| Project/Area Number |
05404088
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| Research Category |
Grant-in-Aid for General Scientific Research (A)
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| Allocation Type | Single-year Grants |
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| Research Field |
神経・脳内生理学
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| Research Institution | Gunma University |
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Principal Investigator |
OZAWA Seiji Gunma University, School of Medicine, Professor, 医学部, 教授 (40049044)
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| Co-Investigator(Kenkyū-buntansha) |
TSUZUKI Keisuke Gunma University, School of Medicine, Lecturer, 医学部, 講師 (60222139)
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| Project Period (FY) |
1993 – 1995
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| Project Status |
Completed (Fiscal Year 1995)
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| Budget Amount *help |
¥19,600,000 (Direct Cost: ¥19,600,000)
Fiscal Year 1995: ¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1994: ¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 1993: ¥14,000,000 (Direct Cost: ¥14,000,000)
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| Keywords | Glutamate receptor / Hippocampal neuron / Patch clamp / Reverse transcription / Polymerase chain reaction / AMPA receptor / GluR2 subunit / Ca^<2+> permeability / 単一ニューロン / ホールセルパッチクランプ / PCR / パッチクランプ法 / PCR法 / サブユニット |
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| Research Abstract |
The glutamate receptor channels in the mammalian central nervous system are classified into three subtypes : NMDA,AMPA and kainate receptors. Among them, AMPA receptors are the principal mediators of fast excitatory neurotransmission. AMPA receptors are formed by various combinations of four subunit proteins, GluRl, GluR2, GluR3 and GluR4. To trace the molecular basis underlying differences in the functional properties of AMPA receptors in rat hippocampal neurons, we coupled whole-cell patch-clamp recordings with reverse transcription (RT) followed by polymerase chain reaction amplification (patch-clamp RT-PCR method).
We have previously found two distinct types of AMPA receptors in cultured rat hippocampal neurons. In type I neurons, AMPA receptors displayd an outward rectification and little Ca^<2+> permeability. In contrast, type II neurons had AMPA receptors that displayd strong inward rectification and high Ca^<2+> permeability. The patch-clamp RT-PCR method revealed that only the
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GluRl and GluR4 subunits were expressed and that no GluR2 subunit was detected in type II neurons. In contrast, GluR2 was expressed at a high level together with GluR1, and occasionally with GluR3 or GluR4, in type I neurons. We further applied this technique for analyzing subunit compositions of AMPA receptors in various types of neurons in rat hippocampal slices. Type II neurons carrying AMPA receptors with strong inward rectification and high Ca^<2+> permeability were found in non-pyramidal interneurons in the hippocampus. In all type II neurons tested (16 neurons) in slices, either GluR1, GluR3 or GluR4 was expressed, but no GluR2 expression was detected.
Thus, It is likely that the presence or absence of GluR2 determines rectification properties and Ca^<2+> permeability in native AMPA receptors in hippocampal neurons both in cultures and slices. The patch-clamp RT-PCR method will be useful for a wide range of physiological studies to elucidate the molecular basis of functional properties at the single cell level. Less
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