No Muscle Is an Island: Integrative

SPECIAL COMMUNICATIONS
Perspectives on Muscle Fatigue
JANE A. KENT1, NIELS KRTENBLAD2,3, MICHAEL C. HOGAN4, DAVID C. POOLE5, and TIMOTHY I. MUSCH5
1
Department of Kinesiology, University of Massachusetts, Amherst MA; 2Institute of Sports Science and Clinical Biomechanics,
University of Southern Denmark, Odense, DENMARK; 3Department of Health Sciences, Mid Sweden University, Östersund,
SWEDEN; 4Department of Medicine, University of California, San Diego, CA; and 5Department of Kinesiology, Kansas State
University, Manhattan, KS

ABSTRACT
KENT, J. A., N. KRTENBLAD, M. C. HOGAN, D. C. POOLE, and T. I. MUSCH. No Muscle Is an Island: Integrative Perspectives on
Downloaded from http://journals.lww.com/acsm-msse by BhDMf5ePHKav1zEoum1tQfN4a+kJLhEZgbsIHo4XMi0hCywCX1AWnYQp/IlQrHD3i3D0OdRyi7TvSFl4Cf3VC1y0abggQZXdtwnfKZBYtws= on 03/22/2021

Muscle Fatigue. Med. Sci. Sports Exerc., Vol. 48, No. 11, pp. 2281–2293, 2016. Muscle fatigue has been studied with a variety
approaches, tools and technologies. The foci of these studies have ranged tremendously, from molecules to the entire organism. Single
cell and animal models have been used to gain mechanistic insight into the fatigue process. The theme of this review is the concept that
the mechanisms of muscle fatigue do not occur in isolation in vivo: muscular work is supported by many complex physiological systems,
any of which could fail during exercise and thus contribute to fatigue. To advance our overall understanding of fatigue, a combination of
models and approaches is necessary. In this review, we examine the roles that neuromuscular properties, intracellular glycogen, oxygen
metabolism, and blood flow play in the fatigue process during exercise and pathological conditions. Key Words: BIOENERGETICS,
GLYCOGEN, OXYGEN, PERFUSION, OXIDATIVE PHOSPHORYLATION, GLYCOLYSIS

M
uscle fatigue, defined as the fall in maximal force
or power production in response to contractile
activity, by definition arises at least in part from
failure of cross-bridge cycling within the muscle cells.
However, this contractile machinery relies on multiple sys-
tems to support its work, including the nervous, vascular,
and energy systems. Thus, as illustrated in Figure 1, failure
at any of the sites upstream from the cross-bridges can and
does contribute to the development of fatigue (54). As such,
no muscle operates in isolation in vivo. It is well understood
at this time that fatigue is complex and task-specific, making
the use of both human studies and animal models important. The
focus of this review is to provide examples of how integrating a
variety of approaches and techniques can provide a more com-
plete understanding of muscle fatigue. Although sometimes re-
sults appear contradictory or inconsistent, the wide array of
conditions, protocols, and models used to study fatigue con-
tributes to the advancement of this exciting field of inquiry.
Using Fatigue to Probe the Integration of
Physiological Systems In Vivo
Integrative approaches to studying muscle
fatigue. As so well by implemented by the work of Brenda
Address for correspondence: Jane A. Kent, Ph.D., Department of Kine-
siology, Totman 160A, University of Massachusetts, Amherst MA 01003;
E-mail: jkent@kin.umass.edu.
Submitted for publication January 2016.
Accepted for publication May 2016.
0195-9131/16/4811-2281/0
MEDICINE & SCIENCE IN SPORTS & EXERCISEÒ
Copyright Ó 2016 by the American College of Sports Medicine
DOI: 10.1249/MSS.0000000000001052
Bigland-Ritchie and colleagues in the 1970s and 1980s
(10,11,91), in vivo studies of skeletal muscle fatigue in
humans benefit from an integrated approach that captures
information about the neural and contractile properties of
muscle during a variety of contraction protocols. For ex-
ample, using both voluntary and electrically stimulated
contractions in a given fatigue protocol allows comparison
of the entire pathway of force production with those events
occurring distal to the point of stimulation (see Neural ac-
tivation, below); methods such as transcranial magnetic
stimulation and EMG have been essential to advancing our
understanding of these neural processes that contribute to
muscle fatigue (54). This approach was expanded in the late
1980s and 1990s by the addition of measures of intracellular
energy metabolism using in vivo magnetic resonance spec-
troscopy (MRS) (4,55,67). With addition of this technique,
the role of energy production and the inhibition of force
production by its by-products (Fig. 1, Bioenergetics) became
possible in vivo. Because studies of human muscle fatigue
generally involve noninvasive measures, an approach that
tackles the question of fatigue from many different directions
can be used to infer much about the causes of contraction-
induced loss of maximal muscle force or power.
Neural activation. Neural activation of the muscle in-
volves transmission of the signal to contract from the brain
to the muscle_s transverse tubules and sarcoplasmic reticu-
lum (SR), with subsequent release of calcium into the cy-
tosol by the SR and initiation of cross-bridge cycling. A
combination of voluntary and stimulated muscle contrac-
tions can be used to evaluate changes in ‘‘central’’ and
‘‘peripheral’’ activation of muscle, with those distinctions
largely established by the location of the stimulation elec-
trodes. For example, a greater decrease in maximal voluntary

2281

Copyright © 2016 by the American College of Sports Medicine. Unauthorized reproduction of this article is prohibited.
SPECIAL COMMUNICATIONS

FIGURE 1—Pathway of force production. A simplified, conceptual schematic illustrating some of the sites that may contribute to fatigue during
muscular work in vivo. The signal to contract arises from the central nervous system (Central Activation) and involves the recruitment and rate coding
of motor units to modulate force production. The next step, peripheral activation, involves the transmission of AP across the NMJ and along the muscle
membrane, where the signal then enters the transverse tubules and initiates contraction in the process of ECC. Bioenergetic processes provide the
energy to support contractile function, and accumulation of by-products from these processes can inhibit force production. Delivery of oxygen and
removal of by-products can support bioenergetics and delay fatigue. Adapted from Kent-Braun (53).

force compared with the force produced by a separate
supramaximal stimulation during a fatiguing contraction pro-
tocol provides evidence of ‘‘central’’ fatigue, or fatigue arising
from sources proximal to the point of stimulation (10,11,52).
An increase in force in response to superimposition of a
stimulated contraction during a maximal voluntary contrac-
tion can be used to detect incomplete voluntary activation of
the muscle (52). Sophisticated techniques including trans-
cranial magnetic stimulation of the brain and spinal cord and
complex EMG electrode arrays have also been developed to
parse out more specifically the sites of failure in the brain and
spinal cord that may contribute to fatigue (54).
Peripheral activation can be evaluated by changes in the
m-wave or compound muscle action potential (AP), which is
recorded by surface EMG electrodes placed over the muscle.
The m-wave is the electrical response to a single, supramax-
imal stimulation applied to the motor nerve just proximal to
its innervation point at the muscle. During fatigue, changes in
the amplitude of the compound muscle AP reflect an im-
pairment of propagation at the neuromuscular junction (NMJ)
or muscle membrane. Typically, failure at this site is un-
common during voluntary contractions except during very
high-intensity, prolonged contraction protocols (52), although
high-frequency stimulation protocols can induce this type of
peripheral ‘‘electrical’’ fatigue (37). In summary, much at-
tention remains focused on clarifying the complex contribu-
tions of neural activation to muscle fatigue in vivo.
Muscle bioenergetics. Muscular work must be sup-
ported by the ready supply of energy for adenosine triphosphate
(ATP)ases functioning at the sarcolemma, SR and cross-
bridges. The increased energy demand of contraction will re-
sult in the accumulation of metabolites known to induce
contractile failure, including inorganic phosphate (Pi) and
hydrogen ion (H+; (54). Phosphorus MRS can be used to
quantify noninvasively and continuously (with ~2–10 s tem-
poral resolution) the changes in these and other energy me-
tabolites in the cytosol of working muscle (4,52,55,62).
Various protocols have been used to probe the associations
between fatigue and metabolic inhibition of contraction, and
the results are consistent with those observed at the cellular
(32) and molecular (24) levels. More recently, use of proton
MRS has shown that intracellular PO2 is maintained well above
critical PO2 during single muscle group contractions (63), and
thus oxygen availability does not appear to be a factor per se
in fatigue under conditions of unrestricted blood flow (BF).
Contractile properties. The same stimulated contrac-
tions that are used to quantify changes in peripheral activa-
tion of the muscle can be used to indirectly evaluate events
such as excitation–contraction coupling (ECC) and calcium
kinetics within the muscle during fatigue. Decreases in force
in response to supramaximal stimulation of the motor nerve
following a contraction protocol indicate the magnitude of
peripheral fatigue, or that arising from changes at the NMJ,
muscle membrane or within the myocytes themselves (67).
Further, a decrease in stimulated twitch or tetanic force in the
absence of a decrease in the amplitude of the m-wave sug-
gests that the failure of force production is occurring distal to
the sarcolemma (52). Changes during stimulated contractions

