Robert Linsenmeier - US grants

The oxygen supply to the retina is crucial in maintaining visual function, but retinal oxygenation is often disturbed by vascular occlusion or disease. Management of these conditions could conceivably profit from knowledge of the normal oxygen supply, the changes that occur with vascular occlusion, and the potential for restoring oxygenation. The proposed work is designed, therefore, to further the basic knowledge of oxygen supply and consumption within the mammalian retina. Retinal oxygen supply is complex in higher mammals, since two circulatory beds nourish the retina. Measurements of vitreal oxygen tension have been made previously, but these do not allow an accurate prediction of the oxygen tension in the outer part of the retina, around the photoreceptors. The next step in understanding retinal oxygen supply is therefore to make intraretinal measurements in a mammal, the cat, that has a dual retinal circulation. The specific aims are to 1) define the normal profile of oxygen tension as a function of depth in the retina, 2) determine how this profile changes as a function of light adaptation and retinal eccentricity, 3) use this experimental data to derive information concerning oxygen consumption as a function of depth, 4) determine how the profiles and retinal function change during graded hypoxia, and 5) develop a mathematical model of retinal oxygenation applicable to disturbances of retinal oxygen supply. To accomplish these goals, intraretinal oxygen tension will be measure with oxygen microelectrodes, and retinal function will be assessed with the electroretinogram recorded intraretinally. Data obatined from the experimental measurements will be used in conjunction with diffusion theory to develop a mathematical model of oxygen supply. In the long term this work could be extended to the primate retina, and could contribute to an understanding of the neovascularization that occurs in diabetes and retrolental fibroplasia.

The experiments proposed in this application are designed to investigate the micro-environment of the intact retina anesthetized cats with regard to the normal delivery and utilization of oxygen, and the consequences and possible treatment of imppaired oxygen delivery. Local measurements of oxygen, glucose and blood flow will be made with microelectrodes in the retina or in the vitreous near the retina. The electroretinogram will be recorded as an index of the functional state of the retina. In other experiments choroidal blood flow will be recorded. The advantage of this preparation is that normal circulation and physiological relationships can be preserved. The project has several portions: 1) The influence of neural (sympathetic and parasympathetic) regulation of the choroidal circulation on oxygenation of the distal retina will be determined. 2) A glucose-sensitive microelectrode capable of measuring glucose concentration in the distal retina will be developed in order to evaluate the contribution of glycolysis to energy metabolism in the intact retina, and to study changes in glycolysis during light and dark adaptation and during hypoxemia. 3) A technique for measuring local retinal blood flow, using hydrogen as a tracer, will be developed and used to study the mechanisms responsible for the normal regulation of blood flow in the microvasculature of the retinal circulation. 4) In vitro studies will be performed in order to understand the mechanisms by which perfluorocarbon artificial blood substitutes are capable of increasing retinal oxygen tension. 5) A new experimental method to study retinal oxygenation during reversible retinal occlusion will be developed and used. 6) The consequences of long term experimental hypoxia (moderate elevation of intraocular pressure) on retinal metabolism and electrical function will be studied, as a means of assessing the role that retinal hypoxia itself may play in disease. These studies are expected to be relevant for several diseases with circulatory components, including vascular occlusive diseases, glaucoma and others in which a metabolic component is thought to be important, such as diabetic retinopathy. The cat will be the animal of choice for these studies, because of the similarity of its retina and its retinal vasculature to those in humans. For this work, there would be no significant advantage in using primates.

