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ANIMAL GROWTH, PHYSIOLOGY, AND REPRODUCTION |
* Department of Animal Sciences and Industry, Kansas State University, Manhattan 66506;
and
Intervet Inc., Millsboro, DE 19966; and
Cactus Research Ltd., Amarillo, TX 79116
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Abstract |
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 Top Abstract INTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONLITERATURE CITED |
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Key Words: bovine • heifer • ractopamine • skeletal muscle • steroidal implant
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INTRODUCTION |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONLITERATURE CITED |
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). Ractopamine administration has resulted in more variable results in heifers than in steers (Schroeder et al., 2003b). Ractopamine binds to ß-adrenergic receptors (AR), which are membrane-bound receptors with 7 transmembrane domains (Mills, 2002a). Once the agonist binds, important enzymes undergo phosphorylation (Mills, 2002a). The receptor-mediated response can be altered by the density of receptors present in tissues (Spurlock et al., 1994; Mills, 2002b). The density of ßAR in adipose tissue has been shown to decrease with RAC administration; however, skeletal muscle ß-AR density remains unchanged in pigs (Spurlock et al., 1994). The potential change in receptor number is a factor that can alter the response to ß-AR agonists. Three subtypes of ß-AR are found in cattle, with ß2-AR being the most abundant in skeletal muscle. Ractopamine is believed to elicit its growth-promoting response through the ß1-AR (Smith et al., 1990; Moody et al., 2000).
Anabolic steroid compounds are used in the feedlot industry to increase growth rate and improve feed efficiency. Steroidal implants improve ADG, feed efficiency, and total lean tissue deposition (Johnson et al., 1996; Pampusch et al., 2003). One of the mechanisms by which increased muscle growth is achieved with implants is through increased production of muscle IGF-I, a stimulator of skeletal muscle growth (Johnson et al., 1998; Dunn et al., 2003; Pampusch et al., 2003). The purpose of these experiments was to investigate the effects of RAC administration in combination with implant strategy or variable days on feed (DOF) on feedlot heifer performance and the expression of mRNA for IGF-I and the ß-AR subtypes.
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MATERIALS AND METHODS |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONLITERATURE CITED |
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Exp. 1
Animals. English x Continental heifer calves (n = 1,147), with an initial BW of 282 ± 3 kg, were used in a 2 x 2 factorial arrangement of treatments, with main effects of implant strategy (Revalor-200 vs. Revalor-IH + Finaplix-H, all from Intervet Inc.) and RAC (0 vs. 200 mg/d, Elanco Animal Health, Greenfield, IN). Treatments were randomly assigned to each pen (n = 16), with approximately 75 heifers per pen, as follows: 1) initial Revalor-200 [200 mg of trenbolone acetate (TBA) and 20 mg of estradiol-17ß, R200] without or 2) with RAC (200 mg/d, R200-RAC), 3) initial Revalor-IH (80 mg of TBA and 8 mg of estradiol-17ß) and reimplanted (d 58) with Finaplix-H (200 mg of TBA, RF) without or 4) with RAC (200 mg/d, RF-RAC). Heifers were fed 3 times daily and allowed ad libitum access to feed.
The finishing diet was a steam-flaked corn-based diet (Table 1). Ractopamine was administered in the last 28 d of the experiment, and all heifers received melengestrol acetate [(MGA), 0.4 mg/d, Pfizer Inc., Kalamazoo, MI]. Ractopamine was hand-weighed, mixed with the basal diet, and given in 3 meals to provide 200 mg/heifer daily, and MGA was included in the top ration. For determination of final BW, pens were weighed and the industry standard 4% pencil shrink was applied. Heifers were slaughtered 182 ± 5 d after the initial implantation. Heifers were transported 200 km to a commercial slaughter facility (National Beef, Liberal, KS). Carcass characteristics were obtained from chilled carcasses 24 h after slaughter.
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Real-Time Quantitative PCR.
