Erik De Schutter - US grants

This research project will use a detailed, complex compartmental model of a cerebellar Purkinje cell to explore how changes in Ca2+ concentration ([Ca2+]), caused by inflow through Ca2+ channels and by release from internal stores, can affect dendritic excitability and synaptic plasticity in a large neuron. The Purkinje cell has been used as an experimental model for the investigation of metabotropic receptors, Ca2+ release from cytoplasmic stores evoked by Ca2+ and inositol triphosphate, and synaptic plasticity. At present, the functional role of these mechanisms is unclear and their interactions are poorly understood. By simulating a detailed, realistic model which integrates all these processes, we expect to make predictions on how such [Ca2+]- regulating mechanisms can interact with synaptic inputs in vivo. This research will not only contribute to our understanding of how the Purkinje cell functions within the cerebellar network, but will also provide insights in general mechanisms that control neural firing properties and learning in cortex and hippocampus. We will expand an existing detailed compartmental model of the Purkinje cell to include Ca2+ diffusion, buffering, pumps and uptake and release by internal stores in all dendritic compartments. The model will be tuned to reproduce high-resolution fura-2 Ca2+ imaging data, obtained in another laboratory. Subsequently, we will also include simple models of metabotropic receptor-activated Ca2+ release from internal stores and [Ca2+]-controlled synaptic plasticity of granule cell and inhibitory inputs. We will simulate changes in [Ca2+] evoked by diverse synaptic inputs. These simulations will be used to predict spatial and temporal changes in [Ca2+] under conditions of synaptic inputs in vivo, and to examine the interactions between the climbing fiber and parallel fiber systems at the level of [Ca2+]. We will examine the effects of [Ca2+] on the variability of amplification by Ca2+ currents of granule cell synaptic inputs. Subsequently, we will use the model to examine how metabotropic receptor activated Ca2+ release from internal stores could modulate synaptic integration by Purkinje cells. We expect that activation of metabotropic receptors might have a pronounced effect on dendritic excitability. We will also use the model to investigate the possible role of the Na+/Ca2+ exchanger in the prolonged depolarizations observed after pharmacological activation of the metabotropic receptor. Finally, we will use the model to examine which [Ca2+] profiles are necessary to induce long term depression (LTD) and other forms of synaptic plasticity. From these results, a simple model of synaptic plasticity will be built and used to explore which combinations and temporal patterns of climbing fiber and parallel fiber inputs could be expected to cause synaptic plasticity.

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Two-photon fluorescent imaging experiments have shown a much larger intracellular anomalous diffusion in CA1 pyramidal neuron dendrites than in cerebellar Purkinje cell dendrites. The most likely explanation at present is a previously undetected, but significant, difference in the structure of the dendritic cytoskeleton between these two neuron types. This hypothesis will be investigated using ultrastructural methods

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. PURKINJE CELL DENDRITIC SPINE DENSITY FROM CORRELATED LIGHT AND ELECTRON MICROSCOPY This project is an international collaboration between Dr. E. De Schutter (OIST, Japan) and Dr. F. Santamaria (Department of Biology, University of Texas at San Antonio, USA, fidel.santamaria@utsa.edu) It was previously shown that spines are an important cause of anomalous diffusion in Purkinje cell dendrites, due to trapping of molecules in dendritic spine heads (Santamaria et al. 2006). More recently we repeated this study, comparing Purkinje and CA1 pyramidal cells. We combined flash photolysis of caged Rhodamine dextran and two-photon fluorescent microscopy to track changes in concentration gradients along the dendrites of both cell types. We confirmed our previous findings and could show in pyramidal cells that the degree of anomalous diffusion depended upon spine density. However, there is no consensus on the spine density of Purkinje cells in the literature. We wish to use serial tomography to determine an accurate measure of spine density in rat Purkinje cells and to compare the values generated using diffraction-limited light microscopy.