2282 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.
in the rates of force development and, more commonly, re-
laxation also support the interpretation of contraction-induced
changes in calcium kinetics or cross-bridge cycling (11).
In the next three sections, we describe three studies that
applied different combinations of the measures described
above to address distinct scientific questions. In the first case,
muscle fatigue was used to perturb homeostasis in individuals
with multiple sclerosis (MS), to evaluate the cardiovascular
response to contractions in this population (70). In another
example of the utility of an integrative approach to evaluating
fatigue mechanisms, an in silico study of fatigue that built
upon studies in vivo by manipulating known factors in fatigue
is described (16). Finally, our third example illustrates how
understanding the bioenergetic contributions to fatigue can
explain the fatigue resistance of older muscle under isometric
conditions in vivo, and reveal new information about age-
related changes in muscle metabolism.
Fatigue as a means of understanding multiple
physiological systems: MS. Fatigue protocols can also
be used to stress the intact organism to evaluate the inte-
gration of various physiological systems in pathological
conditions. For example, in response to a report that per-
sons with MS had an inadequate blood pressure response
to isometric forearm contractions (76), a fatigue study was
conducted during which simultaneous measures of isometric
force, beat-by-beat heart rate and blood pressure, intracellular
energy metabolites and pH, neural activation and ratings of
perceived exertion were obtained (70). The hypothesis tested
was that the blunted pressor response in MS observed by
Pepin et al. (76) was in fact appropriate to a blunted metabolic
response to the contraction protocol, as had been observed
previously in this population (57). Although endurance time, and
changes in heart rate, perceived exertion and neural activa-
tion were similar in MS and controls during the sustained
submaximal isometric contraction, the pressor response and
changes in [Pi] and pH were blunted in MS compared with
controls. Further, the changes in mean arterial pressure were
associated with the changes in both [Pi] and pH. Notably, the
MS group had a normal response to the cold pressor test and
Valsalva maneuver, both nonexercise tests of autonomic
function. Thus, this integrative fatigue paradigm revealed that
the blunted pressor response in MS was appropriate to the
smaller metabolic perturbation during fatigue, and not a result
of cardiovascular dysautonomia in this cohort.
Integrative, in silico study of fatigue. Due to the com-
plexity and interrelationships between systems that support
muscle force production, the study of fatigue in vivo is some-
what limited to indirect measures. Recently, a computational
model was developed to address this limitation (16,17). In-
formation in the literature from in vivo studies of neural ac-
tivation, contractile characteristics, and energy metabolism
were combined with de novo experimental observations to
develop the model. This relatively comprehensive model used
an integrated approach, including some preexisting compo-
nents (33), to evaluate the sites of failure during fatigue of
human skeletal muscle. This approach allows manipulation of
INTEGRATIVE PERSPECTIVES ON FATIGUE
Copyright © 2016 by the American College of Sports Medicine. Unauthorized reproduction of this article is prohibited.
specific variables representing explicit physiological func-
SPECIAL COMMUNICATIONS
tions to determine their independent effects on fatigue; once
developed, the model was applied to the question of age-
related fatigue resistance.
The approach involved development of distinct modules
reflecting relevant physiological processes including motor
unit behavior, bioenergetics, contractile properties, calcium
and force-generation kinetics, and biomechanical properties
(Fig. 2) (16,17). The modules were populated with data from
the literature, and once the forward dynamics simulations
were initiated, no further input was given to the model. The
output from the model agreed well with reports from in vivo
studies (62), both in young muscle and when adjusted to
reflect age-related changes in neuromuscular properties (16).
Changes in phosphocreatine and pH reflected changes in
experimental data, including age-related differences. Fatigue
was well correlated with the concentration of diprotonated
Pi, in both age groups. Analysis of the model output in re-
sponse to various manipulations of the modules indicated
that the predominant effect of age was that of a decreased
reliance on glycolytic ATP production in the older muscle; a
result that agrees well with available in vivo data (62). Thus,
a modular, computational approach to the study of fatigue is
feasible and allows independent manipulation of various
mechanisms of fatigue. Future applications in aging and
pathological conditions should prove useful in augmenting
in vivo studies and our overall understanding of the mecha-
nisms of fatigue.
Age-related differences in acidosis during mus-
cle contractions. On balance, the literature is clear in
showing that older muscle fatigues less than young during
isometric contraction protocols (19). Under these conditions,
the primary source of this difference in fatigue appears to be
the fact that older muscle develops markedly less intra-
cellular acidosis than young muscle during a variety of
contraction protocols (20,56,62). Concomitantly, muscle in
healthy older adults relies more on oxidative metabolism to
meet the ATP needs of the cell during contraction (20,62).
To determine whether this difference in energetic response
was related to an inability of older muscle to increase gly-
colytic flux to the same extent as young muscle, acidosis and
ATP production from the creatine kinase reaction, glycolysis
and oxidative phosphorylation were determined in young
and older adults during contraction protocols in which BF to
the lower leg was left intact or occluded (62). The results of
this study showed that, although older muscle relied relatively
less on glycolytic ATP production during free-flow condi-
tions compared with young, this difference was abolished
during ischemia, indicating no impairment on the part of
the older muscle to produce energy from glycolysis when
necessary. Notably, the age-related difference in fatigue ob-
served during free-flow conditions were also found during
ischemic contractions; thus differences in fatigue were not
attributable to differences in oxygen availability. In a follow-
up study, the availability of glycogen and intracellular oxygen
as substrates for energy metabolism was examined using
Medicine & Science in Sports & Exercised 2283
SPECIAL COMMUNICATIONS
FIGURE 2—Computational model of fatigue. Illustration of the modules in a computational model of fatigue that correspond to physiological events in
the neuromuscular system. Simulations can be a useful approach to understanding the interrelationships between different processes involved in
repeated muscle contractions. Adapted from Callahan et al. (16,17).
carbon-13 and proton spectroscopy, respectively (Tonson et al,
personal communication). Once again, age-related differences
in acidosis and fatigue during intermittent maximal contrac-
tions were observed, despite similar [glycogen] at baseline and
rapid equilibration of cytosolic PO2 well above critical PO2
(63) in both age groups during the contraction protocol. Thus,
by combining approaches, it was demonstrated that age-
related differences in intracellular acidosis were not a result
of differences in substrate availability, at least in the form of
glycogen and oxygen.
Role of Glycogen in Skeletal Muscle ECC
and Function
As noted in section 2, mechanistic studies of contraction-
induced loss of maximal muscle force or power in skeletal
muscle have benefited from a whole-body, integrative ap-
proach. However, this approach does not tackle the question
of fatigue at the isolated whole muscle or cellular levels, which
can help explain processes observed in vivo. Indeed, single
fiber studies have provided invaluable knowledge about fa-
tigue mechanisms in the complex systems that support mus-
cle force, for example, myosin–actin interactions, regulation
of cytosolic Ca2+ by the SR, the force–Ca2+ relationship and
the role of cellular metabolism (1,72,89). In the following
sections, we discuss how one of these important systems,
cellular glycogen metabolism, affects muscle fatigue.
2284 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.
Compartmentalized energy utilization and pro-
duction in the muscle cell. Skeletal muscle is faced with
challenging problems related to metabolic regulation during
exercise, where energy turnover nearly instantly can increase
several hundred-fold during high intensity exercise and more
than 20-fold during more prolonged aerobic exercise (85).
This is achieved without substantial and potentially deleteri-
ous decreases in global myocellular ATP concentration,
demonstrating a remarkable precision of adjusting the rate of
ATP generating processes to the energy requirements (44).
However, the muscle cell forms many microenvironments
with restricted diffusional access of metabolites and high
ATPase activity during muscle contraction, which may result
in some locations within the cell where the metabolic envi-
ronment differs from the measured global energy status.
The sequence of events leading to muscle contraction is
known as ECC (Fig. 3) and a number of the complex steps in
ECC and relaxation are either directly or indirectly depen-
dent on the energy level of the muscle fiber. Both the ion
pumping at the transverse tubular (t)-system and sarcolemma
(Na+/K+-ATPase) as well as the myosin ATPases at the
contractile apparatus and the SR Ca2+-ATPase are directly
dependent on ATP (Fig. 3, steps 1, 5, and 6) and are affected
by either low (ATP) or high by-products of metabolism
(e.g., ADP, Pi). Further, some of the ion channels involved
in muscle activation and relaxation are indirectly regulated
by the energy status of the cell, including the SR Ca2+
http://www.acsm-msse.org
 SPECIAL COMMUNICATIONS
FIGURE 3—Skeletal muscle ECC. The sequence of complex events leading to the muscle contraction, known as ECC, starts with AP depolarizing the
sarcolemma and the t-tubule membrane (i.e., excitation), which induces conformational changes of the voltage sensors in the triad junction (steps 1–2).
The activation of the voltage sensors mediates an opening of the Ca2+ release channels (RyR1) in the sarcoplasmic reticulum (SR) and a subsequent 10-
fold to 20-fold increase in the cytosolic [Ca2+], which in turn initiates the cross-bridge cycle and force generation in the muscle fiber (steps 3–5). The
ensuing buffering and pumping or Ca2+ back to the luminal side of the SR, leads to muscle relaxation (step 6). Low muscle glycogen impair the SR
Ca2+ release, which in turn decreases the AP-activated increase in cytosolic [Ca2+] and hence the contractile activation. Further, low glycogen affects
the SR Ca2+ uptake and Na+/K+-pump activity.

