Equipment for brain slice patch-clamp electrophysiology

大脑中神经元之间的交流是由突触传递控制的，突触前化学神经递质的释放及其与突触后神经元上的细胞表面受体的结合介导了突触传递。我们实验室的研究集中在一类被称为烟碱型乙酰胆碱受体的配体门控离子通道上。我们的长期目标是研究尼古丁受体在调节中枢神经系统神经传递中的基本功能。我们开发了一种新的方法来理解尼古丁受体在大脑中介导的信号。我们用基因工程技术改造了基因敲入小鼠，在其中我们融合了一种荧光蛋白和α4尼古丁受体亚单位。据我们所知，这是第一个用荧光蛋白标记离子通道的敲入小鼠。α4β2烟碱受体是中枢神经系统主要的烟碱受体亚型。第一次，我们有能力在亚微米分辨率下准确地量化中枢神经系统神经元中受体的亚细胞分布。这一重要信息将使我们能够准确地针对这些神经元进行电生理学，以确定含有尼古丁受体的α4亚单位的功能。此外，我们还产生了α4尼古丁受体基因功能获得突变的敲入小鼠。这种突变使受体对尼古丁或乙酰胆碱的激活敏感50倍，并表达4倍以上的尼古丁反应，从而促进了α4尼古丁受体的记录。

Communication between neurons in the brain is governed by synaptic transmission, which is mediated by the presynaptic release of chemical neurotransmitters and their binding to cell surface receptors on the postsynaptic neuron. The research in our lab focuses on a class of ligand-gated ion channels called nicotinic acetylcholine receptors. Our long term goal is to examine the basic function of nicotinic receptors in modulating neurotransmission in the CNS. We have developed a novel approach to understanding nicotinic receptor mediated signaling in the brain. We have genetically engineered knock-in mice in which we fused a fluorescent protein to the alpha4 nicotinic receptor subunit. To our knowledge this is the first knock-in mouse in which an ion channel has been tagged with a fluorescent protein. Alpha4 beta2 nicotinic receptors constitute the major nicotinic receptor subtype in the CNS. For the first time, we have the ability to quantify accurately at submicron resolution the subcellular distribution of the receptor in neurons in the CNS. This important information will allow us to precisely target those neurons for electrophysiology to ascertain the function of alpha4 subunit containing nicotinic receptors. Furthermore, we have also generated knock-in mice with a gain-of-function mutation to the alpha4 nicotinic receptor gene. This mutation renders the receptor 50 fold more sensitive to activation by nicotine or acetylcholine and expresses four fold greater nicotinic responses thus facilitating the recording of alpha4 nicotinic receptors.