[unreadable] DESCRIPTION (provided by applicant): The long-term objectives of this work are to understand aspects of the retinal microenvironment related to oxygen and pH, and how these relate to energy metabolism and function of the mammalian retina in vivo. While this work will be done in animals, it is particularly relevant to blinding diseases that affect the relationships between the circulation and retinal neurons in humans. During the next project period, our main interests are in diabetic retinopathy, retinal detachment, and retinal arterial occlusive disease. However, the results will also provide fundamental information that may be relevant to other types of retinal dysfunction. The proposed work will be done primarily on intact anesthetized cats, since their retina provides a good model for much of the human retina. The techniques are primarily to use oxygen and pH sensitive microelectrodes to map out retinal oxygen levels, pH and electrical activity (the electroretinogram) with high spatial and temporal resolution, as we have done previously under other experimental conditions. Following the measurements, mathematical modeling of diffusion will be used to extract metabolic parameters that are not apparent from the measurements alone, and to perform simulations of situations that may not be amenable to experimentation. Some measurements of retinal histology will also be made. The project has 5 specific aims. 1) We will study intraretinal oxygenation following photocoagulation, because the mechanism by which photocoagulation blocks neovascularization is still unclear. 2) We will use information about oxygenation after photocoagulation from specific aim 1 to create an appropriate two-dimensional diffusion model of this situation, with the hope of providing a better rationale for the density and size of lesions designed to treat retinopathy. 3) We will study retinal oxygenation in the detached retina in order to understand the basis for the protective effect of hyperoxia in retinal detachment, which has been shown recently in cats. 4) We will study pH after retinal arterial occlusion, to understand the potential role of acidosis in damaging the retina. 5) We will investigate the influence of anesthesia on the metabolic measurements we make, and will study metabolic differences between the cat and primate retinas.

TThe objective of this engineering education project is to advance both basic and applied understanding of how to prepare engineering graduates to effectively and efficiently contribute to America's leadership in technological innovation. The PIs propose to study the role that computational and analytical abilities play in innovation in the context of a conceptual framework that has recently emerged in the engineering education literature: adaptive expertise.

Adaptive expertise is an emerging area of research on learning that has shown promise in providing enhanced understanding of transfer of knowledge issues. It is a critical area of research that directly relates to U.S. global competitiveness through improving understanding of what is required to train innovative and efficient problem solvers who can transcend narrow disciplinary fields. The PIs plan to perform fundamental research with the intent to inform the practice of teaching and learning. They hypothesize that the general model of adaptive expertise can be applied specifically to characterize the attributes of efficiency and innovation in the context of developing CADEX. Therefore, they plan to focus on basic research to understand the nature of CADEX and to define the efficiency and innovation axes in terms of the underlying cognitive or affective attributes of each. This approach allows them to uncover how innovation and efficiency may be conceived more broadly, while at the same time enables us to define terms specifically in the context of CADEX. The advantage of combining fundamental and applied research is that the PIs not only identify cognitive aspects of computational adaptive expertise and potentially effective instructional strategies, they will also verify and test their findings in actual educational settings.

The research will benefit society by providing recommendations for instructional and assessment strategies to develop competencies required for technological innovation. These are essential skills in today's competitive economy and thus will enhance the United States' industrial competitiveness by producing graduates poised with cognitive strategies for developing efficient and effective design solutions.

CBET-0757969
Linsenmeier

This project will support the 3rd Biomedical Engineering Summit Meeting will be held in June 15-17, 2008 and will be the first conference focusing on biomedical engineering education since 2005. The Whitaker Foundation, which funded the first two summits, is no longer available to carry on this work. The Council of Chairs of Biomedical Engineering and Bioengineering (CoC) and the Biomedical Engineering Division of the American Society for Engineering Education (ASEE BED) are cosponsoring this third summit.

Biomedical engineering continues to develop as an undergraduate major. This conference will bring together a diverse group for an extended discussion of best practices. The 3rd Biomedical Engineering Summit Meeting is designed as a working meeting to foster progress in biomedical engineering education. Attendance will be limited, but will be open to all current and emerging biomedical engineering and bioengineering academic programs in the US (about 100 programs; two individuals per institution), and will include representatives from industry and the field of learning science. Minority serving institutions will be particularly encouraged to attend, and selected international universities will also be invited. The conference objectives are to 1) share best practices in curriculum and pedagogy, 2) nurture and expand a community in which relevant individuals across the country and around the world know each other and their educational work, 3) exchange ideas about curriculum in a setting where biomedical engineering education rather than research takes center stage, 4) assist newer programs using knowledge gained by more mature programs, and refresh the ideas of established programs, 5) enhance the likelihood that minority serving institutions will adopt biomedical engineering as a major, and 6) discuss the core curricula and areas of specialization that will enable biomedical engineering students to best prepare for industry, medicine, or graduate study.