release channel (25). The highly organized muscle cell forms
many compartments and thereby may have microenviron-
ments with high ATPase activity and restricted diffusional
access of metabolites. Thus, skeletal muscle cells represent
an example of spatial organization where energy production
from energy stores and energy-dependent steps in ECC are
not located in close proximity. Still, within this organization,
glycogen particles are distributed in local depositions and
thereby serve as an efficient energy store for the different
steps in ECC.
Muscle glycogen. Glycogen is the carbohydrate energy
store for ATP production and located primarily within muscle
and liver cells. Glycogen can be observed by electron mi-
croscopy as spherical particles, widely distributed within the
cell in close proximity to the sites of energy consumption,
creating optimal conditions for efficient regulation (72,75).
Conventionally, three distinct subcellular localizations of
glycogen have been defined: i) intermyofibrillar glycogen,
which is located between the myofibrils in close proximity to
SR and mitochondria; ii) intramyofibrillar glycogen, which is
located within the myofibrils interspersed among the con-
tractile filaments most often in the I-band of the sarcomere;
and iii) subsarcolemmal glycogen, which is clustered just
beneath the sarcolemma primarily next to mitochondria,
lipids, and nuclei (72). In relative terms, intermyofibrillar
glycogen is the major site of glycogen deposition, constitut-
ing approximately 75% of the cell_s total store of glycogen,
whereas intramyofibrillar and subsarcolemmal glycogen each
account for 5%–15% of total glycogen. The glycogen particles
are connected with proteins involved in glycogen metabolism
and form a dynamic carbohydrate–protein complex compris-
ing a subcellular compartment that has the ability to respond to
the metabolic requirements of the cell (29,88). Moreover, it
has been demonstrated that these glycogen particles can be
associated physically with the SR (29,88).
Glycogen and muscle function. In line with the
glycogen particle distribution, studies from the single fiber
to the whole body level collectively indicate that steps in
muscle ECC and relaxation are affected by glycogen levels,
which may link low glycogen levels with a decreased mus-
cle function (18,73,75). Thus, there is now compelling evi-
dence that low muscle glycogen and/or glycolytic-derived
energy are associated with SR Ca2+ release and reuptake,
and Na+/K+-pump function (Fig. 3, steps 1, 3, 4 and 6; for
review see (72,75). This may also explain the importance of
glycogen as a fuel during exercise, which is a fundamental
concept in exercise physiology. Some of the first studies
using the needle biopsy technique, in the late 1960s, dem-
onstrated that there is a strong correlation between muscle
glycogen content and endurance capacity during prolonged
cycling exercise (9), and an inability to continue exercise
when glycogen stores are limited (42). These observations
have subsequently been confirmed numerous times and it is
now well established that glycogen oxidation is of major
importance for ATP regeneration during both prolonged
exercise (91 h), and also during high-intensity intermittent
exercise (39). In this context, it is noteworthy that endurance
and high-intensity exercise training not only improve

INTEGRATIVE PERSPECTIVES ON FATIGUE Medicine & Science in Sports & Exercised 2285

