Effect of Ischemic Preconditioning on

Endurance Performance Does Not

Surpass Placebo
JEANN L. SABINO-CARVALHO1,2, THIAGO R. LOPES2,3,4, TIAGO OBEID-FREITAS2,4, THIAGO N. FERREIRA1,2,
JOSE E. SUCCI5, ANTONIO C. SILVA2,4, and BRUNO M. SILVA1,2,4
1
Graduate Program in Translational Medicine, Federal University of Sao Paulo, Sao Paulo, BRAZIL; 2Laboratory of Exercise
Physiology, Olympic Center of Training and Research, Sao Paulo, BRAZIL; 3Sao Paulo Association for Medicine Development,
Sao Paulo, BRAZIL; 4Department of Physiology, Federal University of Sao Paulo, Sao Paulo, BRAZIL; and 5Department of
Surgery, Federal University of Sao Paulo, Sao Paulo, BRAZIL

ABSTRACT
SABINO-CARVALHO, J. L., T. R. LOPES, T. OBEID-FREITAS, T. N. FERREIRA, J. E. SUCCI, A. C. SILVA, and B. M. SILVA.
Effect of Ischemic Preconditioning on Endurance Performance Does Not Surpass Placebo. Med. Sci. Sports Exerc., Vol. 49, No. 1,
pp. 124132, 2017. Purpose: Recent studies have reported ischemic preconditioning (IPC) can acutely improve endurance exercise
performance in athletes. However, placebo and nocebo effects have not been sufficiently controlled, and the effect on aerobic metabolism
parameters that determine endurance performance (e.g., oxygen cost of running, lactate threshold, and maximal oxygen uptake [VO2max])
has been equivocal. Thus, we circumvented limitations from previous studies to test the effect of IPC on aerobic metabolism parameters and
endurance performance in well-trained runners. Methods: Eighteen runners (14 men/4 women) were submitted to three interventions, in
random order: IPC; sham intervention (SHAM); and resting control (CT). Subjects were told both IPC and SHAM would improve
performance compared to CT (i.e., similar placebo induction), and IPC would be harmless despite circulatory occlusion sensations (i.e.,
nocebo avoidance). Next, pulmonary ventilation and gas exchange, blood lactate concentration, and perceived effort were measured during
a discontinuous incremental test on a treadmill. Then, a supramaximal test was used to verify the VO2max and assess endurance
performance (i.e., time to exhaustion). Results: Ventilation, oxygen uptake, carbon dioxide output, lactate concentration, and perceived
effort were similar among IPC, SHAM, and CT throughout the discontinuous incremental test (P 9 0.05). Oxygen cost of running, lactate
threshold, and VO2max were also similar among interventions (P 9 0.05). Time to exhaustion was longer after IPC (mean T SEM, 165.34 T
12.34 s) and SHAM (164.38 T 11.71 s) than CT (143.98 T 12.09 s; P = 0.02 and 0.03, respectively), but similar between IPC and
SHAM (P = 1.00). Conclusions: IPC did not change aerobic metabolism parameters, whereas improved endurance performance. The
IPC improvement, however, did not surpass the effect of a placebo intervention. Key Words: ISCHEMIC PRECONDITIONING,
AEROBIC EXERCISE, ERGOGENIC AID, PLACEBO EFFECT

I
schemic preconditioning (IPC; brief cycles of ischemia
followed by reperfusion) is well known to induce pro-
tection against ischemiareperfusion injury (9,26,36).
Adenosine triphosphate (ATP) sparing (36) and increase of
electron mitochondrial flux (9) are some of the mechanisms
responsible for the IPC protection. In addition, IPC abolishes
the endothelial function impairment induced by ischemia
reperfusion (26), which is mediated by the autonomic nervous
system (26). The beneficial effects of IPC on mitochondria
APPLIED SCIENCES
and blood vessels, if translated to exercising muscles, could,
supposedly, enhance efficiency of ATP use and resynthesis
by the aerobic metabolism, as well as could increase skeletal
muscle blood flow, leading to higher oxygen delivery and
metabolites removal. The hypothetical effects of IPC on mi-
tochondrial and vascular function have, consequently, driven
studies that investigated the ergogenic effect of IPC on en-
durance performance (19). A meta-analysis recently reviewed
8 studies in this area (38), and concluded the IPC induces a

small beneficial effect on endurance performance. Of note,

however, no study has sufficiently controlled placebo and
nocebo effects (37).
Address for correspondence: Bruno Moreira Silva, P.T., Ph.D., Botucatu St
862, Biomedical Sciences Building, 5th Floor, Sao Paulo, SP04023-062, Previous studies have compared the IPC to a sham inter-
Brazil; E-mail: silva.bruno@unifesp.br. vention where cuffs have been inflated at low pressure and
Submitted for publication June 2016. subjects have not been informed about the possible effect of
Accepted for publication August 2016. IPC on endurance performance (2,3,1214,37). Such ap-
0195-9131/17/4901-0124/0 proach can possibly bias the interpretation about the IPC
MEDICINE & SCIENCE IN SPORTS & EXERCISE effect on endurance performance, because IPC induces pain
Copyright 2016 by the American College of Sports Medicine during circulatory arrest and relief during reperfusion (25),
DOI: 10.1249/MSS.0000000000001088 which do not occur during the low cuff inflation. Thus, the net

