Structural and Functional Alterations of Skeletal Muscle Microvasculature in Dystrophin-Deficient mdx Mice.

Duchenne muscular dystrophy (DMD) is a progressive neuromuscular disease, caused by an absence of dystrophin, inevitably leading to death. Although muscle lesions are well characterized, blood vessel alterations that may have a major impact on muscle regeneration remain poorly understood. Our aim was to elucidate alterations of the vascular network organization, taking advantage of Flk1(GFP/+) crossed with mdx mice (model for human DMD where all blood vessels express green fluorescent protein) and functional repercussions using in vivo nuclear magnetic resonance, combining arterial spin-labeling imaging of perfusion, and (31)P-spectroscopy of phosphocreatine kinetics. For the first time, our study focused on old (12-month-old) mdx mice, displaying marked chronic muscle lesions, similar to the lesions observed in human DMD, in comparison to young-adult (3-month-old) mdx mice displaying only mild muscle lesions with no fibrosis. By using an original approach combining a specific animal model, state-of-the-art histology/morphometry techniques, and functional nuclear magnetic resonance, we demonstrated that the microvascular system is almost normal in young-adult in contrast to old mdx mice, displaying marked microvessel alterations, and the functional repercussions on muscle perfusion and bioenergetics after a hypoxic stress vary depending on stage of pathology. This original approach clarifies disease evolution and paves the way for setting up new diagnostic markers or therapeutic strategies.

Quantitative NMRI and NMRS identify augmented disease progression after loss of ambulation in forearms of boys with Duchenne muscular dystrophy.

Quantitative NMRI and (31)P NMRS indices are reported in the forearms of 24 patients with Duchenne muscular dystrophy (DMD) (6-18 years, 14 non-ambulant) amenable to exon 53 skipping therapy and in 12 age-matched male controls (CONT). Examinations carried out at 3 T comprised multi-slice 17-echo measurements of muscle water T2 and heterogeneity, three-point Dixon imaging of fat fraction in flexor and extensor muscles (FLEX, EXT), and non-localised spectroscopy of phosphate metabolites. We studied four imaging indices, eight metabolic ratios combining ATP, phosphocreatine, phosphomonoesters and phosphodiesters, the cytosolic inorganic phosphate (Pia ) and an alkaline (Pib) pool present in dystrophic muscle, and average pH. All indices differed between DMD and CONT, except for muscle water T2 . Measurements were outside the 95th percentile of age-matched CONT values in over 65% of cases for percentage fat signal (%F), and in 78-100% of cases for all spectroscopic indices. T2 was elevated in one-third of FLEX measurements, whereas %pixels > 39 ms and T2 heterogeneity were abnormal in one-half of the examinations. The FLEX muscles had higher fat infiltration and T2 than EXT muscle groups. All indices, except pH, correlated with patient age, although the correlation was negative for T2 . However, in non-ambulant patients, the correlation with years since loss of ambulation was stronger than the correlation with age, and the slope of evolution per year was steeper after loss of ambulation. All indices except Pi/gATP differed between ambulant and non-ambulant patients; however, T2 and %pixels > 39 ms were highest in ambulant patients, possibly owing to the greater extent of inflammatory processes earlier in the disease. All other indices were worse in non-ambulant subjects. Quantitative measurements obtained from patients at different disease stages covered a broad range of abnormalities that evolved with the disease, and metabolic indices were up to 10-fold above normal from the onset, thus establishing a variety of potential markers for future therapy.

Skeletal muscle quantitative nuclear magnetic resonance imaging follow-up of adult Pompe patients.

