| Project/Area Number |
16370033
|
|---|
| Research Category |
Grant-in-Aid for Scientific Research (B)
|
| Allocation Type | Single-year Grants |
|---|
| Section | 一般 |
|---|
| Research Field |
Animal physiology/Animal behavior
|
|---|
| Research Institution | Tokushima Bunri University (2006)
Hokkaido University (2004-2005) |
|---|
Principal Investigator |
ITO Etsuro Tokushima Bunri University, Kagawa School of Pharmaceutical Sciences, Professor, 香川薬学部, 教授 (80203131)
|
|---|
| Co-Investigator(Kenkyū-buntansha) |
MATSUO Ryota Tokushima Bunri University, Kagawa School of Pharmaceutical Sciences, Lecturer, 香川薬学部, 講師 (40334338)
KOBAYASHI Suguru Tokushima Bunri University, Kagawa School of Pharmaceutical Sciences, Research Associate, 香川薬学部, 助手 (50325867)
SADAMOTO Hisayo Tokushima Bunri University, Kagawa School of Pharmaceutical, Sciences Associate, 香川薬学部, 助手 (70374220)
MITA Koichi Tokushima Bunri University, Kagawa School of Pharmaceutical Sciences, Associate Professor, 香川薬学部, 助教授 (50281845)
SAITO Tamao Hokkaido University, Graduate School of Science, Research Associate, 大学院理学研究院, 助手 (30281843)
|
|---|
| Project Period (FY) |
2004 – 2006
|
| Project Status |
Completed (Fiscal Year 2006)
|
| Budget Amount *help |
¥15,400,000 (Direct Cost: ¥15,400,000)
Fiscal Year 2006: ¥4,700,000 (Direct Cost: ¥4,700,000)
Fiscal Year 2005: ¥4,600,000 (Direct Cost: ¥4,600,000)
Fiscal Year 2004: ¥6,100,000 (Direct Cost: ¥6,100,000)
|
|---|
| Keywords | single cell / learning / memory / CREB / real-time PCR / RNAi / 行動変容 / ELISA法 / 酵素サイクリング法 / 抗体カラム / 抗体 / real-time PCR / 学習記憶 / 分子動力学 / モノアラガイ |
|---|
| Research Abstract |
Gene expression is differently regulated in every cell even though the cells are included in the same tissue. For this reason, we need to measure the amount of mRNAs in a single cell to understand transcription mechanism better. We therefore developed a procedure for determining the exact copy numbers of mRNAs within a single cell. We first isolated the cerebral giant cell of Lymnaea stagnalis as this neuron plays a key role in the process of memory consolidation of a learned behavior of feeding. We then determined the copy numbers of mRNAs for the cyclic AMP-responsive element binding proteins (CREBs) by quantitative real-time PCR. Next, we still do not know whether the constitutively expressed forms of CREB are enough or the newly synthesized forms are required for the synaptic enhancement. In addition, if the newly synthesized forms are needed, we must determine the time for translation of CREB from its mRNA. We applied the methods of RNA interference and real-time PCR to CREB in the cerebral giant cells. The cerebral giant cells employ a CREB cascade for the synaptic enhancement to neurons such as the B1 motoneurons. We injected the siRNA of the activator type of CREB (CREB1) or the repressor type (CREB2) into the cerebral giant cells and examined the changes in amplitude of EPSP recorded in the B1 motoneurons. The changes in the amounts of CREB1 and CREB2 mRNAs were also examined in the cerebral giant cells. The EPSP amplitude was suppressed 15 min after injection of CREB1 siRNA, whereas that was augmented 60 min after injection of CREB2 siRNA. In the latter case, the decrease in the amount of CREB2 mRNA was confirmed by real-time PCR. Our results showed that the de novo synthesized forms of CREB are required within tens of minutes for the synaptic enhancement in memory consolidation.
|