124

Copyright 2016 by the American College of Sports Medicine. Unauthorized reproduction of this article is prohibited.
effect on subjects_ expectancy about the interventions is
unpredictable, such that subjects may think the IPC is either
beneficial (i.e., placebo) or harmful (i.e., nocebo). Placebo
and nocebo effects, in turn, could per se modify exercise
performance, according to randomized, controlled, and double-
blinded studies (4,6,11). One of these studies, for example,
showed a placebo supplement ingested during a cycling time
trial enhanced performance by 4.3% (11). On the other hand,
another study showed negative information provided about
ingestion of a capsule reduced sprint performance by 1.6%
(4). Therefore, the isolated effect of IPC on endurance per-
formance remains unclear, due to possible biases from pla-
cebo and nocebo effects.
The effect of IPC on aerobic metabolism parameters
that determine endurance performance has been equivocal
(2,12,14,17,18,24,38), perhaps, in part, because subjects
have not been told about the potential ergogenic effect of
IPC. The reason for that is that uncertainty about an inter-
vention increases individual variability in endurance per-
formance (11), which may reduce precision of performance
outcomes, and so, could possibly mask a small effect of IPC
on both aerobic metabolism parameters and endurance per-
formance. The varied effect of IPC on the maximal oxygen
uptake (VO2max) (2,1214,24), particularly, may still be at-
tributed to the lack of strict criteria to confirm its measure-
ment. Previous studies have used traditional criteria based
on attainment of VO2 plateau and a given level of RER, HR,
capillary blood lactate concentration, and perceived effort
(2,12,14). However, VO2 plateau is observed in only ap-
proximately 50% of well-trained subjects during a continu-
ous incremental test (30), and other criteria may be
insufficient to confirm the VO2max attainment (34). Thus, a
verification protocol has been developed to confirm the
VO2max identification (30), and its use would be helpful to
elucidate the effect of IPC on the VO2max.
The aim of this study was to investigate the effect of IPC
on aerobic metabolism parameters and endurance perfor-
mance in well-trained runners circumventing limitations
from previous studies. To dissect the placebo effect, we
compared IPC with a sham intervention (SHAM), and
resting control (CT). One of the researchers told the sub-
jects that IPC and SHAM would similarly improve perfor-
mance compared with CT (i.e., similar placebo induction),
and IPC would be harmless despite circulatory occlusion
sensations (i.e., nocebo avoidance). In addition, we used a
verification protocol to confirm the VO2max (30). We hy-
pothesized that 1) IPC would improve aerobic metabolism
parameters compared to SHAM and CT, and 2) both IPC
and SHAM would improve endurance performance com-
pared with CT, but the IPC improvement would surpass the
SHAM effect (i.e., placebo effect).
METHODS
Subjects. Twenty subjects volunteered to participate.
However, during the study, a man was excluded due to a
ISCHEMIC PRECONDITIONING AND AEROBIC EXERCISE
Copyright 2016 by the American College of Sports Medicine. Unauthorized reproduction of this article is prohibited.
muscle injury, and a woman was excluded due to symptoms
related to menstruation. Among the 18 subjects that com-
pleted all the study_s visits, 14 were men (mean T SEM; age =
22.3 T 0.9 yr; VO2max = 66.4 T 1.2 mLIkgj1Iminj1; body fat =
9.1% T 0.8%) and four women (age = 24.0 T 2.5 yr;
VO2max = 56.7 T 1.8 mLIkgj1Iminj1; body fat = 23.6% T
0.7%). All subjects were nonsmokers and were not taking
medications, supplements, treating orthopedic injuries, or
presented history of chronic diseases. They had been training
6 times per week, for 5.0 T 0.5 yr, with supervision of a coach
graduated in physical education. Moreover, they had been
competing in official middle and long distance races. The
ethics committee of the Federal University of Sao Paulo
(process 610.367) approved the study, and all subjects signed
an informed consent form before participating in the study.
Study design. The study was randomized, placebo-
controlled and nocebo-controlled, and crossed-over. Sub-
jects visited the laboratory four times. Visit 1 was used for
familiarization with the interventions and execution of a
continuous incremental test. In addition, in this visit, one of
the researchers told subjects that IPC and SHAM would
similarly improve performance compared to CT, and IPC
would be harmless despite circulatory occlusion sensations.
