Heart Rate and Breathing Effects on Attention and Memory
Phase: NA
Conditions: Aging, Alzheimer Disease, Age-related Cognitive Decline
Sponsor: University of Southern California

Study Protocol 
 
Title: Heart rate and Breathing Effects on Attention 
and Memory (HeartBEAM) 
 
NCT number: NCT05602220 
 
IRB approval date: 5/15/2023 
 
Uploaded date: 12/08/2025 
 
 

Research Background 
 
As we age, the parasympathetic system declines as the sympathetic system becomes 
hyperactive. In our previous clinical trial  (Nashiro et al., 2023; Yoo et al., 2023), we 
found that increasing parasympathetic activity through daily practice of slow-paced 
breathing significantly decreased plasma amyloid-β (Aβ) in healthy younger and older 
adults (Min et al., 2023). In healthy adults, higher plasma Aβ is associated with greater 
risk of Alzheimer’s disease (AD). Our primary goal of the current trial is to extend our 
initial findings regarding effects of slow-paced breathing on Aβ. Our secondary 
objectives are to examine the effects of daily slow-paced breathing on brain structure 
and the rate of learning. 
 
Research Objectives 
 
The aim of the study is to examine how daily paced breathing affects plasma Aβ levels 
and the rate of learning in older adults. Healthy adults aged 50-70 who meet all eligibility 
criteria will be invited to this study. Participants will be randomly assigned to one of the 
two conditions: 1) Daily memory and attention training followed by a paced breathing 
protocol designed to increase relaxation or 2) Daily memory and attention training 
followed by a paced breathing protocol to increase alertness. Participants will be asked 
to complete pre and post intervention cognitive testing online, engage in 10 weeks of 
daily brain training (starting Week 2) and 9 weeks of paced breathing (starting Week 3) 
at home. They will also be asked to come in for lab visits on Weeks 2, 7 and 12 to 
provide blood and urine samples to assess amyloid beta levels and to complete 
magnetic resonance imaging scans to assess perivascular space volume. 
 
 
 

Methods and Design 
 
Overview of the study 
Timeline 
Lab Visit 
Home Tasks 
Week 1 
online consent, home cognitive testing 
Week 2 
(Visit 1) EEG tasks, questionnaires 
(Visit 2) MRI, blood & urine 
samples, cognitive tasks, 
questionnaires, give heart rate 
monitoring device  
(Training Week 1) cognitive training + 
resting pulse measures (~40 min/day) 
Week 3 
  
(Training Week 2) cognitive + breathing 
training (~60 min/day) 
Week 4 
(Training Week 3) cognitive + breathing 
training (~60 min/day) 
Week 5 
  
(Training Week 4) cognitive + breathing 
training (~60 min/day) 
Week 6 
(Training Week 5) cognitive + breathing 
training (~60 min/day) 
Week 7 
(Visit 3) MRI, blood & urine 
samples, questionnaires 
(Training Week 6) cognitive + breathing 
training (~60 min/day) 
Week 8 
(Training Week 7) cognitive + breathing 
training (~60 min/day) 
Week 9 
  
(Training Week 8) cognitive + breathing 
training (~60 min/day) 
Week 10 
  
(Training Week 9) cognitive + breathing 
training (~60 min/day) 
Week 11 
  
(Training Week 10) cognitive + breathing 
training (~60 min/day) 
Week 12 
(Visit 4) EEG task, questionnaires 
(Visit 5) MRI, blood & urine 
samples, cognitive tasks, 
questionnaires, return heart rate 
monitoring device 
home cognitive testing 
 

Week 1 - Home Tasks: 
After signing the consent form, participants will be asked to complete questionnaires 
and cognitive tasks online. We will email them a link to a website (custom programmed) 
that will administer a combination of the tasks listed below daily for a week. In the email, 
we will give them the following instructions: “Before you begin, please make sure all 
distractions are removed (i.e. TV, radio, phone and tablets are all turned off). While you 
complete the tasks, please do not eat, snack or drink; please do not leave your screen 
throughout the experiment (and if you have to, try to do it after you finish the task at 
hand); please wear glasses if needed.” Participants’ email will be their username and, 
on the first day, they will create a password for the website app that administers the 
tasks and questionnaires, which will take about 45 minutes each day. The information in 
parentheses below indicates domains of the tasks. 
 
