| Budget Amount *help |
¥14,300,000 (Direct Cost: ¥14,300,000)
Fiscal Year 2002: ¥4,000,000 (Direct Cost: ¥4,000,000)
Fiscal Year 2001: ¥10,300,000 (Direct Cost: ¥10,300,000)
|
|---|
| Research Abstract |
The aim of the present research project was to clarify the role of inositol lA5-trisphosphate (IP_3) in the induction oflong tem depression, especially focusing on the spatio-temporal aspects of IP_3 signaling. To achieve this end, we have developed novel technologies that enable us to visualize the intracellular dynamics of IP_3 in cerebellar Purkinje neurons. As an IP_3 imaging probe, we employed GFP-PHD which is a fusion protein consisting of GFP and pleckstrin homology domain of phospholipase C δ1. We first proved that IP_3 can be visualized in cultured Purkinje neurons which is transduced with GFP-PHD. Furthermore, by directly injecting Sindbis virus vector containing cDNA encoding GFP-PHD into the mouse brain, we successfully introduced GFP-PHD into Purkinje neurons in cerebellar slice. The transduction efficiency of GFP-PHD was found to be high after dissection of cerebellar slices. Using these novel techniques, we found that AMPA receptor activation induces intracellular accumu
… More
lation of IP_3 in cultured Purkinje neuron. AMPA receptor-mediated IP_3 production was not a mere epiphenomenon, because AMPA receptor activation through climbing fiber stimulation also elicited IP_3 production in Purkinje neuron in cerebellar slice preparation. Our findings are thus important in an understanding of the mechanism of LTD induction.
The aim of the present research project was to clarify the role of inositol lA5-trisphosphate (IP_3) in the induction oflong tem depression, especially focusing on the spatio-temporal aspects of IP_3 signaling. To achieve this end, we have developed novel technologies that enable us to visualize the intracellular dynamics of IP_3 in cerebellar Purkinje neurons. As an IP_3 imaging probe, we employed GFP-PHD which is a fusion protein consisting of GFP and pleckstrin homology domain of phospholipase C δ1. We first proved that IP_3 can be visualized in cultured Purkinje neurons which is transduced with GFP-PHD. Furthermore, by directly injecting Sindbis virus vector containing cDNA encoding GFP-PHD into the mouse brain, we successfully introduced GFP-PHD into Purkinje neurons in cerebellar slice. The transduction efficiency of GFP-PHD was found to be high after dissection of cerebellar slices. Using these novel techniques, we found that AMPA receptor activation induces intracellular accumulation of IP_3 in cultured Purkinje neuron. AMPA receptor-mediated IP_3 production was not a mere epiphenomenon, because AMPA receptor activation through climbing fiber stimulation also elicited IP_3 production in Purkinje neuron in cerebellar slice preparation. Our findings are thus important in an understanding of the mechanism of LTD induction. Less
|
