2014 Fiscal Year Annual Research Report
Mechanisms of memory consolidation in sleep
| Project/Area Number |
26285161
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| Research Institution | University of Tsukuba |
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Principal Investigator |
PAVLIDES C 筑波大学, 人間系, 教授 (50712808)
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| Co-Investigator(Kenkyū-buntansha) |
小川 園子 筑波大学, 人間系, 教授 (50396610)
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| Project Period (FY) |
2014-04-01 – 2017-03-31
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| Keywords | sleep / memory / hippocampus / amygdala / fear conditioning |
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| Outline of Annual Research Achievements |
The proposed studies were aimed at determining possible gene cascades which may be occurring specifically in sleep following a learning experience for the long-term consolidation of memory. Male, Long-Evan rats were implanted with guide-cannulae bilaterally aimed at the CA1 field of the dorsal hippocampus. After 1-week recovery, animals were trained in a single-trial contextual fear conditioning following which their sleep/wake behaviors were monitored. Animals were either allowed to sleep or kept awake for 4h. On average, animals started sleeping after 2h. At the start of sleep, they were injected 3μl either with the PKA inhibitor rp-cAMPs or aCSF and their sleep/wake behavior was monitored for the remainder of the 4h period. Animals that were kept awake, were injected at a similar time as the sleep animals either with the PKA activator sp-cAMPs or aCSF. The animals were tested for fear memory on the following day. We found that animals injected with aCSF and kept awake for 4h (n=12) following training, showed an impaired fear memory versus aCSF-injected sleep animals (n=10) (repeated-measures ANOVA, F1,15=13.09, p<0.001). We also found that administration of rp-cAMPs suppressed fear memory in sleep animals (n=5), while the animals kept awake and injected with sp- cAPMs (n=2) showed similar freezing level to aCSF-injected sleep animals. The present results suggest that following learning, PKA activation is involved in the memory consolidation during sleep in a time window of 0-4h post-training.
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| Current Status of Research Progress |
Current Status of Research Progress
2: Research has progressed on the whole more than it was originally planned.
Reason
There is a lot of progress being currently made on the proposed studies, although a number of problems had to be initially overcome. The aim of the studies were to determine the role of PKA activation, specifically in sleep following a learning experience, on long-term memory consolidation. This requires the bilateral cannulation of animals for drug injections (PKA inhibitor/activator/control) into the hippocampus. A couple of problems that had to be overcome were: first, precisely implant the cannulas into the hippocampus. Make sure that the drugs (3 ml) were injected at the precise moment, without uncontrolled leakage; Second, the effective drug dose had to be determined. Although some information existed in the literature, a low dose had no effect while a higher dose produced epileptic activity. An intermediate dose had to be selected; Third, drugs had to be injected when the animals first went to sleep and they had to remain in sleep for a specified amount of time. In the event the animals woke prior to this, the experiment was terminated. Since these animals were fear conditioned just before, they could not be re-conditioned again so they could not be included in the study; Fourth, initially, sleep was assessed visually, since we did not have the electrophysiological set up for EEG/ EMG recording. This has now been setup, although the system still has to be tested; Fifth, the students undertaking the study had to be trained in surgery, sleep recordings, drug injections, behavioral testing, etc. All of this has been currently accomplished.
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| Strategy for Future Research Activity |
Previous studies have reported that the hippocampus is involved in contextual fear conditioning and that it is involved in a time-limited manner - i.e., during the first 4 hours or so following a learning experience. We, therefore, chose to determine whether cAMP/PKA is indeed involved in the consolidation process during this time scale. Once it is established that this is the case, we plan to investigate the time course of cAMP/PKA activation along with the brain structures involved for different types of memory - i.e., contextual vs cued conditioning which involve the hippocampus and amydgala respectively. We could then proceed to perform drug manipulations of these structures at more specific time points to determine that indeed we can differentially affect fear memory. Positive results in these experiments will provide strong evidence that these gene cascades are intricately involved with long- term memory consolidation and that this is specifically occurring off-line, during sleep for the long-term storage of memory.
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| Causes of Carryover |
A main reason that not all the funds have been used, thus far, is that we planned to build electrophysiological equipment along with purchasing software for the recording of sleep behaviors. These are custom build, which is not guaranteed to function without testing. Thus, initially a setup was built and is being tested before more elaborate and duplicate equipment is ordered. The drugs being used are expensive, but do not have an infinite shelf life. Once we are ensured that the drugs are working, more of these will be ordered. Initially, a person who could perform surgeries, was not identified. Currently, such a person has joined the team and is being paid a part-time salary. During this year we plan to publish some of the results, which will also require publication fees, etc.
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| Expenditure Plan for Carryover Budget |
As discussed above, during the 2015 budget year, we will be purchasing further recording equipment, including commutators and cables, testing cages, fear conditioning setup, etc. to speed up the collection of data. This will also require the purchase of more animals, histology supplies, etc. Given the additional results from the ongoing studies, publication fees, presentations at meetings, etc. will also be increased.
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Research Products
(3 results)