1) 
N-back task (memory) 
2) 
Verbal learning memory task (memory)  
3) 
Face-name recognition task (memory) 
4) 
Pattern separation task (memory) 
5) 
Task switching task (executive function) 
6) 
Flanker task (executive function) 
7) 
Stop-signal task (executive function) 
8) 
Spatial orientation task (visuospatial function) 
9) 
The Reading the Mind in the Eyes Test (emotion) 
10) Profile of Mood States (emotion) 
11) Verbal fluency task (language)  
12) Daily Inventory of Stress Events (emotion) 
13) Depression Anxiety Stress Scales – Short Form (emotion) 
14) Positive and Negative Affect Schedule (emotion) 
15) The Five Facet Mindfulness Questionnaire (emotion) 
16) Physical activity ratings (physical activity; note that this questionnaire will be 
asked only at pre-intervention) 
17) Epworth Sleepiness Scale (sleep) 
18) Karolinska Sleepiness Scale (sleep) 
19) Memory for emotional state in the past week and prediction of mood for next 
week (memory) 
20) Demographic information 
21) Emotional memory task (emotion and memory) 
 
If participants are unable to complete the cognitive/questionnaire module for a particular 
day, they will be asked to pick up that module of the study where they left off the next 
day. They can complete that leftover module and start and complete a new module the 
same day. However, they will not be allowed to start more than one module per day. If 
participants miss more than one out of 6 days of cognitive testing, they will be asked to 
leave the study. 
 
 
 

Week 2 - Lab Visit 1 (around 2 hours): 
Participants will be asked to complete the following cognitive tests and questionnaires at 
the Emotion & Cognition Lab: 
• Trail Making Test (TMT) 
• Animal fluency test 
• Multilingual Naming Test (MINT) 
• Functional Activities Questionnaire (FAQ) 
• Montreal Cognitive Assessment (MoCA) 
• Sleep questionnaire 
• EEG tasks 
 
Week 2 - Lab Visit 2 (around 2.5 hours): 
The second visit includes the MRI assessments, which will take place at the USC Dana 
and David Dornsife Cognitive Neuroimaging Center (DNI). In the MRI session, the 
participant will be asked to simply rest (i.e., do nothing without falling asleep) during 
different types of scans.  During the entire MRI session, their heart rate will be 
measured by a pulse oximeter. The total scan time will be an hour. 
 
The rest of the visit will take place at the Emotion & Cognition Lab and the Psychology 
Department (SGM 906), which will take about an hour and half. Phlebotomists will 
collect blood samples up to 30 mL via venipuncture. Note that blood draw volume for 
blood donation in the US is 1 pint (~450 mL); thus, 30 mL should be safe for 
participants. Fasting is not required for the blood draw. Participants will also be asked to 
provide a urine sample (~20mL). Blood pressure will be measured by a blood pressure 
cuff. In addition, participants will be asked to complete the Profile of Mood States, the 
Daily Inventory of Stress Events, and the sleep questionnaire.  
 
At the end of this visit, participants will receive their devices. The devices include a 
laptop with a customized paced breathing app, USB-module and a sensor that clips 
onto the earlobe and measures heart rate.  
 
Week 2-12 (10-week intervention at home): 
After the second lab visit, participants will be asked to start an intervention, which 
involves doing 15-30 minutes of cognitive training and 30 minutes of paced breathing 
practice each day. In the first week of intervention, participants will be asked to do 
cognitive training only followed by measuring their resting heart rate. From the second 
week, they will be asked to do both cognitive training and paced breathing practice (with 
heart rate monitoring via an ear clip) each day. Participants will be randomly assigned to 
one of two conditions. 
 
