Oxidative metabolism of hypertrophic skeletal muscle in the rat

EXPERIMENTAL

Oxidative

JOHN

48, 222-230 (1975)

NEUROLOGY

W.

Departments

Metabolism

CARLO,

R.

STEPHEN

of Neurology

Baltimore,

of Hypertrophic in the Rat

Maryland

University

MAX,

AND

Skeletal

DAVID

H.

Muscle

RIFENBERICK

l

of Maryland School

of Medicine, of Physical Education, of MaryZand, College Park, Maryland 20742 and Pedaktrics, 21201 and

Received

Ukuersity Department

February

21, 1975

The object of this study was to determine whether skeletal muscle adjusts its oxidative metabolism in response to compensatory hypertrophy., ‘We therefore, measured the production of ‘“COZ from glucose-6-Y and B-hydroxybutyrate-3-Y by homogenates of rat plantaris and soleus muscles undergoing compensatory hypertrophy produced by elimination of synergists. There was a decrease in substrate oxidation by hypertrophic muscles. These results are in contrast to the increased oxidative capacity observed in skeletal muscle following endurance training.

INTRODUCTION Extensive biochemical and histochemical studies of the metabolic responses of skeletal muscle to altered usage have revealed that muscle adapts to endurance exercise by increasing its oxidative capacity (1, 2, 5, 11-13, 21-23) and to decreased activity by diminishing its oxidative capacity (4, 19, 20, 24, 25, 27). In contrast, muscle energy metabolism has not been thoroughly investigated in compensatory hypertrophy. The metabolic response of skeletal muscle to compensatory hypertrophy should be documented to further our understanding of muscular metabolic adaptations. In the present study, we have assessedsubstrate oxidation by homogenates of skeletal muscles subjected to compensatory hypertrophy 1 Reprint requests should be sent to Dr. Stephen R. Max, Department of Neurology, University of Maryland School of Medicine, Baltimore, Maryland 21201. We thank Drs. D. H. Clark, C. D. Dotson, and Lois Roeder for invaluable advice, Ms. B. H. Sohmer for expert technical assistance, and Ms. B. Pasko for preparation of the typescript. This research was supported in part by U.S.P.H.S. Grants NS-05077 and HD-06291-03 and the Bressler Reserve Fund. David H. Rifenberick was the recipient of NIH postdoctoral fellowship 1 F02 NS 54205-02. 222 Copyright All rights

s

1975 by Academic Press, Inc. reprcduction in any form reserved.

MUSCLE

IIYPERTROPHY

223

produced by the elimination of synergistic muscles (6, S) This method of causing hypertrophy has recently been used in a number of studies (6, S-10, 14, 16, 17, 2s). Some of our data have been described in a preliminary report (26). MATERIALS

AND

METHODS

The materials used and their sources were: glucose-6-l’C. D,r.-/3-hydroxybutyrate-3J4C, tryptamine bisuccinate-2J*C, PPO, POPOP and hyamine hydroxide, New England Nuclear ; EDTA, CoA, NAD’, NADP’, ADP, ATP, bovine serum albumin, Triton X-100, and Tris-HCl, Sigma; and Toluene, Eastman. Tissue homogenizers (Tenbroeck), rubber septa, and hanging center wells were products of the Kontes Glass Company. Albino, male, Wistar rats, weighing 225-250 g, were anesthetized with chloral hydrate (400 mg/kg, ip). Hypertrophy of soleus and plantaris muscles was produced by tenotomy of synergists (soleus and gastrocnemius, or plantaris and gastrocnemius) (6). Sham-operated contralateral muscles served as controls in all experiments. On days 8, 19, and 41 following tenotomy of synergists, rats were decapitated, and the muscles were excised and weighed. Mincing, homogenization, monoamine oxidase assay, and measurement of l”CO, production from glucose-6-Y and ,&hydroxybutyrate-3-1JC were carried out as previously described (24, 27). Substrate concentrations and specific activities were as follows : glucose-6-14C, 5 mM. 6.34 x loj dpm/qol : ,&hydroxybutyrate-3-14C, 10 mM, 3.06 X lo” dpm/pmol. The data were computed utilizing two reference bases, viz., grams fresh weight and total muscle. Statistical significance of differences between experimental and control preparations was evaluated with the T-test. RESULTS Eight days after tenotomy of synergists there was a significant increase in the fresh weight of plantaris and soleus muscles (Table 1). Tenotomy of gastrocnemius and soIeusmusclescauseda diminution of the oxidation of glucose-6-Y by homogenates of plantaris muscles (Table 2). The oxidation of glucose-6-14Cby homogenates of hypertrophic soleus muscleswas similarly diminished (Table 3). Table 4 shows rates of oxidation of P-hydroxybutyrate-3-14C by homogenates of rat plantaris muscles following tenotomy of soleus and gastrocnemius. There were no significant differences in p-hydroxybutyrate-3-14C oxidation when total muscle activity was determined. On a gram fresh weight basis, however, there was a significant reduction in p-hydroxybutyrate-3-14C oxidation in plantaris muscles on days 8, 19, and 41 (Table 4). Similarly, there was a diminution in ,&hydroxybutyrate-3-14C oxida-

8

41

a Experimental

procedures

as described

in the text.