[unreadable] DESCRIPTION (provided by the applicant): The Third Biomedical Engineering Summit Meeting will be held in June 2008 and will be the first conference focusing on biomedical engineering education since 2005. The Whitaker Foundation, which completely supported the first two summits, is no longer available as a sponsor, and the Council of Chairs of Biomedical Engineering and Bioengineering (CoC) and the Biomedical Engineering Division of the American Society for Engineering Education (ASEE BED) are cosponsoring this third summit. The conference is designed as a working meeting to foster progress in biomedical engineering education at the undergraduate level in a way that is not possible at any other meeting. Attendance will be limited, but will be open to all current and emerging biomedical engineering and bioengineering academic programs in the US (two individuals per institution), and will include representatives from industry and the field of learning science. Minority serving institutions will be particularly encouraged to attend, and selected international universities will also be invited. The conference objectives are to 1) share best practices in curriculum and pedagogy, 2) nurture and expand a community in which relevant individuals across the country and around the world know each other and their educational work, 3) exchange ideas about curriculum in a setting where biomedical engineering education rather than research takes center stage, 4) assist newer programs using knowledge gained by more mature programs, and refresh the ideas of established programs, 5) enhance the likelihood that minority serving institutions will adopt biomedical engineering as a major, and 6) discuss the core curricula and areas of specialization that will enable biomedical engineering students to best prepare for industry, medicine, or graduate study. [unreadable] [unreadable] PUBLIC HEALTH RELEVANCE STATEMENT (provided by applicant): Biomedical engineers are an important part of the workforce that contributes to advances in the increasingly technological field of healthcare in the US and abroad. This conference is designed to foster progress in biomedical engineering education at the undergraduate level. The conference will bring together individuals from academia and industry to discuss the ways in which students can be best prepared with the content knowledge, skills and attitudes to be innovative and adaptive in biomedical engineering careers. [unreadable] [unreadable] [unreadable]

Proposal: 0851930
PI Name: Robert A. Linsenmeier

"This award is funded under the American Recovery and Reinvestment Act of 2009
(Public Law 111-5)."

This three year REU site award will provide an opportunity for undergraduate students to do research focused in the area of bioengineering education. It will build upon the practices established during a successful REU program at the Vanderbilt-Northwestern-Texas-Harvard/MIT (VaNTH) Engineering Research Center for Bioengineering Educational Technologies. Mentors for this REU site program will be selected from Northwestern University, Vanderbilt University, Massachusetts Institute of Technology or the University of Texas at Austin. There are very few opportunities for engineering students to learn about educational pedagogy or to do research in engineering education.

The objectives of the program are: (1) to expose students to quantitative and qualitative aspects of research in engineering education, and to expose them to research methods more broadly; (2) to have each student work with a mentor to perform a project that contributes to the development of curricular materials for bioengineering using learning science principles, and/or understand how to evaluate educational innovations for there impact on student learning; and (3) to contribute to real ongoing research projects. Students may also work on middle school and high school outreach projects involving biomedical engineering concepts. In addition to the research projects, students will receive training in communications and ethics in the context of research and advice about preparing for and applying to graduate school.

The Principal Investigator will recruit undergraduate students from bioengineering programs, other fields of engineering and science, or education. Special emphasis will be placed on the recruitment of minority students to enhance the possibility that such students will become excited about research and be motivated to pursue graduate study.