Copyright © 2016 by the American College of Sports Medicine. Unauthorized reproduction of this article is prohibited.
 performance, but also increase muscle glycogen content
SPECIAL COMMUNICATIONS
(39). However, the link between glycogen depletion and the
development of fatigue, as well as the precise mechanism(s)
whereby muscle glycogen affects the series of events that
ultimately result in fatigue, are not fully understood.
Muscle glycogen can delay fatigue by maintaining energy
transduction during aerobic exercise, as the maximum rate
of ATP production is much higher for muscle glycogen than
for blood glucose or fat oxidation (85). Further, glycogen
may be important by producing tricarboxylic acid cycle in-
termediates permitting the maintenance of oxidative me-
tabolism (85). Low glycogen per se may limit glycogen
phosphorylase activity and hence the overall glycolytic rate.
Although the in vitro Km values of phosphorylase for gly-
cogen are G3 mM glucosyl units and the cellular concen-
tration is usually 9100 mM glucosyl units, the Km of
phosphorylase for glycogen is substantially higher in vivo
than in vitro (51). Interestingly, depletion of glycogen par-
ticles may be localization-dependent, with a higher relative
utilization of intramyofibrillar glycogen content (75).
The important role of muscle glycogen is also apparent
when studying its effect on the steps of ECC, which is con-
sistent with the understanding of muscle glycogen as having
rapid mobilization, being the precursor of glycolytic derived
ATP, having a high ATP production rate and that glycogen
and the glycolytic enzymes are widely distributed within the
cell in close proximity to the energy consumption locations.
Muscle glycogen and SR function. Studies from the
levels of SR vesicles, mechanically skinned and intact ro-
dent single fibers, and human studies have pointed to a
modulating role of glycogen availability on SR Ca2+ han-
dling (18,26,73,74). A reduction in [glycogen] within the
myofibrils is correlated with impaired SR Ca2+ release both
in human muscle after prolonged exercise and in skinned rat
muscle fibers after repeated tetanic stimulation (73,74).
Furthermore, it was recently shown that intact single mouse
muscle fibers undergoing repeated tetanic stimulations ex-
hibit a steep decrease in tetanic cytosolic [Ca2+] when
intramyofibrillar glycogen within the myofibrils reaches low
levels (71). Thus, the results of several recent studies support
a link between glycogen depletion within myofibrils and
decreased SR Ca2+ release. At present, little is known about
the precise mechanism(s) that link low glycogen levels in
the muscle with an impaired SR Ca2+ release rate.
Possible effects of glycogen on the ryanodine
receptor. In skeletal muscle, Ca2+ is released from the SR
Ca2+ stores via specific Ca2+ channels (RyR1 isoform in skel-
etal muscle, Fig. 3, step 3). The RyR1 channels are located at
the junctional SR of the SR-t tubule triad, which ensures effi-
cient Ca2+ release to the contractile proteins. The RyR channels
are modulated by numerous factors and in intact muscle, RyR1
interacts with multiple molecules and metabolites and is regu-
lated by various cellular processes (e.g., phosphorylation and
oxidation). During exercise and with glycogen depletion, the
main physiological modulators of RyR are conceivably: 1)
Ca2+ (both cytosolic and SR luminal), 2) protein phosphorylation,
2286 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.
3) the redox state, and 4) the cellular energy status (for review,
see Dhar-Chowdhury et al. (25).
With respect to the role of glycogen affecting the SR Ca2+
release, low glycogen may lead to local changes in meta-
bolic status of the compartmentalized cell, especially in the
triad region with RyR localization. This may lead to in-
creased free [Mg2+] and decreased free [ATP], which within
the physiological range of changes in these metabolites, are
strong regulators of the RyR1 (13). Further, clear evidence
has accumulated demonstrating that glycolysis preferentially
regulates membrane ion transport mechanisms. In line with
this concept, physiological data directly support the concept
of a regulatory role of glycolysis on SR membrane proteins,
as glycolytic enzymes are associated with these membranes
(38). Indeed, it was found that especially the glycolytic in-
termediate, fructose 1,6-bisphosphate, increased the opening
probability of RyR channels (38). However, low glycogen
has also been demonstrated to modulate the Ca2+ release rate
in isolated vesicles without restricted metabolic space and
during resting metabolic conditions. This result may indicate
a crucial role of the metabolic machinery associated with the
SR in maintaining endogenous metabolism. In line with this,
analysis of the RyR protein sequence reveals that it contains
many phosphorylation sites. The capacity of protein kinase
A (PKA) and Ca2+-calmodulin-dependent kinase II (CaMKII,
fast twitch fibers) to activate the RyR1 channel has been
reported (31), although studies in skinned and intact skeletal
muscle fibers have not supported this idea (12). However,
the impact of phosphorylation and/or dephosphorylation on
single RyR channel behavior and the role of glycogen and
energy status is at present not fully unraveled.
Oxygen and Muscle Fatigue
The previous two sections have described how an inte-
grative approach can elucidate the sources of fatigue in vivo,
and how muscle glycogen metabolism is essential to sup-
porting contractile function at the cellular level. In this sec-
tion, we turn to the role of oxygen in the development of
fatigue. Although numerous factors play into the fatigue
process that occurs in skeletal muscle during exercise (highly
dependent on the type of exercise), one of the more studied
factors has been that of oxygen availability to the working
muscle. It has been well appreciated that reduced oxygen
availability to exercising muscle has profound consequences
on muscle fatigue. However, it remains uncertain as to the
precise mechanisms by which O2 availability may affect the
fatigue process. Certainly, when the exercise is of a very high
intensity, the oxygen availability to the respiring mitochon-
dria may be insufficient for the demand for ATP, resulting in
an instability of the cellular metabolic homeostasis and lead-
ing to changes that induce fatigue. However, at submaximal
exercise intensities, the role of O2 availability in the fatigue
process becomes a bit more curious. Increases in oxygen
uptake and ATP utilization rise in unison until V̇O2max is
reached, and at that point the demand for more ATP cannot
http://www.acsm-msse.org
be met by increases in mitochondrial respiration or oxygen
delivery. This tight coupling of ATP demand by the working
muscle to the ATP supply from the mitochondria only be-
comes uncoupled at the highest exercise intensities (see re-
view by Sahlin, (84). The manner in which O2 availability
may affect the fatigue process at less than the highest work
intensities remains puzzling, and may have important conse-
quences for human health, for example as in the hypoxia that
develops with living at altitude, in some lung and heart dis-
eases, and during the process of human aging.
Since the early work of Krogh (61), researchers have
attempted to measure and model the diffusion of oxygen to
the interior of cells. Although the importance of O2 for
maintaining energy balance within cells became appreciated
with the delineation of oxidative phosphorylation, it has
been postulated that the metabolism and function of many
cell types can be affected by intracellular O2 tensions at
levels well above those considered rate limiting for mito-
chondrial function (83). This field of research has been dif-
ficult to reconcile due to the problems involved in the
precise measurement of intracellular PO2. Whereas some of
the methods for studying intracellular oxygenation and its
effects on cell function have had methodological constraints,
appropriate models to study these questions have also been a
difficulty. There are limited data in skeletal muscle attempting
to measure intracellular PO2 values. For example, it is ex-
ceedingly difficult to remove the variability and heterogeneity
of the microcirculation and fiber population from estimates or
measurements of intracellular PO2. Whole muscle measure-
ments of myoglobin saturation, using MRS technology (79)
are able to at least measure the mean PO2 from a large
number of cells. The consensus from this work suggests that
while intracellular PO2 drops to very low levels during
moderate- to high-intensity exercise, the PO2 remains above
that which limits mitochondrial respiration. And although
knowing the PO2 within a working muscle fiber is important
for understanding mitochondrial function, it has become just
recently appreciated that the mitochondria can form a retic-
ulum throughout the myofiber and the utilization of O2 in
one area of the myofiber may transfer the energy charge to
another area of the fiber closer to the ATP demand (35).
Rumsey and colleagues (83) suggested that there is not
simply a minimal ‘‘critical’’ value for [O2] within the cell
below which oxidative phosphorylation becomes compro-
mised, but rather a range of O2 values that influence both
the metabolic and respiratory state of the cell. The regulation
of tissue respiration and metabolism has generally been
considered to be controlled by the levels of the substrates
needed to rephosphorylate ADP through oxidative phos-
phorylation, principally [ADP], [Pi], the [ATP]/[ADP][Pi]
ratio, and the redox ratio of NADH/NAD (see review by (5).
It has been demonstrated (45) that the levels of some of these
proposed regulators may be influenced by the amount of O2
available for tissue respiration, even when the O2 available
is above that considered ‘‘critical’’ for tissue respiration. It
was demonstrated (40,46), using 31P-MRS, that the factors
INTEGRATIVE PERSPECTIVES ON FATIGUE
Copyright © 2016 by the American College of Sports Medicine. Unauthorized reproduction of this article is prohibited.
thought to regulate tissue respiration and metabolism during
SPECIAL COMMUNICATIONS
work (ADP, Pi, phosphocreatine, etc.) may be modulated by
an interaction with tissue oxygenation levels. It was shown
that, depending on the degree of tissue oxygenation, quite
different amounts of these proposed regulators were required
to attain similar levels of V̇O2, suggesting that the mito-
chondrial sensitivity to these proposed regulators of respira-
tion and metabolism could be altered by tissue oxygenation
levels. Additionally, Hogan and Welch (47) demonstrated
that at similar V̇O2 during low arterial PO2 versus normal
arterial PO2 conditions, there is a greater intracellular muscle
[lactate] and muscle lactate efflux in the hypoxemic condi-
tion. It has been postulated that this is a result of an increase
in the rate of glycolysis brought on in part by the changes
(due to PO2) in the concentration of some of those substrates
listed above (ADP, Pi, Ca2+) that regulate glycolysis, and that
these changes can alter the development of fatigue. This
would occur independent of any O2 limiting situation.
As with the factors that regulate muscle metabolism, the
factors that cause reduced tension development and muscle
fatigue interact in a complex fashion (2,89). It is likely that
various changes in the intracellular milieu can directly affect
cross-bridge function or inhibit SR Ca2+ kinetics, both of
which will reduce tension production. Accumulation of H+
and Pi has both been implicated in these processes, although
the role of changes in pH remains controversial. Increases in
[Pi] have been linked to decreased contractility in isolated
muscle fibers (15) and decreases in SR Ca2+ release (6).
Figure 4 (from Hogan et al. (46); using 31P-MRS) shows the
FIGURE 4—Influence of inspired oxygen on muscle work and inor-
ganic phosphate. Relationship of muscle Pi concentration (means T SE;
relative to resting values) and work rate for each of three different FIO2
conditions. Last open symbol for each FIO2 condition was when num-
ber of subjects was 4, whereas solid symbols represent mean exhaustion
time point for all six subjects. *Significantly different from other FiO2
conditions at this work rate, P G 0.05. Adapted from Hogan et al. (46).
Medicine & Science in Sports & Exercised 2287
 effect of different PO2 on one of these critical modulators of
SPECIAL COMMUNICATIONS
cellular function (including fatigue)—intracellular Pi. It can
be seen that at the same submaximal work intensity (rates of
ATP utilization and mitochondrial respiration will be the
same for any of the varied PO2), the intracellular Pi is in-
creased with decreasing PO2, and this may have influenced
the earlier onset of fatigue in the low PO2 conditions. In
