Regulation and Neural Function of Atypical PKC

非典型PKC的调节和神经功能

基本信息

批准号：
6910853
负责人：
TODD C SACKTOR
金额：
$ 29.54万
依托单位：
SUNY DOWNSTATE MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
1997
资助国家：
美国
起止时间：
1997-04-01 至 2006-06-30
项目状态：
已结题

项目摘要

The molecular events underlying synaptic plasticity and behavioral memory are very complex, but can be divided into 2 fundamentally distinct phases: induction, which initiates the plasticity, and maintenance, which sustains it. We have focused on the enzyme protein kinase C (PKC), which has been implicated in both phases of plasticity, by examining the family of PKC isoforms in hippocampal slices during increases in synaptic strength caused by high-frequency tetanic stimulation (long-term potentiation, LTP). Our previous work has established that one isoform, the atypical PKCzeta, is the dominant PKC maintaining LTP. The activation of zeta is through the protein synthesis- dependent formation of its independent catalytic domain, PKMzeta. The critical role of PKMzeta in LTP maintenance, however, does not exclude the possibility that atypical PKC may also have an important early role in LTP induction. While our previous work focused on PKMzeta, additional atypical PKC forms, including the other atypical isozyme, PKCiota, need to be considered. The major objective of this proposal is to examine the role of atypical PKC in LTP induction. The evidence pointing to this hypothesis is as follows: 1) During LTP induction, there is an early activation of atypical PKC by translocation to membrane. 2) Whole-cell perfusion of a dominant negative inhibitor of atypical PKC into CA1 pyramidal cells prevents the early potentiation during LTP induction. 3) Perfusion of active atypical PKC causes synaptic potentiation that occludes the early potentiation induced by tetanic stimulation. Thus our first aim is to determine the mechanisms of activation of atypical PKC during LTP induction. While the late activation in maintenance is through protein synthesis of PKMzeta, the early activation of atypical PKC is through translocation of PKCiota to membrane. Second, we will examine whether the initial activation of atypical PKC during LTP induction leads to early potentiation of synaptic transmission, and the transition to PKMzeta in LTP maintenance. Third, we will examine the direct physiological effects of the different atypical PKC isoforms in synaptic transmission using whole-cell recording, and during behavior in Drosophila. Initial results show that transgenic expression of PKMzeta in flies consolidates transient memory into long-term memory. These 3 aims will provide fundamental knowledge on the role of atypical PKCs in neuronal signaling, and may lead to unifying, evolutionarily conserved molecular mechanisms for synaptic plasticity and behavioral memory.

突触可塑性和行为记忆背后的分子事件非常复杂，但可以分为两个基本不同的阶段：诱导（启动可塑性）和维持（维持可塑性）。我们通过检查海马切片中高频强直刺激引起的突触强度增加期间的 PKC 亚型家族，重点研究了与可塑性的两个阶段有关的蛋白激酶 C (PKC)。，LTP）。我们之前的工作已经确定一种亚型，即非典型 PKCzeta，是维持 LTP 的主要 PKC。 Zeta 的激活是通过其独立催化结构域 PKMzeta 的蛋白质合成依赖性形成来实现的。然而，PKMzeta 在 LTP 维持中的关键作用并不排除非典型 PKC 也可能在 LTP 诱导中发挥重要早期作用的可能性。虽然我们之前的工作重点是 PKMzeta，但还需要考虑其他非典型 PKC 形式，包括其他非典型同工酶 PKCiota。该提案的主要目的是研究非典型 PKC 在 LTP 诱导中的作用。支持这一假设的证据如下： 1) 在 LTP 诱导过程中，非典型 PKC 通过易位到膜而被早期激活。 2) 将非典型 PKC 的显性失活抑制剂全细胞灌注到 CA1 锥体细胞中，可防止 LTP 诱导过程中的早期增强。 3) 活性非典型 PKC 的灌注引起突触增强，从而阻断强直刺激诱导的早期增强。因此，我们的首要目标是确定 LTP 诱导过程中非典型 PKC 的激活机制。维持的后期激活是通过 PKMzeta 的蛋白质合成，而非典型 PKC 的早期激活是通过 PKCiota 易位到膜上。其次，我们将检查 LTP 诱导过程中非典型 PKC 的初始激活是否会导致突触传递的早期增强，以及 LTP 维持过程中向 PKMzeta 的转变。第三，我们将使用全细胞记录检查不同非典型 PKC 同工型在突触传递中以及果蝇行为过程中的直接生理效应。初步结果表明，果蝇中 PKMzeta 的转基因表达将短暂记忆巩固为长期记忆。这三个目标将提供关于非典型 PKC 在神经元信号传导中的作用的基础知识，并可能导致突触可塑性和行为记忆的统一的、进化上保守的分子机制。