Adult late-onset Pompe disease is most often a slowly progressive limb-girdle and spine extensor muscle dystrophy, due to defective lysosomal acid maltase. With the exception of the few patients who present with a dramatically accelerated clinical course, standard diagnostic imaging fail to detect and evaluate disease progression between two successive visits. In muscle dystrophy of very rapid evolution, like the Duchenne disease, quantitative NMR imaging has successfully demonstrated its capacity to objectivate both disease activity and degenerative changes progression over short follow-up periods. The purpose of this retrospective monocentric open-label study was to investigate whether quantitative NMR imaging can monitor disease progression in adult Pompe patients despite its very slow nature. Quantitative imaging of Pompe patients succeeded in demonstrating that muscle fatty infiltration increased on average by 0.9%/year, with the hamstring and adductor muscles showing the fastest degradation. Muscle water T2 mapping revealed that 32% of all muscles had abnormally high T2 in at least one of two successive examinations. When muscle water T2 was abnormal, fatty degenerative changes were further increased by 0.61%/year. Enzyme replacement therapy resulted in 0.68%/year slowdown of the muscle fatty infiltration, in both muscles with normal and high T2s.

Forelimb treatment in a large cohort of dystrophic dogs supports delivery of a recombinant AAV for exon skipping in Duchenne patients.

Duchenne muscular dystrophy (DMD) is a severe muscle-wasting disorder caused by mutations in the dystrophin gene, without curative treatment yet available. Our study provides, for the first time, the overall safety profile and therapeutic dose of a recombinant adeno-associated virus vector, serotype 8 (rAAV8) carrying a modified U7snRNA sequence promoting exon skipping to restore a functional in-frame dystrophin transcript, and injected by locoregional transvenous perfusion of the forelimb. Eighteen Golden Retriever Muscular Dystrophy (GRMD) dogs were exposed to increasing doses of GMP-manufactured vector. Treatment was well tolerated in all, and no acute nor delayed adverse effect, including systemic and immune toxicity was detected. There was a dose relationship for the amount of exon skipping with up to 80% of myofibers expressing dystrophin at the highest dose. Similarly, histological, nuclear magnetic resonance pathological indices and strength improvement responded in a dose-dependent manner. The systematic comparison of effects using different independent methods, allowed to define a minimum threshold of dystrophin expressing fibers (>33% for structural measures and >40% for strength) under which there was no clear-cut therapeutic effect. Altogether, these results support the concept of a phase 1/2 trial of locoregional delivery into upper limbs of nonambulatory DMD patients.

Multiparametric functional nuclear magnetic resonance imaging shows alterations associated with plasmid electrotransfer in mouse skeletal muscle.

BACKGROUND: In vivo gene electrotransfer is frequently used in preclinical gene therapy. Many studies have attempted to optimize protocols efficiency at the same time as reducing muscle damage. Most of them have reported histological evidence of muscle degeneration and completion of regeneration within 15 days. The functional consequences have rarely been addressed, which may reflect the lack of appropriate techniques. Yet, it is important to characterize the changes induced by the procedure itself because it may interfere with therapy. We used multiparametric functional (mpf)-nuclear magnetic resonance (NMR) imaging to evaluate mice hindlimb muscle after electrotransfer of an empty plasmid. METHODS: NMR experiments were performed in a 4T Bruker magnet. Arterial spin labeling imaging of perfusion and blood oxygenation level dependent contrast and (31) P spectroscopy of phosphocreatine kinetics and pH were simultaneously acquired from the mice hindlimb during 2 min of electrically stimulated exercise and recovery. RESULTS: After 15 days, hindlimb cross-sectional area decreased by 10% compared to control mice. Specific force-time integral and end-exercise pH were identical in both groups, whereas oxidative capacities increased. Perfusion values doubled, and oxygenation significantly decreased. Histology revealed: (i) degeneration/regeneration; (ii) a decrease in type IIb fibers and an increase in type I and IIa fibers; and (iii) increased capillary density. CONCLUSIONS: In this model, loss in muscle mass was accompanied by important alterations of perfusion and bioenergetics. Fifteen days after electrotransfer, this was correlated with fiber type shift, capillary bed remodeling and degeneration/regeneration. mpf-NMR provides new insights into the functional consequences of standard electrotransfer and represents a powerful tool for optimization and longitudinal assessment of preclinical gene therapy protocols.