Then, on visits 2, 3, and 4, subjects were exposed to IPC,
SHAM, or CT, in random order, and next submitted to a
discontinuous incremental test and a supramaximal test.
Visits occurred at the same time of day for a given subject.
Interval between visits was 7 d. Subjects were asked to ar-
rive fully hydrated, ingest a light meal 2 h before the tests,
and not to ingest caffeine or alcoholic beverages for 24 h.
Training intensity was reduced on the day before the testing
sessions to allow adequate recovery from training, and
subjects were assessed only if they reported score of at least
15 (i.e., good recovery) in the total quality recovery scale
(23). Only one subject_s visit had to be postponed 1 d to
allow adequate recovery.
Ischemic preconditioning, sham, and control. IPC
was performed with subjects in supine position, using cus-
tomized cuffs (16). One cuff was placed in each thigh, as
close to the groin as possible. Cuffs had two independent
chambers installed in series. Each chamber was 36 cm long
and 17.5 cm wide. Together, they covered at least 80% of
the thigh_s circumference. Each cuff had a Velcro strap that,
in most cases, surrounded the tight at least two times. Cuffs
were inflated to 220 mm Hg for 5 min and deflated to 0 mm
APPLIED SCIENCES
Hg for 5 min (2). When one cuff was inflated, the other was
deflated (2). Four pressure cycles were performed on each
leg (2), thus the IPC procedure lasted 40 min. Circulatory
occlusion was confirmed with a vascular Doppler (Doppler
vascular 610B, MEDMEGA, Brazil) at the posterior tibial
artery. No pulse was detected in any subject while cuffs
were inflated.
The SHAM procedure consisted of simulating the ad-
ministration of therapeutic ultrasound. Physical therapists
commonly use therapeutic ultrasound in an attempt to
accelerate the healing process of musculoskeletal injuries,
Medicine & Science in Sports & Exercised 125
 supposedly, in part, because it increases deep tissues tem-
perature (15). All subjects had been treated with ultrasound
sometime before the study, and all had a positive belief that
it could aid the treatment of musculoskeletal injuries. Nev-
ertheless, before the study, subjects had little or none infor-
mation about the mechanisms involved on the ultrasound
therapeutic effects. In our study, one of the researchers
(J.L.S.) informed the subjects that the beneficial effects of
ultrasound application could not only heal musculoskeletal
injuries but also enhance exercise performance. If subjects
had doubts or were interested to know more details about the
possible mechanisms involved, the ultrasound heating effect
(15) was used to support the researcher_s explanation.
The ultrasound device was plugged in a power socket,
turned on, and its front lights were on. However, the start
button to deliver the ultrasound was not pushed, and subjects
did not notice that. The subjects lie down before and during
the ultrasound application. The ultrasound device was
placed close to the subjects_ leg and its panel was facing the
researcher that conducted the procedure (J.L.S.). Before
applying the ultrasound, this researcher pushed buttons on
the ultrasound to simulate the adjustment of settings and, at
last, always pushed the timer button, which emitted a sound.
Ultrasound gel was applied on the skin, and the ultrasound
probe was rubbed over the muscles of the anterior thigh,
posterior thigh, anterior leg, and posterior leg. The sham
ultrasound application was done for 5 min in each region,
alternating the application between limbs, which totaled
40 min. Noteworthy, active administration of therapeutic ul-
trasound does not provoke any relevant sensation, than the
sensations provoked by gel application and probe movement
on the skin. Thus, it was impossible for the subjects to notice
that the device was not delivering ultrasound.
The CT procedure consisted of lying supine for 40 min.
Subjects were not allowed to sleep during this period. After
the interventions administration, subjects stood on a tread-
mill to be instrumented for the exercise tests. At the end of
the instrumentation, on visits 3 and 4, or only on visit 4, one
of the researchers (J.L.S.) asked the following question: Do
you think your performance today will be equal, better, or
worse than the performance on the previous visit(s)? Sub-
jects were instructed to compare their performance versus
the CT day. In detail, if the CT was used on visit 2, subjects
compared the IPC and SHAM versus CT (e.g., IPC better
than CT). If the CT was used on visit 4, subjects were asked
APPLIED SCIENCES
