Inhibition of carbonic anhydrase (CA) is
associated with a lower plasma lactate concentration
([La
]pl)
during fatiguing exercise. We hypothesized that a lower
[La]pl
may be associated with faster O2
uptake (
O2) kinetics during constant-load exercise. Seven men performed cycle ergometer exercise during control (Con) and acute CA inhibition with acetazolamide (Acz,
10 mg/kg body wt iv). On 6 separate days, each subject performed 6-min
step transitions in work rate from 0 to 100 W (below ventilatory threshold,
<ET)
or to a O2 corresponding to
~50% of the difference between the work rate at
ET and peak
O2
(>ET).
Gas exchange was measured breath by breath. Trials were interpolated at
1-s intervals and ensemble averaged to yield a single response. The mean response time (MRT, i.e., time to 63% of total exponential increase) for on- and off-transients was determined using a two- (<ET) or a
three-component exponential model
(>ET).
Arterialized venous blood was sampled from a dorsal hand vein and
analyzed for
[La]pl.
MRT was similar during Con (31.2 ± 2.6 and 32.7 ± 1.2 s for on
and off, respectively) and Acz (30.9 ± 3.0 and 31.4 ± 1.5 s for on and off, respectively) for work rates
<ET. At
work rates >ET, MRT
was similar between Con (69.1 ± 6.1 and 50.4 ± 3.5 s for on and
off, respectively) and Acz (69.7 ± 5.9 and 53.8 ± 3.8 s for on and off, respectively). On- and off-MRTs were slower for
>ET than
for <ET
exercise.
[La]pl
increased above 0-W cycling values during
<ET and
>ET exercise but was lower at the end of the transition during Acz (1.4 ± 0.2 and 7.1 ± 0.5 mmol/l for
<ET and
>ET,
respectively) than during Con (2.0 ± 0.2 and 9.8 ± 0.9 mmol/l
for <ET
and >ET,
respectively). CA inhibition does not affect
O2 utilization at the onset of
<ET or
>ET
exercise, suggesting that the contribution of oxidative phosphorylation
to the energy demand is not affected by acute CA inhibition with Acz.
 |
INTRODUCTION |
DURING THE ADJUSTMENT to an abrupt increase in exercise
intensity, pulmonary O2 uptake
(O2) increases with an
initial rapid rise (phase 1) followed by an exponential increase (phase
2) to a new steady state (phase 3) if the exercise is of moderate
intensity [i.e., below the ventilatory threshold
(<ET)].
For exercise intensities that engender a sustained lactic acidosis
(i.e., >ET), an additional increase in O2
of delayed onset leads to a steady-state O2, which, if attained at
all, is higher than predicted from the
<ET
relationship between O2 and
work rate (2, 6, 40, 41). During the transition, the time constant
(
) for phase 2 kinetics reflects the
O2 at the muscle and,
therefore, the contribution of oxidative phosphorylation to ATP
resynthesis (1, 4, 5, 18). Because oxidative phosphorylation does not
meet the total energy requirements (i.e.,
O2 deficit), the breakdown of
high-energy phosphate stores [phosphocreatine (PCr)] and
increases in anaerobic glycolysis provide the balance of energy necessary for maintaining ATP turnover (2).
For moderate-intensity exercise, where a steady-state
O2 is attained, Cerretelli
and colleagues (11, 12) reported that the half time for
O2 kinetics during the
on-transition was slowed when accompanied by an accumulation of blood
lactate (i.e., the early lactate concept). This suggested that
increases in blood lactate concentration
([La]) during
the on-response associated with step increases in work rate reflected
increases in ATP production via anaerobic glycolysis and caused a
slowing of O2 utilization by the
muscle (11, 12). Casaburi and co-workers (8) also demonstrated a
slowing of the O2 on-response
during constant-load exercise not associated with sustained increases
in blood [La].
These researchers suggested that transient increases in
La production may slow the
rate of O2 utilization by muscle
early in exercise and, thereby, affect
O2 kinetics (8). These
findings are consistent with earlier demonstrations of a slower
O2 on-response and greater
O2 deficit during the adjustment
to moderate exercise under hypoxic than under normoxic conditions (23).
The slower O2 kinetics during
hypoxia were associated with an increased reliance on anaerobic energy
sources indicated by the lower muscle PCr and higher blood and muscle
[La] at the end
of exercise (23). Thus the slowing of
O2 kinetics during the
on-response appears to be associated with increases in muscle and blood
[La].
For step increases in work rate resulting in sustained increases in
blood [La],
O2 kinetics become more
complex as an additional slow increase in
O2 of delayed onset becomes
evident. This O2 slow
component has been shown to be highly correlated to the magnitude and
rate of change of blood
[La] (32).
However, a mechanism directly linking the
O2 slow component to the
lactic acidosis or a related metabolic event has not been established
(for review see Refs. 16 and 39). There is evidence to suggest that
overall O2 kinetics are
slowed in association with increases in end-exercise blood
[La] (2).
Interestingly, if the O2
response is modeled so that the kinetics of phase 2 (i.e., rapid
exponential rise) and phase 3 (i.e., slow component) are determined
independently, the relationship between
O2 kinetics and blood
[La] is
unclear. Compared with exercise intensities
<ET, the
phase 2 O2 on-response has
been shown to be slowed (29) or similar (2, 8), despite significant
increases in blood
[La].
