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ClinicalScience( 1986) 70,435-441
435
Thermogenesis in human skeletal muscle as measured by direct
microcalorimetryand muscle contractile performance during
B-adrenoceptor blockade
BIRGER FAGHER, HANS LIEDHOLM*, MARIO MONTI A N D ULRICH MORITZt
Departments of’lnternal Medicine, *Clinical Pharmacology and i- Physical Medicine, University Hospital, Lund, Sweden
(Received 1OJuly/4 November 1985; accepted 2 December 1985)
Summary
1. The influence of /3-adrenoceptor-blockade
on skeletal muscle was studied in ten healthy
males with propranolol, atenolol and pindolol randomly given for 8 days each in a cross-over double
blind test. After 7 days on each drug, muscle function was tested by an isokinetic dynamometer.
Thermogenesis in biopsy samples taken from
vastus lateralis muscle after a low grade exercise
was studied after 8 days on each drug by direct
calorimetry with a perfusion microcalorimeter.
2. Before drug administration, a median heat
production rate of 0.67 mW/g of muscle was
measured. This value was significantly reduced by
25% during propranolol, but no significant change
was found during atenolol or pindolol administration.
3. Peak torque decline during isokinetic endurance test changed significantly in knee flexor but
not in extensor muscles, from 15% to 27% after
propranolol and from 15% to 23% after pindolol.
Maximum dynamic strength was unaltered.
4. Our data suggest that blockade of sympathetic &receptors decreases thermogenesis in
human skeletal muscle and impairs isokinetic
endurance.
Key words: /3-adrenoceptor blockade, heat production, isokinetic contraction, microcalorimetry,
skeletal muscle, thermogenesis.
Abbreviations: HR,,, heart rate after 30 repeated
knee contractions; WE,,, rated perceived exertion.
Correspondence: Dr Birger Fagher, Department of
Internal Medicine, University of Lund, 221 85 Lund,
Sweden.
Introduction
Evidence indicating a thermogenic effect mediated
by /3-receptors has arisen from different sources.
Temperature regulation in resting subjects
exposed to cold conditions was shown to be suppressed by the non-selective /3-blocker oxprenolol
[l]. The thermogenic response to insulin and
glucose infusion, as determined by indirect calorimetry, decreased after intravenously administered
propranolol, whereas the basal metabolic rate
after an overnight fast was unchanged [2,3]. In the
postprandial state, propranolol has also been
shown to reduce the thermogenesis, measured as
the inhibitory effect on the increase in metabolic
rate that normally occurs after food intake [4]. PIselective blockers have not been studied in a similar way.
Microcalorimetric technique has been usefully
applied to different cell systems in the last 15 years
(see, e.g., [5, ,6]). Recording of heat production by
direct calorimetry has not been used, however, in
the study of metabolic effects of /3-blockers. Our
purpose was to evaluate thermogenesis of human
skeletal muscle with a microcalorimeter to ascertain whether medication with various P-adrenergic
blockers could have an inhibiting influence. Three
drugs with widely different pharmacodynamic
profiles were chosen: propranolol (non-selective),
atenolol (p,-selective) and pindolol (non-selective
with partial P,-agonist activity). Furthermore,
muscular strength and dynamic endurance were
studied by an isokinetic technique [7]. It also
seemed appropriate to see if an altered heat production might possibly be linked to changes in the
perception of fatigue, since impaired physical performance and muscle fatigue during /3-adrenergic
blockade have been a matter of discussion and
research in recent years [8-161.
436
B. Fagher et al.
Experimental design
Three drugs were randomly administered for 8
days in a cross-over double blind study: propranolo1 (80 mg twice daily), atenolol (50 mg twice
daily) and pindolol (5 mg twice daily). T h e three
drugs were given as specially prepared identical
capsules which were counted for assessment of
compliance. Each subject received all three drugs
subsequently with at least 1 week intervening
between the study periods. Muscle function was
tested after 7 days in each period. Muscle biopsies
and blood for drug analyses were taken after 8
days on each drug, at the same hour in the morning as the muscle function tests, 2.5 h after drug
intake and after a 12 h fast. For comparison, all
measurements were also performed before the
first medication period. T h e subjects were encouraged not to change their physical activities
and habits during the study. They were informed
as to the purpose and possible risks of the investigation and they gave their consent. T h e study was
approved by the Ethics Committee of the University of Lund.