on this visit to compare their expected performance versus
the previous visits, and later responses were inverted for sta-
tistical analyses (e.g., the response CT worse than IPC was
analyzed as IPC better than CT). These questions were used to
assess the placebo induction and nocebo avoidance. The same
researcher conducted all exposures to IPC, SHAM, and CT
(J.L.S.). This researcher was also responsible to inform the
subjects that IPC and SHAM would similarly improve perfor-
mance compared with CT, and IPC would be harmless despite
circulatory occlusion sensations. Moreover, this researcher
asked the question about performance expectancy. Just one
126 Official Journal of the American College of Sports Medicine
Copyright 2016 by the American College of Sports Medicine. Unauthorized reproduction of this article is prohibited.
researcher executed these methods, according with a system-
atic ritual, to standardize the placebo induction and nocebo
avoidance.
Exercise tests. Exercise tests were performed on a
treadmill (Super ATL, Inbrasport, Brazil), with grade set at
1% to approximate the energy expenditure of outdoor run-
ning (39). The continuous incremental test, conducted on
visit 1, provided data to define the discontinuous incre-
mental test, used on visits 2, 3, and 4. The continuous in-
cremental test consisted of baseline at 8 kmIhj1 for 3 min,
1 kmIhj1 increase in velocity per minute until voluntary
exhaustion, and recovery at 5 kmIhj1 for 5 min.
The discontinuous incremental test started with 6 min of
baseline at velocity 1 kmIhj1 lower than the velocity where
the ventilatory threshold was identified on the continuous
incremental test, which aimed to obtain steady state VO2
data for the oxygen cost of running interpretation (39). After
that, the discontinuous incremental test consisted of 3 min at
velocity 2 kmIhj1 higher than the baseline velocity,
followed by 1 kmIhj1 increase in velocity per stage until
voluntary exhaustion (Fig. 1). Each stage lasted 3 min and
was followed by a 30-s break for blood sampling. At the end
of each stage, the treadmill velocity was reduced to zero,
and, as soon as subjects could, they stepped off the treadmill
belt. They stood for blood sampling and then velocity was
increased to the next stage. When subjects reached exhaus-
tion, treadmill velocity was reduced to zero and subjects
recovered standing on the treadmill for 3 min. Next, they
recovery walking for 7 min at 5 kmIhj1.
After the recovery period from the discontinuous incre-
mental test, a supramaximal exercise test was started (Fig. 1).
The treadmill velocity was firstly set for 2 min at 60% of the
velocity of last completed stage on the previous discontin-
uous incremental test [i.e., peak velocity (Vpeak)]. Next,
velocity was set at 0.5 kmIhj1 higher than Vpeak until vo-
litional exhaustion (30). Subjects were blinded about the
treadmill velocity and all other data collected during the
study. The researcher that gave verbal encouragement during
the test was always the same for a given subject and was
blinded about the test results and the intervention that was
administered. All tests were conducted at temperature be-
tween 21-C and 23-C, and relative humidity between 45%
and 65%.
Measurements. Ventilation and pulmonary gas ex-
change were recorded breath by breath throughout the
exercise tests using a metabolic analyzer (Quark CPET,
Cosmed, Italy). O2 and CO2 analyzers were calibrated
according to the manufacturer_s specifications, using ambi-
ent air and gases with known concentration (16% O2 and 4%
CO2). The flow meter was calibrated using a 3-L syringe.
Electrocardiogram was continuously recorded (Powerlab,
AD Instruments, Australia) to quantify HR (LabChart, AD
Instruments, Australia). Twenty-five microliters of blood
were draw from the earlobe during the breaks of the
discontinuous incremental test. A vasodilator ointment was
used to arterialize blood samples (Finalgon, Boehringer
http://www.acsm-msse.org
FIGURE 1Illustration of the study_s experimental procedures conducted on visits two, three, and four. We firstly exposed subjects exposed to IPC,
SHAM, or CT, in random order. Then, we conducted a discontinuous incremental test and a supramaximal test. The baseline velocity was individ-
ualized. Thus, the absolute velocities presented in the Y axis are merely an example.
Mannheim, Germany). Blood was collected using heparinized
and calibrated capillaries, and then stored in Eppendorfs
containing 50 KL of 1% NaF (i.e., anticoagulant), until
analysis of lactate concentration (YSI 1500 SPORT, Yellow
