Molecular Physiology of Bicarbonate Transport in the Brain

大脑中碳酸氢盐运输的分子生理学

• 批准号: 8321009
• 负责人: Walter F Boron
• 金额: $ 48.69万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1982
• 资助国家: 美国
• 起止时间: 1982-04-01 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8321009
• 关键词: Acid-Base Equilibrium; Acids; Affect; Astrocytes; Autistic Disorder; Bicarbonates; Binding; Biological Assay; Biological Models; Blood - brain barrier anatomy; Brain; Breathing; Calcineurin; Cell Line; Cell membrane; Cells; Cerebrospinal Fluid; Coupled; Cytoplasmic Protein; Data; Deletion Mutation; Detergents; Disease; Electrophysiology (science); Endothelial Cells; Environmental air flow; Epilepsy; Exons; Extracellular Fluid; Family; Genes; Goals; Hippocampus (Brain); Individual; Insecta; Ion Channel; Knock-out; Knockout Mice; Label; Libraries; Mediating; Membrane; Membrane Proteins; Memory; Modification; Molecular; Mus; Neurons; Neurotransmitter Receptor; Oocytes; Pharmaceutical Preparations; Physiological; Physiology; Play; Positioning Attribute; Property; Protein Binding; Protein phosphatase; Proteins; RNA Splicing; Recombinant Proteins; Recombinants; Regulation; Research; Respiratory Acidosis; Role; Screening procedure; Seizures; Site; Sodium Bicarbonate; Structure; Structure of choroid plexus; Sudden infant death syndrome; Surface Plasmon Resonance; Synaptic Transmission; System; Testing; Tissues; Variant; Work; Xenopus oocyte; base; blind; insight; neuronal excitability; novel; protein B; protein phosphatase inhibitor-2; receptor; respiratory; solid state; tool; trafficking; uptake; Acid-Base Equilibrium; Acids; Affect; Astrocytes; Autistic Disorder; Bicarbonates; Binding; Biological Assay; Biological Models; Blood - brain barrier anatomy; Brain; Breathing; Calcineurin; Cell Line; Cell membrane; Cells; Cerebrospinal Fluid; Coupled; Cytoplasmic Protein; Data; Deletion Mutation; Detergents; Disease; Electrophysiology (science); Endothelial Cells; Environmental air flow; Epilepsy; Exons; Extracellular Fluid; Family; Genes; Goals; Hippocampus (Brain); Individual; Insecta; Ion Channel; Knock-out; Knockout Mice; Label; Libraries; Mediating; Membrane; Membrane Proteins; Memory; Modification; Molecular; Mus; Neurons; Neurotransmitter Receptor; Oocytes; Pharmaceutical Preparations; Physiological; Physiology; Play; Positioning Attribute; Property; Protein Binding; Protein phosphatase; Proteins; RNA Splicing; Recombinant Proteins; Recombinants; Regulation; Research; Respiratory Acidosis; Role; Screening procedure; Seizures; Site; Sodium Bicarbonate; Structure; Structure of choroid plexus; Sudden infant death syndrome; Surface Plasmon Resonance; Synaptic Transmission; System; Testing; Tissues; Variant; Work; Xenopus oocyte; base; blind; insight; neuronal excitability; novel; protein B; protein phosphatase inhibitor-2; receptor; respiratory; solid state; tool; trafficking; uptake

DESCRIPTION (provided by applicant): Electroneutral Na+coupled HCO-3 transporters (nNCBTs), in the SLC4 family, mediate - HCO3 uptake and (along with Na-H exchangers, NHEs) play important roles in regulating - the intracellular pH (pHi) of neurons. nNCBTs also contribute to HCO3 secretion by choroid plexus and blood-brain barrier endothelial cells. Thus, nNCBTs are in a position to have profound effects on ion channels and on neurotransmitter receptors and transporters, thereby influencing neuronal excitability and synaptic transmission. Preliminary data show that truncating an autoinhibitory domain (AID) from the cytosolic N terminus (Nt) of nNCBTs increases transport more than 2 fold. The cytoplasmic protein IRBIT-which binds to IP3 receptors and inhibits Ca release-also interacts with the Nt of nNCBTs, nullifying inhibition by the AID. A third protein, the phosphatase CnAB, appears to bind to the Nt of one of the nNCBTs and profoundly inhibit transport. These three players (AID, IRBIT, and CnAB) may be the most powerful known proteins controlling acid-base transport. Certain nNCBTs splice variants may lack the AID or binding domains for IRBIT or CnAB. The major goal of this proposal is to understand the control of nNCBTs by the AID and IRBIT (and also by CnAB), and how these interactions modulate neuronal function. The four aims: (1) Use functional assays in oocytes to elucidate the above interactions. (2) Use binding assays to detect the interactions. (3) Determine the contribution of the nNCBTs to pHi regulation in hippocampal (HC) vs. medullary-raphi (MR) neurons (some of which are chemosensitive), and the effect of pHi changes on electrophysiological parameters. This aim exploits mouse knockouts of the three nNCBTs as well as mice with fluorescent-labeled neurons to aid in identification. (4) To express, solubilize, and characterize several ancestral nNCBT proteins. The proposed work will clarify the molecular mechanism and regulation of the nNCBTs, and the role they play in neuronal function. The research could have important implications for understanding the control of ventilation, SIDS, seizure disorders, and perhaps autism.

描述（由申请人提供）：slc4家族中的电负立Na+耦合HCO -3转运蛋白（NNCBTS），介导-HCO3摄取-HCO3摄取，（与Na -h -H交换器，NHES）在调节中起着重要作用 - 神经元的细胞内pH（PHI）。 NNCBT还有助于脉络丛和血脑屏障内皮细胞的HCO3分泌。因此，NNCBT可以对离子通道，神经递质受体和转运蛋白产生深远影响，从而影响神经元的兴奋性和突触传播。初步数据表明，NNCBTS的胞质N末端（NT）截断自身抑制域（AID）会增加运输超过2倍。细胞质蛋白IRBIT与IP3受体结合并抑制Ca释放与NNCBTS的NT相互作用，从而通过AID抑制了NNCBT的NT。第三个蛋白质磷酸酶CNAB似乎与其中一种NNCBT的NT结合并深刻抑制转运。这三个玩家（AID，IRBIT和CNAB）可能是控制酸碱运输的最强大的蛋白质。某些NNCBT剪接变体可能缺乏IRBIT或CNAB的辅助或结合域。该提案的主要目的是了解辅助和IRBIT（以及CNAB）对NNCBT的控制，以及这些相互作用如何调节神经元功能。这四个目的：（1）在卵母细胞中使用功能测定来阐明上述相互作用。 （2）使用绑定测定来检测相互作用。 （3）确定NNCBT对海马（HC）与髓质 - 拉斐（MR）神经元（其中一些是化学敏感）的贡献，以及PHI变化对电生理参数的影响。该目标利用了三个NNCBT的小鼠敲除以及具有荧光标记神经元的小鼠以帮助识别。 （4）表达，溶解和表征几种祖先NNCBT蛋白。提出的工作将阐明NNCBT的分子机制和调节，及其在神经元功能中的作用。这项研究可能对了解控制通风，小晶月，癫痫发作甚至自闭症具有重要意义。