Material and methods
Subjects
Twelve healthy habitually active and nonsmoking males were recruited for the study. None
of the subjects could be classified as an athlete.
Their age was 18-38 years, their height
171-195 cm and their weight 60-85 kg. They
took no medication. One subject was withdrawn
from the study because of a respiratory tract infection and another subject was excluded owing to
non-compliance. T h e other ten subjects had 100°/o
compliance according to the number of capsules
returned.
Tests of contractile performance
Maximum dynamic strength [7, 171 was
measured in knee extensor and flexor muscles
with an isokinetic dynamometer (Cybex 11; Lumex
Inc., New York, U.S.A.). T h e angular velocity
during maximum dynamic contraction was controlled at a pre-set level and the velocities studied
were 90°/s and 180"/s. Measurements were performed with the subject seated with the hips fixed
in 90" flexion. T h e leg was attached to the lever
arm of the dynamometer and the centre of the
dynamometer's axis of rotation was aligned with
the anatomical axis of rotation of the knee joint.
T h e methodological variation of maximum
strength measurements was previously determined to be about 10% [17].T h e torque produced
by the muscle contraction was calculated and
expressed in Nm (force x the length of the lever
arm). T h e maximum value of three consecutive
contractions was used for calculation.
Local muscle fatigue was assessed by 30
repeated maximal isokinetic contractions of the
knee extensor and flexor muscles at an angular
velocity of 90"/s [ 181. Dynamic endurance was calculated as the peak torque decline, expressed as a
percentage, from the mean of the three initial
contractions and the mean of the three final
contractions. Between the contractions there was
an approximately 1 s relaxation period. T h e coefficient of variation was previously reported to be
3.2% at an angular velocity of 180°/s [18]. Rated
perceived exertion for the endurance test (RPE,,)
was studied by asking the volunteers to estimate
leg effort on a scale from 6 to 20, according to
Borg [ 191. Heart rate was measured before and at
the end of the endurance test (HR,,).
Blood pressure, heart rate and exercise test
O n the eighth day on each drug, immediately
after drug intake, heart rate and blood pressure
were measured after at least 10 min rest in the
supine position. T h e mean of three successive
readings was taken. Diastolic blood pressure was
indicated by Korotkoff phase V. Mean arterial
blood pressure was calculated by adding one-third
of the pulse pressure to the diastolic blood pressure. A mercury sphygmomanometer with appropriate cuffs was used. To evaluate the influence of
the different drugs, a submaximal exercise test was
thereafter performed on an electrically braked
bicycle ergometer (Siemens-Elema). T h e volunteers worked for 4 min on each of the following
work loads: 50W, 100 W, 1 5 0 W and 200W.
Heart rate was measured by auscultation with a
stethoscope after 4 min on each load. All exercise
tests were supervised by the same person (H.L.).
Afterwards, the subjects were advised to rest
before the muscle biopsy, which was preceded by
a new, light exercise.
Muscle biopsy
O n four occasions, before and after 8 days on
each drug, a muscle biopsy was performed. After
local anaesthesia of the skin, the volunteers performed a 4 min exercise with a work load of 50 W.
Immediately after the termination of the exercise,
while recording the time lag, a biopsy was taken in
the supine position from the middle portion of the
vastus lateralis muscle by use of the needle technique of Bergstrom [20]. All biopsies were performed by the same person (B.F.). T h e muscle
P-Receptor blockade and muscle thermogeiiesis
specimens were collected and handled in icechilled Krebs-Ringer-phosphate
buffer (0.12
molA NaCl, 0.005 molA KCI, 0.001 molA CaCl,,
0.016 molA Na,HPO,, 0.001 molA MgSO,, pH
7.40) supplemented with glucose (8.25 mmolA)
and insulin (0.1 unit/ml). Fibre bundles with a
diameter of 0.5-1 mm and a resting length of about
4-7 mm were teased away, carefully blotted on filter paper and quickly weighed. Specimens with a
weight of about 50 mg were used in the calorimetric
experiments (Table 1).
