Quantitative MRI water content mapping of porcine intervertebral disc during uniaxial compression.

BACKGROUND: Intervertebral disc (IVD) diseases are major public health problem in industrialized countries where they affect a large proportion of the population. In particular, IVD degeneration is considered to be one of the leading causes of pain consultation and sick leave. The aim of this study was to develop a new method for assessing the functionality of IVD in order to diagnose IVD degeneration. METHODS: For this purpose, we have designed a specific device that enables to mechanically load porcine IVD ex vivo in the 4.7-Tesla horizontal superconducting magnet of a magnetic resonance (MR) scanner. Proton density weighted imaging (ρH-MRI) of the samples was acquired. FINDINGS: The post-processing on MR images allowed (1) to reconstruct the 3D deformation under a known mechanical load and (2) to infer the IVD porosity assuming an incompressible poroelastic model. INTERPRETATION: This study demonstrates the ability to follow the change in morphology and hydration of an IVD using MR measurements, thereby providing valued information for a better understanding of IVD function.

In vivo and in vitro investigations of heterozygous nebulin knock-out mice disclose a mild skeletal muscle phenotype.

Nemaline myopathy is the most common congenital skeletal muscle disease, and mutations in the nebulin gene account for 50% of all cases. Recent studies suggest that the disease severity might be related to the nebulin expression levels. Considering that mutations in the nebulin gene are typically recessive, one would expect that a single functional nebulin allele would maintain nebulin protein expression which would result in preserved skeletal muscle function. We investigated skeletal muscle function of heterozygous nebulin knock-out (i.e., nebulin(+/-)) mice using a multidisciplinary approach including protein and gene expression analysis and combined in vivo and in vitro force measurements. Skeletal muscle anatomy and energy metabolism were studied strictly non-invasively using magnetic resonance imaging and 31P-magnetic resonance spectroscopy. Maximal force production was reduced by around 16% in isolated muscle of nebulin(+/-) mice while in vivo force generating capacity was preserved. Muscle weakness was associated with a shift toward a slower proteomic phenotype, but was not related to nebulin protein deficiency or to an impaired energy metabolism. Further studies would be warranted in order to determine the mechanisms leading to a mild skeletal muscle phenotype resulting from the expression of a single nebulin allele.

Capsiate administration results in an uncoupling protein-3 downregulation, an enhanced muscle oxidative capacity and a decreased abdominal fat content in vivo.

OBJECTIVES: The involvement of skeletal muscle mitochondrial uncoupling protein-3 (UCP3) in the control of energy expenditure in skeletal muscle and at the whole-body level is still a matter of debate. We previously reported that UCP3 downregulation is linked to an enhanced mitochondrial energy metabolism in rat skeletal muscle as a result of acute capsiate treatment. Here, we aimed at investigating noninvasively the effects of chronic capsiate ingestion on metabolic changes occurring in exercising gastrocnemius muscle and at the whole-body level. METHODS: We used an original experimental setup allowing a complete noninvasive investigation of gastrocnemius muscle function in situ using 31-phosphorus magnetic resonance spectroscopy. Whole-body fat composition was determined using magnetic resonance imaging and UCP3 gene expression was measured by quantitative real-time RT-PCR analysis. RESULTS: We found that a 14-day daily administration of capsiate (100 mg kg(-1) body weight) reduced UCP3 gene expression and increased phosphocreatine level at baseline and during the stimulation period in gastrocnemius muscle. During muscle stimulation, pH(i) showed a larger alkalosis in the capsiate group suggesting a lower glycolysis and a compensatory higher aerobic contribution to ATP production. Although the capsiate-treated rats were hyperphagic as compared to control animals, they showed a lower weight gain coupled to a decreased abdominal fat content. CONCLUSION: Overall, our data indicated that capsiate administration contributes to the enhancement of aerobic ATP production and the reduction of body fat content coupled to a UCP3 gene downregulation.

Downregulation of uncoupling protein-3 in vivo is linked to changes in muscle mitochondrial energy metabolism as a result of capsiate administration.