Springs Instruments, USA). Rating of perceived exertion was
assessed during the continuous and discontinuous incremen-
tal tests using 010 Borg scale. Time to exhaustion was used
to assess endurance performance. Time was recorded using a
digital stopwatch. The chronometer was started when subjects
achieved the target supramaximal velocity, and stopped when
subjects asked to interrupt the test (i.e., volitional exhaustion),
despite strong verbal encouragement. Time to exhaustion was
recorded from all subjects, but only data obtained at the same
supramaximal velocity, for a given subject, were used for sta-
tistical analyses. As a result, for this variable, the sample size
was 15. Due to technical problems, the sample size for peak
ventilation, peak pulmonary gas exchange, and peak HR were
reduced to 16. In addition, blood samples were only obtained
from the first 14 subjects that participated in the study.
Data analyses. Pulmonary ventilation and gas exchange
were filtered to exclude aberrant breaths (two standard de-
viations from the mean of a 15-breath window). Oxygen cost
of running was calculated as the average VO2 of the last 2 min
of baseline of the discontinuous incremental test, divided by
the corresponding velocity in kilometers per minute (39). Two
experienced investigators independently identified the venti-
latory threshold using 20-s mean data. When there was no
agreement on the independent identification, investigators
reviewed the identification together. The following criteria
were used (1): 1) break point in CO2 output (VCO2) plotted
versus VO2; and 2) increase in ventilatory equivalent of O2
(VE/VO2) plotted versus time, without increase in ventilatory
equivalent of CO2 (VE/VCO2) plotted versus time. Lactate
threshold was determined using a mathematical model that
identified the point of intersection between two linear re-
gressions that yielded the least sum of squared differences
between the observed lactate values and the fitted values (31).
Peak lactate concentration in the discontinuous test was
ISCHEMIC PRECONDITIONING AND AEROBIC EXERCISE
Copyright 2016 by the American College of Sports Medicine. Unauthorized reproduction of this article is prohibited.
obtained from the blood sample collected after the last com-
pleted stage. Peak lactate concentration in the supramaximal
test was the highest value obtained from blood samples col-
lected at exhaustion and the third minute of recovery.
The highest 20-s mean VO2 during the exercise tests was
considered the peak oxygen uptake (VO2peak). If VO2peak of
the discontinuous and supramaximal tests was within the
tolerance of measurement error (i.e., 2%), the VO2max was
assumed to be achieved (30), and the highest value was
used for statistical analyzes. In our study, the VO2max
was confirmed in all cases. Beat by beat HR was reduced to
20-s means, and peak HR was the highest value during each
test. VO2, VCO2, lactate concentration, and perceived ef-
fort during the discontinuous test were linearly interpolated
to compare data throughout the test among the in-
terventions (Origin, Microcal, USA). VO2 and VCO2 in-
terpolation was done using 20-s mean data. The highest
common velocity among the interventions was considered
100% and, based on that, interpolated data corresponding
to 70%, 75%, 80%, 85%, 90%, and 95% were identified
and used for statistical analyses.
Statistical analyses. Sample size was calculated con-
sidering the endurance performance as the main endpoint
in one-way repeated measures ANOVA (G*Power 3.1
Dusseldorf University, Germany), as well as considering a
mean difference of 3% in endurance performance among
interventions (2,3,14), P value at 0.05 and power at 0.80. As a
APPLIED SCIENCES
result, the sample size calculation indicated that 15 subjects
would be necessary to conduct the study. All variables
showed normal distribution in the ShapiroWilk test. One-
way repeated measures ANOVA was used to compare IPC,
SHAM, and CT. The GreenhouseGeisser correction was
used to adjust ANOVA results, whenever sphericity was vi-
olated in the Mauchly test. The Bonferroni post hoc was used
when significant F values were found. Results are presented
as mean T SEM. All analyses were two-tailed, and statistical
significance was accepted for P G 0.05. Statistical analyses
were done using the software STATISTICA (Statsoft, USA).
Medicine & Science in Sports & Exercised 127
 RESULTS TABLE 2. Physiological, metabolic, and perceptual data at peak of the discontinuous
incremental test and the supramaximal test.
The subject_s best time is reported and compared with the n IPC SHAM CT P
