Timothy S. Moerland - US grants

A cellular/biochemical study of diabetic myopathy in fast and slow twitch skeletal muscle or rats will be conducted over five years. Insulin dependent diabetes will be induced in WKY rates by injection of streptozotocin. The specific objectives are to (1) quantify the functional capacity of the fast-twitch extensor digitorum longus (ed1) and slow-twitch soleus muscles of both acutely diabetic (4 days after treatment) and long- term diabetic rats (>5 weeks) to generate and maintain tension, and measure the specific energetic costs of contractile activity. The energy cost of contraction will be measured by determining oxygen consumption and lactate evolution after tetanic stimulation, and the time-course of aerobic recovery metabolism will be determined. Maximum twitch and tetanic force will be measured. (2) Determine the nature and extent of changes in the myosin isoenzyme composition of edl and soleus of diabetic rats. Myosin isoenzymes will be analyzed by nondenaturing and SDS gel electrophoresis. (3) Determine the relative capacity of muscle from diabetic rats to maintain intracellular concentrations to controls. In vivo and in vitro experiments will use 3lP-nuclear magnetic resonance to monitor changes in high-energy phosphates during graded consequence of motoneuron atrophy. Diabetic neuropathy will be treated with aldose reductase inhibitors and by dietary supplementation of myo-inositol. Conduction velocity of the sciatic nerve will be measured and correlated with the extent of diabetic myopathy in the soleus and edl of treated rats, as determined by measurements of contractile function. Results of the proposed studies will provide information that could prove useful for designing effective treatments for the prevention of diabetic myopathy in humans. This information also should contribute to our basic understanding of the mechanisms and functional significance of normal adaptation in muscle.

This is a proposal to refurbish and upgrade an NMR spectrometer system to be used for high resolution multinuclear studies of liquids. The spectrometer will be used for a wide range of studies including metabolism in marine invertebrates, molecular conformation in gramicidin, and interaction of metals with organic polymers. A new magnet will be obtained that has higher field, better homogeneity, and superior stability than the previous system. The spectrometer console will be rebuilt to accommodate the different resonant frequency of this new magnet. This will be a very cost effective way to provide the required instrumental capacity for these studies.

9808120 Moerland This three year project by Dr. Moerland is a comprehensive study of the effects of temperature on intracellular diffusion, which is a process of fundamental importance to all cells. Diffusion of small water-borne molecules, including those with key roles in energy metabolism, will be examined in muscle by PFG-NMR, a technique that is closely related to clinical magnetic resonance imaging. The sensitivity of diffusion to naturally relevant changes in temperature will be measured. Diffusion measurements will be compared with the internal structure of muscle cells, as measured by electron microscopy, to identify those feature of the cellular architecture that determine patterns of intracellular diffusion. Experiments will be performed on select muscles that represent a range of cell structure from fish and invertebrates. This project will make a threefold contribution: First, it will contribute to our basic understanding of the effects of temperature on intracellular diffusion in all cell types. Second, it will contribute to our overall understanding of the mobility of energetically important compounds in muscle. Third, it will provide fundamental information about the structural properties of the living cytoplasm. Further, this project will contribute to the training and development of scientific investigators, including undergraduate and graduate students.

Most cells are small, with dimensions <100mm along the shortest axis. Small size promotes a high surface area to volume ratio and short intracellular diffusion distances, both of which are thought to be necessary design features of cells. However, cells of some organisms are nearly 10-fold larger than the norm. In some crustaceans, muscle fibers from juvenile animals have "normal" dimensions (<100mm), but as the animals grow, some of these fibers may exceed 600mm. Attaining such a large size while maintaining function should be difficult or impossible, which raises the question: What are the rules and tradeoffs that govern cell size?
The effects of developmental increases in cell size will be examined in isolated fast-twitch muscle fibers of the blue crab, Callinectes sapidus. Size-associated changes in muscle structure, metabolism, and contraction will be examined using methods that include electron microscopy, chemical assays, nuclear magnetic resonance and polarographic measurements of oxygen consumption. In addition, a reaction-diffusion mathematical model will be generated to aid interpretation of experimental data. This study constitutes a novel view of cellular energetics that is broadly applicable. Once the rules are established for an extreme model system, the approach can be applied to traditional models. This will be particularly useful for studying the impact of energetic challenges like hypoxia, exercise regimes and developmental processes, all of which are inherently linked to cell size.
This is a collaboration of biologists and chemical engineers. The lead institution (UNCW) emphasizes undergraduate education, and undergraduate students will work alongside graduate students and the PIs. The research environment at the lead institution will be enhanced by the project's multi-disciplinary approach, and students will benefit from facilities and expertise at the collaborating institution. At the collaborating institution (FSU), the work will also involve training of undergraduate and graduate students. Mathematical modeling will form the basis for the dissertation of at least one graduate student, and undergraduate participation will be sought for all phases of the project.