Technology for functional study of cells and circuits in large postmortem brains ex vivo

项目总结 哺乳动物的大脑可以说是最复杂的生物结构。研究细胞功能和 绘制大脑中的神经连接图是更好地了解健康和疾病中的大脑的关键任务。 由于实验纬度和同步的固有限制，这在活体中尤其具有挑战性 可以接触到同一动物体内的多个大脑区域。这些缺点阻碍了多式联运的审问 多突触电路和中尺度连接。尤其重要的是，这些实验上的不足之处 与大脑和颅骨解剖的复杂性成比例增长，阻碍了向更大的哺乳动物的转化。 这笔拨款通过优化和验证一流的神经技术BrainEx来解决这些任务 体外条件下死后大型哺乳动物脑的分子和细胞功能的恢复， 常温条件下。我们特别建议继续在猪脑中优化BrainEx，同时 验证了BrainEx系统作为电生理新实验平台的有效性， 在完全隔离的、完整的和有功能的大型哺乳动物脑中进行连接和成像研究。确实有 这一应用程序的四个主要区别方面：(1)实施开发的新方法以 恢复脑宏微循环，延长死后脑的细胞活力 常温条件下，研究人员可以(2)同时追踪连接并表征 通过化学和基于载体的技术跨越无数大脑区域的细胞功能和形态， 包括活体手术方法无法触及的区域；(3)研究多突触远程回路和 皮层网络电活动；以及(4)在体外大鼠中进行功能性PET和CT成像研究 哺乳动物的大脑。这种方法代表了一种新的工具，可以更彻底地研究结构和 复杂电路及其内部单元的功能。这项技术的广泛传播将使调查人员 组织培养或活体方法无法提供的跨物种实验优势。

PROJECT SUMMARY The mammalian brain is arguably the most complex biological structure. Investigating cellular functions and mapping neural connections in the brain are critical tasks to better understand the brain in health and disease. This is particularly challenging in vivo due to the inherent limitations in experimental latitude and simultaneous access to multiple brain regions within the same animal. These shortcomings hinder multimodal interrogation of multi-synaptic circuits and mesoscale connectomics. Of particular importance, these experimental inadequacies grow in proportion to the complexity of the brain and cranial anatomy, impeding translation to larger mammals. This grant addresses these tasks by optimizing and validating a first-in-class neurotechnology called BrainEx for the restoration of molecular and cellular functions of the postmortem large mammalian brain under ex vivo, normothermic conditions. We specifically propose to continue optimizing BrainEx in porcine brains, while validating the efficacy of the BrainEx system as a new experimental platform for electrophysiological, connectomic, and imaging studies in the fully isolated, intact, and functional large mammalian brain. There are four major distinguishing aspects of this application: (1) implementation of novel approaches developed to restore cerebral macro- and microcirculation and extend cellular viability of the postmortem brain under normothermic conditions such that researchers can (2) simultaneously trace connections and characterize cellular function and morphology by chemical and vector-based techniques across myriad brain regions, including areas inaccessible to in vivo surgical approaches; (3) investigate multisynaptic long-range circuitry and cortical network electrical activity; and (4) perform functional PET and CT imaging studies in the ex vivo large mammalian brain. This methodology represents a new tool for more thorough investigation of the structure and function of complex circuits and the cells within them. Wide distribution of this technology will grant investigators experimental advantages across species not afforded by tissue culture or in vivo approaches.