corresponding world record in Table 1. On average, men and Incremental test
women_s best time were, respectively, 123% T 3% and 130% T VO2, mLIkgj1Iminj1 16 64.9 T 1.7 64.7 T 1.8 64.6 T 1.4 0.92
HR, bpm 16 190 T3 191 T3 189 T 3 0.10
3% of the corresponding world record (i.e., ~23% and ~30% RER, a.u. 16 1.01 T 0.01 1.02 T 0.01 1.03 T 0.01 0.42
above the world record). Score on the total quality recovery [La], mmolILj1 14 7.5 T 0.5 7.5 T 0.4 7.3 T 0.5 0.82
Borg, a.u. 18 8.8 T 0.4 8.7 T 0.5 9.2 T 0.4 0.26
scale was similar among interventions (IPC: 16.5 T 0.3 a.u. vs Supramaximal test
SHAM: 16.3 T 0.3 a.u. vs CT: 16.3 T 0.39 a.u., P = 0.52). VO2, mLIkgj1Iminj1 16 64.8 T 1.6 64.6 T 1.8 63.4 T 1.3 0.17
Occlusion pressure during IPC was recorded in the first four HR, bpm 16 189 T3 188 T3 189 T 3 0.71
RER, a.u. 16 1.00 T 0.01 1.00 T 0.02 1.00 T 0.01 0.80
subjects that participated in the study, and it varied between 140 [La], mmolILj1 14 9.3 T 0.6 8.9 T 0.5 9.4 T 0.5 0.37
and 150 mm Hg. The majority of the subjects expected per- Data are mean T SEM. Note that no difference was found for any variable among IPC,
formance would better after IPC than CT (better, 83%; equal, SHAM, and CT.
17%; worse, 0%) and after SHAM than CT (better, 78%; equal,
22%; worse, 0%). The percentage of subjects who responded
performance after IPC did not surpass the effect of a placebo
performance would be better versus CT was statistically similar
intervention, which refuted the ergogenic effect specifically
between IPC and SHAM (83% vs 78%; P = 0.41).
mediated by the IPC; and 2) the improvement in perfor-
Peak data were similar among interventions in the dis-
mance after IPC and SHAM was probably mediated by
continuous and supramaximal tests (Table 2). Ventilation,
neural mechanisms related to placebo induction, instead of
VO2, VCO2, lactate concentration, and perceived effort were
change in metabolic responses.
similar among IPC, SHAM, and CT throughout the discon-
The present study sought to induce similar placebo effect
tinuous test (P 9 0.05; Fig. 2). Oxygen cost of running,
on the IPC and SHAM interventions, and to avoid a possible
lactate threshold, and VO2max were also similar among in-
nocebo effect related to circulatory occlusion sensations.
terventions (P 9 0.05; Fig. 3). Noteworthy, VO2 was similar
Only one of the researchers interacted with the subjects to
between the fourth and sixth minute of baseline within all
induce placebo and avoid nocebo, according with a sys-
interventions (IPC: fourth minute, 39.70 mLIkgj1Iminj1 vs
tematic ritual, as the interlocutor interaction with the sub-
sixth minute, 39.66 mLIkgj1Iminj1, P = 0.98; SHAM:
jects can modulate the placebo effect in clinical studies (7).
fourth minute, 39.74 mLIkgj1Iminj1 vs sixth minute,
Most subjects, in fact, expected IPC and SHAM would im-
39.85 mLIkgj1Iminj1, P = 0.95; CT: fourth minute,
prove performance compared with CT, which indicates that,
39.78 mLIkgj1Iminj1 vs sixth minute, 39.93 mLIkgj1Iminj1,
in general, the placebo induction and nocebo avoidance was
P = 0.93). These results indicate that VO2 was at steady
successful. However, the positive expectancy about the in-
state. Time to exhaustion was longer after IPC (mean T
terventions effect was not unanimous. The lack of unanimity
SEM, 165.34 T 12.34 s; Fig. 4) and SHAM (164.38 T 11.71 s)
may be explained by the question used in the study, because
than CT (143.98 T 12.09 s; P = 0.02 and 0.03, respectively),
subjects were asked to compare their expected performance
but similar between IPC and SHAM (P = 1.00).
with the performance on previous visits. The response to this
question may rely not only on the researcher_s explanation
about the possible effect of interventions but also on any
DISCUSSION
factor that may modulate the subject well-being and moti-
We circumvented limitations from previous studies to test vation on the experiment day. In addition, the effect of pla-
the effect of IPC on aerobic metabolism parameters and cebo induction on endurance performance also depends on
endurance performance in well-trained runners. The data the subjects_ personality (6), which may explain part of the
showed no effect of IPC on aerobic metabolism parameters. between-subject variability on expectancy about the in-
Time to exhaustion, on the other hand, was longer in the IPC terventions effect.
and SHAM versus CT, but similar between IPC and SHAM. A recent meta-analysis reported a small beneficial effect
Therefore, the results indicate that 1) the improvement in of IPC on endurance performance (effect size, 0.51; 90%
APPLIED SCIENCES

