AMPK and substrate availability regulate creatine transport in cultured cardiomyocytes

Abstract

Darrabie MD, Arciniegas AJ, Mishra R, Bowles DE, Jacobs DO, Santacruz L. AMPK and substrate availability regulate creatine transport in cultured cardiomyocytes. Am J Physiol Endocrinol Metab 300: E870 -E876, 2011. First published March 1, 2011; doi:10.1152/ajpendo.00554.2010.-Profound alterations in myocellular creatine and phosphocreatine levels are observed during human heart failure. To maintain its intracellular creatine stores, cardiomyocytes depend upon a cell membrane creatine transporter whose regulation is not clearly understood. Creatine transport capacity in the intact heart is modulated by substrate availability, and it is reduced in the failing myocardium, likely adding to the energy imbalance that characterizes heart failure. AMPK, a key regulator of cellular energy homeostasis, acts by switching off energy-consuming pathways in favor of processes that generate energy. Our objective was to determine the effects of substrate availability and AMPK activation on creatine transport in cardiomyocytes. We studied creatine transport in rat neonatal cardiomyocytes and HL-1 cardiac cells expressing the human creatine transporter cultured in the presence of varying creatine concentrations and the AMPK activator 5-aminoimidazole-4-carboxamide-1-␤-D-ribonucleoside (AICAR). Transport was enhanced in cardiomyocytes following incubation in creatine-depleted medium or AICAR. The changes in transport were due to alterations in Vmax that correlated with changes in total and cell surface creatine transporter protein content. Our results suggest a positive role for AMPK in creatine transport modulation for cardiomyocytes in culture. myocardial creatine content; cardiac energy metabolism; adenosine 5=-monophosphate-activated protein kinase; cardiac failure CREATINE (Cr) and its phosphorylated form phosphocreatine (PCr), together with creatine kinases (CK), comprise a system that helps maintain ATP stores in cardiac and skeletal myocytes (23). During cardiac hypertrophy and heart failure, the myocardium exhibits marked changes in energy metabolism, including changes in substrate utilization, as well as in the content of high-energy metabolites. The PCr/ATP ratio decreases (9,19

Hypoxia decreases creatine uptake in cardiomyocytes, while creatine supplementation enhances HIF activation

“…1999; Darrabie et al. 2011). Characterization of kinetics of Cr transport in transduced RNCMs subjected to hypoxia for 12 h revealed a significant decrease in V max (from 90.31 ± 7.07 to 30.76 ± 5.24 nmol/mg of protein) and K m (from 30.76 ± 5.24–7.20 ± 1.11  μ mol/L) compared with controls ( n  = 3, t ‐test P  < 0.05).…”

“…2011). Therefore, we tested the effect of pharmacological AMPK activation by 0.5 mmol/L AICAR beginning 24 h before exposure to hypoxia.…”

“…Recently, we (Darrabie et al. 2011) and others (Li et al. 2010) have demonstrated that activation of AMP‐activated kinase (AMPK), the cell's energy master regulator (Hardie et al.…”

“…1988; Darrabie et al. 2011), whereas decreases in Cr content increase Cr uptake (Loike et al. 1988; Darrabie et al.…”

“…1988; Darrabie et al. 2011). These changes in Cr transport capacity are secondary to an increase or decrease in V max , with no alteration in K m .…”