Inhibition of carbonic anhydrase (CA) with an acute administration of
acetazolamide (Acz) compared with control is associated with lower peak
blood [La]
after short-term high-intensity exercise (21). The metabolic consequence of this Acz-induced lowering of blood
[La] during
moderate- and heavy-intensity constant-load exercise has not been
investigated. The lower blood
[La] after CA
inhibition may be due to a decreased rate of
La production, possibly by
an increased flux of pyruvate through the pyruvate dehydrogenase
complex and the subsequent oxidation of pyruvate. If the rate of
pyruvate oxidation is increased, the kinetics of
O2 may be expected to be
faster after CA inhibition. Alternatively, CA inhibition may lead to
acid-base changes on the extracellular surface of the sarcolemma that
may impair the efflux of La
from the muscle to the blood (13). Under this condition,
O2 kinetics would not be
expected to be altered by CA inhibition. We hypothesized that CA
inhibition after an acute administration of Acz would be associated
with a lower blood
[La] as a
consequence of a faster rate of adjustment of
O2 (i.e., phase 2) at the
start of moderate- and heavy-intensity exercise, suggesting that CA
inhibition was associated with a greater flux through pyruvate
dehydrogenase and not with an impaired efflux of
La. In addition, we
hypothesized that the O2 slow
component during heavy exercise would be reduced after Acz-induced CA
inhibition in accordance with the lower blood
[La].
| |
METHODS |
Subjects.
Seven healthy men participated in the study. The experimental protocol
and all possible risks associated with participation in the study were
outlined, and informed consent was obtained from each subject. The
study was approved by the University's Review Board for Health
Sciences Research Involving Human Subjects.
Material and methods.
The subjects were studied on six separate occasions during control
(Con) conditions and after acute Acz administration. The subjects
reported to the laboratory after consuming only a light meal and
abstaining from exercise and beverages containing caffeine for 
12 h
preceding the test. The exercise tests were performed at the same time
of the day for each subject. To facilitate blood sampling and Acz
administration, on one occasion during the Con studies and before each
of the Acz studies the subjects rested supine while a percutaneous
Teflon catheter (Angiocath, 21 gauge) was placed into a dorsal hand
vein. The blood was arterialized by wrapping the hand and forearm in a
heating pad. After 15 min of rest, a blood sample was drawn, then Acz
was infused (10 mg/kg over a 3-min period). The administration of Acz
was randomly ordered during the six visits. A placebo was not
administered, because in our experience the side effects of Acz
administration, although producing minor discomfort, are still
noticeable by the subject. After an additional 30 min of rest (15 min
supine and 15 min seated upright), a blood sample was drawn, and the
subject moved to the cycle ergometer. Breath-by-breath measurements of
gas exchange and ventilation were made throughout the exercise
protocol; blood samples were obtained on one occasion during each of
the two conditions at specific times during exercise (0, 15, 30, and 45 s and 1, 1.5, 2, 3, 4, and 6 min) and recovery (30 s and 1, 2, 4, and 6 min).
Preliminary testing of each subject consisted of an incremental
exercise test (work rate was increased as a ramp function at 25 W/min)
to volitional fatigue on an electromagnetically braked cycle ergometer
(model H-300-R, Lode) for the determination of ET and peak
O2
(O2 peak). The
highest O2 averaged over a
20-s interval was taken as
O2 peak. The
ET
was defined as the O2 at
which there was a systematic increase in the ventilatory equivalent for
O2
(E/O2,
where
E
is ventilation) and end-tidal PO2,
with no concomitant increase in the ventilatory equivalent for
CO2
output (E/CO2)
or decrease in end-tidal PCO2.
For the determination of O2
kinetics, subjects performed two step transitions to an absolute work
rate <ET
(100 W) and one transition to a work rate
>ET. At
>ET, the
work rate assigned was estimated to elicit a
O2 corresponding to ~50%
of the difference between the
O2 at
ET and
O2 peak:
ET + [(O2 peak 
ET) · 0.5].
Each step transition was 6 min in duration, with 6 min of loadless
cycling between each bout. The exercise protocol was performed during
three laboratory visits for each condition, resulting in six
repetitions for the moderate-intensity exercise and three repetitions
for the heavy-intensity exercise.
Inspired and expired airflow and volumes were measured during the
exercise test by a low-resistance, low-dead-space (90 ml) bidirectional
turbine and volume transducer (model VMM-110, Alpha Technologies); the
volume signal was calibrated before each test with a syringe of known
volume (990 ml). Respired gases were sampled continuously at the mouth
(1 ml/s) by a mass spectrometer (model MGA-1100, Perkin-Elmer) for
determination of the fractional concentrations of
O2,
CO2, and
N2. The mass spectrometer was
calibrated with precision-analyzed gas mixtures. Analog signals from
the mass spectrometer and turbine transducer were sampled every 20 ms
and stored on disk for later analysis. Breath-by-breath computations for O2,
CO2,
E, and
end-tidal PO2 and
PCO2 were performed after accounting
for delays in the analysis system and fluctuations in lung gas stores
in the computer algorithms (7). Corrections for temperature and water
vapor were made for conditions measured near the mouth. Heart rate was
monitored using an electrocardiogram with the electrodes placed in a
modified V5 configuration.
