P J Reeds

Stimulation of muscle growth by clenbuterol: lack of effect on muscle protein biosynthesis

1. Young rats were offered to appetite a semi-synthetic diet either alone or containing the beta 2-selective agonist clenbuterol (4-amino-alpha[t-butylamino)methyl]-3,5-dichlorobenzyl alcohol). 2. In female rats (starting weight 116g) the presence of the drug at daily doses greater than 10 micrograms/kg body-weight per d increased the growth of skeletal and cardiac muscle but had no stimulatory effect on the growth of the liver, gastrointestinal tract and kidney. 3. Male rats (starting weight 53 g) received clenbuterol at a daily oral dose of 200 micrograms/kg body-weight per d. Animals were slaughtered after 0, 4, 8, 11, 18, 21 and 25 d of treatment. At 4, 11, 21 and 25 d muscle protein synthesis was measured by the method of Garlick et al. (1980). Although clenbuterol increased the rate of protein and RNA accretion in gastrocnemius and soleus muscles, protein synthesis was not increased. 4. The results suggested that the drug had a rapid, perhaps direct, inhibitory effect on protein degradation. It is concluded that the growth-promoting effect of clenbuterol may be specific to muscle and that the drug may act in a novel manner which circumvents the physiological mechanisms responsible for the control of muscle growth.

Whole body and tissue protein synthesis in cattle

1. The daily rates of synthesis of protein by the whole body and by the individual tissues were determined in two Hereford x Friesian heifers (236 kg and 263 kg live weight), and a dry Friesian cow (628 kg live weight). 2. The rate of whole-body protein synthesis (g protein/d) was estimated from the total flux through the blood of [3H]leucine and [3H]tyrosine following infusion at a constant rate for 8 h. 3. The fractional rates of protein synthesis (ks) in the tissues (g synthesized/d per g tissue protein) were obtained after slaughter of the animals at the end of the infusion period. The fractional rate of protein synthesis was calculated assuming that the specific radioactivity of free tyrosine in either the blood (to give ks, b) or the tissue homogenate (to give ks, h) defined closely the specific radioactivity of the amino acid precursor for protein synthesis. total protein synthesis (As, b or As, h; g/d) in an individual tissue was calculated as the product of ks, b (or ks, h) x protein content. 4. Based on the total leucine flux, i.e. without correction for oxidation, 1.6 kg protein were synthesized daily in the heifers; for the cow this value was 2.0 kg/d. 5. The sum of the daily total synthesis in the major tissues (muscle+bone+brain, gastrointestinal tract (GIT), liver, hide) gave values of 1.4-1.9 kg/d based on As, b, and 2.2-3.0 kg/d based on As, h. 6. percentage contributions of the individual tissues to the total protein synthesis were similar in all three animals, for example based on As, h muscle was 12-16; carcass (muscle+bone+brain) 32-33; GIT 38-46; liver 7-8; skin 14.21%. 7. The contribution of muscle to total synthesis estimated from the leucine flux was 19-22%; this value is in agreement with those calculated on the same basis for other species. 8. The energy cost of protein synthesis was estimated to account for a maximum of 30% of heat production.

Protein turnover in growing pigs. Effects of age and food intake

1. Measurements were made of the nitrogen and energy balances of pigs of 30, 60 and 90 kg given a conventional diet at various daily rates. 2. Body protein synthesis was estimated from the irreversible loss of leucine from the blood following the infusion of [1-14C]leucine, and from the oxidation of the labelled amino acid. 3. Protein synthesis (g/d) increased by 2.17 for each 1 g increase in daily protein accretion and by 1.55 for each 1 g increase in daily protein intake. 4. At 30 kg, pigs close to energy equilibrium synthesized 270 g protein daily compared with 406 g and 512 g when their ration supplied twice and three times their maintenance requirement. 5. There was a close correlation between the daily urinary excretion of urea + ammonia and total amino acid catabolism estimated from the catabolism of leucine, but the latter underestimated the observed excretion by 2.5 g N/d. 6. The results imply that protein turnover accounts for only a proportion of the heat production associated with protein deposition.

The effect of a growth promoting drug, clenbuterol, on fibre frequency and area in hind limb muscles from young male rats

The effect of dietary administration of clenbuterol on soleus and extensor digitorum longus muscles was studied after 4 and 21 days. Both muscles showed an increase in wet weight with no significant change in total fibre number. After 4 days fibre cross-sectional areas were increased in soleus, but not in extensor digitorum longus, and after 21 days there was a change in fibre frequencies in extensor digitorum longus but not soleus muscles.

Effects of changes in the intakes of protein and non-protein energy on whole-body protein turnover in growing pigs.