Real-time quantitative PCR was used to measure IGF-I, ß1-AR, ß2-AR, and ß3-AR gene expression relative to the quantity of 18S rRNA in total RNA isolated from muscle tissue. Measurement of the relative quantity of cDNA was performed by using TaqMan Universal PCR Master Mix (Applied Biosystems, Foster City, CA), 900 nM of the appropriate forward and reverse primers, 200 nM of appropriate TaqMan detection probe, and 1 µL of the cDNA mixture. The bovine-specific IGF-I, ß1-AR, ß2-AR, and ß3-AR forward and reverse primers and TaqMan detection probes (Table 2) were synthesized by using published GenBank sequences. Commercially available eukaryotic 18S rRNA primers and probes were used as an endogenous control (Applied Biosystems; Genbank Accession No. X03205). The ABI Prism 7000 detection system (Applied Biosystems) was used to perform the assay by using the recommended thermal cycling variables from the manufacturer (50 cycles of 15 s at 95°C and 1 min at 60°C). The 18S rRNA endogenous control was used to normalize the expression of IGF-I, ß1-AR, ß2-AR, and ß3-AR.
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Exp. 2
Animals.
English x Continental yearling heifers (n = 2,077), with an initial BW of 285 ± 1 kg, were used in a 2 x 3 factorial arrangement of treatments to evaluate the effects of RAC administration and DOF on feedlot heifer performance and carcass characteristics. Treatments consisted of 3 slaughter dates of 129, 150, and 170 d. Within each slaughter date, heifers received RAC (0 or 200 mg/d, Elanco Animal Health) for the last 28 d of feeding. All heifers received an initial Revalor-IH (80 mg of TBA and 8 mg of estradiol-17ß, Intervet Inc.) implant and were reimplanted on d 56 with Revalor-200 (200 mg of TBA and 20 mg of estradiol-17ß, Intervet Inc.), and all heifers received MGA (0.4 mg/d, Pfizer, Inc.). Heifers were fed 3 times daily and allowed ad libitum access to feed. The finishing diet was a steam-flaked corn-based diet (Table 3). For determination of final BW, pens were weighed and the industry standard 4% pencil shrink was applied. Heifers were transported 113 km to a commercial slaughter facility (Tyson Fresh Meats Inc., Amarillo, TX). Carcass characteristics were obtained from chilled carcasses 24 h after slaughter.
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RESULTS AND DISCUSSION |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONLITERATURE CITED |
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Performance data for Exp. 1 are presented in Table 4. The RF-implanted group had an increased (P = 0.05) percentage of USDA Choice carcasses as well as a tendency (P = 0.06) for a greater marbling score compared with the R200 group; however, there were no other differences between the implant groups. Ractopamine administration increased (P < 0.05) ADG and G:F in heifers (Table 4). Ractopamine also increased (P < 0.05) HCW and the LM area, decreased (P < 0.05) 12th rib fat depth, and improved (P < 0.05) yield grades in heifers (Table 4). Similarly, Schroeder et al. (2003b) reported increased ADG, feed efficiency, and HCW in heifers that received 200 mg/d of RAC. Our current results are also similar to a study in which feedlot heifers implanted with Revalor-H (140 mg of TBA and 14 mg of estradiol-17ß) 60 d before the initiation of RAC administration had improved ADG, carcass-adjusted ADG, and carcass-adjusted G:F (Walker et al., 2006).
