The Journal of Nutrition Vol. 128 No. 2 February 1998, pp. 360S-363S

Nutrition, Development and Efficacy of Growth Modifiers in Livestock Species¹

Alan W. Bell², Dale E. Bauman, Donald H. Beermann, and Robert J. Harrell

Department of Animal Science, Cornell University, Ithaca, NY 14853-4801

	ABSTRACT

Abstract Introduction References

Somatotropin (ST) and synthetic -adrenergic agonists (-AA) are growth-modifying agents that increase the rate and sometimes, the efficiency of protein deposition in lean tissues of livestock species. The ST-induced increase in muscle protein deposition is effected by a relatively modest increase in protein synthetic rate. This is possibly mediated by the endocrine influence of marked increases in circulating IGF (insulin-like growth factor)-I, and other ST-dependent components of the IGF system; mediation by locally expressed IGF-I may also occur. Increased muscle protein accretion in animals treated with -AA seems to be directly mediated by binding of the synthetic agonist to muscle -1 or -2 receptors, leading to increased muscle protein synthesis, possibly accompanied or followed by decreased protein degradation. This response is transient, due to down-regulation of -adrenergic receptors. Maximal responses of muscle protein accretion to both ST and -AA are attenuated by feeding inadequate levels of total protein or specific, limiting amino acids. For ST, but not -AA, this effect in growing pigs is partially offset by increased efficiency of utilization of absorbed amino acids for protein deposition, with predictable consequences for dietary protein and amino acid requirements. Both ST and -AA are less efficacious in promoting muscle protein deposition in very young animals. For ST, this is related to postnatal development of the somatotropic axis; a mechanistic explanation for the similar lack of effect of -AA is lacking. In both cases, this phenomenon must be considered against the very high inherent capacity and efficiency of lean tissue protein accretion in the neonate.

	INTRODUCTION

Abstract Introduction References

During the past 15 years, there has been unprecedented research interest in a diverse group of compounds that have been shown to affect the rate and/or composition of growth in farm livestock and other species, including humans in some cases. These compounds are generally described as metabolic (or growth) modifiers. They include the anabolic steroids, some of which are approved for commercial use in selected species in the United States, somatotropin (ST)³ and a group of synthetic phenethanolamine derivatives, also described as -adrenergic agonists (-AA), which chemically and pharmacologically resemble the natural catecholamines. For information on the chemical identity and mechanisms of action of these growth modifiers, and a detailed summary of their published effects on growth performance before 1994, the reader is referred to NRC (1994).

The present review will focus on two types of growth modifiers, the somatotropins (porcine, pST and bovine, bST) and selected -AA, which have been extensively studied in growing swine and cattle. Recent insights into mechanisms of action involving control of nutrient partitioning between lean and fat will be discussed, focusing on regulation of protein metabolism in support of the growth of skeletal muscle. The main themes of nutritional and developmental modulation of efficacy of actions on rate, composition and efficiency of growth will then be addressed.

	MECHANISMS OF ACTION

Somatotropin.  Treatment of swine with exogenous pST causes spectacular increases of up to 90% in rate of protein accretion in lean tissues, including muscle; somewhat smaller but still impressive responses are seen in ruminants (NRC 1994). We have used the arteriovenous difference/blood flow technique in combination with isotope dilution to study the simultaneous, in vivo effects of ST on protein synthesis and degradation in hindlimb (predominantly muscle) tissues (Boisclair et al. 1994). These studies confirmed that the chronic protein anabolic effect of ST is achieved exclusively through promotion of protein synthesis, with no discernible effect on protein degradation, and highlighted the relatively subtle stimulation of synthesis (~10%) required to effect a much greater increase (~40%) in net protein accretion (Boisclair et al. 1994).

