Benjamin Levine, M.D.
Professor of Medicine                          
Division of Cardiology                                              

Craig Crandall, M.D.
Associate Professor of Medicine
Division of Cardiology

The Multi System Effect of Exercise Training/Nutritional Support During Prolonged Bed Rest Deconditioning: An Integrated Approach to Countermeasure Development for the Heart, Lungs, Muscle and Bones

Sustained exposure to microgravity leads to adaptive changes in the cardiovascular and musculoskeletal systems that may result in substantial morbidity. For example cardiovascular deconditioning may lead to orthostatic hypotension and syncope. Atrophy of skeletal muscle will diminish work capacity and may lead to muscle injury. Bone demineralization increases the risk of kidney stone formation and may reduce bone strength increasing the risk of fracture. Bone resorption may be particularly severe after long duration space flight with uncertain recovery. Despite in depth study, the optimal countermeasure for each system has not yet been defined. More importantly, previous work has focused predominantly on one organ system at a time, ignoring the interaction among systems, and preventing the development of a specific countermeasure for an individual astronaut that might be effective for the heart, muscles and bones. The global objective of this proposal is to test an integrated countermeasure that will be effective against cardiovascular deconditioning, skeletal muscle atrophy, and bone demineralization, and that ultimately can be applied practically aboard the International Space Station or a mission to Mars. Hypotheses and
Specific Aims: Hypothesis 1: An "optimized" exercise training program combining dynamic plus intermittent resistance exercise can prevent the cardiovascular atrophy and deconditioning associated with prolonged bed rest. Hypothesis 2: This dynamic plus resistance exercise training program, when combined with potassium magnesium citrate supplementation will attenuate the increased risk for stone formation, and diminish bed rest induced bone loss to a greater extent than the effect of exercise training or supplementation alone. Hypothesis 3: This dynamic plus resistance exercise training program during bed rest will also attenuate structural and functional alterations in skeletal muscle induced by prolonged bed rest, thereby preserving strength and endurance. To test these hypotheses, we propose to accomplish the following specific aims: Specific Aim 1: To perform an exercise countermeasure using rowing ergometry combined with resistance training to obtain the most intensive stimulus to cardiac hypertrophy in the shortest period of time. The functional importance of cardiac atrophy for orthostatic tolerance after prolonged bed rest will be determined from invasive measurements of ventricular performance and compliance (Frank Starling and LV pressure/volume curves), and non invasive imaging techniques to measure the dynamic component of diastole. A novel oral volume loading strategy will also be applied just prior to orthostatic tolerance testing. Specific aim 2: To assess the effect of exercise training combined with potassium magnesium citrate (KMgCit) supplementation in preventing microgravity induced increases in bone resorption, urinary calcium excretion, and risk of stone formation. These specific aims will be accomplished by precise metabolic control and evaluation, plus non invasive evaluation of bone structure and function (bone quality by ultrasound). Specific Aim 3: To demonstrate the effectiveness of dynamic and resistance exercise training in attenuating the loss of structure and functional capacity of skeletal muscle during prolonged bed rest. This aim will include measures of whole muscle size and function (magnetic resonance imaging/spectroscopy), functional exercise testing (strength and endurance), biochemistry (enzyme activities, ubiquitin proteasome pathway induction), and histology (muscle fiber type and morphometry, and capillary density).

This proposal addresses the high priority area of integrative physiology, by testing a countermeasure at readiness level (CMRL) 6. The hypotheses and specific aims of the project will address Bioastronautics Critical Path Roadmap Risks #s 1,4,5,6,13,14, and 23, and critical questions 1e,f, i;4b; 5e,j;6a d,f h,k m,o,p;13b d,g k,p r,t;and 23a,b.