Splitting of Pi and other ³¹P NMR anomalies of skeletal muscle metabolites in canine muscular dystrophy.

Many anomalies exist in the resting (31) P muscle spectra of boys with Duchenne muscular dystrophy (DMD) but few have been reported in Golden Retriever muscular dystrophy (GRMD), the closest existing animal model for DMD. Because GRMD is recommended for preclinical evaluation of therapies and quantitative outcome measures are needed, we investigated anomalies of (31) P NMRS in tibial cranial and biceps femoris muscles from 14 GRMD compared to 9 control (CONT) dogs. Alterations observed in DMD children - low phosphocreatine and high phospho-monoesters and -diesters - were all found in GRMD but increased pH was not. More surprisingly, inorganic phosphate (Pi) appeared to present a prominent splitting with an enhanced Pi(b) resonance at 0.3 ppm downfield of Pi(a) . Assuming that both resonances are Pi, the pH for Pi(a) in GRMD corresponded to a physiological intracellular pH(a) (6.97 ± 0.05), while pH(b) approached the extracellular range (7.27 ± 0.10) and correlated with pH(a) in GRMD (R(2) = 0.65). Both Pi(a) and Pi(b) were elevated compared to CONT and Pi(a) increased with age for GRMD (R(2) = 0.48, p < 0.001). Magnetisation transfer experiments between γATP and Pi were conducted to better characterise Pi pools. Equal T1 relaxation times for Pi(b) and Pi(a) did not support a mitochondrial origin of Pi(b) . We suggest that Pi(b) could originate from degenerating hypercontracted cells that have a leaky membrane and inadequate cell homeostasis and pH regulation. Pi(b) showed minimal chemical exchange in all dogs, while the exchange rate of Pi(a) was reduced in GRMD and might extraneously reflect low glycolytic activity in DMD. Taken together, the ensemble of (31) P NMRS alterations identifies muscle dysfunction and could provide useful biomarkers of therapeutic efficacy. Furthermore, among these, two might relate more specifically to dystrophic processes and merit further investigation: one is the existence of the enhanced alkaline Pi(b) pool; the other, mechanisms by which membrane disruption might increase phosphodiesters in dystrophy.

Abnormal response to cortical activation in early stages of Huntington disease.

BACKGROUND: We wished to identify noninvasive in vivo biomarkers of brain energy deficit in Huntington disease. METHODS: We studied 15 early affected patients (mean motor United Huntington Disease Rating Scale, 18 ± 9) and 15 age- and sex-matched controls. We coupled (31)phosphorus nuclear magnetic resonance spectroscopy with activation of the occipital cortex in order to measure the relative concentrations of adenosine triphosphate, phosphocreatine, and inorganic phosphate before, during, and after visual stimulation. RESULTS: In controls, we observed an 11% increase in the inorganic phosphate/phosphocreatine ratio (P = .024) and a 13% increase in the inorganic phosphate/adenosine triphosphate ratio (P = .016) during brain activation, reflecting increased adenosine diphosphate concentrations. Subsequently, controls had a return to baseline levels during recovery (P = .012 and .022, respectively). In contrast, both ratios were unchanged in patients during and after visual stimulation. CONCLUSIONS: (31)Phosphorus nuclear magnetic resonance spectroscopy could provide functional biomarkers of brain energy deficit to monitor therapeutic efficacy in Huntington disease.

Antioxidants and aging: NMR-based evidence of improved skeletal muscle perfusion and energetics.

