The long range goal of this application is to understand the function of the polymeric matrix, present in secretory granules, in the mechanism of regulated secretion. As a hypothesis, the investigator proposes that the matrix regulates the amount and time course of release after membrane fusion, by acting as a cation exchanger with specific electrical and dynamic properties. The application contains three specific aims. 1. To identify the sources of counterions for ion exchange. Projects are aimed at determining the role of ion transport through the granule membrane as a source of counterions. By RT-PCR and immunocytochemistry attempts will be made to identify the ionic pathways of the mast cell granule membrane. Patch clamp and amperometry will be used to determine the role of H+-ATPases, serotonin transporters and cation channels in regulating magnitude and time course of serotonin release. 2. To examine determining the movement of secretory products and counterions through the matrix. By means of patch clamp, amperometry and pulse-laser imaging, Dr. Fernandez will study the parameters that regulate the movement of ions through the matrix and their effect on the secretory response. 3. To detect smart hydrogels in sub-micron secretory granules. Atomic Force Microscopy will be employed to determine the presence of smart hydrogels in synaptic vesicles and chromaffin granules, with the long range goal eventually to understand their function-related plastic properties.
| Oberhauser, A F; Fernandez, J M (1996) A fusion pore phenotype in mast cells of the ruby-eye mouse. Proc Natl Acad Sci U S A 93:14349-54 |