Craig Crandall, Ph.D.
Effects of Hyperthermia on Human Baroreflex Function (NIH)
The effects of hyperthermia on human baroreflex control of blood pressure are unknown. Hyperthermia increases sympathetic activity in humans evidenced by increases in cardiac output, heart rate, splanchnic and renal vascular resistances, and muscle sympathetic nerve activity. Since baroreceptor control of these variables is explained by a sigmoidal relationship between changes in these efferent variables relative to changes in blood pressure, the functional reserve to further increase these variables during a hypotensive challenge will be reduced in hyperthermia if the respective baroreflex curves are not adjusted. Thus, the following hypothesis will be tested: hyperthermia alters baroreceptor control of blood pressure in humans. Studies have shown that hyperthermia attenuates a-adrenoceptor responsiveness in both whole animals and isolated vessels. Parallel studies have not been conducted in humans. If a-adrenoceptor responsiveness in humans is likewise attenuated in this environment, baroreceptor adjustments to hyperthermia will be less effective in maintaining pressure since end organ vascular responses will be attenuated. Therefore, studies will be conducted to test the hypothesis that hyperthermia decreases a-adrenoceptor responsiveness in humans. A primary function of the baroreflex is to maintain blood pressure to adequately perfuse the cerebral circulation. The cerebral circulation has a wide autoregulatory range in which large changes in perfusion pressure result in no change in cerebral blood flow. This curve shifts to higher pressures during sympathetic stimulation, which effectively increases the lower limit of cerebrovascular autoregulation to higher pressures. As previously mentioned hyperthermia increases sympathetic activity, and therefore may increase the lower limits of cerebrovascular autoregulation to high perfusion pressures. Such an occurrence will predispose the individual to syncope during a hypotensive challenge. Thus the following hypothesis will be tested: hyperthermia shifts the cerebrovascular autoregulatory curve resulting in impaired autoregulation of cerebral blood flow to decreases in perfusion pressure. To address these issues integrated and individual baroreflex functions will be assessed in normothermia and hyperthermia as will a-adrenoceptor responsiveness in both skin and muscle through intra-arterial and local administration of a-adrenoceptor agonists. Finally steady state and dynamic cerebrovascular autoregulation will be assessed in normothermia and hyperthermia using transcranial Doppler. Following the completion of this work important information will be provided regarding the effects of hyperthermia on blood pressure regulation in humans.

Craig, Crandall, Ph.D.
Mechanisms of Skin Cooling to Improve Orthostatic Tolerance (NASA)
Post-space flight orthostatic hypotension/intolerance occurs in 25 to 66% of crew members upon returning to a 1 G environment. The mechanism(s) causing this response are not completely understood. Identification of countermeasures to reduce the incidence of orthostatic intolerance associated with space flight is paramount to NASA's mission. One such countermeasure may be skin surface cooling. In light of this, three specific objectives will be accomplished by the proposal work: 1) Identify an optimal skin surface cooling paradigm that causes the largest increase in autonomic responses (i.e. stroke volume, blood pressure, sympathetic nerve activity, etc.) without causing shivering or altering motor function. 2) Identify the mechanisms by which skin surface cooling increases the aforementioned autonomic responses resulting in improved tolerance to orthostatic stress. 3) Identify whether skin surface cooling is an effective countermeasure to improve orthostatic tolerance in men and women following simulated microgravity exposure using the head-down tilt bed rest model. Upon completion of the proposed studies important information will be provided that will be beneficial for both operational and safety concerns for astronauts, as well as to individuals who suffer from idiopathic orthostatic intolerance.

Benjamin Levine, M.D.
Aging, Fitness, and Failure: Mechanisms of Diastolic Dysfunction (NIH)
Aging is associated with alterations in left ventricular (LV) relaxation and compliance, even in the absence of manifest co-morbid conditions such as coronary artery disease or hypertension. This "diastolic dysfunction" may result in an elevation of LV filling pressure and cause signs and symptoms of congestive heart failure (CHF) despite the presence of preserved systolic function. This problem is critical for elderly CHF patients in whom nearly 1/2 may have predominantly diastolic dysfunction as the cause of their heart failure. The broad objective of this proposal is to determine the precise alterations of diastolic function associated with normal, healthy aging in humans, and compare these changes with those associated with CHF due to diastolic dysfunction. The hypotheses to be tested include: 1) Normal aging is associated with alterations in both relaxation and chamber compliance of the LV leading to impaired ventricular filling compared to healthy young adults. Moreover, such abnormalities will be substantially reduced in fit compared to sedentary healthy elderly subjects; 2) Patients with CHF and preserved LV systolic function have more pronounced abnormalities of LV chamber compliance and relaxation which contribute to the development of CHF; 3) Exercise training in both the healthy aged, as well as patients with CHF and diastolic dysfunction will improve abnormalities of diastolic function and will be an effective therapy for this disease. To test these hypotheses, we propose to accomplish the following specific aims: a) To measure the static component of diastole directly by constructing ventricular function (Starling) and LV pressure/volume curves in well, sedentary and fit elderly adults. The dynamic component of diastole will be assessed using state-of-the-art imaging techniques including: tissue Doppler imaging; color Doppler M-mode echo; and magnetic resonance imaging with myocardial tagging. b) To select a group of patients with CHF but preserved systolic function, and to quantify left ventricular relaxation and compliance using the same methods; c) To repeat the specific measures of diastolic function after a prolonged (one year) endurance exercise training program in both the sedentary elderly, and patients with CHF and diastolic dysfunction. These studies will result in a comprehensive understanding of the effect of normal aging and physical conditioning on LV diastolic function, and will identify the specific abnormalities of diastole which lead to CHF in the absence of contractile dysfunction. The precise dose of exercise necessary to restore normal diastolic function will be identified and will allow specific exercise prescription for these populations