Condition 1: Slow-paced breathing condition (a paced breathing protocol designed 
to increase relaxation) 
Each day, participants will be guided to do slow paced breathing at paces between 10-
15 seconds per breath. Participants will receive continuous biofeedback in the form of a 

score that is higher the more that their heart rate shows strong oscillatory activity at a 
slow breathing frequency. This is described as the ‘relaxation’ condition to participants. 
 
Condition 2: Random-paced breathing condition (a paced breathing protocol to 
increase alertness) 
Each day, participants will be guided to do random-paced breathing at paces between 
4-6 seconds per breath. Participants will receive continuous biofeedback in the form of a 
score that is higher the less that their heart rate shows strong oscillatory activity at a 
slow breathing frequency. This is described as the ‘alertness’ condition to participants. 
 
Participants in both conditions will also play Lumosity cognitive training games daily. 
Lumosity games are commercial cognitive training games designed to improve various 
core cognitive skills including memory, attention, processing speed, spatial orientation, 
mental flexibility, reasoning, and problem-solving skills 
(https://www.lumosity.com/en/brain-games/). 
 
Lumosity provides a platform for researchers who aim to better understand human 
cognition. They agreed to provide services to our research team. Our liaison at 
Lumosity, Nicole Ng, has consulted with us on some study design decisions but will not 
be involved in the collection or analysis of the data. In addition, Lumosity will not have 
access to identifiable data and will make the data collected on their platform available to 
us, as described below:  
Data requested from the Lumosity database is available in a securely de-identified 
format. There is sufficient data meeting the criteria provided by the investigator to 
complete a thorough and robust analysis appropriate for the research question. 
 
The app will assign each participant a Lumosity email. This Lumosity email will be 
stored in our app database along with the study participant ID. 
 
Participants will play a list of games shown below and we will be getting final scores for 
each game each time they played it via email. Participants will have free access to the 
Lumosity games for the duration of the study. At the end of the study when they return 
the devices, their access to Lumosity will terminate. If they wish to continue with 
Lumosity after the completion of the study, they would need to sign up for an account 
and pay for the service.  
 
Game Name 
Category 
Type 
Description (taken from 
lumosity.com/en/brain-games/) 

Familiar Faces Memory 
Episodic memory 
(long-term memory for 
face-name pairs) 
Play the role of a waiter and remember 
your customers’ names and orders in order 
to earn higher tips and job promotions. 
Face-Name Recall: Face-name recall 
involves being able to pay attention to and 
remember names and physical 
characteristics. 
Tidal treasures Memory 
Episodic memory 
(binding action to item 
across rounds) 
Keep choosing unique ocean treasures 
from a tide pool as more and more items 
wash up. 
 
Objects will be more difficult to tell apart as 
they accumulate in the tide pool -- so you’ll 
need to remember more details about each 
item on your growing list of treasures as 
you spend more time at the shore. 
  
Working Memory: Working memory is used 
for temporarily storing and manipulating 
information. 
Memory 
Match 
Memory 
2-back working 
memory 
In Memory Match, you must quickly 
determine whether a flashcard symbol 
matches the one presented 2 times 
previously.  
 
Quickly memorizing and comparing the 
symbols challenges your working memory. 
This game also makes you continually 
update your working memory with new 
information.  
 
Working Memory: Working memory is used 
for temporarily storing and manipulating 
information. 