1. The relationships between the intakes of protein and of non-protein energy (NPE), nitrogcn retention and body protein synthesis have been studied in female pigs weighing 30 and 35 kg. 2. Four animals were assigned to three regimens and given a conventional (basal) diet supplemented with fat, carbohydrate or protein. After 1 week, measurements of N excretion in urine and faeces (7 d collection) and gaseous exchange (3–4 d) were made. At the end of the balance period a solution of [l- 14 C]leucine was infused at a constant rate. Body protein synthesis was then calculated as the difference between the apparent irreversible loss of blood leucine and the loss of 14 C in expired air. The animals were then offered the basal diet without supplement for 10 d and the measurements of N retention, energy retention and protein synthesis were repeated. 3. The intakes of metabolizable energy (ME; MJ/kg body-weight (W) 0.75 per d) were 1.75 for fat, 1.58 for carbohydrate, 1–25for protein and 1.18 for the basal diet; corresponding intakes of apparently digestible N (ADN; g N/kgW 0.75 per d) were 2.30,2.31,4.35 and 2–17. Daily N retention, which during the period of basal feeding was 13.6 g was increased by between 3.4 and 7.2 g by the supplements. Daily fat deposition was also increased in the animals that received the diets supplemented with carbohydrate and fat. 4. The rate of leucine catabolism was significantly reduced in the animals receiving the diets that were supplemented with W Eand increased by the addition of protein to the diet. 5. When based on the specfic radioactivity of blood leucine both the synthesis and breakdown of body protein (per unit metabolic body-weight) were increased by 30% in the animals receiving the high-protein diet but the increases in protein synthesis associated with the addition of carbohydrate (+14%) and fat (+12%) were much less marked. Consideration of these results together with previous observations (Reeds et al. 1980) suggested that body protein synthesis(g N/d) increased by 0.88 for each g increase in daily ADN and by 0.93 for each MJ increase in daily ME intake. 6. Comparison of the results obtained with the animals given high-carbohydrate diets and those given high-protein diets suggested an increase in heat production of 14 KJ/g of additional fat deposition. A similar comparison of animals receiving the high-protein and basal diets suggested a heat increment of 23.5KJ/g additional protein deposition. The changes in heat production and protein synthesis in the animals given the protein supplement were compatible with a heat increment of 5.3 KJ/g additional protein synthesized. Because of the large proportion of heat production associated with the deposition of fat this could not be confirmed with either of the other supplements, but it is possible that the energy cost of protein accretion varies with the relative proportions of protein and NPE in the diet.

Effects of the amount and quality of dietary protein on nitrogen metabolism and protein turnover of pigs

1. The interrelation between protein accretion and whole-body protein turnover were studied by varying the quantity and quality of protein given to growing pigs. 2. Diets with 150 or 290 g lysine-deficient protein/kg were given in hourly meals, with or without lysine supplementation, to female pigs (mean weight 47 kg). 3. After the animals were adapted to the diets, a constant infusion of [14C]urea was given intra-arterially for 30 h, during the last 6 h of which an infusion of [4,5-3H]leucine was also infused at a constant rate. At the same time, yeast-protein labelled with 15N was given in the diet for 50 h. 4. The rate of urea synthesis was estimated from the specific radioactivity (SR) of plasma urea. The rate of leucine flux was estimated from the SR of plasma leucine. The irrevocable breakdown of leucine was estimated from the 3H-labelling of body water. Total N flux was estimated from the 15N-labelling of urinary urea. 5. Addition of lysine to the low-protein diet significantly increased N retention, with a substantial reduction in leucine breakdown, but there was no significant change in the flux of leucine or of total N. 6. Increasing the quantity of the unsupplemented protein also increased N retention significantly, with concomitant increases in leucine breakdown and in the fluxes of leucine and of total N. 7. It is concluded that a doubling of protein accretion brought about by the improvement of dietary protein quality is not necessarily associated with an increased rate of whole-body protein turnover.

Inhibition and Reversal of Denervation-Induced Atrophy by the Beta-Agonist Growth Promoter, Clenbuterol

Dietary administration of the growth promoter, clenbuterol, ameliorated denervation-induced atrophy in rat soleus muscles. In acutely denervated muscles the drug inhibited the appearance of atrophy, and in chronically denervated muscles the atrophy was almost fully reversed. Responses in slow twitch oxidative fibres were particularly marked.

Clenbuterol, a beta agonist, induces growth in innervated and denervated rat soleus muscle via apparently different mechanisms

Dietary administration of the anabolic agent, clenbuterol, has already been shown to inhibit or reverse denervation-induced atrophy in rat soleus muscles. We now show that the ameliorative effects of clenbuterol in denervated rat muscles are due principally to a large increase in protein synthesis. This results from both an increase in protein synthetic capacity and a normalised translational efficiency. The responses of innervated and denervated muscles are therefore fundamentally different, the changes in denervated muscles being reminiscent of the classical pleiotypic response of cells to growth factors.

The effect of β-agonists and antagonists on muscle growth and body composition of young rats (Rattus sp.)

1. The addition of the beta-selective adrenergic agonist clenbuterol to the diet was associated with an increase in the protein and RNA of skeletal and cardiac muscle, a reduction in fat deposition and an increase in energy expenditure. 2. Neither propranolol nor atenolol blocked the effect of clenbuterol on muscle protein but both reduced its effect on cardiac and fat mass and energy expenditure. 3. Five other beta-agonists were tested. All increased the interscapular brown fat mass and lowered body fat but only two increased skeletal muscle protein. 4. It is concluded that the anabolic and anti-lipogenic actions of certain beta-agonists are mechanistically distinct.

The Effect of the Anabolic Agent, Clenbuterol, on Overloaded Rat Skeletal Muscle

The dietary administration of clenbuterol to young male rats has been shown to produce a muscle specific hypertrophic growth response. This paper demonstrates that the combined effect of drug treatment and hypertrophic stimulus induced by tenotomy produced an additive effect on muscle growth. This effect was demonstrated in terms of both muscle composition (protein and RNA) and fibre size.