Benjamin Levine, M.D.
CARDIAC ATROPHY AND DIASTOLIC DYSFUNCTION DURING AND AFTER LONG DURATION SPACEFLIGHT: FUNCTIONAL CONSEQUENCES FOR ORTHOSTATIC INTOLERANCE AND RISK OF CARDIAC ARRHYTHMIAS (NASA)
Cardiac atrophy appears to develop during spaceflight or its ground based analogues, leading to diastolic dysfunction and orthostatic hypotension. Such atrophy also may be a potential mechanism for the cardiac arrhythmias recently identified in some crew members after long duration exposure to microgravity aboard the Mir space station. Recent work by the PI has suggested that cardiac atrophy may be progressive, without a clear plateau over at least 12 weeks of bed rest, and thus may be a significant limiting factor for extended duration space missions. The global objective of this proposal is to quantify the extent and time course of cardiac atrophy and identify its mechanisms. The functional consequences of this atrophy also will be determined for cardiac filling dynamics, orthostatic tolerance, and arrhythmia susceptibility both in space on the International Space Station, and following return to earth. Three specific aims will be addressed: 1) To determine the magnitude of cardiac atrophy associated with long duration spaceflight, and to relate this atrophy to measures of physical activity and cardiac work in flight. Magnetic resonance imaging will be performed pre-and post-flight as the most accurate means of measuring cardiac mass, and cardiac ultrasound will be performed in-flight to determine the time course and pattern of progression of atrophy in space; 2) To determine the functional importance of this atrophy for orthostatic tolerance and the regulation of stroke volume by using a combination of classical, invasive cardiovascular physiology to measure the static component of diastole, in conjunction with novel, non-invasive imaging techniques to measure the dynamic component of diastole; 3) To identify changes in ventricular conduction and repolarization during and after long duration spaceflight, and relate these to changes in cardiac mass. After completion of this study, the clinical manifestations of cardiac atrophy during long duration space flight will be defined clearly, and its significance for diastolic function and orthostatic tolerance will be elucidated, thus supporting the application of specific countermeasures currently being developed by the PI in parallel ground based experiments. Information will be obtained regarding ventricular conduction and repolarization that may provide insight into the risk for cardiac arrhythmias. The information obtained from these spaceflight experiments also will be relevant for patients after prolonged confinement to bed rest, as well as conditions that alter cardiac stiffness such as congestive heart failure, ischemic heart disease, and normal aging.

Rong Zhang, Ph.D.
Dynamic Cerebral Autoregulation in Hypertension (American Heart Association)
Specific aims: Chronic hypertension causes both structural and functional changes in cerebral circulation and is an important risk factor for stroke. However, little is known about dynamic regulation of cerebral blood flow in hypertensive patients. This project is to explore:
1) Whether dynamic cerebral autoregulation is altered in essential hypertension;
2) 2) whether altered cerebral hemodynamics in patients with mild to moderate hypertension is reversible with antihypertensive therapy; and
3) 3) what role sympathetic nerve activity plays in dynamic cerebral autoregulation in hypertension.
Methods: Dynamic cerebral autoregulation will be evaluated in both controls and in patients with mild to moderate hypertension before and after antihypertensive therapy. Effects of sympathetic nerve activity on cerebral circulation will be evaluated under orthostatic stress and by ganglionic blockade. Changes in cerebral blood flow will be measured by transcranial Doppler in the middle cerebral artery with simultaneous recording of systemic pressure. Dynamic cerebral autoregulation will be quantified by using both the linear transfer function and the non-linear Volterra model method for analysis of beat-to-beat changes in cerebral blood flow velocity and arterial pressure.
Objectives: Upon successful accomplishment of this project, the following specific questions will be answered: 1) does the autoregulatory effects of cerebrovascular bed on oscillations in cerebral blood flow reduce with severity of hypertension? 2) are the changes in cerebral hemodynamics in essential hypertension reversible with antihypertensive therapy? 3) does the augmented sympathetic activity in hypertensive patients contribute to changes in cerebral autoregulation?