356.96 569.58 464.68

f f f

73.02 77.44 132.35

f f f

8 8 7

8 19 41

nmols/hr/muscle Ctl

292.36 310.68 438.55

in the text.

82.83 103.47 110.94 Data

TABLE

are means

f



P

MUSCLES

2

P

f

0.22

0.14 0.17

experimental

f f f

Exp

control.)

0.18

0.11 0.14

; Ctl,

control.)

1537.25 2126.56 1262.98

nmols/hr/gfw

f f f

0.02

0.02 0.02

243.39 335.84 334.48

HYPERTROPHY~

Ct1

f

ff

Soleus fresh weight) Ct1

COMPENSATORY

Ctl,

0.05

(grams

OF SYNERGISTS~

0.02 0.04

191.04 206.94 219.97

FOLLOWING

995.07 847.41 856.84

oxidation

MUSCLES

f

ff

EXP

TENOTOMY

experimental;

FOLLOWING

SD. (Exp,

Substrate

NS
1

SD. (Exp,

OF RAT PLANTARIS

0.08

0.03 0.02

Data

0.39 f

0.23 f 0.27 f

Exp

No. of rats

AND SOLEUS

Plantaris fresh weight) Ct1

BY HOMOGENATES

as described

OF GLUCOSE-6J4C

procedures

0.05

0.50

f

0.02 0.08

0.29 f 0.37 f

J+P

(grams

OF RAT PLANTARIS

ow

Days after tenot-

OXIDATION

a Experimental

88

No. of rats

198

omy

Days after tenot-

WEIGHTS

TABLE


P

5

<0.05

d E s R

0 $
P

E I&

procedures

procedures

173.89 135.26 152.59

as described

f f f

7 8 6

8 19 41

a Experimental

EXP

No. of rats

Days after tenotow

625.91 841.66 1097.64

79.03 42.77 54.19

as described

OXIDATIONOFP-HYDROXYBUTYRATE-3-W

a Experimental

f f f

8 5 6

8 19 41

238.60 162.46 144.59

EXP

No. of rats

GLUCOSE-~-~~C

Days after tenotov

OXIDATIONOF

f f f Data

60.08 19.14 76.88

in the text.

856.57 857.25 1040.25

nmols/hr/muscle

NS
P

f

4

SD. (Exp,

are means

NS NS NS

P

f

Ctl,

349.64 146.07 275.27 control.)

2046.74 1963.30 2023.61

nmols/hr/gfw

COMPEN~ATOKY

f f f experimental;

2110.19 2286.39 2137.96

EXP

Ctl,

548.88 206.51 288.41

control.)

3650.47 3200.86 2733.09

nmols/hr/gfw

MUSCLESFOLLOWINGCOXPENSATORY

oxidation

SD. (Exp,

Substrate

f f f

Exp

experimental;

1294.97 937.52 630.71

oxidation

MUSCLESFOLLOWING

Substrate

3

RAT PLANTARIS

TABLE

are means

224.56 114.87 298.81

Data

f f f

Ctl

TABLE OF RAT SOLEUS

BYHOMOGENATESOF

in the text.

279.82 279.36 358.15

nmols/hr/muscles Ct1

BY HOMOGENATES

f 724.35 f 290.02 f 303.38

Ct1

f 427.77 f 223.53 f 32.35

Ctl


P


P

HYPERTROPHY"

HYPERTROPHY"