addition, recent evidence has pointed to the role of intra-
cellular [Ca2+] as a significant factor in the fatigue process,
and there is good evidence that release of Ca2+ from the SR
complex may be inhibited by changes in the other intracel-
lular constituents mentioned previously (including Mg2+) as
the ATP requirements become excessive (1).
Finally, numerous investigations have also implicated
oxygen radical formation during intense muscle contractions
as a causative agent in the fatigue process (86). However,
there remains a paucity of information related to the manner
in which intracellular oxygen tension affects reactive oxygen
species (ROS) production during contractile activities. It has
been known from the early 1980s that ROS are generated by
working skeletal muscle (23). ROS are generated in low
amounts within resting skeletal muscle, and it is generally
agreed that ROS production increases with contractile ac-
tivity. Since ROS generation is dependent on the metabo-
lism of O2, processes related to intracellular oxygenation are
of critical importance to this field of research. However, the
oxygen dependence of ROS generation is poorly understood
(21), and it has only recently become evident (22) that
hypoxia can paradoxically induce ROS generation—thereby
potentially affecting the process of fatigue. Ferreira and Reid
(30) have postulated that muscle force production is sensi-
tive to the muscle redox status, with inhibition of force at
either a very reduced or a very oxidized muscle redox state.
At this time, after years of investigation, we still know
very little about intracellular O2 levels in muscle at rest or
during the conditions of increased mitochondrial respiration
caused by contractile activity and how this modulates ROS
formation and the other factors that can induce fatigue. In
terms of contractile function, it should be clear that the
changes in the intracellular metabolic state induced by al-
tered oxygenation states can have a profound impact on
cellular function. Increases in Pi, ADP, H+, ROS and other
metabolites that occur with altered oxygenation state, inde-
pendent of respiratory rate, can seriously impair the activity
of the contractile and ion pump processes. In this way, cell
function can be compromised by the altered metabolic state
of the cell that results from altered O2 availability, even
when there is enough O2 available for the required respira-
tory rate.
Is There a Disconnect between Human and Animal
Fatigue Models?
In this final section of our review, we focus on the role of
BF in muscle fatigue, to provide some historical perspective
on how studies of both humans and animal models have
2288 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.
contributed to our understanding of fatigue over the years.
Almost without exception, experimental evidence gathered
in animal species proves valuable to furthering our knowl-
edge of human physiology and the basis for relieving the
burden of human disease. Comparative sequence analysis
has revealed genetic homologies across phenotypically di-
verse species and, as the genes and diseases are the same, so
is the therapeutic treatment. Insertion of a human disease
gene into the mouse genome causes that disease in the
mouse which then becomes indispensable as a model for
helping to understand and treat the disease in humans. It
should be noted that engineered animal models of disease
that produce a phenotype similar to that found in humans
may not faithfully reproduce the mechanistic bases for that
disease. As a result, therapeutic interventions developed
from such animal models often fail in human trials because
the models are too narrowly focused on a single pathological
pathway. This overengineered approach, combined with a
lack of diversity of animal models (i.e., reliance on mice
versus larger animals), has slowed drug discovery.
Given the above, it is difficult to understand a defensible
scientific approach to solving any physiological or medical
problem that does not garner relevant evidence from both
humans and animal species. This section highlights the se-
quence of discovery across species for several major scien-
tific advances as they relate directly to our present knowledge
of muscle fatigue and especially the role of O2 delivery and
metabolism in those processes.
The alleviation of fatigue and exercise intolerance in
health and particularly in endemic chronic diseases such as
heart failure and diabetes remains a cornerstone principle
of modern medicine and National Institutes of Health fund-
ing strategies. For rhythmic contractions, muscle perfor-
mance is inextricably connected to the sustained and adequate
supply of oxygen. Consequently, any discussion of fatigue
and fatigue mechanisms is not complete without consider-
ation of muscle BF and O2 supply and their potential to im-
pact contractile performance.
In 1987, the ACSM Annual Meeting provided a forum (87)
for luminaries in the field to pose sentinel questions regarding
what was then known regarding skeletal muscle(s) BF during
exercise. One impetus for this symposium was the, then re-
cent, article by Andersen and Saltin (3), documenting BF
in the human quadriceps approximately 2.5 LIminj1Ikgj1.
This value was far greater than found previously by indi-
cator washout/dilution, venous occlusion plethysmography,
or bolus infusion of cold saline in humans (3,49,58) and
raised major concerns regarding control of the exercise hy-
peremia and muscle metabolism itself. Cross-species com-
parisons featured in that symposium included the rat and dog
as well as human studies and highlighted the primary role
played by research on animal muscle(s) in past and future
scientific discovery.
What are the upper limits to skeletal BF during
exercise? As demonstrated for the heart (left ventricle,
96 LIminj1Ikgj1) (65) and skin (2–3 LIminj1Ikgj1) (81), it
http://www.acsm-msse.org
was appreciated that organ BF could reach extraordinarily
high levels. However, despite the existence of similar values in
individual rodent muscles, skepticism regarding the radioac-
tive microsphere technique precluded mainstream acceptance
of these values. Specifically, in rats running at 75 mIminj1,
several muscles including the red gastrocnemius and vastus
intermedius reached BF in excess of 3 LIminj1Ikgj1 (64).
Furthermore, Musch and colleagues (69) recorded similarly
BF in the locomotor muscles of exercise trained foxhounds
running maximally on the treadmill. In 1993, Richardson and
colleagues (80) considered that the prolonged duration of the
Andersen and Saltin (3) knee extension protocol may have
constrained both work rate at fatigue and maximal BF. Using
the same techniques as Andersen and Saltin during a more
rapidly work rate-incremented knee extensor exercise test in
trained cyclists, Richardson et al. (80) found that BF in-
creased to approximately 4 LIminj1Ikgj1, the highest mea-
sured to date in human muscle(s). These values in humans led
to widespread acceptance of the very high animal BF gathered
using microspheres. Today, the microsphere technique has
measured some of the greatest BF in skeletal muscle (rats
running at 96 mIminj1, vastus intermedius, 6.8 LIminj1Ikgj1)
and also the diaphragm (costal diaphragm, 5.5 LIminj1Ikgj1)
(78). As well as establishing the extraordinary vascular ca-
pacity of skeletal muscle(s), Barclay and Stainsby_s (7) ex-
periments using their classic canine gastrocnemius-plantaris
model made it evident that, during maximal contractions,
skeletal muscle oxidative capacity was limited by O2 delivery
and not mitochondrial metabolism as was subsequently proven
for human muscle by Richardson et al. (79).
How heterogeneous is BF among/within muscles
during exercise? As discussed in Poole et al. (77), animal
muscle may be more highly stratified with respect to fiber
type(s) than their human counterparts. This property means
that animals may be ideal models for investigating skeletal
muscle functional vascular and metabolic control during
exercise as related to distinct fiber types and their recruit-
ment during different exercise intensities and durations.
Thus, Laughlin and Armstrong (64) determined that, in the
resting rat, the soleus muscle supported a BF some sixfold that
of the white vastus lateralis and 2.5-fold that of the red gas-
trocnemius. However, during high speed running at 75 mIminj1
the red gastrocnemius BF increased to 1.4- and 7.5-fold that of
the soleus and white vastus lateralis, respectively. Although
many of the highly oxidative muscles were recruited at rela-
tively slow running speeds their low oxidative counterparts
(such as the white gastrocnemius) required far greater speeds.
These investigations also dispelled the notion that muscles
comprised of slow oxidative (type I) fibers sustained higher
peak BF (and metabolic rates) than fast twitch oxidative
(type IIa) muscles. Human muscles may possess very dif-
ferent fiber types both across muscles (e.g., %type I; soleus
88%, rectus femoris [RF], 36%) and also, to a lesser extent,
with respect to depth from the skin (deeper muscle has more
type I fibers; (48). In 2000, Kalliokoski and colleagues (50)
using H2O15 positron emission tomography (PET) identified
INTEGRATIVE PERSPECTIVES ON FATIGUE
Copyright © 2016 by the American College of Sports Medicine. Unauthorized reproduction of this article is prohibited.
a distinct heterogeneity of BF among the thigh extensors
SPECIAL COMMUNICATIONS
during submaximal exercise. Specifically, BF increased from
the vastus lateralis and RF (most superficial) toward the
deeper vastus medialis and vastus intermedius (~2-fold that of
the VL). This is the precise pattern expected based upon
Laughlin and Armstrong_s (64) data obtained in rats almost
two decades earlier!
Rat muscles composed of type I fibers sustain a higher O2
delivery/O2 utilization ratio during contractions compared
with type IIs, such that their microvascular PO2 (which
constitutes the O2 pressure driving blood–myocyte O2 flux)
is greater and falls more slowly after the onset of contrac-
tions (8,66). Based on these data and that of Kalliokoski
et al. (50), Koga and colleagues (59) hypothesized that
the deeper thigh muscles should deoxygenate more slowly
after exercise onset. Using high-power Time Resolved
Spectroscopy–near-infrared spectroscopy the mean response
time for the deeper RF deoxygenation was indeed substan-
tially slower than its superficial region (i.e., superficial RF,
37 s versus deep RF, 65 s, P G 0.05). This finding calls into
question the interpretation of more superficial NIRS mea-
surements as representative of the muscle as a whole.
What is the role of nitric oxide in the exercise
hyperemia? Early human studies investigating the role of
nitric oxide (NO) in regulating the skeletal muscle BF re-
sponse to exercise produced conflicting results (49). Whereas
some investigators (27,34) found a significant role for NO in
the hyperemic response through NO synthase (NOS) block-
ade, others (28,90) did not. The reasons for these conflicting
results remain unclear, but they may have been associated with
differences in experimental design and methodologies used to
measure BF (i.e., venous occlusion plethysmography, Doppler
ultrasound, constant-infusion thermodilution technique) along
with varied exercise paradigms (i.e., arm versus leg exercise;
exercising at different work intensities) used in these in-
vestigations. In addition, unlike research using animals it is
challenging to achieve full NOS blockade in humans.
Boushel et al. (14) have demonstrated that the hyperemic
responses to incremental knee extension exercise in both
the vastus lateralis and vastus medialis are reduced in in-
dividuals subjected to both NOS and cyclooxygenase
blockade. Similarly, Heinonen et al. (41) using PET scan-
ning showed that BF was reduced in the working quadriceps
femoris muscle by 13% during NOS and cyclooxygenase
blockade and if one examines closely the representative
cross-sectional PET BF images produced in that study it is
evident that the reductions in BF occurred in the muscles
that were located very close to the femur (the more highly
oxidative muscles). These studies provided compelling evi-
dence that NO bioavailability plays a significant role in the
hyperemic response to leg exercise in humans. They also
suggest that NO production and bioavailability make a
greater contribution to the hyperemic response of the deep
(and more oxidative) muscles found in the human thigh.
Interestingly, the results of Heinonen et al. (41) are very
similar to those produced by Hirai et al. (43) nearly 2 decades
Medicine & Science in Sports & Exercised 2289
 ago. Specifically, Hirai et al. measured BF with the radioactive However, as found in rats, BF to both the vastus intermedius
SPECIAL COMMUNICATIONS