There is considerable circumstantial evidence that the actions of ST on protein accretion in skeletal muscle and other lean tissues are mediated by the insulin-like growth factor (IGF) system (Florini et al. 1996). However, the specific mechanisms involved are unclear, in several important aspects. First, the relative influence of systemic vs. local sources of IGF and IGF-binding proteins (IGFBP) on muscle protein metabolism and growth has not been determined. Treatment with ST of swine and ruminants during later stages of growth causes pronounced increases in plasma levels of IGF-I and its major binding protein, IGFBP3 (Coleman and Etherton 1991, Hodgkinson et al. 1991, Walton and Etherton 1989). The primary tissue source of this systemic response is most likely the liver, in which ST specifically regulates transcription of IGF-I, IGFBP3 and the third component of the ternary binding complex, the acid-labile subunit (Rotwein et al. 1997). Other tissues, including adipose, may also contribute to the ST-induced increase in circulating IGF-I (Coleman et al. 1994, Wolverton et al. 1992). However, in some studies (Brameld et al. 1996, Duffy et al. 1992), but not in all (Coleman et al. 1994, Grant et al. 1991), expression of IGF-I in muscle itself has been shown to respond to ST, which raises the possibility that at least part of the protein anabolic effect of ST is mediated by local actions of the IGF system. The significance of observed variations in response among anatomical muscles, species and stage of development is presently unclear.

A second area of mechanistic uncertainty concerns the degree to which effects of IGF-I mimic those of ST on muscle protein turnover in vivo. For example, Oddy and Owens (1996) found that short-term (4 h), close-arterial infusion of recombinant human IGF-I into the hindlimb of growing lambs improved protein accretion in infused tissues by reducing protein catabolism without effect on protein synthesis. This response is consistent with previous observations of reduced whole-body proteolysis in lambs (Douglas et al. 1991) and humans (Laager et al. 1993) acutely administered IGF-I. However, it is notable that in humans treated with low doses of IGF-I for 5-7 d, a moderate increase in apparent protein synthesis was observed (Mauras and Beaufrere 1995). Part of the lack of a protein synthetic response to acute treatment may be due to the accompanying decrease in circulating levels of essential amino acids (Oddy and Owens 1996). Further studies are required to determine whether the response could be elicited by infusing supplementary amino acids with the IGF-I, and whether there is an adaptive return to normal aminoacidemia with longer-term IGF treatment. It is notable that plasma amino acid concentrations are little changed by chronic treatment with ST and its attendant increase in plasma IGF-I (Boisclair et al. 1994).

Finally, in relation to one of the themes of this symposium, it is not known to what degree the effects of ST on muscle protein metabolism are mediated by altered tissue responses to other endocrine factors, especially insulin, either directly or via modulation of the expression or actions of IGF system components. Future investigations should include application of the hyperinsulinemic amino acid clamp approach described elsewhere in these proceedings (Davis et al. 1998), analogous to our previous applications of the glucose clamp technique to demonstrate the attenuation by ST of insulin's effects on glucose production and utilization in growing pigs and steers (Dunshea et al. 1992b and 1995).

-adrenergic agonists.  Like ST, the best-studied synthetic -AA, including clenbuterol, cimaterol, ractopamine and L-644,969, have multiple actions on various aspects of nutrient metabolism that lead to increased lean and decreased fat deposition in meat animals and other species. However, in contrast to ST, these effects are generally more pronounced in ruminants than in swine and the positive responses in lean tissue protein accretion are largely confined to skeletal muscle (NRC 1994). It has also become clear that the effects on muscle protein metabolism are mediated directly through binding of the agonist to specific -1- or -2-adrenergic receptors in muscle, and that the initially marked responses become attenuated through down-regulation of these receptors.

We recently obtained convincing evidence for the direct action of -AA on muscle protein accretion in vivo by close-arterial infusion of cimaterol into a single hindlimb for 21 d in growing steers (Byrem et al. 1997). Net protein accretion in the treated limb was estimated to have increased by 65% compared with that in the contralateral, saline-infused limb, on the basis of measurements of net uptake of amino acids by each limb. This remarkable response was corroborated by direct confirmation of differences in weight and protein content of hindlimb muscles when animals were slaughtered at the end of the study. The experiment also confirmed the transient nature of the anabolic response, which peaked at 14 d, but was greatly attenuated by 21 d of treatment. It is generally accepted that this phenomenon is due to desensitization of the -adrenergic receptor (Hausdorff et al. 1990), consistent with the reduction in -1 receptor density in longissimus dorsi muscle of pigs treated with ractopamine for 3 wk (Sainz et al. 1993).