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The gene expression results for the ß-AR are shown in Figures 1 to 3. There was no effect (P > 0.10) of treatment on the expression of ß1-AR mRNA (Figure 1); however, there was a tendency for RAC feeding to increase (P = 0.10) ß2-AR mRNA (Figure 2). For ß3-AR mRNA, there was an implant by RAC interaction (P = 0.05), with RAC leading to a numerical increase (P = 0.28) in ß3-AR mRNA in heifers implanted with RF, but a decrease (P < 0.05) in heifers implanted with R200 (Figure 3). Oral administration of ß-AR agonists has resulted in altered mRNA expression of ß-AR similar to that in the current study (Winterholler et al., 2007). Our results indicate that RAC may increase the expression of ß2-AR, which has been observed in other studies (Winterholler et al., 2007). Ractopamine administration had a tendency to increase the expression of ß2-AR mRNA in semimembranosus muscle tissue of yearling steers at the time of slaughter (Winterholler et al., 2007). The altered expression of skeletal muscle ß3-AR mRNA resulting from the implant strategy is intriguing; however, its relevance in the current study has not been established. Despite the lack of effect of implant on ß1-AR mRNA, the R200 group had reduced variability compared with the RF group. The R200 group was not reimplanted, whereas the RF group was reimplanted on d 58. Reimplantation may have resulted in greater variability in ß1-AR mRNA. Additionally, the expression of ß2-AR mRNA was 10-fold greater than that of both ß1- and ß3-AR, indicating that ß2-AR is the most abundant in bovine skeletal muscle. The tendency for up-regulation of ß2-AR mRNA in the current study is different from the report of Spurlock et al. (1994). The number of ß2-AR in adipose tissue of pigs was decreased at 1, 8, and 24 d following RAC administration, whereas the number in the skeletal muscle was unchanged. However, it has been suggested that the response to ß-AR agonists in muscle tissue may be different from the response observed in adipose tissue (Mills, 2002b). The species differences in receptor populations should also be considered when comparing previously published results (Mills, 2002b). There is also evidence suggesting that RAC administration increases the proportion of white muscle fibers in pigs (Aalhus et al., 1992). Altered muscle fiber distribution may affect the expression of ß-AR mRNA; however, muscle fiber distribution was not analyzed in the current study. These results indicate that RAC may alter the expression of ß2-AR mRNA; however, further research is needed to determine the specificity of RAC for specific receptor subtypes in bovine skeletal muscle.
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There was an implant by RAC interaction (P = 0.03) on IGF-I mRNA, with RAC leading to a decrease (P < 0.05) in IGF-I mRNA for heifers implanted with RF but a numerical increase (P = 0.15) in IGF-I mRNA in heifers implanted with R200 (Figure 4). This response is intriguing because of the lack of interaction on performance and carcass characteristics in the study. The changes in IGF-I mRNA may be a response that occurred late in the feeding period, closer to slaughter when samples were taken. This may not have allowed for an interaction to occur on performance. However, the interaction does raise questions regarding the potential implant and RAC interactions that can occur. The 2 growth promotants have different modes of action; however, the potential exists for interactions to occur between their respective pathways. Anabolic steroids enhance muscle growth, increase the rate of gain, and improve feed efficiency (Johnson et al., 1996; Pampusch et al., 2003). Steers implanted with a combined TBA-estradiol-17ß implant expressed greater muscle IGF-I mRNA production (Dunn et al., 2003; Pampusch et al., 2003). Biopsy samples of LM contained IGF-I mRNA levels that were 2.4-fold greater in implanted steers 28 d after implantation (Dunn et al., 2003). In contrast, ß-AR agonists do not appear to affect plasma concentrations of IGF-I or muscle IGF-I mRNA production (Dawson et al., 1993; Grant et al., 1993). Those studies are in contrast to the altered expression of IGF-I mRNA shown in the current study. However, Walker et al. (2007) reported that administration of RAC to Holstein steers resulted in reduced IGF-I mRNA, which is in agreement with our results for the RF-implanted group.
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In Exp. 2, RAC administration improved (P < 0.05) feed efficiency but had no other effects on performance or carcass characteristics (Table 5). There was no (P > 0.10) interaction between RAC administration and DOF. There was a decrease (P < 0.05) in ADG and G:F as DOF increased (Table 5). There was an increase (P < 0.05) in HCW, LM area, and marbling score as DOF increased (Table 5). Similarly, there were increases (P < 0.05) in 12th rib fat and calculated yield grade and a decrease in the percentage of KPH fat as DOF increased. The lack of response to RAC administration in this study is in contrast to that of Winterholler et al. (2007), who reported significant effects of RAC administration on performance and carcass characteristics in yearling steers regardless of DOF. Because the response to RAC administration appears to be more pronounced in steers than in heifers (Schroeder et al., 2003a,b), it is possible that these differences may be associated with endogenous hormone status between the 2 genders.