We sought to examine the potential role of oxidative stress on skeletal muscle function with advancing age. Nuclear magnetic resonance (NMR) was employed to simultaneously assess muscle perfusion (arterial spin labeling) and energetics ((31)P NMR spectroscopy) in the lower leg of young (26 + or - 5 yr, n = 6) and older (70 + or - 5 yr, n = 6) healthy volunteers following the consumption of either placebo (PL) or an oral antioxidant (AO) cocktail (vitamins C and E and alpha-lipoic acid), previously documented to decrease plasma free radical concentration. NMR measurements were made during and after 5 min of moderate intensity (approximately 5 W) plantar flexion exercise. AO administration significantly improved end-exercise perfusion (AO, 50 + or - 5, and PL, 43 + or - 4 ml x 100 g(-1) x min(-1)) and postexercise perfusion area under the curve (AO, 1,286 + or - 236, and PL, 866 + or - 144 ml/100 g) in older subjects, whereas AO administration did not alter hemodynamics in the young group. Concomitantly, muscle oxidative capacity (time constant of phosphocreatine recovery, tau) was improved following AO in the older (AO, 43 + or - 1, and PL, 51 + or - 7 s) but not the young (AO, 54 + or - 5, and PL, 48 + or - 7 s) group. These findings support the concept that oxidative stress may be partially responsible for the age-related decline in skeletal muscle perfusion during physical activity and reveal a muscle metabolic reserve capacity in the elderly that is accessible under conditions of improved perfusion.

Quantitative, dynamic and noninvasive determination of skeletal muscle perfusion in mouse leg by NMR arterial spin-labeled imaging.

Because mouse may relatively easily be genetically tailored to develop equivalent of human muscular diseases or to present controlled alterations of mechanisms involved in vasoregulation, it has become the prevalent species to explore such questions. However, the very small size of the animals represents a serious limitation when evaluating the functional consequences of these genetic manipulations. In this context, the recourse to arterial spin labeling (ASL) nuclear magnetic resonance (NMR) methods in which arterial water spins act as an endogenous and freely diffusible tracer of perfusion is tempting but challenging. This article shows that despite the small size of the animal, mouse muscle perfusion may be measured, at rest and in conditions of reactive hyperemia, using saturation inversion recovery sequence, a pulsed ASL variant, combined with NMR imaging. Baseline perfusion values in the mouse leg were 17+/-11 ml.min(-1).100 g(-1) (n=11) and were comparable to microsphere data from the literature. Under ischemia, leg perfusion was 1.2+/-9.3 ml.min(-1).100 g(-1) (n=11). The difference observed between basal and ischemic measurements was statistically different (P=.0001). The temporal pattern of hyperemia in mouse muscle was coherent with previously published measurements in humans and in rats. The mean peak perfusion was 62+/-24 ml.min(-1).100 g(-1) (n=6) occurring 48+/-27 s after the end of occlusion. In conclusion, this study demonstrated the ability of ASL combined to NMR imaging to quantify skeletal muscle perfusion in mice legs, both at rest and dynamically.

Longitudinal functional and NMR assessment of upper limbs in Duchenne muscular dystrophy.

OBJECTIVE: To explore the value of nuclear magnetic resonance (NMR) and functional assessments for follow-up of ambulatory and nonambulatory patients with Duchenne muscular dystrophy (DMD). METHODS: Twenty-five 53-skippable patients with DMD were included in this study; 15 were nonambulatory at baseline. All patients underwent clinical and functional assessments every 6 months using the Motor Function Measure (MFM), hand grip and key pinch strength, MoviPlate, and NMR spectroscopy and imaging studies. RESULTS: Upper limb distal strength decreased in nonambulatory patients over the period of 1 year; ambulatory patients showed improvement during the same period. The same applied for several NMRS indices, such as phosphocreatine/adenosine triphosphate, which decreased in older patients but increased in younger ambulatory patients. Fat infiltration in the upper limbs increased linearly with age. Almost all NMR and functional assessment results correlated. CONCLUSIONS: Our results underscore complementarity of functional and NMR assessments in patients with DMD. Sensitivity to change of various indices may differ according to disease stage.

MRS Evidence of Adequate O2 Supply in Human Skeletal Muscle at the Onset of Exercise.