Calorimetric measurements
Total metabolic activity in muscle was monitored by measurement of heat production rate by
perfusion microcalorimetry with a 0.7 ml flowthrough vessel. We have recently described the
technique in detail [21]. Measurement was made at
37°C and pH 7.4. A flow of fresh buffer medium
(see above) was pumped through the reaction vessel at a rate of 5 ml/h and close to the resting
muscle fibre bundles in order to keep p H constant. The thermal power generated by the muscle
was calculated from the observed power-time
curve and expressed in units of mW/g wet weight.
The power values reported are mean values during the second hour after the start of the calorimetric experiments. T h e coefficient of variation
for the method was previously determined to be
4.2%. Time intervals from biopsy to the start of
the calorimetric experiments are shown in Table 1.
Drug analysis
Venous blood for analysis of P-blockers was
sampled both before drug intake and immediately
after the muscle biopsy. T h e blood was collected
437
in heparinized Venoject tubes, centrifuged and
stored at - 20°C until analysis. Previously
described methods were used for determination of
propranolol[22], atenolol[23] and pindolol[24].
Statistical procedures
Data are presented as medians and interquartile
ranges unless otherwise noted. Differences
between groups have been tested by Wilcoxon's
non-parametric test for paired data. Correlation
coefficients were calculated according to Spearman's rank test. P values of less than 0.05 were
considered as significant.
Results
Muscle heat production
Before drug administration, the median heat
production rate in muscle after the ergometric
exercise was 0.67 mW/g with an interquartile
range of 0.53-0.75 (Table 1). Neither before nor
during medication was any correlation found
between heat production and the time lag between
the termination of the exercise and the biopsy.
After administration of propranolol, the muscle
heat production was 0.46 mW/g (interquartile
range 0.28-0.68), which is 25% lower than the
premedication level, P= 0.02 (Fig. 1).The corresponding values after atenolol and pindolol were
0.62 mW/g ( - 4%) and 0.53 mW/g ( - 13%),
respectively. These two values were not significantly different from the pre-drug value. Representative calorimetric curves, obtained from one
subject before and after medication with propranolol, are shown in Fig. 2.
TABLE
1. Premedication characteristics and changes (A) after P-adrenoceptor blockade with propranolol,
atenolol and pindolol
Results are median values. Interquartile range is given in parentheses; total range for drug plasma levels.
Significance versus initial value: **P< 0.02, ***P< 0.01.
Pre-drug values
Drug plasma levels (nmol/l)l'
Mean arterial pressure (mmHg)
82 (79-86)
(A%)
Time of biopsy after exercise (min)
1.3 (1.0-1.3)
Muscle sample weight (mg)
50.1 (49.8-50.4)
Start of calorimetric measurement
19 (17-22)
after biopsy (min)
Heat production in muscle (mW/g) 0.67 (0.53-0.75)
(A%)
?At the time of biopsy.
-
Propranolol
194 (131-530)
- 11 ( - 11 to - 7)***
1.3 (1.0-1.2)
50.1 (49.6-50:s)
20 (20-24)
0.46 (0.28-0.68)**
-25(-41t0 - 4 )
Atenolol
Pindolol
1313 (713-1838)
73 (40-133)
13 ( - 13 to - 11)*** - 5 ( - 9 t 0 3 )
1.3 (1.1-1.5)
1.3 (1.2-1.31
49.4 (46.1-49:7)
48.8 (39.7-49:7)
22 (19-25)
25 (18-30)
0.62 (0.53-0.72)
- 4 ( -27 to 31)
0.53 (0.40-0.63)
- 13 (-40 to 0)
B. Fagher et al.
438
Contractile performance
Results of the muscle function measurements
are summarized in Table 2. After B-blockade,
maximum dynamic strength of the knee extensor
and flexor muscles did not change significantly
from the basal premedication value.
T h e median peak torque decline before medication was 29% in knee extensor muscles and 15%
in knee flexor muscles. T h e change in dynamic
endurance of the extensor muscles was not significant for any of the drugs, although significant
changes were found in the flexor muscles after
both propranolol, from 15% to 25% ( P < 0.02)
c,
10
I-
"
0
U
1
2
3
4
5
Time (h)
-100 J
P=O.O2
Propranolol
NS
Atenolol
NS
Pindolol
FIG. 1. Change ( O h ) from initial value in muscle
heat production (vastus lateralis) after 8 days of
B-adrenoceptor blockade in ten healthy subjects.