Arterialized venous blood was drawn into syringes containing lithium
heparin, mixed, placed in a slurry of ice and water, and analyzed after
a short delay. Whole blood samples (200 µl) were analyzed (at
37°C) for plasma pH and the plasma
[La]
([La]pl)
using selective electrodes (StatProfile 9 Plus Blood Gas-Electrolyte Analyzer, Nova Biomedical Canada); the electrodes were calibrated before each test and at regular intervals during analysis.
Data analysis.
Breath-by-breath data obtained during the step increases in work rate
were linearly interpolated at 1-s intervals, time aligned, and ensemble
averaged to provide a single response for each subject. The model
utilized to describe the kinetic response (Fig.
1) provides an estimate of the baseline
(G0), gains
(G1,
G2, and
G3), time delays
(TD1,
TD2, and
TD3), and time constants
(1,
2, and
3). For step changes in work
rate <ET,
the kinetic parameters for the on- and off-transitions in work rate
were determined as a function of time
(t) using a two-component
exponential model
![⋅ [1 − <IT>e</IT><SUP>−(<IT>t</IT> − T D<SUB>2</SUB>)/&tgr;<SUB>2</SUB></SUP>] ⋅ <IT>u</IT><SUB>2</SUB>](/content/vol85/issue4/fulltext/1384/img002.gif)
|
(1)
|
![<A><AC>V</AC><AC>˙</AC></A><SC>o</SC><SUB>2 off</SUB> = G<SUB>0</SUB> + G<SUB>1</SUB> ⋅ [<IT>e</IT><SUP>−(<IT>t</IT> − T D<SUB>1</SUB>)/&tgr;<SUB>1</SUB></SUP>] ⋅ <IT>u</IT><SUB>1</SUB> + G<SUB>2</SUB> ⋅ [<IT>e</IT><SUP>−(<IT>t</IT> − T D<SUB>2</SUB>)/&tgr;<SUB>2</SUB></SUP>] ⋅ <IT>u</IT><SUB>2</SUB>](/content/vol85/issue4/fulltext/1384/img003.gif)
|
(2)
|
where
u1 = 0 for
t < TD1,
u1 = 1 for
t > TD1,
u2 = 0 for
t < TD2, and
u2 = 1 for
t > TD2. For step changes in work rate
>ET, the
kinetic parameters for the on- and off-transitions in work rate
were determined using a three-component exponential
model
![⋅ [1 − <IT>e</IT><SUP>−(<IT>t</IT> − T D<SUB>2</SUB>)/&tgr;<SUB>2</SUB></SUP>] ⋅ <IT>u</IT><SUB>2</SUB> + G<SUB>3</SUB> ⋅ [1 − <IT>e</IT><SUP>−(<IT>t</IT> − T D<SUB>3</SUB>)/&tgr;<SUB>3</SUB></SUP>] ⋅ <IT>u</IT><SUB>3</SUB>](/content/vol85/issue4/fulltext/1384/img005.gif)
|
(3)
|
![⋅ [<IT>e</IT><SUP>−(<IT>t</IT> − T D<SUB>2</SUB>)/&tgr;<SUB>2</SUB></SUP>] ⋅ <IT>u</IT><SUB>2</SUB> + G<SUB>3</SUB> ⋅ [<IT>e</IT><SUP>−(<IT>t</IT> − T D<SUB>3</SUB>)/&tgr;<SUB>3</SUB></SUP>] ⋅ <IT>u</IT><SUB>3</SUB>](/content/vol85/issue4/fulltext/1384/img007.gif)
|
(4)
|
where
u1 = 0 for
t < TD1,
u1 = 1 for
t > TD1,
u2 = 0 for
t < TD2,
u2 = 1 for
t > TD2,
u3 = 0 for
t < TD3, and
u3 = 1 for t > TD3. Model parameters were
determined by least-squares nonlinear regression, in which the best fit
was defined by minimization of the residual sum of squares. The overall
time course of the response was determined from the mean response time
(MRT), which is calculated from a weighted sum of TD and for each
component. The MRT is equivalent to the time required to achieve
~63% of the difference between
G0 and the new steady-state value.
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Fig. 1.
Characteristics of 2- and 3-component exponential models used to
describe O2 uptake
(O2) kinetics during exercise
and recovery. During moderate-intensity exercise, a 2-component model
was used to fit data from baseline of loadless cycling with parameter
estimates corresponding to those in Eqs.
1 (exercise) and 2 (recovery). During-heavy intensity exercise, a 3-component model was
used to fit data from baseline of loadless cycling with parameter
estimates corresponding to those in Eqs.
3 (exercise) and 4 (recovery). G0, baseline; G,
gains; TD, time delay; , time constant.
EEO2, end-exercise
O2. TLT, mean
response time (time to reach 63% of steady-state response).
|
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The magnitude and slope of the
O2 slow component were
determined as the difference between the
O2 at the end of exercise and the O2 at 3 min of
exercise
[
O2(6-3 min)],
O2 at 3 min was taken as
the mean O2 from 10 s
before and 10 s after 3 min, and the end-exercise
O2 was taken as the
mean O2 during
the last 20 s of exercise. The kinetics of the slow component were
described by the parameter estimates derived from the three-component exponential model fit. Similarly, the change in
[La]pl
{[La](6-3 min)}
was determined as the difference between
[La]pl
at the end of exercise and at 3 min of exercise.
Statistics.