PURPOSE: At exercise onset, intramuscular oxidative energy production responds relatively slowly in comparison with the change in adenosine triphosphate demand. To determine whether the slow kinetics of oxidative adenosine triphosphate production is due to inadequate O2 supply or metabolic inertia, we studied the kinetics of intramyocellular deoxygenation (deoxy-myoglobin (Mb)) and metabolism (phosphocreatine (PCr)) using proton (1H) and phosphorus (31P) magnetic resonance spectroscopy in six healthy subjects (33 ± 5 yr). METHODS: Specifically, using dynamic plantarflexion exercise, rest to exercise and recovery were assessed at both 60% of maximum work rate (moderate intensity) and 80% of maximum work rate (heavy intensity). RESULTS: At exercise onset, [PCr] fell without delay and with a similar time constant (τ) at both exercise intensities (approximately 33 s). In contrast, the increase in deoxy-Mb was delayed at exercise onset by 5-7 s, after which it increased with kinetics (moderate τ = 37 ± 9 s; heavy τ = 29 ± 6 s) that was not different from τPCr (P > 0.05). At cessation, deoxy-Mb recovered without time delay and more rapidly (τ = ∼20 s) than PCr (τ = ∼33 s) (P < 0.05). CONCLUSIONS: Using a unique combination of in vivo magnetic resonance spectroscopy techniques with high time resolution, this study revealed a delay in intramuscular deoxygenation at the onset of exercise and rapid reoxygenation kinetics upon cessation. Together, these data imply that intramuscular substrate-enzyme interactions, and not O2 availability, determine the exercise onset kinetics of oxidative metabolism in healthy human skeletal muscles.

Evaluation of muscle glycogen content by 13C NMR spectroscopy in adult-onset acid maltase deficiency.

Muscle glycogen storage was measured by in vivo, natural abundance 13C nuclear magnetic resonance spectroscopy in distal and proximal lower limb segments of patients suffering from adult-onset acid maltase deficiency. Interleaved T1-weighted acquisitions of glycogen and creatine served to quantify glycogen excess. For acid maltase deficient patients (n=11), glycogen:creatine was higher than controls (n=12), (1.20+/-0.39 vs. 0.83+/-0.18, P=0.0005). Glycogen storage was above the normal 95% confidence limits in at least one site for 7/11 patients. The intra-individual coefficient of reproducibility was 12%. This totally atraumatic measurement of glycogen allows repeated measurement at different muscle sites of acid maltase deficient patients, despite selective fatty replacement of tissue. This could provide an additional parameter to follow the development of disease in individual patients, including in the perspective of forthcoming therapeutic trials. It may also offer an appropriate tool to study the role of glycogen accumulation in progression of the pathology.

Ultrafast multiplanar determination of left ventricular hypertrophy in spontaneously hypertensive rats with single-shot spin-echo nuclear magnetic resonance imaging.

OBJECTIVE: To determine whether left ventricular hypertrophy can be correctly evaluated in hypertensive rats with a new nuclear magnetic resonance (NMR) imaging modality that is relatively simple to operate and provides results of constant quality while offering a high signal-to-noise ratio. DESIGN Left ventricular mass as calculated from the NMR imaging analysis was compared with the actual left ventricular mass measured by gravimetry. METHODS: Single-shot ultrafast spin-echo (SSFSE) imaging of hearts of Wistar-Kyoto rats and spontaneously hypertensive rats was performed at 4 T. Left ventricular mass was determined by using Simpson's rule on stacks of images acquired in systole and diastole. RESULTS: SSFSE NMR imaging performed in systole or in diastole evaluated and quantified left ventricular hypertrophy in hearts of spontaneously hypertensive rats very similarly to gravimetry. The left ventricular mass as determined by NMR was in good accordance with the actual left ventricular weight (SEE: 30.39 and 35.86 mg for the systolic and diastolic NMR acquisitions, respectively). CONCLUSION: Using an SSFSE sequence, high-quality NMR images of the rat heart can be generated very reliably with sufficient contrast and temporal and spatial resolution, and allow precise, non-invasive and fast characterization of left ventricular hypertrophy in a hypertensive rat model.

