Impact of Ergot Alkaloid and Estradiol 17B on Whole-Body Protein Turnover and Expression of mTOR Pathway Proteins in Muscle of Cattle

OF THESIS IMPACT OF ERGOT ALKALOID AND ESTRADIOL 17B ON WHOLE-BODY PROTEIN TURNOVER AND EXPRESSION OF MTOR PATHWAY PROTEINS IN MUSCLE OF CATTLE Beef cattle consuming endophyte infected tall fescue typically exhibit reduced performance in terms of both decreased dry matter intake (DMI) and growth rates. It has been suggested that lower concentrations of circulating IGF-1 (an important stimulator of the mTOR pathway and ultimately protein synthesis), as observed with consumption of ergot alkaloids, contribute to reduced growth rates. The objective of the current study was to determine if fescue-derived alkaloids decrease muscle protein synthesis though inhibitory action on the mTOR pathway via a direct effect on signal proteins, and if these negative effects can be alleviated by implantation with anabolic agents. Thirty-two Holstein steers were used in a 2x2 factorial design, and treatments consisted of intramuscular administration of bromocriptine (vehicle or 0.1 mg/kg BW) and a subdermal estradiol implant (with or without). Throughout the 35-day experiment, steers were fed a corn silage-based diet, with intake restricted to 1.5 times maintenance energy requirement. Bromocriptine injections were given every three days for 34 days. On days 27 through 32, steers were moved to metabolism stalls for urine collection and whole-body protein turnover was determined using a single pulse dose of [15N] glycine into the jugular vein on day 28. On day 35, muscle samples were collected from the musculus obliquus externus abdominis before (basal state) and 60 mins after (stimulated state) an i.v. glucose challenge (0.25 g glucose/kg). Blood samples were collected at regular intervals before and after glucose infusion for determination of circulating concentrations of glucose and insulin. Bromocriptine reduced insulin and glucose clearance following the glucose challenge, indicating decreased insulin sensitivity and possible disruption of glucose uptake and metabolism in the skeletal muscle. This suggests that fescue-derived alkaloids are detrimental to growing cattle in terms of overall glucose homeostasis and energy metabolism. Conversely, analysis of whole-body protein turnover demonstrated that bromocriptine does not appear to affect protein synthesis or N retention and western immunoblot analysis of skeletal muscle showed that it did not affect abundance of S6K1 or 4E-BP1, so does not appear to inhibit activation of the mTOR pathway or protein synthesis. Implantation improved N retention, decreased protein turnover, and had no effect on protein synthesis, suggesting that steroidal implants promote protein accretion through unchanged rates of synthesis and decreased degradation, even in the presence of bromocriptine, resulting in improved daily gains. Implanted steers likely experienced increased IGF-1 signaling, but downstream activation of mTOR, S6K and 4E-BP1, and thus increased protein synthesis did not occur as expected. Overall, this data suggests that fescue derived alkaloids do not have a negative impact on muscle protein synthetic pathways, independent of DMI.