microsphere technique in individual muscles or muscle parts of
the hindlimb during moderate treadmill exercise both before
and after NOS and cyclooxygenase blockade. Results demon-
strated that BF was reduced in 16 of the 28 muscles examined
and that the reductions in BF were highly correlated with the
oxidative capacity of the individual muscles. In agreement with
Heinonen et al. (41), the muscles demonstrating the largest
NOS-blockade-induced reductions in BF in the thigh region
of the rat were those located deep and close to the femur.
Can animal studies help unravel the complexities
of vascular control in age muscle? Understanding
how aging affects vascular control and the exercise hyper-
emia in skeletal muscle is central to maintaining health,
exercise tolerance and quality of life. However, without
defining the healthy aging response the interactions between
aging and age-related diseases (i.e., chronic heart failure,
diabetes, chronic obstructive pulmonary disease) cannot be
defined and the most effective countermeasures to exercise
intolerance undertaken.
Animals studies have consistently demonstrated that work-
ing muscle BF is not different (either at submaximal or
maximal treadmill running speeds) in young (2–3 yr) versus
old (10–14 yr) beagles (36) or young adult (12 months) and
senescent (24 months) rats (60). In addition, Musch et al.
(68) found that although total hindlimb muscle BF might be
unaffected by age per se during submaximal treadmill ex-
ercise in mature (6–8 months) and senescent (27–29 months)
rats, there was a profound redistribution within and among
the 28 different muscles or muscle parts examined. Specifi-
cally, in the old rats BF was increased in eight highly gly-
colytic muscles but reduced in six highly oxidative muscles.
Moving ahead 10 yr, PET scanning measurements of
exercising knee extensor BF revealed no difference in total
muscle flow in young (26 T 6 yr) and old (77 T 6 yr) men.
and vastus medialis were significantly higher in the old men
(82). In addition, in the old subjects, there was a markedly
greater heterogeneity of intramuscular flow found within
each of the muscles. These findings, initially in rats and
subsequently in humans, reveal a complex effect of aging on
skeletal muscle vascular control and provide irrevocable
evidence that animal studies are indispensable to unravelling
the impact of aging on muscle vascular control and hence
age-related exercise intolerance.
Conclusions. Perusal of the Nobel prize-winning sci-
ence over more than a century reveals an effective symbiosis
between animal and human research leading to major ad-
vances in physiology and medicine (Fig. 5). This is nowhere
more true than for progress in understanding the funda-
mental mechanisms of fatigue and the role of muscle(s) BF
and O2 delivery in facilitating or limiting exercise tolerance.
Animal experimentation has yielded basic scientific infor-
mation, unraveled vasomotor control pathways and framed
key questions that can then be addressed ethically in
humans. As dealt with above, this symbiosis has been es-
pecially effective for discovering: 1) The upper limits of
muscle perfusion. 2) The extraordinary degree of BF hetero-
geneity among and within muscles that permits exquisite
matching of O2 delivery to V̇O2 during exercise. 3) The es-
sential role of NO in vasomotor control. 4) How aging impacts
the exercise hyperemia. These provide just a few of a multitude
of examples where investigative science (and knowledge of
fatigue mechanisms) has advanced through judicious use of
both animal and human models.
OVERALL CONCLUSIONS
Integrative approaches to the study of fatigue can provide
unique information about the interaction of physiological