Whole-body muscle MRI in 20 patients suffering from late onset Pompe disease: Involvement patterns.

To describe muscle involvement on whole-body MRI scans in adult patients at different stages of late-onset Pompe disease. Twenty patients aged 37 to 75 were examined. Five were bedridden and required ventilatory support. Axial and coronal T1 turbo-spin-echo sequences were performed on 1.5T or 3T systems. MRI was scored for 47 muscles using Mercuri's classification. Whole-body scans were obtained with a mean in-room time of 29 min. Muscle changes consisted of internal bright signals of fatty replacement without severe retraction of the muscles' corpus. Findings were consistent with previous descriptions of spine extensors and pelvic girdle, but also provided new information on recurrent muscle changes particularly in the tongue and subscapularis muscle. Moreover thigh involvement was more heterogeneous than previously described, in terms of distribution across muscles as well as with respect to the overall clinical presentation. Whole-body MRI provides a very evocative description of muscle involvement in Pompe disease in adults.

Investigating glycogenosis type III patients with multi-parametric functional NMR imaging and spectroscopy.

Debranching enzyme deficiency (Glycogen storage disease (GSD) type III) causes progressive muscle wasting myopathy. A comprehensive nuclear magnetic resonance study involving spectroscopy (NMRS) and imaging (NMRI) evaluated status and function of calf muscles in 18 GSDIII patients. At rest, (31)P NMRS showed elevated pH and accumulation of anomalous phosphomonoesters, (13)C NMRS quantified excess glycogen accumulation and NMRI demonstrated progressive fat replacement that paralleled muscle weakness. Multi-parametric functional NMR, performed at recovery from a single bout of aerobic exercise, simultaneously assessed oxidative phosphorylation from (31)P NMRS, muscle perfusion and BOLD, a marker of blood oxygenation, from arterial spin labeled NMRI, and oxygen uptake from deoxymyoglobin proton NMRS. While blocked glycogenolysis caused inadequate substrate supply to the mitochondria, combined measurements suggested that altered perfusion was also responsible for impaired post-exercise phosphocreatine recovery and could contribute to exercise intolerance in GSDIII. These non-invasive investigations provide new indices to quantify the progression of GSDIII.

Multiparametric NMR-based assessment of skeletal muscle perfusion and metabolism during exercise in elderly persons: preliminary findings.

BACKGROUND: Aging is associated with a decline in exercise capacity that may be attributable to maladaptations in both skeletal muscle perfusion and metabolism; yet very little is known regarding the real-time, within-muscle interplay between these parameters during physical activity. Therefore, we utilized an unique nuclear magnetic resonance sequence to concomitantly examine changes in lower leg skeletal muscle perfusion and metabolism. METHODS: In young (26+/-5 years, n=6) and older (70+/-5 years, n=6) healthy volunteers, arterial spin labeling measurements of muscle perfusion were combined with 31 Phosphorous (31P) nuclear magnetic resonance spectroscopy to monitor high-energy phosphate metabolites during and after 5 minutes of moderate-intensity (approximately 5W) plantar flexion exercise. RESULTS: Compared with young, end-exercise perfusion was diminished in older participants (43+/-10 mL/100 g/minute, old; 60+/-7 mL/100 g.minute, young), accompanied by greater phosphocreatine (PCr) depletion (-28%+/-12%, old; -19%+/-7%, young) and elevated inorganic phosphate/PCr (0.41+/-0.2, old; 0.24+/-0.09, young); yet the time constant for PCr recovery (tau, an index of muscle oxidative capacity) was similar between groups (51+/-17 seconds, old; 48+/-7 seconds, young). CONCLUSIONS: Together, these preliminary data provide evidence of an age-related decline in tissue perfusion and increased "metabolic stress" during exercise but demonstrate that overall oxidative capacity in the elderly does not appear negatively affected by this relatively hypoperfused state.