DESCRIPTION (provided by applicant): Diabetic retinopathy is one of the most common causes of retinal disease in adults. Many molecular and histological changes have been demonstrated, and it is now recognized that neural as well as vascular problems exist. The most important contributor to vascular dysfunction is the elevation of vascular endothelial growth factor (VEGF), which causes vascular leakage and angiogenesis. Unfortunately, we still do not know what underlies the increase in VEGF. Hypoxia and acidosis are both known regulators of VEGF, but the magnitude and time course of changes in retinal oxygen and pH are not known in diabetes. More information about these parameters will allow a better understanding of disease progression, and ultimately may lead to an understanding of pathways that can be targeted before VEGF causes damage. We will make intraretinal recordings of PO2 and pH in rats with streptozotocin (STZ)-induced diabetes, and in rats with acidosis in the absence of diabetes. To place these changes in the context of other events, we will also measure important vascular parameters (blood flow, leakage, molecular signals). SPECIFIC AIM 1: Oxygen in diabetic retinopathy. Retinal tissue hypoxia has been implicated in the increased VEGF that is found in both the background and proliferative phases of retinopathy. There is considerable indirect support for the existence of hypoxia in diabetic retinopathy, but very limited direct evidence. We will characterize intraretinal oxygenation in rats with up to 1 year of diabetes. Our hypothesis is that inner retinal PO2 will decrease over this time. We will also investigate changes in the regulation of retinal oxygen in diabetics. Our hypothesis is that changes in regulation of PO2 will be a more sensitive indicator of changes in the microenvironment and occur earlier than changes in baseline PO2. We found increased PO2 in the retina in an earlier study at 12 weeks of diabetes. Our hypothesis is that this was due to increased blood flow, which we will now investigate. SPECIFIC AIM 2: Acidosis in diabetic retinopathy. VEGF is regulated by acidosis in some tissues but the role of pH in controlling VEGF in the adult retina is unknown. We will characterize retinal pH in rats with up to 1 year of diabetes. Our hypothesis is that the inner retina will be more acidic than normal for at least a few months. We also hypothesize that molecular mechanisms come into play to reduce acidosis, such as increased carbonic anhydrase and acid removal mechanisms. These will be evaluated with mRNA and protein measurements. SPECIFIC AIM 3: Is retinal acidosis sufficient to upregulate VEGF? Finding that the diabetic retina is acidotic will open the possibility that it is one of the factors that regulates VEGF, but will not tell us whether it is important. Consequently, we will produce acidosis in the absence of diabetes, via carbonic anhydrase inhibition. We will first establish a treatment protocol that yields intraretinal pH values similar to diabetes, and then evaluate changes in VEGF and leakage. Our hypothesis is that acidosis alone will be sufficient to increase VEGF, produce molecular changes that underlie leakage, and cause leakage itself.

This project will study the adaptation, implementation, and dissemination of best practices in experiential learning in the core middle years of the engineering curriculum where students take the bulk of technical fundamentals. The study will involve two series of junior-year fundamental core engineering courses; one taught at Northwestern University's Department of Biomedical Engineering and the other taught at the University of Florida's Department of Electrical and Computer Engineering. The course sequences use similar tools and teach similar topics geared towards building skills needed for success in the engineering workplace or graduate school. The differences are in methods of delivery and the types of experiential learning modules employed.

Using a mixed method approach, the main goals of this proposal are to 1) assess each course sequence in terms of How People Learn and assess how student experiences and outcomes are linked to those attributes of each course and 2) enhance and expand effective experiential learning modules for broader adoption and implementation. The project will evaluate student learning preferences, student engagement, retention of material in subsequent courses in their respective sequences, transferrable skills between courses and learning of course concepts. Both schools have highly selective engineering programs, but the student demographics differ. These differences give the opportunity to consider more demographic factors in assessing the courses and in designing materials to take advantage of diversity. The knowledge gained will guide future design or development of new interventions and their dissemination strategies.