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, which showed no effect of RAC (200 mg/d) on 12th rib fat, LM area, USDA yield grade, or USDA quality grade in heifers. The implant strategy used in that study was not as aggressive as in Exp. 2 of the current study, with all heifers receiving an initial Revalor-IH and being reimplanted on d 56 with Revalor-200, indicating that aggressive implant strategies used in combination with RAC administration may interfere with the expected response to the ßAR agonist. There are differences in implant release patterns and payout times, and these can result in variability of animal performance (Preston, 1999). At the initiation of RAC feeding in Exp. 1, the R200 group had been implanted for 154 d and the RF group had been implanted with Finaplix-H for 96 d; under these conditions, the payout steroids from the implants at the time of RAC initiation was likely low. Thus, the administration of RAC resulted in a stimulation of the growth rate and feed efficiency. In contrast, the heifers in Exp. 2 were on a more aggressive implant strategy, with an initial Revalor-IH followed by Revalor-200 on d 56. This led to heifers being implanted with Revalor-200 for 65, 66, and 86 d, respectively, at RAC initiation. These heifers did not respond to RAC administration as did the heifers in Exp. 1, suggesting that the anabolic status of the heifer caused by steroid payout from the implant may be an important determinant in predicting the response to RAC in feedlot heifers. General Conclusions
The variability in performance responses to RAC in the present studies suggests a possible interaction between steroid hormone implants and RAC administration. These inferences come from 2 different studies in heifers with disparate implant strategies. More research comparing different implant strategies in heifers fed RAC is needed to aid in our understanding of the potential interaction of these 2 growth-promoting strategies in heifers.
Ractopamine and steroidal implants are believed to have different mechanisms of action; however, there are possibilities of some interactions between the 2. Ractopamine is a ß-AR agonist that binds to the ß-AR and signals through second messenger systems such as cyclic adenosine monophosphate, which leads to enzyme phosphorylation (Mersmann, 1998; Mills, 2002b). There are different proposed mechanisms of action for implants, with more recent attention on local effects on skeletal muscle such as the up-regulation of genes such as IGF-I, as well as increased circulating IGF-I (Dunn et al., 2003; Pampusch et al., 2003). Hormones in steroid-containing implants bind to their receptors located in the nucleus of cells and act as transcription factors for genes (Falkenstein et al., 2000). The 2 growth promotants appear to have different mechanisms of action; however, recent evidence suggests that steroid hormones can work through second messenger systems such as cyclic adenosine monophosphate and intracellular calcium, similar to that of RAC (Falkenstein et al., 2000). These are often referred to as nongenomic actions of steroid hormones. These potential nongenomic actions have been observed in numerous tissues, including bovine skeletal muscle satellite cells (Sissom et al., 2006). The nongenomic actions of steroid hormones could be a point of interaction between RAC and steroid hormones because of potentially similar modes of action.
Steroid hormones have also been shown to affect the sensitivity and number of ß-AR in numerous tissues (Engstrom et al., 2001; Malo and Puerta, 2001). In the rat myometrium, estradiol benzoate treatment resulted in desensitization of the ß2-AR function (Engstrom et al., 2001). Estrogen administration to ventricular myocytes was accompanied by reduced protein expression of the ß1-AR (Kam et al., 2004). In a similar manner, estradiol and progesterone reduced the density of ß3-AR in brown adipocytes (Malo and Puerta, 2001). These data demonstrate that steroid hormones can differentially affect ß-AR, which needs to be investigated further in livestock species that have received steroid hormone implants.
The data obtained from our studies aid in our understanding of the mechanism of action of ß-agonists used as growth promotants in the feedlot industry. It can also help in our understanding of the potential for interactions among the use of steroidal implants, management strategies, and ß-agonists. As suggested by our data, the implant strategy as well as the sex of the animal should be considered when trying to determine proper management strategies. This knowledge will aid in our ability to improve the efficiency of lean tissue deposition in beef cattle.
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Footnotes |
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2 Corresponding author: bjohnson{at}ksu.edu
Received for publication September 28, 2006. Accepted for publication May 10, 2007.
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LITERATURE CITED |
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TopAbstractINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONLITERATURE CITED |
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-actin mRNA and insulin-like growth factor-I mRNA in pigs during feeding and withdrawal of ractopamine. J. Anim. Sci. 71:3319–3326.[Abstract]This article has been cited by other articles:
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