Although it has been suggested that the skeletal muscle mitochondrial uncoupling protein-3 (UCP3) is involved in regulating energy expenditure, its role is still poorly understood. In the present study, we aimed at investigating noninvasively, using magnetic resonance techniques, metabolic changes occurring in exercising muscle as a result of capsiate treatment, which has been previously linked to UCP3 upregulation. We showed that capsiate ingestion strongly reduced UCP3 gene expression in rat gastrocnemius muscle. This large underexpression was accompanied by a significant increase in the rate of mitochondrial ATP production and phosphocreatine level both at rest and during muscle stimulation. Similarly, the stimulation-induced ATP fall and ADP accumulation were significantly less after capsiate administration than in untreated rats. The larger oxidative ATP production rate could not be explained by a proportional decrease in the anaerobic component, i.e., glycolysis and phosphocreatine breakdown. In addition, the mechanical performance was not affected by capsiate administration. Finally, the plasma free fatty acid (FFA) level increased in capsiate-treated rats, whereas no significant change was observed after muscle stimulation in the control group. Considering the corresponding enhanced UCP3 mRNA expression occurring in the control group after muscle stimulation, one can suggest that changes in FFA level and UCP3 mRNA expression are not mechanistically correlated. Overall, we have shown that capsiate administration induced a UCP3 downregulation coupled with an increased mitochondrial ATP synthesis, whereas the muscle force-generating capacity was unchanged. This suggests that a decrease in muscle efficiency and/or additional noncontractile ATP-consuming mechanisms result from UCP3 downregulation.

In vivo MR investigation of skeletal muscle function in small animals.

In vivo 31P-MRS investigations have been widely used in small animals to study skeletal muscle function under normal and pathological conditions. Paradoxically in these studies, the benefit provided by 31P-MRS in terms of non-invasiveness is lost because of the utilization of experimental setups that integrate invasive devices for inducing muscle contractions and for measuring mechanical performance. These traditional methodologies, which require surgical preparations, have obvious limitations regarding repeatability in the same animal. The purpose of this review is to highlight the technical aspects of the in vivo MR investigations of skeletal muscle function in small animal models. We will more particularly address the issue related to the invasiveness of different procedures used so far in order to show finally that a further step into non-invasiveness can be achieved, in particular with the support of muscle functional 1H-MRI.

Functional investigations of exercising muscle: a noninvasive magnetic resonance spectroscopy-magnetic resonance imaging approach.

Muscle fatigue, which is defined as the decline in muscle performance during exercise, may occur at different sites along the pathway from the central nervous system through to the intramuscular contractile machinery. Historically, both impairment of neuromuscular transmission and peripheral alterations within the muscle have been proposed as causative factors of fatigue development. However, according to more recent studies, muscle energetics play a key role in this process. Intramyoplasmic accumulation of inorganic phosphate (P(i)) and limitation in ATP availability have been frequently evoked as the main mechanisms leading to fatigue. Although attractive, these hypotheses have been elaborated on the basis of experimental results obtained in vitro, and their physiological relevance has never been clearly demonstrated in vivo. In that context, noninvasive methods such as 31-phosphorus magnetic resonance spectroscopy and surface electromyography have been employed to understand both metabolic and electrical aspects of muscle fatigue under physiological conditions. Mapping of muscles activated during exercise is another interesting issue which can be addressed using magnetic resonance imaging (MRI). Exercise-induced T2 changes have been used in order to locate activated muscles and also as a quantitative index of exercise intensity. The main results related to both issues, i.e. the metabolic and electrical aspects of fatigue and the MRI functional investigation of exercising muscle, are discussed in the present review.

Combined in situ analysis of metabolic and myoelectrical changes associated with electrically induced fatigue.