TABLE 1. Subject_s best time in official races.

Men (n = 14) Women (n = 4)
Meters World Record (s) n Best Time (s) Best Time (%) World Record (s) n Best Time (s) Best Time (%)
800 100.91 4 114.04 T 1.91 113.01 T 1.89 115.03
1500 206.00 2 252.20 T 14.48 122.43 T 4.97 230.07
3000* 473.63 1 518.00 T 0.00 109.37 T 0.00 538.81 2 679.70 T 29.70 126.15 T 3.90
5000 757.35 4 1020.50 T 11.47 134.75 T 3.70 851.15 1 1160.00 T 00.00 136.29 T 0.00
10,000 1577.53 3 2076.67 T 65.67 131.64 T 3.40 1771.78 1 2340.10 T 00.00 132.08 T 0.00
Data are mean T SEM of absolute and percent values. Percent values were calculated in relation to the corresponding world record. If a subject had time recorded in more than one type of
race, the time that was closer to the world record was reported.
*With steeplechase.

128 Official Journal of the American College of Sports Medicine http://www.acsm-msse.org

Copyright 2016 by the American College of Sports Medicine. Unauthorized reproduction of this article is prohibited.
FIGURE 2Data are mean T SEM of the oxygen uptake (VO2; panel A; n = 16), carbon dioxide output (VCO2; panel B; n = 16), lactate concentration
([La]; panel C; n = 14), and ratio of perceived effort (panel D; n = 18) throughout the discontinuous incremental test. The highest common velocity
among the interventions was considered 100%, and, based on that, interpolated data corresponding to 70%, 75%, 80%, 85%, 90%, and 95% were
identified and used for statistical analyses. Note that no difference was found for any variable among IPC, SHAM, and CT.

confidence interval, 0.350.67), based on the analysis of eight
studies (38). Some of these studies specifically investigated
the effect of IPC on time to exhaustion during constant
workload tasks (3,13,24), which is similar to the endurance
assessment used in the present study. IPC prolonged time to
exhaustion during rhythmic handgrip exercise (3), cycling at
heavy intensity (24), and cycling at maximal intensity (13).
However, these and other studies about the effect of IPC on
endurance performance did not sufficiently control placebo
and nocebo effects, which complicate the interpretation about
the isolated effect of IPC (37). In this sense, Marocolo et al.
(28) attempted to exclude the placebo effect to test the ergo-
genic effect of IPC on a 100-m time trial and a bout of lower
limb resistance exercise (29). Authors told subjects_ IPC and
low-pressure cuff inflation would improve performance ver-
sus resting CT. They found both interventions improved
performance compared with CT, but there was no difference
between IPC and low-pressure cuff inflation, which does not
support the ergogenic effect specifically mediated by the IPC.
A recent study from our group, on the other hand, com-
pared the effect of IPC, a sham intervention, and low cuff
inflation on swimming performance in a repeated sprint
swimming task (16). The sham intervention consisted of
short-time circulatory occlusion, which approximated signs
and symptoms of IPC, without attaining enough duration
that could possibly induce ergogenic effect. Similarly to the
present study, a researcher told subjects that IPC and a sham
intervention would improve performance compared with the
CT condition, and that IPC and sham intervention would
cause absolutely no harm. Most subjects in fact believed IPC
and sham intervention would improve performance. At the
end, the IPC enhanced swimming performance, whereas the
sham intervention did not, which supported the ergogenic
effect specifically mediated by the IPC.
In the present study, data did not support that IPC per se
mediated ergogenic effect on endurance performance. The
findings herein presented are therefore in contrast to those from
the recent study of our group (16). Of note, however, in the
present study, we applied IPC once, 10 min before the per-
formance assessment. In the former study, we applied IPC on
three occasions (48 h, 24 h, and 30 min) before the perfor-
mance assessment, which may have summed early and late
effects of IPC and amplified the IPC effect (26,40). In addition,
the exercise mode was different (running vs swimming), as
well as the testing protocol (constant velocity at supramaximal
intensity vs repeated sprints). Consequently, these characteris-
tics may be related to the distinct effect of IPC between these
studies, which should be further investigated to enable the use
of IPC to enhance exercise performance.
Multiple mechanisms are known to regulate endurance
performance, including determinants of oxygen delivery and
utilization (20), and mechanisms that modulate perceived
effort (27). Nevertheless, three variables summarize most of
APPLIED SCIENCES
the contribution of aerobic mechanisms to endurance per-
formance (20,22), that is oxygen cost of running, lactate
threshold, and VO2max (20,22). In our study, oxygen cost of
running was unaffected by IPC and, as far as we know, no
study has reported reduced oxygen cost at submaximal ex-
ercise intensities (2,14,18,24,32). Altogether, the present
and previous findings suggest that IPC does not change the
efficiency of the aerobic metabolism during whole body
exercise performed at submaximal intensity. The lack of IPC
effect on the aerobic metabolism during exercise is some-
what surprising, because IPC induces ATP sparing (36) and

ISCHEMIC PRECONDITIONING AND AEROBIC EXERCISE Medicine & Science in Sports & Exercised 129

Copyright 2016 by the American College of Sports Medicine. Unauthorized reproduction of this article is prohibited.

