AGAT knockout mice provide an opportunity to titrate tissue creatine content

Abstract

Creatine (Cr) plays an important role in muscle energy homeostasis via the Cr phosphokinase reaction (ATP + Cr ↔ ADP + CrP). Previous experimental models designed to investigate the importance of Cr and CrP content in skeletal muscle have used competitive inhibitors of Cr transport such as β-guanidinoproprionic acid (β-GPA) (Shoubridge et al. 1985) or have utilized whole body gene knockout of guanidine acetate methyltransferase (GAMT) (Schmidt et al. 2004), the second of two enzymes required for endogenous Cr biosynthesis. In both these models skeletal muscle Cr and CrP content is essentially absent, but both models are imperfect examples of blocking energy supply from the Cr phosphokinase reaction. For instance, β-GPA can be phosphorylated and utilised for energy partly compensating for a lack of PCr. Similarly, in the GAMT knockout model, guanidinoacetate (GAA) accumulates and this molecule can also be phosphorylated and utilised for energy by skeletal muscle. Recently, Skelton et al. (2011) generated a whole body Cr transporter knockout mouse model which resulted in depletion of skeletal muscle and brain Cr content. The utility of this model for precise regulation of skeletal muscle Cr content is currently questionable given that muscle may be able to synthesise its own Cr in situations where Cr content is compromised (McClure et al. 2007). In an article in this issue of The Journal of Physiology, the problem of the unwanted side reactions from β-GPA and GAMT knockout models was ingeniously resolved by Nabuurs et al. (2013) by generating a whole body arginine-glycine amidinotransferase (AGAT) gene knock out mouse model. AGAT catalyses the first reaction involved in Cr synthesis and when eliminated no endogenous Cr biosynthesis is possible. The only available source of Cr for these knockout animals is from the diet. Furthermore, no accumulation of GAA and its subsequent effects occur because GAA is produced by AGAT. With this model the researchers were able to manipulate the Cr and CrP content of skeletal muscle and the brain by simply altering the period in which the mice were fed a Cr free diet. The findings reported by Nabuurs et al. (2013) clearly demonstrate that skeletal muscle devoid of Cr and CrP is severely affected. In summary, they found that Cr deficient muscle was characterised by atrophy and decreased force production, and exhibited major metabolic disturbances including decreased resting ATP content, elevated inorganic phosphate levels, increased intramyocellular lipid droplets and mitochondrial dysfunction. Importantly, all these muscle disturbances could be reversed reasonably rapidly when the AGAT knockout mice were fed a diet supplemented with Cr. Another interesting part of the work reported by Nabuurs et al. (2013) was the research into the rates of Cr accumulation and loss from skeletal muscle and brain in the AGAT knockout mice. Their results clearly showed that Cr accumulation into Cr deficient skeletal muscle was surprisingly rapid occurring within 24 h, whilst brain Cr stores took up to 20 days to fully recover. On the flip side, the rate of Cr loss from skeletal muscle and brain in AGAT knockout mice pre-loaded with Cr and then starved of dietary Cr appeared to be similar taking several months to be completely lost from these tissues. The work by Nabuurs et al. (2013) has important implications for future research into the effects of Cr content on the function of various organs. For the first time there is an animal model in which tissue Cr content can be easily manipulated from depleted to fully loaded without the interference of side reactions. For example, the AGAT knockout mouse model could be used to titrate Cr and CrP content in brain to establish dose–function relationships for this organ. In addition, the AGAT knockout model could be employed to investigate the effect of manipulating Cr content on other Cr containing tissues such as the heart and testes. In summary, the AGAT knockout mouse model will provide an important tool to advance our current knowledge of the effects of Cr content on the in vivo regulation and function of Cr containing tissues.