Electrical muscle stimulation (Mstim) at a low or high frequency is associated with failure of force production, but the exact mechanisms leading to fatigue in this model are still poorly understood. Using 31P magnetic resonance spectroscopy (31PMRS), we investigated the metabolic changes in rabbit tibialis anterior muscle associated with the force decline during Mstim at low (10 Hz) and high (100 Hz) frequency. We also simultaneously recorded the compound muscle mass action potential (M-wave) evoked by direct muscle stimulation, and we analyzed its post-Mstim variations. The 100-Hz Mstim elicited marked M-wave alterations and induced mild metabolic changes at the onset of stimulation followed by a paradoxical recovery of phosphocreatine (PCr) and pH during the stimulation period. On the contrary, the 10-Hz Mstim produced significant PCr consumption and intracellular acidosis with no paradoxical recovery phenomenon and no significant changes in M-wave characteristics. In addition, the force depression was linearly linked to the stimulation-induced acidosis and PCr breakdown. These results led us to conclude that force failure during 100-Hz Mstim only results from an impaired propagation of muscle action potentials with no metabolic involvement. On the contrary, fatigue induced by 10-Hz Mstim is closely associated with metabolic changes with no alteration of the membrane excitability, thereby underlining the central role of muscle energetics in force depression when muscle is stimulated at low frequency. Finally, our results further indicate a reduction of energy cost of contraction when stimulation frequency is increased from 10 to 100 Hz.

In vivo reduction in ATP cost of contraction is not related to fatigue level in stimulated rat gastrocnemius muscle.

1. We tested whether the reduction in ATP cost of contraction during in vivo stimulation of rat gastrocnemius muscle was related to fatigue level. 2. Muscles (n = 44) were electrically stimulated to perform 6 min repeated isometric contractions at different frequencies; one non-fatiguing protocol (stimulation at 0.8 Hz) and five fatiguing protocols (2, 3.2, 4, 5.2 and 7.6 Hz) were used. Anaerobic and oxidative ATP turnover rates were measured non-invasively using (31)P-magnetic resonance spectroscopy. 3. At the onset of the stimulation period, no signs of fatigue were measured in the six protocols and ATP cost of contraction did not differ significantly (P = 0.45) among protocols (mean value of 1.76 +/- 0.11 mM (N s)(-1)). 4. For the six protocols, ATP cost of contraction was significantly reduced (P < 0.05) at the end of the stimulation period when compared with the initial value. This reduction did not differ significantly (P = 0.61) among the five fatiguing protocols (averaging 35 +/- 3 % of initial value), whereas isometric force decreased significantly as stimulation frequency increased. No significant correlation (P = 0.87, r(2) = 0.01) was observed between isometric force and ATP cost of contraction at the end of the stimulation period. In addition, this reduction was significantly lower (P < 0.05) for the non-fatiguing protocol (67 +/- 9 % of initial value) when compared with the fatiguing protocols. 5. These results demonstrate that (i) the reduction in ATP cost of contraction during in vivo stimulation of rat gastrocnemius muscle is not related to the fatigue level; (ii) surprisingly, this reduction was significantly larger during the fatiguing protocols compared with the non-fatiguing protocol.

Time-dependent and indirect effect of inorganic phosphate on force production in rat gastrocnemius exercising muscle determined by 31P-MRS.

The relationship of inorganic phosphate (P(i)) and its diprotonated form (H(2)PO(4)(-)) to isometric force (F) was analyzed non-invasively using 31P-magnetic resonance spectroscopy. Rat gastrocnemius muscles were electrically stimulated at six different frequencies in order to produce different levels of fatigue. A curvilinear relationship was demonstrated between force production and [P(i)] and [H(2)PO(4)(-)] accumulation. [P(i)] and [H(2)PO(4)(-)] were correlated with F at the end of the stimulation period but not when F was maximal at the early stage of the stimulation period. Interestingly, the respective [P(i)] and [H(2)PO(4)(-)] did not differ significantly between these two stages demonstrating that [P(i)] and [H(2)PO(4)(-)] cannot be considered as direct effectors of fatigue. This time-dependent and indirect effect of [P(i)] and [H(2)PO(4)(-)] on force production might be mediated by calcium ions.