Calcium sensors for molecular fMRI

 描述(由申请人提供)：神经活动微创直接读数的发展是当今神经科学面临的最大挑战之一。我们最近的工作表明，使用对神经递质释放标志敏感的磁共振造影剂，可以对分子水平的现象进行高分辨率功能磁共振成像(FMRI)。更有价值的贡献将是创造适用于细胞内神经信号过程的分子功能磁共振成像的钙传感器。使用这些传感器进行的功能成像将结合MRI的非侵入性和全脑覆盖范围，以及已建立的光学钙神经成像技术的分子特异性和广泛适用性。钙依赖功能磁共振成像将是分析动物神经回路的一项突破性技术，在人类中具有潜在的长期应用。这项技术与超高场磁共振扫描仪和细胞标记技术相结合，可以实现细胞分辨率。然而，实现这一进步的一个主要障碍是创造有效的钙依赖磁共振造影剂。这项建议描述了创造适合于分子功能磁共振成像的新型磁共振钙探针的策略，以及在动物身上验证该方法的初步实验。创新包括膜渗透探针本身的合理设计，以及体内钙成像和基因靶向应用的方法。在目标1中，我们合成了基于顺磁性细胞渗透性芳香族络合物的磁共振钙传感器，并在体外对其进行了表征。在目标2中，我们形成了钙探针的乙酰甲氧基衍生物，并首先在细胞培养中验证它们，然后在大鼠身上使用体感刺激范式进行验证。如有必要，目标2的结果将指导探测器的进一步改进。在目标3中，我们通过选择性酯酶使钙传感器适应于细胞内的捕获，这将促进探针在基因靶向细胞中的积累。这项技术将提供一种方法，对大脑中动态钙水平进行特定细胞类型的功能成像，在某些情况下还提供个别细胞特定的功能成像。该项目的潜在影响和研究团队的初步成果使这一下一代神经成像项目特别适合根据RFA-NS-14-007为大脑倡议提供资金。

 DESCRIPTION (provided by applicant): The development of minimally invasive direct readouts of neural activity is one of the greatest challenges facing neuroscience today. Our recent work has shown that it is possible to perform high resolution functional magnetic resonance imaging (fMRI) of molecular-level phenomena using MRI contrast agents sensitive to hallmarks of neurotransmitter release. An even more valuable contribution would be the creation of calcium sensors suitable for molecular fMRI of intracellular neural signaling processes. Functional imaging performed with these sensors would combine the noninvasiveness and whole-brain coverage of MRI with the molecular specificity and broad applicability of established optical calcium neuroimaging techniques. Calcium-dependent fMRI will be a breakthrough technique for analysis of neural circuits in animals, with potential longer term applications in humans. The technique could achieve cellular resolution in conjunction with ultrahigh field MRI scanners and cell labeling techniques. A major hurdle in realizing this advance is the creation of effective calcium-dependent MRI contrast agents, however. This proposal describes strategies for creating novel MRI calcium probes suitable for molecular fMRI, as well as initial experiments that validate the approach in animals. Innovations include the rational design of membrane permeable probes themselves as well as approaches for in vivo calcium imaging and genetically targeted applications. In Aim 1, we synthesize MRI calcium sensors based on paramagnetic cell-permeable aromatic chelates and characterize them in vitro. In Aim 2, we form acetomethoxy derivatives of the calcium probes and validate them first in cell culture and then in rats, using a somatosensory stimulation paradigm. Results of Aim 2 will direct further refinement of the probes, if necessary. In Aim 3, we adapt the calcium sensors for intracellular trapping by selective esterases that will promote probe accumulation in genetically targeted cells. This technique will provide a means for cell type-specific and in some cases individual cell-specific functional imaging of dynamic calcium levels in the brain. Potential impact of the project and preliminary achievements of the research team make this next-generation neuroimaging project particularly suitable for BRAIN Initiative funding under RFA-NS-14-007.