Probing presynaptic receptor function with two-photon uncaging, Ca2+ imaging and photobleaching

通过双光子解笼、Ca2 成像和光漂白探测突触前受体功能

• 批准号: G0600368/1
• 负责人: Dmitri Rusakov
• 金额: $ 55.3万
• 依托单位: University College London
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2007
• 资助国家: 英国
• 起止时间: 2007 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=G0600368%2F1
• 关键词: Probing; presynaptic; receptor; function; two

Synapses that transmit information among neurons are modulated in a highly diverse manner by an interplay of intrinsic and extrinsic factors. This contributes both to the richness of information processing by brain circuits and to their stability, which can however break down in disease states such as epilepsy. Several methods are available to study rather indirectly how chemical receptors interact with calcium signals within individual neurotransmitter-releasing nerve processes. However, they cannot be readily integrated with one another: some can only be used with living neurons, while others require the tissue to be frozen or chemically treated. We propose to achieve a much more detailed insight into the function of individual synapses by incorporating a novel method to manipulate the local concentration of chemically active substances with laser-evoked excitation of molecules. This method relies on hitting individual electrons with two photons in very rapid succession, thus evoking a highly localised and brief change in the state of the atoms. Such localised manipulations are necessary to study synapses, because they normally function on a space- and time-scale of microns and milliseconds. We will apply this method to several topical questions that are relevant to normal brain function and to epilepsy. Specifically, we will investigate how the spatial distribution of neurotransmitter receptors and different proteins mediating calcium movements into nerve processes interact with one another. We will also ask whether some changes that occur in epilepsy are caused by alterations in the local expression of these receptors. The work will shed light on how neuronal signalling underlies cognitive processes and how abnormal synaptic function contributes to seizures.

在神经元之间传递信息的突触受到内在和外在因素相互作用的高度多样性的调节。这有助于大脑回路处理信息的丰富性及其稳定性，但在癫痫等疾病状态下可能会崩溃。有几种方法可以间接地研究化学受体如何与单个神经递质释放神经过程中的钙信号相互作用。然而，它们不能容易地彼此整合：有些只能与活的神经元一起使用，而另一些则需要将组织冷冻或化学处理。我们建议实现一个更详细的洞察到单个突触的功能，通过将一种新的方法来操纵局部浓度的化学活性物质与激光诱发的分子激发。这种方法依赖于用两个光子快速连续地撞击单个电子，从而引起原子状态的高度局部化和短暂的变化。这种局部操作对于研究突触是必要的，因为它们通常在微米和毫秒的空间和时间尺度上发挥作用。我们将把这种方法应用于与正常脑功能和癫痫有关的几个主题问题。具体来说，我们将研究神经递质受体的空间分布和不同的蛋白质介导的钙运动到神经过程相互作用。我们还将询问癫痫中发生的某些变化是否是由这些受体的局部表达改变引起的。这项工作将揭示神经元信号是如何构成认知过程的基础，以及异常的突触功能是如何导致癫痫发作的。