NS, Not significant. Wilcoxon's test for paired
data.
FIG. 2. Representative measurements of thermal
power ( P )on biopsy samples from vastus lateralis
muscle, obtained from one subject before and
after medication with propranolol. Sample
ampoule inserted into the microcalorimeter 1, and
taken out t. P,= mean value during the second
hour of registration.
TABLE
2. Peak torque values of the knee extensor (Ext) and flexor (Flex) muscles before and the change (A)
after P-adrenoceptor blockade in healthy men are shown in (a) dynamic endurance and rated perceived
exertion (RPE,,) after 30 knee contractions are given in (b)
Results are median values. Interquartile range is given in parentheses. Significance versus initial value:
*P< 0.05, **P < 0.02.
(a)
Angular
velocity
Maximum strength
Pre-drug values
(Nm)
90"h
18o"/s
Ext 163 (127-167)
Flex 97 (77-108)
Ext 111 (92-125)
Flex 79 (67-83)
Propranolol
(A%)
Atenolol
(A%)
Pindolol
(A%)
0 ( - 9 to 10)
9 ( - 2 to 26)
10(-2to23)
15 (13 to 27)
9 ( - 9 to 19)
5 ( 2 to 27)
6 ( - 15 to 16)
8 (3 to 12)
1 ( - 6 to 20)
8 ( - 9 to 29)
10 ( - 15 to 25)
6 ( - 10 to 18)
Dynamic endurance (peak torque decline in "/o)
Pre-drug values
Propranolol
Atenolol
Pindolol
Ext 29 (19-35)
Flex 15 (13-21)
(Score) 16 (15-17)
(A)
29 (28-36)
25 (23-30)**
16.5 (15-19)
l(0-1.8)
30 (26-39)
15 (11-26)
17 (16-17)
l(0-1.8)
29 (25-34)
23 (16-26)*
17 (16-18)
1.5 (0-2.0)*
t At the last of 30 contractions.
/3- Receptor blockade and muscIe thermogenesis
439
150- ( a )
-. 140-
3.9 130110120-
;;i
8 1003 90-
-2
2
80706050-
/
I
I
J
P <0.02
Propranolol
NS
Atenolol
(i)
P <0.05
Pindolol
FIG. 3. Dynamic endurance (peak torque decline
during 30 repeated contractions) tested in knee
flexor muscles after B-adrenoceptor blockade.
Results are expressed as change from initial
value.
and pindolol, from 15% to 23% ( P < 0.05) (Fig. 3).
The force decline in knee flexor muscles showed
no correlation with the decrease in heat production of the vastus lateralis muscle.
Perceived exertion
Six subjects reported increased REP,, values
after each medication period. However, the
change in WE,, was statistically significant for
pindolol only, with a median increase of 1.5 Borg
scale units ( P < 0.05) (Table 2). Changes in the
perception of muscular effort during /3adrenoceptor blockade did not correlate with
changes in dynamic endurance or in muscle heat
production.
Heart rate and blood pressure
Resting heart rate decreased significantly
during treatment with propranolol and atenolol
( P < 0.01), and with pindolol in the sitting position
only (P<O.O5) (Fig. 4, Table 1). Heart rate was
also significantly reduced by all three drugs during
the ergometric exercise as well as after the endurance test. However, pindolol was less effective
than propranolol and atenolol, especially at low
grade ergometric work (50-100 W). Resting mean
arterial blood pressure is shown in Table 1. Diastolic blood pressure was significantly lowered by
all three drugs, whereas the systolic blood pressure was lowered by atenolol only ( P < 0.05).
(ii)
(iii)
Rest 50 100 150 200 Post- Rest HR,,
(supine)
exercise 2’ (sitting)
Work load (W)
NS * *** *** *** ***
* **+
*** *** *** * NS ***
*** *** *** NS ** ***
*** *
*** NS
FIG.4. Comparison of the effects of propranolol
( O ) , atenolol ( x ) and pindolol ( A ) on heart rate
during ergometric exercise ( a ) and after 30
repeated knee contractions (HR,,) ( b ) with the
pre-drug state (0).NS, Not significant, *P < 0.05,
**P= 0.02, ***P< 0.01. Wilcoxon’s test for paired
data. (i) Pindolol versus untreated. (ii) Pindolol
versus atenolol. (iii)Pindolol versus propranolol.