Kinetic parameter estimates were analyzed using a two-way
repeated-measures ANOVA for Con vs. Acz and on- vs. off-transitions as
the main effects. Blood data were analyzed for condition and time
effects using a two-way repeated-measures ANOVA. A significant F ratio was further analyzed using
Student-Newman-Keuls post hoc analysis.
O2(6-3 min)
and
[La](6-3 min)
were analyzed using Student's paired
t-test. Statistical significance was
accepted at P < 0.05. Values are means ± SE.
| |
RESULTS |
Subjects.
The physical characteristics and peak values for the ramp exercise test
are presented in Table 1. The individual
work rates for the constant-load exercise tests are presented in Table
2. All subjects completed six repetitions
at the <ET
work rate and three repetitions at the
>ET work
rate for each condition. The work rates utilized during the
constant-load tests were 100 W (73.5 ± 2.9%
ET and 42.2 ± 1.1% O2 peak)
and 237 ± 8.6 W (138.4 ± 2.1%
ET and 79.7 ± 1.2%
O2 peak) for
the <ET and >ET exercise
intensities, respectively.
[La]pl
and acid-base status.
The mean group responses for the
[La]pl
change during step increases in exercise intensities
<ET and
>ET are
presented in Fig. 2. The experimental
design required that
<ET and
>ET protocols be conducted within a single testing session, and therefore pre- and postinfusion values for
[La]pl
were determined at rest before
<ET exercise
only. Preinfusion values for
[La]pl
were similar between Con and Acz (0.8 ± 0.1 and 0.9 ± 0.2 mmol/l, respectively); infusion of Acz had no effect on resting [La]pl
(0.8 ± 0.1 and 0.8 ± 0.1 mmol/l in Con and Acz, respectively; Fig. 2A).
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Fig. 2.
Group response (mean ± SE) of plasma lactate concentration
([lactate])
during moderate- (A) and
heavy-intensity (B) exercise during
control period and after acute acetazolamide administration. Data are
plotted as a function of time including preinfusion (Pre), postinfusion
at rest (R), and during loadless cycling (0 W). Dotted lines, onset of
exercise and recovery.
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|
During moderate-intensity exercise
<ET,
[La]pl
increased above loadless cycling values during Con
(P < 0.05, t 7.5 min) and Acz
(P < 0.05, t 9 min; Fig.
2A). At 2 min after the onset of
exercise,
[La]pl
was higher during Con than during Acz
(P <0.05), a difference that
persisted until 2 min after the end of the exercise bout. Even at this
moderate-intensity exercise, end-exercise
[La]pl
was higher during Con than during Acz (2.0 ± 0.2 and 1.4 ± 0.2 mmol/l, respectively). The
[La]pl
returned to resting values during the recovery period for Con
(P > 0.05, t = 18 min) and Acz conditions
(P > 0.05, t 13 min).
With the onset of
>ET
exercise,
[La]pl
increased above loadless cycling values during Con
(P < 0.05, t 7.5 min) and Acz (P < 0.05, t 8 min) and remained elevated for
the remainder of the protocol (Fig.
2B).
[La]pl
was higher (P < 0.05) during Con
than during Acz by 2 min of exercise and remained higher for the
duration of exercise. End-exercise
[La]pl
was higher (P < 0.05)
during Con than during Acz (9.8 ± 0.9 vs. 7.1 ± 0.5 mmol/l). During recovery,
[La]pl
remained elevated above loadless cycling values in both conditions, with Con
[La]pl
remaining higher (P < 0.05) than
Acz.
To determine the effect of CA inhibition on
O2 kinetics, the confounding
influence of the metabolic acidosis that occurs with chronic Acz
administration was avoided by acute administration of Acz. Plasma pH,
determined 30 min after infusion at rest, was not affected by the
infusion of Acz (7.434 ± 0.006 and 7.429 ± 0.007 for
Con and Acz, respectively). During 0-W pedaling, plasma pH was
slightly, but significantly, higher (P < 0.05) during Con than during Acz (7.429 ± 0.007 vs. 7.412 ± 0.008). At the end of moderate-intensity exercise, plasma pH decreased
(P < 0.05) below loadless cycling
values in Con and Acz; end-exercise plasma pH was higher
(P < 0.05) in Con than in Acz (7.413 ± 0.005 vs. 7.390 ± 0.007). Similarly, during heavy-intensity
exercise, plasma pH decreased (P < 0.05) to end-exercise plasma pH values that were higher
(P < 0.05) during Con than during
Acz (7.353 ± 0.011 vs. 7.335 ± 0.012). When the plasma
acid-base status was expressed as the difference in end-exercise plasma
H+ concentration between Con and
Acz, the difference, although significant (P < 0.05), was small (2.1 and 1.9 nmol/l for
<ET and
>ET, respectively).
O2 uptake kinetics.
A summary of the model parameters derived for the
O2 on- and off-response to a
step increase of moderate-intensity exercise is presented in
Table 3. An example of the
O2 response for a single
subject during
<ET exercise
is presented in Fig. 3. The baseline value
for O2 during 0-W pedaling
(G0) was similar between Con and
Acz conditions. The total gain (GT = G1 + G2) in
O2 from baseline to
steady-state exercise was similar in Con and Acz (963 ± 18 and
970 ± 14 ml/min, respectively), resulting in a similar
end-exercise O2 (1,797 ± 41 and 1,777 ± 44 ml/min during Con and Acz, respectively). For Con
and Acz conditions, O2
returned to baseline values within 6 min of recovery.