Ronald G. Haller, M.D.
Evaluation and Treatment of Metabolic Myopathies (Muscular Dystrophy Association and VA Merit Review)
The long term objectives of these studies are to identify optimal ways of managing patients with hereditary disorders of muscle energy metabolism, including patients with mitochondrial myopathies and those with disorders of muscle glycolysis/glycogenolysis. We will attempt to improve energy production relative to energy demand and reduce symptoms of exercise intolerance by augmenting the metabolic capacity of oxidative energy pathways in skeletal muscle by exercise training or by utilizing supplements of fuels or cofactors that bypass the metabolic block or that increase the capacity of alternative energy pathways.
The specific aims are: 1) to evaluate the efficacy of aerobic conditioning in increasing levels of rate limiting enzymes and in improving functional capacity in patients with mitochondrial myopathies attributable to nuclear gene defects, including patients with carnitine palmitoyltransferase II deficiency; 2) to investigate whether treatment with succinate + riboflavin in patients with selective complex I defects or vitamin K + vitamin C in patients with selective complex III defects will effectively bypass the metabolic block; and 3) in McArdle disease and muscle PFK deficiency we will investigate whether supplements of pyruvate, alanine, or lactate will augment aerobic power by increasing the availability of pyruvate for oxidative metabolism and whether supplements of creatine will increase anearobic capacity by increasing muscle availability of phosphocreatine.
Exercise training will consist of sustained (30-40 minutes) moderate (eliciting 60-70% of maximal heart rate) exercise performed at least 4 times per week for 14 weeks. Oxidative capacity will be evaluated utilizing cycle exercise monitoring gas exchange, oxygen transport (cardiac output), oxygen extraction (arteriovenous O2 difference), and levels of blood lactate and pyruvate. Muscle adaptation to training will be evaluated by comparing the level of oxidative and related enzymes in needle biopsy samples obtained before and after training. Nutritional supplements will be administered utilizing double blinded, placebo-controlled methodology. Effectiveness will be evaluated using 31P magnetic resonance spectroscopy to evaluate high energy phosphates in conjunction with arm and leg exercise protocols to evaluate strength, endurance, and metabolic capacity.

Ronald G. Haller, M.D.
Muscle Glycolytic Enzyme Deficiency - Metabolic and Physiologic Effects
The cellular mechanisms by which impaired glycolytic energy production produces muscle weakness, fatigue and injury are poorly understood. Impaired anaerobic glycolysis (substrate-level phosphorylation) is the mechanism of energy limitation usually invoked to explain these symptoms. However, previous studies from our laboratory indicate that severe blocks in glycogenolysis/glycolysis results in substrate-limited oxidative phosphorylation. The goals of this application are 1) to define the level of impaired glycolysis necessary to impair oxidative metabolism; 2) to characterize the biochemical mechanisms by which impaired glycogenolysis limits muscle oxidative phosphorylation; and 3) to define the clinical and physiological consequences of pyruvate-dependent oxidative metabolisms with emphasis on effects on muscle blood flow in exercise.
We propose to study human genetic errors of muscle glycogenolysis and experimental inhibition of glycolysis using metabolic inhibitors in the rate to elucidate the sequence of biochemical and physiologic abnormalities that accompany progressive limitations of glycolytic substrate-level and oxidative phosphorylation. We anticipate that progressive reductions in glycolytic enzyme activity will result in : 1) compensated glycolytic impairment in which glycolytic flux is maintained by increases levels of glycolytic activators (e.g. ADP Pi); 2) limitation of glycolytic flux in which substrate -level phosphorylation and ischemic work capacity are impaired, but pyruvate dependent oxidative metabolism and aerobic exercise capacity are preserved; 3) and marked restriction of glycolytic flux associated with impaired pyruvate-dependent oxidative phosphorylations and marked limitations in aerobic exercise capacity. We further hypothesize that severe restrictions in glycolytic flux impair muscle oxidative metabolism by limiting flux in the tricaboxylic acid (TCA) cycle primarily by limiting pyruvate-dependent anaplerosis to expand the pool of 4 carbon TCA intermediates necessary to 'spark' maximal rates of TCA cycle flux.
Human subjects to be studies include; carriers of muscle phosphorylase and muscle PFK deficiency identified by molecular genetic analysis; patients with complete phosphorylase deficiency; patients with variant phosphorylase deficiency in whom residual enzyme activity remains; and patients with partial glycolytic blocks due to distal defects in glycolysis. In rats, we will assess the level of iodoacetate- mediated glycolytic inhibition necessary to impair pyruvate-dependent oxidative phosphorylation, the cellular expression of substrate-limited oxidative metabolism, and the ability of alternative substrates to repair the metabolic deficit.
Metabolic responses will measured during forearm and leg exercise utilizing 31P magnetic resonance spectroscopy, venous effluent metabolites, muscle oxygenation as assessed utilizing near infrared spectroscopy; and by evaluating metabolite accumulation in needle biopsies. In rate experiment, the effect of iodoacetate-inhibited glycolysis on oxidative metabolism and the relative ability of acetate and lactate to repair the metabolic block will be assessed in resting and stimulated gastrocnemius muscle in which oxidative and anaplerotic incorporation of substrate in the TCA cycle will be monitored by measurement of metabolites in freeze clamped muscle and by an 13C magnetic resonance spectroscopy.

