1990 — 1992 |
Stollberg, Jes |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Mechanisms of Acetylcholine Receptor Clustering @ University of California Irvine
Localization of cell-surface components to synaptic regions is an important problem in developmental neurobiology. It has been shown that lateral migration of diffusely-expressed receptors makes a major contribution to receptor clustering at the developing neuromuscular junction. As electric fields offer a convenient means to induce lateral migration of membrane components, their use constitutes a useful experimental system with which to study receptor localization. In particular, cultures of spherical muscle cells from the African frog Xenopus laevis have been shown to cluster receptors in response to such fields. The clustering continues after the field has been terminated, and is specific for acetylcholine receptors. These observations render this a powerful model system with which to probe the mechanisms underlying receptor clustering, because i) the cultured cells form receptor clusters rapidly in response to precisely measurable stimuli, ii) experiments can be conducted in the absence of neurites, cell-substrate contacts, or exogenous factors, and iii) the cultured cells are geometrically simple, permitting high-resolution quantitation of receptor density. The goal of the proposed study is to examine further the mechanisms responsible for the clustering of receptors in the model system afforded by the Xenopus cultures. The experiments will utilize global and local application of external electric fields to cultured Xenopus muscle cells. Following these manipulations, digital video-microscopic analysis of fluorescently labeled cells will be performed in order to characterize the distribution of acetylcholine receptors and other components. Three broad questions will be addressed: 1. What molecular event(s) trigger the field-induced clustering of acetylcholine receptors? 2. How do other post-synaptic molecules behave in response to electric fields? 3. What are the parameters relevant to competition for receptors between neighboring clusters? Examination of these questions will increase our understanding of the in vivo mechanisms responsible for the developmental formation of synaptic contacts. The questions are intellectually crucial within the context of developmental neurobiology, and also have significance with respect to synapse-based disorders. The fundamental importance of receptor clustering is clear from the devastating effects of diseases such as myasthenia gravis, which reduce the number of available receptors at the junction. Moreover, experimental results and theoretical considerations suggest that the mechanisms by which neurons compete for receptors relate to the developmental refinement of synaptic connections, a topic of great interest with respect to developmental disabilities.
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0.997 |
1995 — 2001 |
Stollberg, Jes |
R25Activity Code Description: For support to develop and/or implement a program as it relates to a category in one or more of the areas of education, information, training, technical assistance, coordination, or evaluation. S06Activity Code Description: To strengthen the biomedical research and research training capability of ethnic minority institutions, and thus establish a more favorable milieu for increasing the involvement of minority faculty and students in biomedical research. |
Mechanisms of Acetylcholine Receptor Aggregation in a Simple Model System @ University of Hawaii At Manoa
Localization of cell-surface components to synaptic regions is an important problem in developmental neurobiology. It has been shown that lateral migration of diffusely-expressed receptors makes a major contribution to receptor clustering at the developing neuromuscular junction. As electric fields offer a convenient means to induce lateral migration of membrane components, their use constitutes a useful experimental system with which to study receptor localization. In particular, cultures of spherical muscle cells from the African frog Xenopus laevis have been shown to cluster receptors in response to such fields. The clustering continues after the field has been terminated, and is specific for acetylcholine receptors. These observations render this a powerful model system with which to probe the mechanisms underlying receptor clustering, because i) the cultured cells form receptor clusters rapidly in response to precisely measurable stimuli, ii) experiments can be conducted in the absence of neurites, cell- substrate contacts, or exogenous factors, and iii) the cultured cells are geometrically simple, permitting high-resolution quantitation of receptor density. The goal of the proposed study is to examine further the mechanisms responsible for the clustering of receptors in the model system afforded by the Xenopus cultures. the experiments will utilize global and local application of external electric fields to cultured Xenopus muscle cells. Following these manipulations, digital video-microscopic analysis of fluorescently labeled cells will be performed in order to characterize the distributions of acetylcholine receptors and other components. Three broad questions will be addressed: 1. What molecular event(s) trigger the field-induced clustering of acetylcholine receptors? 2. How do other post-synaptic molecules behave in response to electric fields? 3. What are the parameters relevant to competition for receptors between neighboring clusters? Examination of these questions will increase our understanding of the in vivo mechanisms responsible for the developmental formation of synaptic contacts. The questions are intellectually crucial within the context of developmental neurobiology, and also have significance with respect to synapse-based disorders. The fundamental importance of receptor clustering is clear from the devastating effects of diseases such as myasthenia gravis, which reduce the number of available receptors at the junction. Moreover, experimental results and theoretical considerations suggest that the mechanisms by which neurons compete for receptors relate to the developmental refinement of synaptic connections, a topic of great interest with respect to developmental disabilities.
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0.997 |
1997 — 2001 |
Stollberg, Jes Kunkel, Dennis |
N/AActivity Code Description: No activity code was retrieved: click on the grant title for more information |
Ultrastructural Organization of Developing Acetylcholine Receptor Aggregates
IBN-9724035 PI: STOLLBERG This research is directed at a better understanding of the molecular events underlying "synaptogenesis" (the formation of synapses during fetal development), which is of fundamental importance because synapses are the conduits through which nerve cells communicate with each other and with other cell types such as muscle fibers. At nerve-muscle synapses the communication depends on the release of acetylcholine from the nerve terminal. This molecule diffuses across a small space between the cells and binds to specialized muscle-cell molecules called "acetylcholine receptors." The binding of acetylcholine to its receptors triggers muscle contraction. Because nerve-muscle synapses must be fast and reliable, the receptors are concentrated in that part of the muscle cell membrane that is adjacent to the nerve terminal. This concentration takes place during fetal development, and many research efforts have been directed at discovering the molecular mechanisms that cause receptors to aggregate at this precise location. This project will examine aggregates of acetylcholine receptors and two related synaptic components at the ultrastructural level. The experiments involve labeling the molecules under study with small gold particles which can be detected using scanning electron microscopy. A sophisticated numerical analysis of label locations is then used to deduce the structure of labeled aggregates to within <1 nanometer. This represents a greater than hundred-fold improvement in resolution over previous experiments with developing receptor aggregates. The successful completion of this work will greatly enhance our understanding of the molecular mechanisms underlying receptor aggregation, and will thus contribute to our understanding of the basic mechanisms by which synapses are formed.
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0.97 |