FIGURE 5—Summary of human and animal model studies of fatigue. Mechanisms of fatigue are not complete without considerations of muscle O2
supply (BF) and its impact on contractile performance. Accordingly, exercise hyperemia has been examined using a variety exercise paradigms and
different methodologies in attempting to discern whether there is a significant disconnect between animal and humans studies. As discussed in the text,
there appears to be no significant disconnect between animal (rat; treadmill exercise) and human (knee extensor exercise) investigations; however,
discoveries made in the rat precede, and, in some cases, may have inspired those found in humans. Overall, both animal and human models contribute
significantly to our scientific understanding of both physiology/pathophysiology, and both models should be considered to optimize scientific discovery.

2290 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.
systems during stresses such as fatigue. Information about
multiple systems informs the overall interpretation of fa-
tigue data, and provides the opportunity to develop novel
hypotheses about how systems function under a variety of
conditions. Solving the complex mystery of fatigue re-
quires both a combination of approaches within a given
study, as well the use of varied models across studies; a full
understanding of fatigue will best be served by commu-
nication among researchers working at all levels. In this
review, we have provided examples of successful ap-
proaches to the study of skeletal muscle fatigue, including
REFERENCES
1. Allen DG, Kabbara AA, Westerblad Hk. Muscle fatigue: the role of
intracellular calcium stores. Can J Appl Physiol. 2002;27(1):83–96.
2. Amann M, Calbet JA. Convective oxygen transport and fatigue.
J Appl Physiol (1985). 2008;104(3):861–70.
3. Andersen P, Saltin B. Maximal perfusion of skeletal muscle in
man. J Physiol. 1985;366(1):233–49.
4. Baker AJ, Kostov KG, Miller RG, Weiner MW. Slow force recov-
ery after long-duration exercise: metabolic and activation factors in
muscle fatigue. J Appl Physiol (1985). 1993;74(5):2294–300.
5. Balaban RS. Modeling mitochondrial function. Am J Physiol Cell
Physiol. 2006;291(6):C1107–13.
6. Balog EM, Fruen BR, Kane PK, Louis CF. Mechanisms of P(i)
regulation of the skeletal muscle SR Ca(2+) release channel. Am J
Physiol Cell Physiol. 2000;278(3):C601–11.
7. Barclay JK, Stainsby WN. The role of blood flow in limiting max-
imal metabolic rate in muscle. Med Sci Sports. 1975;7(2):116–9.
8. Behnke BJ, McDonough P, Padilla DJ, Musch TI, Poole DC.
Oxygen exchange profile in rat muscles of contrasting fibre types.
J Physiol. 2003;549(2):597–605.
9. Bergström J, Hermansen L, Hultman E, Saltin B. Diet, muscle
glycogen and physical performance. Acta Physiol Scand. 1967;
71(2):140–50.
10. Bigland-Ritchie B, Jones D, Hosking GP, Edwards RH. Central and
peripheral fatigue in sustained maximum voluntary contractions of
human quadriceps muscle. Clin Sci Mol Med. 1978;54(6):609–14.
11. Bigland-Ritchie B, Johansson R, Lippold OC, Woods JJ. Con-
tractile speed and EMG changes during fatigue of sustained max-
imal voluntary contractions. J Neurophysiol. 1983;50:313–24.
12. Blazev R, Hussain M, Bakker AJ, Head SI, Lamb GD. Effects of
the PKA inhibitor H-89 on excitation–contraction coupling in
skinned and intact skeletal muscle fibres. J Muscle Res Cell Motil.
2001;22(3):277–86.
13. Blazev R, Lamb GD. Low [ATP] and elevated [Mg2+] reduce
depolarization-induced Ca2+ release in rat skinned skeletal muscle
fibres. J Physiol. 1999;520(1):203–15.
14. Boushel R, Langberg H, Gemmer C, et al. Combined inhibition of
nitric oxide and prostaglandins reduces human skeletal muscle
blood flow during exercise. J Physiol. 2002;543(Pt2):691–8.
15. Bruton JD, Wretman C, Katz A, Westerblad H. Increased tetanic
force and reduced myoplasmic [P(i)] following a brief series of
tetani in mouse soleus muscle. Am J Physiol. 1997;272(3 Pt 1):
C870–4.
16. Callahan DM, Umberger BR, Kent JA. Mechanisms of in vivo muscle
fatigue in humans: investigating age-related fatigue resistance with a
computational model. J Physiol. 2016;594(12):3407–21.
17. Callahan DM, Umberger BR, Kent-Braun JA. A computational model
of torque generation: neural, contractile, metabolic and musculo-
skeletal components. PLoS One. 2013;8(2):e56013.
18. Chin ER, Allen DG. Effects of reduced muscle glycogen concen-
tration on force, Ca2+ release and contractile protein function in
intact mouse skeletal muscle. J Physiol. 1997;498(1):17–29.
INTEGRATIVE PERSPECTIVES ON FATIGUE
Copyright © 2016 by the American College of Sports Medicine. Unauthorized reproduction of this article is prohibited.
a variety of applications to understanding fatigue in aging.
SPECIAL COMMUNICATIONS
Future work that expands upon the approaches described
here and other approaches in use today will continue to
illuminate from the molecular scale to whole-body func-
tion the complex factors involved in the development of
muscle fatigue.
The authors report no conflicts of interest. There are no funding
sources to acknowledge for this review. The results of the present
study do not constitute endorsement by ACSM.
19. Christie A, Snook EM, Kent-Braun JA. Systematic review and
meta-analysis of skeletal muscle fatigue in old age. Med Sci Sports
Exerc. 2011;43:568–77.
20. Christie AD, Tonson A, Larsen RG, DeBlois JP, Kent JA. Human
skeletal muscle metabolic economy in vivo: effects of contraction
intensity, age, and mobility impairment. Am J Physiol Regul Integr
Comp Physiol. 2014;307(9):R1124–35.
21. Clanton TL. Hypoxia-induced reactive oxygen species formation
in skeletal muscle. J Appl Physiol (1985). 2007;102(6):2379–88.
22. Clanton TL, Hogan MC, Gladden LB. Regulation of cellular gas
exchange, oxygen sensing, and metabolic control. Compr Physiol.
2013;102(6):2379–88.
23. Davies KJ, Quintanilha AT, Brooks GA, Packer L. Free radicals
and tissue damage produced by exercise. Biochem Biophys Res
Commun. 1982;107(4):1198–205.
24. Debold EP, Beck SE, Warshaw DM. Effect of low pH on single
skeletal muscle myosin mechanics and kinetics. Am J Physiol Cell
Physiol. 2008;295(1):C173–9.
25. Dhar-Chowdhury P, Malester B, Rajacic P, Coetzee WA. The
regulation of ion channels and transporters by glycolytically de-
rived ATP. Cell Mol Life Sci. 2007;64(23):3069–83.
26. Duhamel TA, Perco JG, Green HJ. Manipulation of dietary car-
bohydrates after prolonged effort modifies muscle sarcoplasmic
reticulum responses in exercising males. Am J Physiol Regul Integr
Comp Physiol. 2006;291(4):R1100–10.
27. Dyke CK, Proctor DN, Dietz NM, Joyner MJ. Role of nitric oxide
in exercise hyperaemia during prolonged rhythmic handgripping in
humans. J Physiol. 1995;488(Pt 1):259–65.
28. Endo T, Imaizumi T, Tagawa T, Shiramoto M, Ando S, Takeshita
A. Role of nitric oxide in exercise-induced vasodilation of the
forearm. Circulation. 1994;90(6):2886–90.
29. Entman ML, Keslensky SS, Chu A, Van Winkle WB. The sarco-
plasmic reticulum-glycogenolytic complex in mammalian fast twitch
skeletal muscle. Proposed in vitro counterpart of the contraction-
activated glycogenolytic pool. J Biol Chem. 1980;255(13):6245–52.
30. Ferreira LF, Reid MB. Muscle-derived ROS and thiol regulation in
muscle fatigue. J Appl Physiol (1985). 2008;104(3):853–60.
31. Fill M, Copello JA. Ryanodine receptor calcium release channels.
Physiol Rev. 2002;82(4):893–922.
32. Fitts RH. The cross-bridge cycle and skeletal muscle fatigue. J Appl
Physiol (1985). 2008;104(2):551–8.
33. Fuglevand AJ, Winter DA, Patla AE. Models of recruitment and
rate coding organization in motor-unit pools. J Neurophysiol.
1993;70(6):2470–88.
34. Gilligan DM, Panza JA, Kilcoyne CM, Waclawiw MA, Casino PR,
Quyyumi AA. Contribution of endothelium-derived nitric oxide to
exercise-induced vasodilation. Circulation. 1994;90(6):2853–8.
35. Glancy B, Hartnell LM, Malide D, et al. Mitochondrial reticulum
for cellular energy distribution in muscle. Nature. 2015;523(7562):
617–20.
Medicine & Science in Sports & Exercised 2291
 36. Haidet GC, Parsons D. Reduced exercise capacity in senescent bea-
SPECIAL COMMUNICATIONS
gles: an evaluation of the periphery. Am J Physiol. 1991;260(1 Pt 2):
H173–82.
37. Hainaut K, Duchateau J. Muscle fatigue, effects of training and
disuse. Muscle Nerve. 1989;12(8):660–9.
38. Han JW, Thieleczek R, Varsányi M, Heilmeyer LM Jr. Compart-
mentalized ATP synthesis in skeletal muscle triads. Biochemistry.
1992;31(2):377–84.
39. Hargreaves M, McConell G, Proietto J. Influence of muscle gly-
cogen on glycogenolysis and glucose uptake during exercise in
humans. J Appl Physiol (1985). 1995;78(1):288–92.
40. Haseler LJ, Richardson RS, Videen JS, Hogan MC. Phosphocre-
atine hydrolysis during submaximal exercise: the effect of FIO2.
J Appl Physiol (1985). 1998;85(4):1457–63.
41. Heinonen I, Saltin B, Kemppainen J, et al. Skeletal muscle blood
flow and oxygen uptake at rest and during exercise in humans: a
pet study with nitric oxide and cyclooxygenase inhibition. Am J