Human skeletal muscle intracellular oxygenation: the impact of ambient oxygen availability.

Intracellular oxygen (O2) availability and the impact of ambient hypoxia have far reaching ramifications in terms of cell signalling and homeostasis; however, in vivo cellular oxygenation has been an elusive variable to assess. Within skeletal muscle the extent to which myoglobin desaturates (deoxy-Mb) and the extent of this desaturation in relation to O2 availability provide an endogenous probe for intracellular O2 partial pressure (P(iO2)). By combining proton nuclear magnetic resonance spectroscopy (1H NMRS) at a high field strength (4 T), assessing a large muscle volume in a highly efficient coil, and extended signal averaging (30 min) we assessed the level of skeletal muscle deoxy-Mb in 10 healthy men (30 +/- 4 years) at rest in both normoxia and hypoxia (10% O2). In normoxia there was an average deoxy-Mb signal of 9 +/- 1%, which, when converted to P(iO2) using an O2/Mb half-saturation (P50) of 3.2 mmHg, revealed an P(iO2) of 34 +/- 6 mmHg. In ambient hypoxia the deoxy-Mb signal rose to 13 +/- 3% (P(iO2) = 23 +/- 6 mmHg). However, intersubject variation in the defence of arterial oxygenation (S(aO2)) in hypoxia (S(aO2) range: 86-67%) revealed a significant relationship between the changes in S(aO2) and P(iO2)(r2 = 0.5). These data are the first to document resting intracellular oxygenation in human skeletal muscle, highlighting the relatively high P(iO2) values that contrast markedly with those previously recorded during exercise (approximately 2-5 mmHg). Additionally, the impact of ambient hypoxia on P(iO2) and the relationship between changes in S(aO2) and P(iO2) stress the importance of the O2 cascade from air to cell that ultimately effects O2 availability and O2 sensing at the cellular level.

New insight into abnormal muscle vasodilatory responses in aged hypertensive rats by in vivo nuclear magnetic resonance imaging of perfusion.

OBJECTIVE: Increased peripheral arterial resistances and decreased maximum vasodilation are characteristic features of chronic hypertension. However, little data are available in the literature regarding the possible alterations in the temporal patterns of vasodilatory responses elicited by various stimuli. DESIGN: This question was addressed by measuring skeletal muscle perfusion using nuclear magnetic resonance imaging combined with arterial spin labeling. METHODS: Ninety-week-old male spontaneously hypertensive (SHR; n = 7) and normotensive Wistar Kyoto (WKY; n = 8) rats were studied, and calf muscle perfusion was measured at rest and during reactive hyperemia following total ischemia of 5 and 30 min duration. RESULTS: Reactive hyperemia profiles differed according to duration of ischemia. In WKY rats, 5 min of ischemia induced a short peak of hyperemia lasting no more than 63 s, while 30 min of ischemia were followed by a prolonged hyperemic response of 261 s. In SHRs, after 5 min of ischemia, peak muscle arterial conductance was decreased to 0.5 +/- 0.3 versus 0.9 +/- 0.3 ml.min(-1).100 g(-1).mm Hg(-1) in the WKY rats (p < 0.05), as expected. After 30 min of ischemia, there was, in addition, a shortening of the hyperemic response duration. Time to post-ischemic half normalization of arterial conductance was 38 +/- 24 s in the SHRs versus 149 +/- 58 s in the WKY rats (p < 0.001). CONCLUSION: In vivo perfusion measurement not only confirmed the existence of a reduced maximum peripheral vasodilation in chronically hypertensive rats, it revealed a blunted hyperemic response after prolonged ischemia in the SHRs, which might be an important contributing factor to the increased sensitivity to ischemia in hypertension.