Word Bubble 
rising 
Language Verbal fluency 
In Word Bubbles, you write words that 
start with the same starting letters. This 
task challenges verbal fluency — your 
ability to rapidly retrieve words from your 
mental vocabulary. More specifically, Word 
Bubbles challenges phonemic fluency, or 
fluency with words that begin with a 
certain letter or sound.  
Verbal Fluency: Verbal fluency is the ability 
to rapidly retrieve words from your mental 
vocabulary. 
Raindrops 
Math 
Arithmetic 
Raindrops uses arithmetic to improve 
problem-solving skills. Play Raindrops to 
become better at mental calculations. You 
might even find yourself volunteering the 
next time someone needs to calculate a tip! 
Numerical Calculation: Numerical 
calculation is the ability to perform simple 
arithmetic operations including addition, 
subtraction, multiplication, and division. 
Penguin 
pursuit 
Attention Spatial orientation + 
speed 
Penguin Pursuit makes you use your spatial 
orientation, your ability to adjust your 
perspective in a mental map, to guide a 
penguin through a maze. When the maze 
rotates, you must rotate your mental map 
of the maze and recalibrate the directions 
to get to the fish.  
Spatial Orientation: Spatial orientation, also 
known as sense of direction, involves being 
aware of the surrounding environment. 

Lost in 
Migration 
Attention Flanker task 
Lost in Migration exercises your attention 
abilities. Games like Lost in Migration can 
boost your focus, helping you avoid 
distraction and improve productivity at 
work.   
Selective Attention: The ability to focus on 
relevant information while ignoring 
irrelevant distractions 
Splitting Seeds Attention Subitizing (visual 
estimation) 
In this game, you evenly divide a pile of 
seeds. To do this quickly, you’ll need your 
subitization skills. Subitization is your ability 
to quickly discern the number of items in a 
small group without having to count.  
Information Processing: Information 
processing is the initial identification and 
analysis of incoming sensory input. 
Pirate passage Reasoning Planning 
Exercise your planning skills by finding the 
route to buried treasure.  
Planning: Planning involves thinking ahead, 
evaluating options, and choosing the best 
course of action. 
Brain Shift 
Flexibility Task Switching 
In this game, you will switch between two 
rules depending on whether the letter-
number pair shows up on the top or 
bottom card.  
The location of the letter-number pair will 
be unpredictable, so you will exercise your 
mental flexibility to answer as quickly as 
possible.  
Task Switching: Task switching is the 
process of adapting to changing 

circumstances, switching from one goal to 
another. 
Ebb and Flow Flexibility Task Switching 
In this game, you shift your focus between 
two details: where the leaves point and 
how they move. This challenges your ability 
to switch between 2 cognitive processes: 
one interprets shape, the other movement. 
When you switch tasks, your brain 
suppresses one of these processes while 
activating the other — and this complex 
process often leads to mistakes.  
Task Switching: Task switching is the 
process of adapting to changing 
circumstances, switching from one goal to 
another. 
Color match 2 Flexibility Stroop[SL34]  
Color Match is a response-inhibition game 
that enhances your flexibility skills.  
Improving these abilities makes it easier to 
resist temptation, control impulses, and 
stay focused on your goals.  
Response Inhibition: Response inhibition is 
the ability to suppress inappropriate 
responses that interfere with goal-directed 
actions. 
 

The app is intended to use for collecting data, providing feedback on their performance 
and sending notifications (i.e., we will send a reminder to practice). We will use the data 
collected by the app as part of our research study. These data mainly involve 
amount/time of practice, the sequence of intervals between heartbeats during the 
practice sessions, and the strategies they were asked to try during each session. We 
will need these data for reporting the success of our intervention and comparison 
condition in our study. These data will not be shared with the FDA. 
 
The feedback will differ between the two conditions, in that the slow-paced breathing 
participants will be trained to try to maximize heart rate oscillations while the random-
paced breathing participants will be trained to try to avoid heart rate oscillations. In each 
condition, they will receive a feedback score that is incremented during the session and 
has a final value at the end of the session. This score will reflect the opposing goals in 
the two conditions, with larger values for higher heart rate oscillations in the slow-paced 
breathing condition and larger values for less oscillatory power in the random-paced 
breathing condition. 
 
Participants in the two conditions will be guided through which breathing paces to use to 
try to optimize their feedback scores each time they practice. During each 15-min 
practice session, participants will be guided through five 3-min segments of randomly 
selected paces from their set of paces. Over time, this set will get smaller, as the app 
gathers average performance of each pace for that participant and compares it to the 
other paces and discards those that are clearly worse than the numerically best pace 
because its 95% confidence interval no longer overlaps with the top ranked pace. Our 
intention is to find paces that work well for each individual to achieve their goal. 
 