Project Summary This proposal seeks to establish a correlation between the metabolism of the inner retina and the oxygen supply from the inner retinal vascular network measured by optical coherence tomography. This work will be done in rats, but will have immediate clinical application. Several important diseases cause damage to the inner retina, including diabetes, glaucoma, and vascular occlusion. Considerable effort has gone into measuring parameters that provide some information about the metabolism of the inner retina, particularly oxygen saturation (sO2) in the retinal vessels and retinal blood flow (RBF), but each alone is insufficient to allow strong conclusions. It has now become possible in animals and humans to measure saturation and blood flow together and combine them to yield the Inner Retinal Metabolic Rate of Oxygen (IR-MRO2), which is the total amount of oxygen extracted from the retinal circulation per unit time. These non-invasive measurements of IR-MRO2 could potentially be predictive of functional damage in disease before other measures, and could be useful in evaluating treatments for those diseases. The most promising way to measure IR-MRO2 is a novel method using visible-light optical coherence tomography (vis-OCT). However, the retina is a unique tissue with a dual blood supply, so further validation is required to understand how IR-MRO2 corresponds to the actual oxygen utilization of the inner retina (IR-QO2). It is often assumed that these two quantities are equal, but this is not true. There are conditions under which the retinal circulation provides an important component of the oxygen needed by the photoreceptors in the outer retina, so IR-MRO2 overestimates IR-QO2, and other conditions under which oxygen derived from the choroid provides oxygen to the inner retina, so that IR-MRO2 underestimates IR-QO2. If the actual correspondence between IR-MRO2 and IR-QO2 is not understood, the great promise of the new methods of measuring IR-MRO2 will be lost due to confusion and false conclusions. It is really IR-QO2 that is needed for understanding the progression of inner retinal changes in diabetes, glaucoma, and vein occlusion. We propose to perform experiments on the same rats with both well-established oxygen-sensitive microelectrode methods and vis-OCT to identify the conditions under which IR-MRO2 can be used to infer IR-QO2 directly, and to provide a clinically useful model to allow a prediction of the deviations between IR-MRO2 and IR-QO2. No study has previously used both imaging and microelectrodes in the same animals, or attempted to provide validation of oximetry methods with other methods. The proposed work is important for any method of characterizing IR- MRO2, not just vis-OCT. The specific aims are 1) to develop an analytical model to relate the inner retinal sO2 and IR-MRO2, measured by vis-OCT, to the metabolic demands of the inner and outer retina in both rats and humans and 2) to validate and refine this model in rats by nearly simultaneous measurement of IR-MRO2 and oxygen consumption derived from microelectrode measurements during air breathing in light and darkness, hypoxia and hyperoxia.

Abstract Understanding retinal metabolism in detail is fundamental to knowing how the retina withstands, compensates for, or suffers from the stresses imposed by systemic changes and disease. This application takes a new approach to understanding glycolytic and oxidative metabolism of the mammalian retina, by recording spatial profiles of oxygen and pH in the isolated rat and mouse retina with microelectrodes, using mathematical models of diffusion to extract information about rates of substrate utilization and waste generation, and employing pharmacology to isolate different processes. This strategy is complimentary to previous in vivo and in vitro approaches, but allows a separation of metabolic events in the inner and outer retina, which has rarely been possible, and cannot be done with measurements of whole tissue metabolism. Microelectrode approaches simultaneously allow recording of transretinal and intraretinal electroretinograms to monitor retinal function. There are three aims. 1) Depth profiles of oxygen will be recorded in order to determine quantitatively how the metabolism of the isolated retina compares to that in vivo, and how the inner and outer retina differ metabolically. Photoreceptors are known to perform high rates of (anaerobic) glycolysis, but the balance between oxidative and glycolytic energy production is not known in the inner retina, and whether this changes depending on glucose supply. Retinal blood flow increases in response to flickering light (neurovascular coupling), but the size of the metabolic change in the inner retina that drives this is unknown, and will be measured here. 2) By recording depth profiles of pH in the isolated retina, the production of acidic waste will be quantified. Glycolysis and oxidative metabolism are very different in acid production. To identify the components of acid production, hypoxia and metabolic poisons that suppress either glycolysis or oxidative metabolism will be used. 3) In some tissues there is evidence that glycolysis and oxidative metabolism are compartmentalized, with a transfer of lactate and/or pyruvate from glycolytically active cells to ones that depend more on oxidative metabolism. This concept has led to the idea that the retina uses a similar strategy, with lactate being produced in Muller cells and shuttled to neurons. The relatively recent availability of selective blockers of monocarboxylate transport, provides an opportunity to evaluate the importance of such transfer quantitatively in both the inner and outer retina. All the information to be gained is fundamental to understanding how the retina changes in disease, and what the capabilities of the inner and outer retina are for oxidative and glycolytic metabolism under different conditions.