226

CARLO,

MAX

AND

RIFENBERICK

tion by homogenates of hypertrophic soleus muscle on the basis of grams fresh weight, but not on the basis of total activity (Table 5). A possible cause of the loss of oxidative activity observed in hypertrophic muscles is reduction in mitochondrial content. That this possibility is unlikely is suggested by the essentially unaltered monamine oxidase activities observed in soleus and plantaris muscles on days 8, 19, and 41 following tenotomy of synergistic muscles (Tables 6 and 7). DISCUSSION Compensatory hypertrophy of skeletal muscle resulting from elimination of synergists has been extensively employed as a model for physiological and histological studies. It has been suggested that such hypertrophy is caused by passive mechanical stretching produced by the relatively unopposed contractions of the antagonistic muscles (10, 18). Hypertrophy of this type is accompanied by an increase in muscle fiber diameter, an increase in sarcoplasmic proteins, and a decrease in contractile proteins 63, 9). The data described in the present report demonstrate that muscular hypertrophy resulting from elimination of synergists is not accompanied by enhanced oxidative activity, in contrast to the results of endurance training (1, 2, 11-13, 21-23). In fact, in many experiments we noted a decrease which may simply reflect dilution of mitochondria in a muscle that is increasing in mass. This conclusion is supported by the essentially unchanged homogenate monamine oxidase activity (Tables 6 and 7). Monoamine oxidase is useful as an estimation of mitochondrial content (25). The decrease in substrate oxidation which we observed (Tables 2-S), however, suggests that compensatory hypertrophy may result in an is impairment of oxidative metabolism. The cause of this impairment unknown. Our findings regarding oxidative metabolism are consistent with those of previous investigators who have studied compensatory hypertrophy and noted decreased contractile strength and diminished intermyofibrillar ATPase activity (10, 14, 16), and with histochemical studies of hypertrophy (3, 8, 10). I n addition, decreased activity of the mitochondrial enzyme succinate dehydrogenase has been reported in muscles from weight lifters compared with those from untrained men (7). It is thus apparent that, although it causes a substantial increase in muscle weight (Table l), this type of muscular activity is of no benefit to muscle in terms of increasing its ability to do work aerobically. We recently described similar results in hypertrophy of the rat hemidiaphragm following unilateral phrenectomy ( 15).

5 6

6 6 6

No. of rats

L1Experimental

41

8 19

after tenoton,>

Days

U Experimental

8

8

No. of rats

19 41

Days after tenotomy

OXIDATION

as described

zk 22.70 f 27.48 f 32.04

Exp

Data

in the text.

172.43 129.86 164.08 The

f f f

Ctl

TABLE

are means

NS NS NS

P

data

20.11 7.30 30.98

5

6

SD.

are means

NS
P

f

f f f

Exp

655.74 497.5 369.75

f f f

Exp

; Ctl,

79.99 36.68 28.37

control.)

704.36 478.02 399.97

f f f

Ctl 72.6 25.38 25.52

P

NS NS NS

P


HYPEKTROPHY=

1592.56 604.31 1365.23

HYPERTROPHY~

f f f

nmols/hr/gfw

control.)

12170.48 8717.79 8282.19

Cd

COMPENSATORY

nmols/hr/gfw

COVPENSATORY

experimental

oxidase

FOLLOWING

; Ctl,

1370.47 898.99 1296.14

FOLLOWING

experimental

7877.32 5405.01 5606.92

SD. (Exp,

Monoamine

MUSCLES

f

MUSCLES

oxidation

(Exp,

SOLEUS

Substrate

OF RAT

TABLE

PLANTARIS

345.14 172.75 225.23

OF RAT

f f f

nmols/hr/muscle

OF HOMOGENATES

in the text.

1334.01 1214.66 1479.97

Cd

BY HOMOGENATES

nmols/hr/muscle

as described

z!= 294.97 f 255.29 f 212.33

190.79 173.67 184.65

ACTIVITY

procedures

MAO

procedures

1096.45 949.19 1331.43

Exp

OF P-H~DKOXYBUTYKATE-3-W

z

%

E m z 2 E 2

x 4

procedures

in the text,

74.54

f 15.64

f

68.77 67.91

13.72 25.83

f

Exp

nmols/hr/muscle

OF HOMOGENATES

as described

71.98

6

41

Q Experimental

72.16 70.05

6 6

No. of rats

ACTIVITY

8 19

Days after tenotomy

MAO

are means

12.98

9.95 15.26

Data

f

f

f

Ctl

7

i

NS

NS NS

P

SD. (Exp,

; Ctl,

f 58.91 f 76.44 f 36.32

Exp 627.97 507.18 419.96

nmols/hr/gfw

f 41.00 f 64.43 f 37.69

Ct1

HYPERTROPHY”

control.)

COMPENSATORY

experimental

360.61

522.26 431.14

oxidase

FOLLOWING

M onoamine

MUSCLES

TABLE OF RAT SOLEUS


P

F4

E z

2 z

E

;

K

F T1 “0

MUSCLE

HYI’ERTROPHY

229

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AND

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28. TURTO, H., S. LINDY, and J. HALNE. ity in work-induced hypertrophy

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