Physiol Heart Circ Physiol. 2011;300(4):H1510–7.
42. Hermansen L, Hultman E, Saltin B. Muscle glycogen during pro-
longed severe exercise. Acta Physiol Scand. 1967;71(2–3):129–39.
43. Hirai T, Visneski MD, Kearns KJ, Zelis R, Musch TI. Effects of NO
synthase inhibition on the muscular blood flow response to treadmill
exercise in rats. J Appl Physiol (1985). 1994;77(3):1288–93.
44. Hochachka PW, Matheson GO. Regulating ATP turnover rates
over broad dynamic work ranges in skeletal muscles. J Appl
Physiol (1985). 1992;73(5):1697–703.
45. Hogan MC, Arthur PG, Bebout DE, Hochachka PW, Wagner PD.
Role of O2 in regulating tissue respiration in dog muscle working
in situ. J Appl Physiol (1985). 1992;73(2):728–36.
46. Hogan MC, Richardson RS, Haseler LJ. Human muscle perfor-
mance and PCr hydrolysis with varied inspired oxygen fractions: a
31P-MRS study. J Appl Physiol (1985). 1999;86(4):1367–73.
47. Hogan MC, Welch HG. Effect of altered arterial O2 tensions on
muscle metabolism in dog skeletal muscle during fatiguing work.
Am J Physiol. 1986;251(2 Pt 1):C216–22.
48. Johnson MA, Polgar J, Weightman D, Appleton D. Data on the
distribution of fibre types in thirty-six human muscles. An autopsy
study. J Neurol Sci. 1973;18(1):111–29.
49. Joyner MJ, Casey DP. Regulation of increased blood flow (hy-
peremia) to muscles during exercise: a hierarchy of competing
physiological needs. Physiol Rev. 2015;95(2):549–601.
50. Kalliokoski KK, Kemppainen J, Larmola K, et al. Muscle blood
flow and flow heterogeneity during exercise studied with positron
emission tomography in humans. Eur J Appl Physiol. 2000;83(4–5):
395–401.
51. Katz A, Westerblad H. Regulation of glycogen breakdown and its
consequences for skeletal muscle function after training. Mamm
Genome. 2014;25(9–10):464–72.
52. Kent-Braun JA. Central and peripheral contributions to muscle
fatigue in humans during sustained maximal effort. Eur J Appl
Physiol Occup Physiol. 1999;80(1):57–63.
53. Kent-Braun JA. Skeletal muscle fatigue in old age: whose advan-
tage? Exerc Sport Sci Rev. 2009;37(1):3–9.
54. Kent-Braun JA, Fitts RH, Christie A. Skeletal muscle fatigue.
Compr Physiol. 2012;2:997–1044.
55. Kent-Braun JA, Miller RG, Weiner MW. Phases of metabolism
during progressive exercise to fatigue in human skeletal muscle.
J Appl Physiol (1985). 1993;75(2):573–80.
56. Kent-Braun JA, Ng AV, Doyle JW, Towse TF. Human skeletal
muscle responses vary with age and gender during fatigue due to
incremental isometric exercise. J Appl Physiol (1985). 2002;93(5):
1813–23.
57. Kent-Braun JA, Sharma KR, Weiner MW, Miller RG. Effects of
exercise on muscle activation and metabolism in multiple sclerosis.
Muscle Nerve. 1994;17(10):1162–9.
2292 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.
58. Klausen K, Secher NH, Clausen JP, Hartling O, Trap-Jensen J.
Central and regional circulatory adaptations to one-leg training.
J Appl Physiol Respir Environ Exerc Physiol. 1982;52(4):976–83.
59. Koga S, Barstow TJ, Okushima D, et al. Validation of a high-
power, time-resolved, near-infrared spectroscopy system for mea-
surement of superficial and deep muscle deoxygenation during
exercise. J Appl Physiol (1985). 2015;118(11):1435–42.
60. Kregel KC. Augmented mesenteric and renal vasoconstriction
during exercise in senescent Fischer 344 rats. J Appl Physiol (1985).
1995;79(3):706–12.
61. Krogh A. The number and distribution of capillaries in muscles
with calculations of the oxygen pressure head necessary for sup-
plying the tissue. J Physiol. 1919;52(6):409–15.
62. Lanza IR, Larsen RG, Kent-Braun JA. Effects of old age on human
skeletal muscle energetics during fatiguing contractions with and
without blood flow. J Physiol. 2007;583(3):1093–105.
63. Lanza IR, Tevald MA, Befroy DE, Kent-Braun JA. Intracellular
energetics and critical PO2 in resting ischemic human skeletal
muscle in vivo. Am J Physiol Regul Integr Comp Physiol. 2010;
299(5):R1415–22.
64. Laughlin MH, Armstrong RB. Muscular blood flow distribution
patterns as a function of running speed in rats. Am J Physiol. 1982;
243(2):H296–306.
65. Manohar M. Transmural coronary vasodilator reserve and flow
distribution during maximal exercise in normal and splenectomized
ponies. J Physiol. 1987;387(1):425–40.
66. McDonough P, Behnke BJ, Padilla DJ, Musch TI, Poole DC.
Control of microvascular oxygen pressures in rat muscles com-
prised of different fibre types. J Physiol. 2005;563(Pt 3):903–13.
67. Miller RG, Giannini D, Milner-Brown HS, et al. Effects of fatiguing
exercise on high-energy phosphates, force, and EMG: evidence for
three phases of recovery. Muscle Nerve. 1987;10(9):810–21.
68. Musch TI, Eklund KE, Hageman KS, Poole DC. Altered regional
blood flow responses to submaximal exercise in older rats. J Appl
Physiol (1985). 2004;96(1):81–8.
69. Musch TI, Haidet GC, Ordway GA, Longhurst JC, Mitchell JH.
Training effects on regional blood flow response to maximal ex-
ercise in foxhounds. J Appl Physiol (1985). 1987;62(4):1724–32.
70. Ng AV, Dao HT, Miller RG, Gelinas DF, Kent-Braun JA. Blunted
pressor and intramuscular metabolic responses to voluntary iso-
metric exercise in multiple sclerosis. J Appl Physiol (1985). 2000;
88(3):871–80.
71. Nielsen J, Cheng AJ, Krtenblad N, Westerblad H. Subcellular dis-
tribution of glycogen and decreased tetanic Ca2+ in fatigued single
intact mouse muscle fibres. J Physiol. 2014;592(9):2003–12.
72. Nielsen J, Krtenblad N. Physiological aspects of the subcellular
localization of glycogen in skeletal muscle. Appl Physiol Nutr
Metab. 2012;38(2):91–9.
73. Nielsen J, SchrLder HD, Rix CG, Ortenblad N. Distinct effects of
subcellular glycogen localization on tetanic relaxation time and
endurance in mechanically skinned rat skeletal muscle fibres.
J Physiol. 2009;587(Pt 14):3679–90.
74. Krtenblad N, Nielsen J, Saltin B, Holmberg HC. Role of glycogen
availability in sarcoplasmic reticulum Ca2+ kinetics in human
skeletal muscle. J Physiol. 2011;589(3):711–25.
75. Krtenblad N, Westerblad H, Nielsen J. Muscle glycogen stores and
fatigue. J Physiol. 2013;591(18):4405–13.
76. Pepin EB, Hicks RW, Spencer MK, Tran ZV, Jackson CG. Pressor
response to isometric exercise in patients with multiple sclerosis.
Med Sci Sports Exerc. 1996;28(6):656–60.
77. Poole DC, Burnley M, Vanhatalo A, Rossiter HB, Jones AM.
Critical power: an important fatigue threshold in exercise physi-
ology. Med Sci Sports Exerc. 2016;48(11):2320–34.
78. Poole DC, Sexton WL, Behnke BJ, Ferguson CS, Hageman KS,
Musch TI. Respiratory muscle blood flows during physiological
and chemical hyperpnea in the rat. J Appl Physiol (1985). 2000;
88(1):186–94.
http://www.acsm-msse.org
79. Richardson RS, Duteil S, Wary C, Wray DW, Hoff J, Carlier PG.
Human skeletal muscle intracellular oxygenation: the impact of
ambient oxygen availability. J Physiol. 2006;571(Pt 2):415–24.
80. Richardson RS, Poole DC, Knight DR, et al. High muscle blood
flow in man: is maximal O2 extraction compromised? J Appl Physiol
(1985). 1993;75(4):1911–6.
81. Rowell LB. Human cardiovascular adjustments to exercise and
thermal stress. Physiol Rev. 1974;54:75–159.
82. Rudroff T, Weissman JA, Bucci M, et al. Positron emission to-
mography detects greater blood flow and less blood flow het-
erogeneity in the exercising skeletal muscles of old compared
with young men during fatiguing contractions. J Physiol. 2014;
592(2):337–49.
83. Rumsey WL, Schlosser C, Nuutinen EM, Robiolio M, Wilson DF.
Cellular energetics and the oxygen dependence of respiration in
cardiac myocytes isolated from adult rat. J Biol Chem. 1990;265(26):
15392–402.
84. Sahlin K. Muscle energetics during explosive activities and po-
tential effects of nutrition and training. Sports Med. 2014;44(2):
167–73.
INTEGRATIVE PERSPECTIVES ON FATIGUE
Copyright © 2016 by the American College of Sports Medicine. Unauthorized reproduction of this article is prohibited.
85. Sahlin K, Tonkonogi M, Söderlund K. Energy supply and muscle
SPECIAL COMMUNICATIONS
fatigue in humans. Acta Physiol Scand. 1998;162(3):261–6.
86. Smith MA, Reid MB. Redox modulation of contractile function in
respiratory and limb skeletal muscle. Respir Physiol Neurobiol.
2006;151(2):229–41.
87. Stainsby WN. Maximal muscle blood flow during contractions in situ
compared to in vivo. Med Science Sports Exerc. 1987;19(2 Suppl):S1.
88. Wanson JC, Drochmans P. Role of the sarcoplasmic reticulum in
glycogen metabolism. Binding of phosphorylase, phosphorylase
kinase, and primer complexes to the sarcovesicles of rabbit skeletal
muscle. J Cell Biol. 1972;54(2):206–24.
89. Westerblad H, Allen DG. Cellular mechanisms of skeletal muscle
fatigue. In: Molecular and Cellular Aspects of Muscle Contraction.
Springer; 2003. pp. 563–71.
90. Wilson JR, Kapoor S. Contribution of endothelium-derived relaxing
factor to exercise-induced vasodilation in humans. J Appl Physiol
(1985). 1993;75(6):2740–4.
91. Woods JJ, Furbush F, Bigland-Ritchie B. Evidence for a fatigue-
induced reflex inhibition of motoneuron firing rates. J Neurophysiol.
1987;58:125–37.
Medicine & Science in Sports & Exercised 2293