The literature on mode of action of the -AA on muscle protein turnover is somewhat conflicting. Some studies have shown clear increases in protein synthesis (Bergen et al. 1989, Claeys et al. 1989) and in the abundance of mRNA for muscle-specific proteins (Grant et al. 1993, Smith et al. 1989); others suggest that most if not all of the increase in net protein accretion is due to reduced protein degradation (Bohorov et al. 1987, Dawson et al. 1991), possibly mediated by reduced activity of calpains and other specific proteolytic systems (Kretchmar et al. 1989, Wang and Beermann 1988). A consensus view is that both arms of protein turnover are affected, but to degrees that vary in temporal pattern.

The effects of -AA on muscle protein metabolism are probably not mediated indirectly by hormonal effects beyond the direct, -receptor-mediated mode of action, as evidenced by the ability of hypophysectomized (Thiel et al. 1987) and severely diabetic rats (McElligot et al. 1987) to respond to treatment.

	PROTEIN NUTRITION AND EFFICACY OF GROWTH MODIFIERS

The ability of growth modifiers such as ST and the -AA to stimulate muscle protein deposition is affected by intake of protein or limiting amino acids, such as lysine. The enhanced protein deposition also has consequences for optimization of dietary requirements for total protein and specific essential amino acids. The best experimental evidence for the effect of growth modifiers on relations between protein/amino acid intake and protein deposition comes from a series of studies on responses to pST and the -AA, ractopamine, in growing swine (NRC 1994).

Campbell (1988) summarized the elegant series of experiments in which he and others defined the effects of energy and protein intake on body protein deposition in pigs at different stages of growth, including effects of sex, genotype and other factors. It is now well established that the pattern of response to dietary protein (or the first-limiting amino acid, lysine) is linear up to a plateau that, if energy is not limiting, is determined by the animal's inherent capacity for protein deposition. The slope of the relationship during the protein-dependent phase is an index of the efficiency of utilization of absorbed amino acids. As illustrated and discussed by Boyd et al. (1991) and NRC (1994), growth modifiers could, hypothetically, increase protein deposition simply by increasing the maximal response plateau without affecting the efficiency of amino acid use. In this scenario, the dietary protein or amino acid requirement would increase in direct proportion to the rate of protein accretion. Alternatively, if the efficiency of amino acid use was increased in addition to maximal protein deposition, the effect of the growth modifier on requirements would be less than in the first scenario. Evidence for both possibilities exists.

Effect of pST.  There is convincing evidence that treatment of growing pigs with pST significantly increases the efficiency of utilization of absorbed amino acids for protein deposition. The magnitude of this effect and its consequences for determination of protein/amino acid requirements seem to be influenced by growth stage, quality of dietary protein, and possibly, sex (NRC 1994). However, most studies have shown an increase in apparent efficiency of use of dietary protein of 25-50% (Boyd et al. 1991, Campbell et al. 1990 and 1991, Caperna et al. 1995, Krick et al. 1993), indicating that pST separately increases the maximal capacity for protein accretion and the efficiency with which amino acids are used for protein accretion. By increasing the slope of the relation between protein deposition and protein intake, increased efficiency of amino acid utilization offsets the effect of treatment on dietary requirements for total protein and lysine (NRC 1994).

The mechanism by which pST improves efficiency of utilization of absorbed amino acids in growing pigs has not been studied in detail but some clues exist. Within a few hours after the first of a course of daily intramuscular injections of pST, growing pigs exhibit a discernible decline in plasma urea nitrogen (PUN) concentration that becomes progressively greater over several days of treatment (Dunshea et al. 1992a). The most likely interpretation is that pST elicits a relatively rapid decline in amino acid catabolism, especially in the liver. This was recently confirmed by observations of marked decreases in hepatic oxidation of lysine, methionine and valine, and of the activity of lysine -ketoglutarate reductase in rats treated with bST for 5 d (Blemings et al. 1996).