Plasma drug concentrations
The drug levels 2.5 h after drug intake are shown
in Table 1.There was no correlation between drug
plasma levels, sampled immediately before and
2.5 h after drug intake, and heat production in
muscle, heart rate or mean arterial blood pressure.
Discussion
Thermogenesis in biopsy samples from vastus
lateralis muscle was found to be significantly decreased by non-selective j3-adrenoceptor blockade but not by p,-selective blockade. Thus, mainly
&receptors seem to be involved in the process.
The lack of thermogenic inhibition after medication with the non-selective /%blocker pindolol
was probably due to the marked partial agonist
activity on j3,-adrenoceptors inherent in this drug.
The degree of receptor stimulant effect of pindolo1 differs from one tissue to another [25]. In
human lymphocytes, for example, only pindolol
was shown to stimulate cyclic AMP generation, in
contrast to three other P-blockers with pharmacologically different properties [26]. As far as we
know, there are no such data for human skeletal
muscle. On the other hand, propranolol has been
shown to produce a decrease in cyclic AMP content in human skeletal muscle [ 121.
440
B. Fagher et al.
Our finding, by direct calorimetry, of a reduced
metabolic heat production in skeletal muscle during propranolol medication, is in agreement with
indirectly obtained data [2-41. T h e causal
mechanisms for this has received little attention. It
seems likely that the adrenergic system is involved
with a direct retardation of glycogenolysis. A decreased activity of glycogen phosphorylase a in
human muscle after propranolol was recently
shown, both at rest and after short intense work
[12]. There is also evidence that the degradation of
glycogen is mediated by &receptors (see e.g.
[13]), down to lactate, leading to energy production and liberation of heat.
To evaluate the effects of B-blockers, the doses
and plasma drug levels have to be taken into consideration. T h e muscle responses were found to
be unrelated to the plasma concentrations of the
drugs. Equipotent doses of propranolol and
atenolol were administered, as judged from the
almost identical effects on heart rate and mean
arterial blood pressure. In quantifying the relative
P-adrenoceptor blocking potency of pindolol,
methodological problems arise when it is compared with drugs devoid of partial agonist activity
[27]. At rest and at low and intermediate work
loads, less reduction in heart rate is to be expected
with pindolol, reflecting the P,-receptor stimulant
effect at low sympathetic tone. At more intense
work, when sympathetic tone is higher, the difference in heart rate in response to atenolol and
propranolol will consequently be less or minimal,
as in the submaximal exercise test in the present
study or at the end of the isokinetic endurance test
(Fig. 4). It is therefore not surprising that pindolol
in circumstances with comparatively less pronounced agonist activity, induced an almost similar change in peak torque decline as propranolol
(Fig. 3). This is again evidence for mainly a p2receptor mediated influence on skeletal muscle, as
P,-selective blockade had no significant effect on
peak torque decline.
P,-Receptor blockade did not affect maximum
voluntary contraction strength whereas, in the isokinetic endurance test, an influence was found on
flexor muscles of the knee. T h e fact that no effect
was found on knee extensor muscles might be
explained by the fibre-type composition, as the
hamstring muscles in man have more glycogenolytic type I1 fibres than the quadriceps muscle
[281.
As judged from the comparison of the perception of discomfort in the exercising muscles
(WE3& analysed and rated according to the Borg
scale, and the objective way of measuring fatigue
as change in peak torque decline, propranolol
gave diverging results. However, as many factors
are responsible for the subjective feeling of fatigue
[9, 291 and the number of subjects studied was
small, the change in clinical scores must be interpreted with great caution.
Acknowledgments
Excellent technical assistance was given by Mrs
Berit Persson, Mrs Helga Fleischer, Mrs Minda
Lundwall and Miss Marianne BCve. T h e study was
in part supported by a grant from Sandoz AB,
Sweden, and also by grants no. 3880 and no. 6587
from the Swedish Medical Research Council. We
thank Sandoz AB, Sweden, for supplying the drug
capsules.
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