FIGURE 3Individual data (dashed lines) and mean T SEM (bars and
whiskers) of the oxygen cost of running (OCR; panel A; n = 18), lactate
threshold (LT; panel B; n = 14), and maximal oxygen uptake (VO2max,
panel C; n = 16) in the IPC, SHAM, and CT. Note that no difference
was found for any variable among interventions.
augment electron mitochondrial flux during prolonged ische-
mia (9). Therefore, the mechanisms that mediate ergogenic
effects of IPC during whole body aerobic exercise, if any,
may not be the same to the protective effects mediated by IPC
during prolonged ischemia and subsequent reperfusion.
Bailey et al. (2) reported that IPC decreased blood lactate
concentration during an incremental treadmill test, which
trended to dislocate the onset of blood lactate accumulation to
APPLIED SCIENCES
higher running velocity. In contrast, Crisafulli et al. (12), de
Groot et al. (14), Jean-St-Michel et al. (21), and Patterson et al.
(33) found no effect of IPC on blood lactate concentration
during varied exercise tests. In the present study, after con-
trolling placebo and nocebo effects, there was no effect of IPC
on the blood lactate concentration throughout exercise inten-
sities, as well as on the lactate threshold. Thus, the present
results supported that IPC did not change blood lactate levels
during an incremental treadmill test in well-trained runners.
The effect of IPC on the VO2max has also been equivocal.
For example, de Groot et al. (14) and Cruz et al. (13)
130 Official Journal of the American College of Sports Medicine
Copyright 2016 by the American College of Sports Medicine. Unauthorized reproduction of this article is prohibited.
reported IPC increased VO2max during cycling tests, but
others did not corroborate this finding (2,12). Nevertheless,
all previous IPC studies did not use strict criteria to confirm
the VO2max. In the present study, therefore, we used a veri-
fication protocol. The protocol successfully confirmed the
VO2max in all subjects, although peak RER was relatively
low and peak perceived effort was not always at score 10.
These findings have been previously reported in discontinuous
incremental tests (30), and restexercise transitions (10). Of
note, the low peak RER has been attributed to increase in en-
dogenous CO2 stores during restexercise transitions (10).
However, without the use of the verification protocol, doubts
about the VO2max attainment could have been raised. Conse-
quently, the verification protocol strengthens the interpretation
that IPC did not change the VO2max in the present study.
Given that IPC and SHAM similarly enhanced endurance
performance, without any change in metabolic responses,
neural mechanisms related to placebo induction could be
responsible for the performance enhancement. In fact, strong
evidences support that neural factors can per se determine
endurance exercise performance, independent from changes
in metabolic responses (27,32), but the mechanisms specif-
ically involved in placebo-induced exercise performance
enhancement are unknown (5). It is possible, however, that
some mechanisms already described in clinical settings may
be involved. For instance, Benedetti et al. (8) manipulated
the meaning of a painful experience from negative to posi-
tive through verbal suggestions. The manipulation resulted
in increased pain endurance, which was mediated by
coactivation of opioid and cannabinoid pathways (8).
One of the study_s limitations was the assessment of en-
durance performance using an open-ended laboratory test
(i.e., the end is not fixed) performed at constant velocity,
instead of a close-ended task (i.e., the end is fixed)
performed at subject_s preferred pace. The latter is nearer to
a real sport event and is more physiologically complex.
Thus, our findings may not be extrapolated to other situa-
tions. Another limitation was the measurement of VO2 at the
lungs and lactate concentration at ear capillaries. These
measurements may not exactly reflect the VO2 and lactate
FIGURE 4Individual data (dashed lines) and mean T SEM (bars and
whiskers) of the time to exhaustion (Tlim; n = 15) in the IPC, SHAM,
and CT. Note that Tlim was higher in the IPC and SHAM versus CT,
but there was no difference between IPC and SHAM.
http://www.acsm-msse.org
concentration at contracting skeletal muscles nor differentiate
metabolic responses between subtypes of skeletal fibers. IPC
was applied just once, 10 min before the performance as-
sessment, which may have not been enough to induce ergo-
genic effects in well-trained subjects. Or, alternatively, the
effect of IPC in well-trained subjects is blunted compared with
less fit subjects, which, for example, has been reported for the
ergogenic effect of nitrate supplementation (35). Furthermore,
IPC may not change metabolic responses, but instead, mod-
ifies neuromuscular mechanisms, such as muscle activation
(13) and/or rate of muscle contraction and relaxation (3),
which deserves further investigation. At last, we assessed
men and women, but our sample size was not enough to
dissect whether gender modulates the effect of IPC.
In conclusion, IPC did not change aerobic metabolism
parameters, whereas improved endurance performance in
well-trained runners. The IPC improvement, however, did
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