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Table 3.
Summary of parameter estimates for model fit of
O2 response at onset and
recovery from moderateintensity exercise below
ET during Con and Acz
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Fig. 3.
O2 on-transient
(left) and off-transient
(right) response for a single
subject during moderate-intensity exercise below ventilatory threshold
during control period (A) and after
acetazolamide administration (B).
 , Breath-by-breath data; solid line, line of best fit. Goodness of
fit for each trial was determined by minimizing residual sum of
squares. A plot of residuals is shown below each model fit.
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At the onset of moderate-intensity exercise,
O2 increased similarly
between Con and Acz, as indicated by the similar MRT (31.2 ± 2.6 and 30.9 ± 3.0 s during Con and Acz, respectively). In addition, no
differences were observed in the model parameters for phase 1 (TD1 and
1) or phase 2 (TD2 and
2) for the
O2 on-response between Con
and Acz. The time course for the
O2 off-response, as indicated
by MRT, was similar to that of the O2 on-response during each
condition. The model parameters (TD and ) for the
O2 off-response were similar
to those for the O2
on-response for Con and Acz. The
O2 deficit
(MRT · GT,
on-transient) was not different between Con and Acz (500 ± 40 and
503 ± 56 ml, respectively). The
O2 debt
(MRT · GT,
off-transient) was similar between conditions (519 ± 17 and 499 ± 25 ml for Con and Acz, respectively) and was not different from
the O2 deficit in either condition.
The model parameters determined for the
O2 on- and off-response to a
step increase to heavy-intensity exercise are summarized in Table
4. An example of the
O2 response for a
single subject during
>ET exercise
is presented in Fig. 4. During 0-W
pedaling, baseline
O2
(G0) was similar between Con and
Acz. At the onset of the step increase in exercise intensity,
O2 increased similarly in Con
and Acz. No differences were observed in gains
(G1,
G2, and
G3) between conditions,
resulting in a similar end-exercise O2 (3,709 ± 111 and 3,735 ± 120 ml/min in Con and Acz, respectively). O2 returned to baseline
values during the 6-min recovery period in Con and Acz.
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Table 4.
Summary of parameter estimates for model fit of
O2 response at onset and
recovery from heavy-intensity exercise above
ET during Con and Acz
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Fig. 4.
O2 on-transient
(left) and off-transient
(right) response for subject in Fig.
3 during heavy-intensity exercise above ventilatory threshold during
control period (A) and after
acetazolamide administration (B).
, Breath-by-breath data; solid line, line of best fit. A plot of
residuals is shown below each model fit.
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O2 increased with similar
kinetics at the onset of the step increase to heavy-intensity exercise,
with no difference between conditions as shown by the similar MRT (69.1 ± 6.1 and 69.7 ± 5.9 s in Con and Acz, respectively). No
differences were observed between Con and Acz for any of the model
parameters (TD and ) during the
O2 on-response. During the
O2 off-transient, MRT was
similar during Con and Acz (50.4 ± 3.5 and 53.8 ± 3.8 s,
respectively); no differences were observed in the model parameters (TD
and ) for the off-response between conditions. The
O2 on-response was slower
(P < 0.05) than the
O2 off-response during Con
and Acz as indicated by the slower MRT and
2. The delayed onset of phase 3 during the on-transient as shown by
TD3 (146.5 ± 16.7 and 121.2 ± 13.4 s in Con and Acz, respectively) was not apparent in the
off-transient, inasmuch as the values for
TD3 (24.8 ± 4.0 and 22.2 ± 3.2 s in Con and Acz, respectively) were similar to those observed for
TD2 (17.0 ± 0.8 and 15.0 ± 1.0 s for Con and Acz, respectively).
Moderate-intensity exercise resulted in a faster
O2 on-response
(P < 0.05) than did heavy-intensity
exercise during Con and Acz, as indicated by the faster MRT and
2 (Tables 3 and 4). During the
off-response, O2 recovered
faster during moderate- than during heavy-intensity exercise during Con
and Acz, as demonstrated by the slower MRT (Tables 3 and 4).
O2 slow component and
[La]pl.
A summary of the O2
slow component and corresponding changes in
[La]pl
during heavy-intensity exercise is presented in Table
5. No differences were observed in the
magnitude [G3 in Table 4 and
O2(6-3 min)
in Table 5] or the time course
(TD3 and
3 in Table 4) over which the
O2 slow component
developed during Con and Acz. The
[La](6-3 min)
and end-exercise
[La]pl
were higher during Con than during Acz; however, the change in
O2 when expressed relative to
the change in
[La]pl
{O2(6-3 min)/[La](6-3 min)}
did not reach statistical significance
(P = 0.055).
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Table 5.
O2(6-3 min),
plasma
[La](6-3 min),
and end-exercise [La]pl during
heavy-intensity exercise above ET for Con
and Acz
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| |
DISCUSSION |
In the present study, constant-load exercise was performed after acute
Acz administration to examine the effect of CA inhibition on the
kinetics of the O2 adjustment
to and from moderate- and heavy-intensity cycling exercise. In
agreement with previous studies (21), we showed that the acute
administration of Acz to inhibit CA was associated with a lowering of
[La]pl
during moderate- and heavy-intensity constant-load exercise and
recovery. However, in contrast to our hypotheses, the results of the
present study demonstrate that 1)
acute CA inhibition with Acz, despite lower
[La]pl,
did not affect the overall time course for
O2 or phase 2 O2 kinetics during moderate-
or heavy-intensity exercise and 2)
the magnitude and time course of the
O2 slow component were independent of the absolute increase in
[La]pl.
