³¹P-magnetization transfer magnetic resonance spectroscopy measurements of in vivo metabolism

Abstract

Magnetic resonance spectroscopy offers a broad range of noninvasive analytical methods for investigating metabolism in vivo. Of these, the magnetization-transfer (MT) techniques permit the estimation of the unidirectional fluxes associated with metabolic exchange reactions. Phosphorus (³¹P) MT measurements can be used to examine the bioenergetic reactions of the creatine-kinase system and the ATP synthesis/hydrolysis cycle. Observations from our group and others suggest that the inorganic phosphate (P(i)) → ATP flux in skeletal muscle may be modulated by certain conditions, including aging, insulin resistance, and diabetes, and may reflect inherent alterations in mitochondrial metabolism. However, such effects on the P(i) → ATP flux are not universally observed under conditions in which mitochondrial function, assessed by other techniques, is impaired, and recent articles have raised concerns about the absolute magnitude of the measured reaction rates. As the application of ³¹P-MT techniques becomes more widespread, this article reviews the methodology and outlines our experience with its implementation in a variety of models in vivo. Also discussed are potential limitations of the technique, complementary methods for assessing oxidative metabolism, and whether the P(i) → ATP flux is a viable biomarker of metabolic function in vivo.

FIG. 1.

The ³¹P-ST technique can be applied in human muscle in vivo to assess the unidirectional fluxes that contribute to phosphate-exchange reactions. Irradiating the γATP peak with a frequency-selective saturation pulse leads to a reduction in the phosphocreatine (ΔPCr) and inorganic phosphate (ΔP_i) signals due to phosphate exchange via CK or the ATP synthesis/hydrolysis cycle.

FIG. 2.

Direct manipulation of genes involved in mitochondrial content and metabolism modulates muscle P_i → ATP flux. Mice with muscle-specific overexpression of PGC1α (mPGC1α), a regulator of oxidative metabolism that increases mitochondrial density and enhances expression and protein content of genes involved in oxidative phosphorylation, exhibit a concomitant increase in P_i → ATP flux compared with wild-type (WT) mice. Muscle P_i → ATP flux is decreased in transgenic mice that overexpress UCP3 (UCP3-TG, D.E.B. and G.I.S unpublished observations), a mitochondrial membrane protein found in skeletal muscle that may uncouple the H⁺ electrochemical gradient across the inner mitochondrial membrane and decrease the efficiency of oxidative phosphorylation. Data obtained from UCP3-TG mice were acquired using the same experimental protocols described in Choi et al. (22). Data adapted from Choi et al. (22), reproduced courtesy of the National Academy of Sciences.

FIG. 3.

Three weeks of high-fat feeding (HFF) in both wild-type (WT) mice and mice with muscle-specific overexpression of PGC1α (mPGC1α) leads to an increase in muscle P_i → ATP flux. Data reproduced from Choi et al. (22), courtesy of the National Academy of Sciences.

FIG. 4.

Both P_i → ATP flux, assessed by ³¹P-ST MRS, and TCA cycle flux, measured using ¹³C-MRS to monitor the metabolism of infused [2-¹³C]acetate, were reduced by ∼40% in the muscle of healthy elderly subjects. Data adapted from Petersen et al. (18), reproduced courtesy of Science.

FIG. 5.

P_i → ATP flux was reduced by ∼30% in healthy, young, lean IR offspring of type 2 diabetic patients (IR offspring) compared with age-BMI-activity–matched IS control subjects (IS subjects). A similar decrease in TCA cycle flux was observed in an equivalent group of IR offspring. Adapted from Petersen et al. (19) and reproduced courtesy of the New England Journal of Medicine. Data reproduced from Befroy et al. (20).

FIG. 6.

Muscle-specific rates of P_i → ATP flux were obtained using a localized ³¹P-ST pulse-sequence to restrict detection to either the gastrocnemius or soleus muscles of the calf. Estimated rates for each muscle compartment are noted in the table.