis). Interactions with group allocation will be examined to explore 
potential moderators.    
In a last exploratory step, we will examine potential associations between the exercise-induced 
changes in memory formation or cognitive function and changes in the other secondary 
outcomes (Q10-Q12). According to the currently proposed contributors to the exercise-
memory interaction, we will correlate (i.e., Pearson product-moment correlations or 
Spearman’s rank correlation) the changes from pre- to post-assessment in procedural and 
episodic memory with the changes in (i) global cognitive function, (ii) working memory, (iii) 
cardiorespiratory fitness, (vi) sleep parameters, and (v) BDNF blood concentration. For 
changes in cognitive function, we will explore associations with the changes in (i) 
cardiorespiratory fitness, (ii) sleep parameters, and (iii) BDNF blood concentration. All 
correlations are performed at group level. To further explore potential candidates that drive the 
exercise-memory interaction, the mediating effect of changes in (i) global cognitive function, 
(ii) working memory, (iii) cardiorespiratory fitness, (vi) sleep parameters, and (v) BDNF blood 
concentration on changes in procedural or episodic memory formation will be analyzed (Cerin 
et al., 2006). For changes in cognitive function, mediating effects of changes in (i) 
cardiorespiratory fitness, (ii) sleep parameters, and (iii) BDNF blood concentration will be 
analyzed. 

 
Statistical analysis plan V1; 21.04.2024 
 
13 
5 
Appendix 
Variable 
Measures 
T1 
INT 
T2 
 
Sociodemographic & physical characteristics 
 
 
 
ST, SC, DV 
Age, biological sex 
X 
 
 
DV 
Years of education 
X 
 
 
DV 
Height, weight, body mass index 
X 
 
X 
DV 
Handedness: Edinburgh Handedness Inventory (Oldfield, 
1971) 
X 
 
X 
 
Medical information & health status 
 
 
 
DV 
Time since diagnosis 
X 
 
 
SC, DV 
Severity of disease (Hoehn & Yahr, 1967) 
X 
 
X 
DV 
Motor & non-motor symptoms: Movement Disorder Society 
Unified Parkinson´s Disease Rating Scale part I & III (Goetz 
et al., 2008) 
X 
 
X 
DV 
Drugs & levodopa equivalent daily dose (LEDD) (Schade et 
al., 2020) 
X 
X 
X 
DV 
Quality of life: Parkinson´s Disease Questionnaire-39 
(Jenkinson et al., 1997) 
X 
 
X 
DV 
Depression. Beck-Depression-Inventory (Beck & 
Beamesderfer, 1974) 
X 
 
X 
DV 
Anxiety: Beck-Anxiety-Index (Beck et al., 1988) 
X 
 
X 
DV 
Self-reported falls in the past 12 months (Schwenk et al., 
2012) 
X 
 
 
SC, DV 
Other disease besides Parkinsonism or acute injuries 
X 
 
 
 
Memory formation 
 
 
 
PO 
SO 
Procedural memory: Visuomotor Serial Targeting Task 
(Ghilardi et al., 2003; Ghilardi et al., 2007; Marinelli et al., 
2017) 
- Explicit global motor learning score 
- Implicit global motor learning score 
- Encoding:  
- explicit component 
- implicit component 
- Consolidation 24 h:  
- explicit component 
- implicit component 
X 
 
X 
SO 
Episodic memory: German version of the Rey Auditory 
Verbal Learning Test (Helmstaedter et al., 2001) 
- Global episodic learning score 
- Encoding 
- Consolidation 20 min 
- Consolidation 24 h* 
X 
 
X 
  
Cognitive function 
 
 
 
SC, DV 
Cognitive status: Montreal Cognitive Assessment 
(Dalrymple-Alford et al., 2010) 
X 
 
X 
SO 
Short-term & working memory: Digit & Spatial Span Test 
forward & backward (Härting et al., 2001) 
X 
 
X 
SO 
Inhibition: Stroop Test (MacLeod, 1991) 
X 
 
X 
SO 
Cognitive flexibility: Trail Making Test (Salthouse et al., 2000) 
X 
 
X 
 
Cardiovascular examination 
 
 
 
DV, SO 
Graded exercise test with spiroergometry (VO2peak) 
X 
 
X 
 
Biomarkers 
 
 
 
SO 
Brain-derived neurotrophic factor (BDNF) blood 
concentration level at rest 
X 
 
X 
  
Sleep 
 
 
 
DV, SO 
Sensor-based sleep (ActiGraph GT9X) for 7 days/nights 
X 
 
X 

 
Statistical analysis plan V1; 21.04.2024 
 
14 
DV, SO 
Disease-related sleep disturbance: Parkinson's Disease 
Sleep Scale–2 (Trenkwalder et al., 2011) 
X 
 
X 
DV, SO 
Sleep quality: Pittsburgh Sleep Quality Index (Monk et al., 
1994) 
X 
X 
X 
DV, SO 
Daytime Sleepiness. Epworth Sleepiness Scale (Johns, 
1991) 
X 
 
X 
  
Physical activity 
 
 
 
DV 
Sensor-based physical activity (ActiGraph GT9X) for 7 
days/nights 
X 
 
X 
DV 
Self-reported physical activity: 
German Physical Activity Questionnaire 50+ (Huy & 
Schneider, 2008) 
International Physical Activity Questionnaire (Craig et al., 
2003) 
X 
X 
X 
T1 = pre-assessment (baseline) within 2 weeks before intervention; T2 = post-assessment within 2 weeks after 
intervention; PO = primary outcome; SO = secondary outcome; SC = screening parameter for inclusion; ST = 
stratification criterion; DV = descriptive variable 
*In addition to the standard retrieval after 20 min, we conduct a recall test after 24 h to investigate the consolidation 
following nocturnal sleep and to be comparable to the procedural memory task.  
6 
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