Exercise was initiated 30 min after an infusion of Acz to examine the
effects of CA inhibition alone without the confounding effects of a
metabolic acidosis that usually develops with prolonged Acz
administration (22, 37). In the present study the small but significant
acidosis that developed was probably not of any physiological
significance. Although CA activity was not measured in the present
study, it has previously been shown that CA inhibition is essentially
complete in most body tissues at Acz doses of 5-20 mg/kg (24). The
Acz dose of 10 mg/kg used in the present study was similar to that in
the work of Swenson and Maren (37), in which they reported that the
erythrocyte isozymes CA I and CA II were ~93.3 and 99.3% inhibited,
respectively, with an Acz dose of 7-10 mg/kg. In addition, we
observed slower CO2 kinetics after Acz administration, indicating that the dose and protocol used in
this study effectively inhibited CA activity (unpublished observations).
Steady-state O2.
Inhibition of CA with an acute infusion of Acz did not affect
steady-state O2 during
loadless cycling or moderate- or heavy-intensity constant-load exercise
(Tables 3 and 4). These results are in agreement with previous studies
that examined the effects of CA inhibition during submaximal exercise
(28, 34, 36). Studies using short-term high-intensity exercise (21, 22)
or progressive exercise (34) report lower
O2 peak with Acz. The
reason for the lower O2
reported by Kowalchuk et al. (21) is not apparent, inasmuch as the
maximum peak power, average power, and total work performed during the
30-s exercise bout in that study were not affected by Acz. However, the
lower O2 peak reported
for progressive maximal exercise may be a function of the reduced
maximal work rate achieved with this type of protocol (34).
Interestingly, Stager and colleagues (36) reported a similar
O2 peak during progressive maximal exercise, despite a significant reduction in peak
work rate after Acz administration. Although there is evidence from
isolated rat muscle preparations that
O2 may be increased by as
much as 27% after CA inhibition, this increase appears to be a
function of the fiber type (i.e., oxidative fibers demonstrate
increased O2 after CA
inhibition) and the amount and activity of the intracellular CA isozyme
(i.e., CA III) (17). Alternatively, the degree of intracellular CA
inhibition may be minor compared with the inhibition of erythrocyte and
extracellular CA isozymes, and thus the effect on
O2 by this mechanism may be
small. Furthermore, several muscle groups of mixed-fiber composition are involved in cycle ergometer exercise in humans, and therefore any
difference in O2 as a result
of CA inhibition may not be detectable on examination of pulmonary
O2.
O2 on-transient kinetics.
The results of the present study demonstrate that the kinetics of the
O2 on-response for
<ET and
>ET exercise
were not affected by CA inhibition, despite reductions in
[La]pl
early in exercise. These findings are in contrast to previous studies
demonstrating a close association between
O2 on-kinetics and early
blood La accumulation (11,
12, 14). An increase in blood
[La] during the
transition may be a consequence of inadequate
O2 delivery or an inability to
utilize O2 and PCr stores at a
rate sufficient to support ATP production without any contribution from
anaerobic glycolysis. Cerretelli and di Prampero (10) proposed a
control model that suggested
O2 on-kinetics were slowed as a consequence of ATP production by anaerobic glycolysis. According to
their control model, the metabolic drive for oxidative phosphorylation was related to a net increase in free creatine (Cr) and a decrease in
PCr. They hypothesized that regeneration of ATP by anaerobic glycolysis
would not only result in La
production but would attenuate the increase in Cr and decrease in PCr,
thereby slowing the O2
on-kinetics by reducing the drive for oxidative phosphorylation.
Although this mechanism is consistent with the observation of slowed
O2 on-kinetics in association with early La accumulation
(11, 12, 14), the lower
[La]pl
during Acz in the present study was not associated with a faster
O2 on-response. If in our
study the lower
[La]pl
during Acz reflects a lower intramuscular
La production, then this
suggests that O2
during the on-transition is independent of
La accumulation and that
the energy derived from anaerobic glycolysis does not spare or slow
O2 utilization. However, if
La production was similar
in Con and Acz conditions but
La efflux from muscle was
impaired during Acz, then the similar O2 kinetics observed for the
on-transitions would be appropriate. Using an isolated muscle
preparation, De Hemptinne et al. (13) demonstrated that during recovery
from an induced propionic acid load the muscle surface pH transiently
acidified (independent of the acid-base status of the bulk solution)
and that the sarcolemmal acidification was magnified in the presence of
Acz. A similar mechanism acting in vivo during Acz may explain the
lower
[La]pl
in this study. Inhibition of the outward-facing sarcolemmal CA IV
isozyme may lead to increased acidification on the extracellular surface as acid equivalents generated during heavy-intensity exercise are transported from the muscle interior. An increased acidification on
the muscle surface may act to inhibit
La efflux, inasmuch as the
monocarboxylate transporter and diffusion of undissociated lactic acid
are slowed by an extracellular acidosis (20). However, this mechanism
remains speculative, inasmuch as muscle
La production during acute
Acz administration has not been examined previously.