During the intervention, each participant’s progress will be facilitated by the app that will 
send reminders when they fail to complete practices. Progress will be monitored by a 
group of researchers; if participants fall behind on practice, we will contact them by 
email, phone, or video calls (e.g., Zoom) for us to check if they have any problems with 
any aspects of the study, such as device and internet issues. There will be no 
opportunities to make up sessions and if participants only complete part of a session 
one day, they will start at the beginning of a session the next time they practice. We 
anticipate high compliance with practice, as in our prior 5-week HRV biofeedback study, 
older participants averaged over 100% of the practice we asked them to complete (and 
by week 3, this was 40 min/day of biofeedback). In that study, we did not ask them to do 
more than 40 min/day so this is an indicator that they enjoyed it or felt they were getting 
something worthwhile from it. Thus, we have some experience indicating that the HRV-
biofeedback is something that participants can do every day. In addition, Lumosity is a 
highly engaging platform and we anticipate that older adults will enjoy the activity.  
 
Week 7 - Lab Visit 3 (around 2 hours): 
Participants will be asked to complete the same assessments as in Visit 2 (i.e., MRI, 
blood draw, urine collection, blood pressure measurement, questionnaires).  
 
Week 12 - Lab Visit 4 (around 2 hours): 

Participants will be asked to complete the same assessments as in Visit 1 except that 
we will not repeat the following cognitive tests: MINT, FAQ and MoCA.  
 
Week 12 - Lab Visit 5 (around 2 hours) 
Participants will be asked to complete the same assessments as in Visit 2. They will 
also be asked to return their devices.  
 
A few days after Visit 2, 3, and 5, participants will be asked to complete the post blood 
draw questionnaire to report any adverse events associated with the blood draw, if any. 
We will report to the IRB if the responses to any of the questions were the most 
negative response option. For example, one of the questions is “do you notice any 
bleeding after your session (on your way home or at home)? The response options are 
1) no bleeding, 2) a little (perhaps a few drops, 3) yes (needed a new bandage). We will 
report to the IRB when the response was #3.   
 
Week 12 - Home Tasks: 
After their last lab visit, participants will be asked to complete daily questionnaires and 
cognitive tasks as they did in the first week of the study. In addition, participants will be 
asked about their experience during this study. Pre and post cognitive data will be 
compared to see if there are any group differences in change.  
 

References 
 
Min, J., Rouanet, J., Martini, A. C., Nashiro, K., Yoo, H. J., Porat, S., Cho, C., Wan, J., 
Cole, S. W., Head, E., Nation, D. A., Thayer, J. F., & Mather, M. (2023). 
Modulating heart rate oscillation affects plasma amyloid beta and tau levels in 
younger and older adults. Scientific Reports, 13(1), 3967. 
Nashiro, K., Min, J., Yoo, H. J., Cho, C., Bachman, S. L., Dutt, S., Thayer, J. F., Lehrer, 
P. M., Feng, T., Mercer, N., Nasseri, P., Wang, D., Chang, C., Marmarelis, V. Z., 
Narayanan, S., Nation, D. A., & Mather, M. (2023). Increasing coordination and 
responsivity of emotion-related brain regions with a heart rate variability 
biofeedback randomized trial. Cognitive, Affective & Behavioral Neuroscience, 
23(1), 66–83. 
Yoo, H. J., Nashiro, K., Min, J., Cho, C., Mercer, N., Bachman, S. L., Nasseri, P., Dutt, 
S., Porat, S., Choi, P., Zhang, Y., Grigoryan, V., Feng, T., Thayer, J. F., Lehrer, 
P., Chang, C., Stanley, J. A., Head, E., Rouanet, J., … Mather, M. (2023). 
Multimodal neuroimaging data from a 5-week heart rate variability biofeedback 
randomized clinical trial. Scientific Data, 10(1), 503.