Bączyk Marcin, Krutki Piotr
Department of Neurobiology, Poznań University of Physical Education;
Department of Neurobiology, Poznań University of Physical Education.
J Vis Exp. 2020 May 11(159). doi: 10.3791/61439.
Intracellular recording of spinal motoneurons in vivo provides a "gold standard" for determining the cells' electrophysiological characteristics in the intact spinal network and holds significant advantages relative to classical in vitro or extracellular recording techniques. An advantage of in vivo intracellular recordings is that this method can be performed on adult animals with a fully mature nervous system, and therefore many observed physiological mechanisms can be translated to practical applications. In this methodological paper, we describe this procedure combined with externally applied constant current stimulation, which mimics polarization processes occurring within spinal neuronal networks. Trans-spinal direct current stimulation (tsDCS) is an innovative method increasingly used as a neuromodulatory intervention in rehabilitation after various neurological injuries as well as in sports. The influence of tsDCS on the nervous system remains poorly understood and the physiological mechanisms behind its actions are largely unknown. The application of the tsDCS simultaneously with intracellular recordings enables us to directly observe changes of motoneuron membrane properties and characteristics of rhythmic firing in response to the polarization of the spinal neuronal network, which is crucial for the understanding of tsDCS actions. Moreover, when the presented protocol includes the identification of the motoneuron with respect to an innervated muscle and its function (flexor versus extensor) as well as the physiological type (fast versus slow) it provides an opportunity to selectively investigate the influence of tsDCS on identified components of spinal circuitry, which seem to be differently affected by polarization. The presented procedure focuses on surgical preparation for intracellular recordings and stimulation with an emphasis on the steps which are necessary to achieve preparation stability and reproducibility of results. The details of the methodology of the anodal or cathodal tsDCS application are discussed while paying attention to practical and safety issues.
体内脊髓运动神经元的细胞内记录为确定完整脊髓网络中细胞的电生理特性提供了“金标准”，相对于传统的体外或细胞外记录技术具有显著优势。体内细胞内记录的一个优点是，这种方法可以在具有完全成熟神经系统的成年动物身上进行，因此许多观察到的生理机制可以转化为实际应用。在这篇方法学论文中，我们描述了结合外部施加恒定电流刺激的这一程序，该刺激模拟脊髓神经元网络内发生的极化过程。经脊髓直流电刺激（tsDCS）是一种创新方法，越来越多地被用作各种神经损伤后康复以及体育领域的神经调节干预手段。tsDCS对神经系统的影响仍知之甚少，其作用背后的生理机制很大程度上尚不清楚。tsDCS与细胞内记录同时应用，使我们能够直接观察运动神经元膜特性的变化以及节律性放电的特征，以响应脊髓神经元网络的极化，这对于理解tsDCS的作用至关重要。此外，当所提出的方案包括根据所支配的肌肉及其功能（屈肌与伸肌）以及生理类型（快肌与慢肌）来识别运动神经元时，它提供了一个机会来选择性地研究tsDCS对脊髓回路中已识别成分的影响，这些成分似乎受到极化的不同影响。所介绍的程序侧重于细胞内记录和刺激的手术准备，重点是实现准备稳定性和结果可重复性所需的步骤。在关注实际和安全问题的同时，讨论了阳极或阴极tsDCS应用方法的细节。