For heavy-intensity exercise at
>ET,
characterizing the O2
on-response is further complicated because of an additional increase in
O2 of delayed onset that
results in a higher asymptotic value for
O2 than that predicted from
the O2-power output relationship for
<ET
exercise (2, 6, 40, 41). Phase 2 kinetics have been reported to be
longer than (19, 29) or similar to (2, 6, 8) the response during
<ET exercise and to be closely associated with increases in end-exercise
[La]pl
(2, 8, 9). In the present study the longer MRT for the
>ET
on-transient demonstrates that the overall rate of adaptation was
slowed compared with
<ET
exercise. In addition, values for 2 indicate that the primary
increase in O2 was also
slowed during the step increase in
>ET exercise
compared with
<ET exercise, despite the gain (i.e.,
G2) projecting to similar
O2/ work rate values
(9.7 ± 0.4 and 10.4 ± 1.1 ml · min1 · W1
for <ET and
>ET,
respectively). This finding is in agreement with the results of
Paterson and Whipp (29), who included only data between 20 s and 3 min
of the on-transient to eliminate any confounding influence that
inclusion of phase 1 or phase 3 would have on determining the model
parameters. They suggested that the longer phase 2 kinetics during
>ET exercise
reflected slowed O2 utilization
and, therefore, would be consistent with increases in muscle and plasma
La. In addition, using
sinusoidal forcing functions to avoid development of the slow
component, Haouzi and co-workers (19) demonstrated a slowing of
O2 kinetics for
>ET
exercise. In contrast to these studies, Barstow and colleagues (2, 6)
argued that the kinetics of the primary rise in
O2 (i.e., phase 2) were invariant across power outputs and end-exercise
[La]pl.
Possible explanations for the differences observed between studies are
unclear. However, a slowing of
O2 kinetics in this exercise
intensity domain is consistent with a slower rate of PCr breakdown (27)
and, therefore, may reflect a slowed drive for oxidative
phosphorylation. As discussed above,
O2 kinetics were similar
between Con and Acz conditions, despite a lower
[La]pl
after Acz. This suggests that
O2 adapts at a rate
independent of an absolute change in
[La]pl
and that this slowing of
O2 kinetics with
heavy-intensity exercise may not be associated with the accumulation of
plasma La, especially if
[La]pl
does not reflect intracellular
La production (see above).
O2 off-transient kinetics.
During the off-transient,
O2 decreased to
loadless cycling values by the end of the 6-min recovery for
<ET and
>ET
exercise, with no difference in kinetics between Con and Acz. For
<ET exercise, symmetry between on- and off-transients was observed as
demonstrated by the similar MRT,
2, and other model parameters. However, although
>ET exercise
resulted in a slower off-transient MRT than
<ET
exercise, the fast recovery phase (i.e., phase 2) was similar between
exercise intensities, as indicated by the similar
2. In addition, recovery from
>ET
exercise was faster than the onset of exercise as indicated by the
faster MRT and 2. These results
are consistent with previous studies demonstrating a slowed recovery
from >ET
exercise compared with recovery from moderate-intensity exercise (29)
but are inconsistent with findings of symmetrical on- and
off-transients during heavy-intensity exercise (3). The similar
off-transient kinetics of phase 2 across exercise intensities found in
this study may reflect the restoration of PCr and venous
O2 stores to preexercise levels.
However, evidence indicates that PCr recovery may be slowed after
high-intensity exercise (27), suggesting that the rate of oxidative
phosphorylation may not reflect the rate of PCr resynthesis after
heavy-intensity exercise.
The O2 slow component is
delayed relative to the onset of exercise
(TD3 = 121.2-146.5 s);
however, during recovery the time delay for the slow recovery phase
(TD3 = 22.2-24.8 s) converged toward TD2 (15.0-17.0 s).
These findings suggest that the kinetic characteristics of the
underlying mechanism of the slow component are different between
exercise and recovery or, perhaps, different mechanisms may underlie
the slow component of O2
during recovery and exercise. Our results clearly demonstrate that
recovery from moderate- or heavy-intensity exercise is not affected by
[La]pl
accumulation, despite the difference in end-exercise and recovery [La]pl
after CA inhibition. The decline in
O2 was temporally (i.e., similar recovery kinetics for Con and Acz) and quantitatively (i.e.,
similar O2 debt for Con and Acz)
similar for each of the respective exercise intensities, which has also
been shown in studies that manipulate blood
[La] by prior
glycogen depletion (35) or leg blood flow occlusion (33) during
exercise. These results cast considerable doubt on the traditional
hypothesis that the "lactacid"
O2 debt (26) is equivalent to the
oxidative removal of La
formed during exercise and the subsequent conversion of
La to glycogen. Further
investigations utilizing on- and off-transients during heavy-intensity
exercise to determine the relationship between oxidative
phosphorylation (i.e., O2
kinetics), PCr degradation-resynthesis, and anaerobic
glycolysis-gluconeogenesis are warranted.
O2 slow component.
Contrary to our hypothesis, the magnitude and time course of the
O2 slow component to the
overall O2 response were not affected by CA inhibition. Although the time course of the slow component has been well described (2, 8) and was shown to arise
predominantly from the exercising muscle (31), the underlying mechanism(s) of the slow component is not clearly evident (see Refs. 16
and 39 for review). The dynamics of the slow component appear to be
related to the magnitude and time course of the increase in blood
[La] (32).
