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
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