Tony G. Babb, Ph.D.
Human Aging: The ventilatory response to even mild exertion is the largest challenge to breathing faced during activities of daily living in humans. In the aged, the exercise ventilatory response to exercise is increased although breathing capacity progressively declines with aging. This dilemma between increased ventilatory demand and decreased ventilatory capacity is marked in the very old, which is one of the most rapidly increasing segments of our population today. Moreover, dyspnea during exercise is one of the most common complaints of the elderly and often prevents older adults from obtaining the exercise they need to maintain a high quality of life. My research in this area has focused on defining the limitations imposed by aging on resting pulmonary function, the exercise ventilatory response, respiratory mechanics during exercise, and exercise tolerance (i.e., age-related limits on expiratory flow, lung volume, and breathing pattern). These investigations have not only extended our basic knowledge of respiratory and exercise physiology in the aged, but have had broad clinical implications on the evaluation of ventilatory limitations during exercise in young and old patients with respiratory limitations or unexplained shortness of breathing during exertion.

As a result of the above studies in the aged, I have extended my focus of research from studying mechanical ventilatory limitations (i.e., limits to function of the respiratory pump) during exercise to a more basic line of investigations focused on age-related changes in respiratory control during exercise. Animal work on respiratory control mechanisms has demonstrated both short and long term modulation of breathing during exercise, which integrates perfectly with my previous findings in aged humans. Briefly, the fundamental goal of this translational work is to investigate the capacity for modulation and plasticity of the exercise ventilatory response in normal and aging human adults, which has never before been tested in humans. Furthermore, I propose to test the hypothesis that short term modulation of the exercise ventilatory response in humans (as in goats) is serotonin-dependent and that increasing central serotonergic function can enhance short term modulation, particularly in the elderly. Preliminary data in these regards are quite exciting. Furthermore, these findings may be extended to studies of several important patient populations where breathing regulation is altered at rest and during exercise (i.e., obesity, lung disease, heart failure patients).

Human Obesity: Obesity is an epidemic problem in the US and is among the most important health challenges of the 21st Century (NIH Obesity Research Report 2004 and Surgeon General's Call to Action, USPHS 2001). Approximately 32% of Americans 20-74 yr are overweight (25 < BMI > 30) and 23% are frankly obese (BMI > 30). Thus, an astounding 55% of Americans are overweight or obese. The major consequences of obesity are increased rates of mortality and morbidity from diabetes, heart disease, hypertension, and metabolic syndrome. Many obese adults experience shortness of breath on exertion and are unable to exercise. However, regular exercise is a major factor in the prevention and treatment of obesity as well as the management of many of its comorbidities. Thus, respiratory limitations, exertional dyspnea, and exercise intolerance is an immensely important national concern. Our studies have addressed these important, but poorly understood and understudied issues. As part of this work we have developed an MR imaging technique in collaboration with the Rogers Center at UT for determining fat distribution in humans, which has increased our ability to investigate the relationships between respiratory limitations, exertional dyspnea, and fat distribution. Furthermore, we have recently initiated a study examining the mechanism of exertional dyspnea in obese adults, which appears to be highly related to the work of breathing and fat distribution.