Study of the calcium dynamics and molecular mechanisms in the dendrite of cerebellar Purkinje cell

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We investigated dynamics and functions of IP3 receptor (IP3R) in the neuronal dendrite. Since IP3R is known to play a pivotal role in synaptic plasticity in the postsynapse, changes in the trafficking of IP3R into/out of dendritic spines would affect the mechanism underlying synaptic plasticity. We compared lateral diffusion efficacy of IP3R with another ER membrane protein, SERCA, in spines of cerebellar Purkinje cells by expressing genes of the ER proteins fused with fluorescent proteins by means of gene gun gene delivery technique and two photon microsoopy. We found that the diffusion efficacy of IP3R was significantly lower than that of SERCA, suggesting a mechanism which enriches IP3R proteins in spines due to their high affinity to the actin cytoskeleton as has been shown in the dendrites of hippocampal pyramidal cells by us previously. In another approach to the IP3R function in the cerebellar cortex, we found that IP3R plays a role in formation of the dendrite of Purkinje cell. It was unexpected that IP3R expressed in the cerebellar granule cells was the key player instead of the highly expressed IP3Rs in the Purkinje. This result shows another important role of IP3R in the cerebellar cortex. We also succeeded to create an IP3-indicator by engineering the IP3-binding part of IP3R with fluorescent proteins, which clearly showed IP3 oscillation during calcium oscillation in the COS-7 cell line. This novel indicator will be a potent tool investigate IP3 dynamics in neuronal dendrites where IP3 and IP3R play important roles in synaptic plasticity.