Proceedings of The Physiological Society
University of Oxford (2011) Proc Physiol Soc 23, PC332
Dihydrotestosterone (DHT) stimulates amino acid uptake in mouse fast-twitch fibre bundles by increasing the expression and activity of the L-type amino acid transporter (LAT) 2
G. Mutungi1, M. M. Hamdi1
1. School of Medicine, University of East Anglia, Norwich, United Kingdom.
In a recent communication to the society we showed that dihydrotestosterone (DHT) stimulates amino acid uptake in isolated intact mouse fast-twitch skeletal muscle bundles without significantly affecting uptake in slow-twitch fibre bundles (Mutungi & Hamdi 2010). This effect, like that which DHT had on force (Hamdi and Mutungi, 2010), was mediated through the epidermal growth factor receptor (EGFR) and involved activation of the mitogen-activated protein kinase (MAPK) pathway. However, little is known about the amino acid transporter(s) stimulated by DHT. Therefore, the primary aim of this study was to investigate the amino acid transporter(s) mediating DHT-induced increase in amino acid uptake in mouse skeletal muscle fibre bundles. All the experiments were performed at room temperature using small fibre bundles isolated from a fast-twitch (edl) and slow-twitch (soleus) muscle of adult female CD1 mice. The mice were killed as recommended by the Animals (Scientific Procedures) Act 1986, UK and all the experiments conformed to the local animal welfare committee guidelines. The fibre bundles were treated with Ringer’s solution containing either 107.9µM ethanol (controls) or 630ρgml-1 DHT (treated) plus 2mM carbon-14 labelled isoleucine (L-(U-14 C) Ile; 3.46µCi ml -1) for 1hr. In some experiments, pharmacological interventions were used to investigate the transporter(s) mediating the effects of DHT on L-(U-14 C) Ile uptake. At the end of the experiments, the fibre bundles were processed for either liquid scintillation counting or Western blotting. Treatment of the muscle fibres with DHT increased the uptake of both isoleucine (Ile) and α-methylaminoisobutyric acid (MeAIB) in the fast-twitch fibre bundles only. Also, it increased the expression of the L-type amino acid transporter LAT2 and led to an increase the phosphorylation of the epidermal growth factor receptor (EGFR) in the same fibre bundles and that of RSK1/2 in both fibres types. However, it had no effect on the phosphorylation of the mitogen- and stress-activated protein kinases (MSK) 1/2. Pre-treating the fibre bundles with MeAIB (a system A amino acid transporter inhibitor) and 2-aminobicyclo-(2,2,1)heptane-2-carboxylic acid (BCH; a system L amino acid transporter inhibitor) significantly (p<0.05) reduced the basal uptake of Ile in both fibre types and completely abolished the DHT-induced increase in Ile uptake. From these findings we suggest that DHT stimulates amino acid uptake in isolated mammalian skeletal muscle fibres by modulating the activity of LAT2 and the sodium-coupled neutral amino acid transporter (SNAT) 2. These effects are mediated through the EGFR and involve an increase in LAT2 expression.
Where applicable, experiments conform with Society ethical requirements