Effect of ractopamine.  Treatment of growing pigs with the -AA, ractopamine, causes appreciable increases in body protein deposition, albeit somewhat less than those achieved with maximal doses of pST (NRC 1994). In the only protein titration study of its type, Dunshea et al. (1993a) found no evidence that any of this response was due to altered efficiency of amino acid utilization in female pigs growing from 60 to 90 kg. Thus, the dietary protein requirement was increased in proportion to the increase in protein deposition, and no response to ractopamine was evident when dietary crude protein concentration was 140 g/kg. The possibility of subtle effects of ractopamine on efficiency of amino acid use cannot be excluded because treatment also caused modest reductions in PUN in growing pigs (Dunshea et al. 1993b).

	PHYSIOLOGICAL DEVELOPMENT AND EFFICACY OF GROWTH MODIFIERS

Most studies on the effects of ST or -AA on lean tissue protein accretion and underlying mechanisms have been done in ruminants and swine approaching market weight, when the capacity for lean growth is waning and the propensity for fattening is markedly increasing. During earlier growth phases, when the inherent capacity for and efficiency of protein deposition are greater (Carr et al. 1977), there is increasing evidence that treatment with growth modifiers is less effective.

Somatotropic axis development and response to ST.  During fetal life in the precocial sheep, the somatotropic axis is not fully engaged in the regulation of lean tissue growth, as shown by the relatively modest effects of hypophysectomy on prenatal muscle and bone development (Broad et al. 1980, Mesiano et al. 1987). Very high concentrations of ST, together with low IGF-I in fetal plasma, are consistent with the limited ability of the fetal liver to bind and respond to ST (Gluckman et al. 1983). Thus, although the fetal liver expresses ST receptor mRNA through much of prenatal life (Klempt et al. 1993), significant abundance of functional receptors is not achieved until after birth in sheep (Breier et al. 1994), calves (Breier et al. 1988) or pigs (Breier et al. 1989). Subsequent patterns of postnatal increase in hepatic binding of ST are associated with progressive increases in plasma IGF-1 in young pigs (Breier et al. 1989).

This picture of developmental increases in hepatic sensitivity and/or responsiveness to ST is entirely consistent with our recent observations in young pigs treated for 4 d with pST at intervals from 10 to 125 d of age (Harrell et al., personal communication). Treatment-induced increments in plasma concentrations of both IGF-I and IGFBP3 were small in pigs weighing <20 kg (~40 d of age), but increased steadily with age thereafter. This pattern of response also correlates well with developmental increases in the ability of exogenous pST to promote body tissue protein deposition in young pigs. In a series of studies at Cornell University, maximal responses in protein deposition were 16, 25 and 74% in female or castrated male pigs of similar genotype weighing 10-20 (Harrell et al. 1997), 20-55 (Krick et al. 1993), and 55-100 kg liveweight (Boyd et al. 1991), respectively. By extrapolation, a minimal response would be expected in pigs younger than those studied at 10-20 kg, which are already growing extremely rapidly and with a very high efficiency of utilization of dietary protein (Carr et al. 1977).

Response to -AA.  As summarized by NRC (1994), effects of -AA on growth performance and carcass composition were much smaller in younger pigs and ruminants than in animals approaching market weight. For example, in young lambs weighing 7-15 kg and fed milk replacer and cimaterol for 3 wk, protein accretion in semitendinosus muscle was minimally affected (+3%) (Williams 1990), compared with the 35% increase observed in weaned lambs of the same genotype, weighing 36-42 kg and treated with cimaterol for 3 wk (O'Connor et al. 1991). It is not known whether the lack of responsiveness of young animals to -adrenergic agonists is due to initially low receptor abundance and/or affinity in skeletal muscle, or to more rapid development of refractoriness to these compounds.

	FOOTNOTES

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