However, increases in blood
[La] by
infusion of L-(+)-lactate (30)
did not increase O2,
suggesting that La in
itself does not have a cause-effect relationship with the slow
component. Our results support this hypothesis, inasmuch as
O2 was similar, despite the
lower
[La]pl
after CA inhibition by 2 min of heavy-intensity exercise. Indeed, the
O2(6-3 min)-
[La](6-3 min)
relationship, although not statistically different (Table 5;
P = 0.055), strongly suggests that
O2 was actually higher relative to the change in
[La]pl
during Acz.
The results of the present study provide evidence against the proposal
made by Wasserman and colleagues (38) that there is a direct link
between the increase in blood
[La] and the
slow component. They suggest that the lactic acidosis and consequent
decrease in arterial pH associated with heavy-intensity exercise result
in a rightward shift in the oxyhemoglobin dissociation curve in the
capillaries of exercising muscles, thereby allowing a greater unloading
of O2 (i.e., Bohr effect) and
aerobic ATP resynthesis (38). In blood the Bohr effect involves the
diffusion of CO2 and
O2 through the erythrocyte
membrane and cytoplasm as well as the generation of
H+ from the intracellular
CO2 hydration-dehydration
reaction. The kinetics of this process are sufficiently fast so that,
under normal conditions, equilibrium is reached within the capillary transit time of 0.75-1 s (25). However, when CA is completely inhibited, the generation of H+
from the hydration of CO2 is
slowed, and therefore the rate of O2 off-loading may also be slowed
(15). Maren and Swenson (25) noted that, in humans, there is an
~360-fold excess of erythrocyte CA activity, such that the Bohr
effect may proceed normally and CO2 hydration is not rate
limiting. We observed that
CO2 kinetics were slowed
during the on-transient, suggesting that
CO2 hydration may be slowed
(unpublished observations). Inasmuch as the
O2 slow component was not
affected by CA inhibition (either in kinetics or magnitude), this
suggests that the Bohr effect is not altered by CA inhibition at the
level of the muscle or, alternatively, the Bohr effect does not
contribute to the development of the O2 slow component.
Summary.
In conclusion, Acz-induced CA inhibition resulted in lower
[La]pl
during moderate- and heavy-intensity exercise but did not affect
O2 on- or off-transition
kinetics at either exercise intensity. Compared with moderate-intensity
exercise, phase 2 and MRT were slowed for
O2 kinetics during the
on-transition to heavy-intensity exercise. Recovery from
heavy-intensity exercise was associated with slower
O2 off-kinetics
with no difference between Con and Acz; the slower off-kinetics were
due to the appearance of a slow recovery phase (i.e., phase 3), since
the kinetics of the fast recovery phase (i.e., phase 2) were similar to
those observed after moderate-intensity exercise. The longer kinetics of the slow phase of recovery were not associated with an elevated [La]pl.
The dissociation between the
O2 slow component and
[La]pl
suggests that mechanisms other than plasma
La appearance or the
associated change in acid-base status determine the time course and
magnitude of the slow component.
The authors thank the subjects who took part in this study. The
technical support of Brad Hansen is greatly appreciated.
Financial support was provided to J. M. Kowalchuk by an operating grant
from the Natural Sciences and Engineering Research Council of Canada.
B. W. Scheuermann was supported by a Natural Sciences and Engineering
Research Council Graduate Scholarship. This research was carried out at
The Centre for Activity and Ageing (affiliated with the School of
Kinesiology, Faculty of Health Sciences, and the Faculty of Medicine at
The University of Western Ontario and The Lawson Research Institute at
the St. Joseph's Health Centre).
Address for reprint requests: J. M. Kowalchuk, School of Kinesiology,
3M Centre, The University of Western Ontario, London, ON, Canada N6A
3K7.
Top
Abstract
Introduction
![<A><AC>V</AC><AC>˙</AC></A><SC>o</SC><SUB>2 on</SUB> = G<SUB>0</SUB> + G<SUB>1</SUB> ⋅ [1 − <IT>e</IT><SUP>−(<IT>t</IT> − T D<SUB>1</SUB>)/&tgr;<SUB>1</SUB></SUP>] ⋅ <IT>u</IT><SUB>1</SUB> + G<SUB>2</SUB>](/content/vol85/issue4/fulltext/1384/img001.gif)
![<A><AC>V</AC><AC>˙</AC></A><SC>o</SC><SUB>2 on</SUB> = G<SUB>0</SUB> + G<SUB>1</SUB> ⋅ [1 − <IT>e</IT><SUP>−(<IT>t</IT> − T D<SUB>1</SUB>)/&tgr;<SUB>1</SUB></SUP>] ⋅ <IT>u</IT><SUB>1</SUB> + G<SUB>2</SUB>](/content/vol85/issue4/fulltext/1384/img004.gif)
![<A><AC>V</AC><AC>˙</AC></A><SC>o</SC><SUB>2 off</SUB> = G<SUB>0</SUB> + G<SUB>1</SUB> ⋅ [<IT>e</IT><SUP>−(<IT>t</IT> − T D<SUB>1</SUB>)/&tgr;<SUB>1</SUB></SUP>] ⋅ <IT>u</IT><SUB>1</SUB> + G<SUB>2</SUB>](/content/vol85/issue4/fulltext/1384/img006.gif)
