Imaging the Xenopus Laevis Sciatic Nerve

The myelin sheath is essential for regulating chemical and electrical signaling in neurons. The sciatic nerve comprises numerous nerve fibers and contains one of the longest myelin sheaths of the human body. Degeneration of the myelin axonal region in either the central or peripheral nervous systems may lead to disabilities of sensorineural pathways and the motor cortex. Multiple sclerosis and Wallerian degeneration are examples of degenerative disorders that affect the brain and spinal cord, leading to complications in movement if the peripheral nervous system (PNS) is affected. The African clawed frog, Xenopus, is useful as a neurophysiological model organism due to its caapcity for neural regeneneration. Moreover, outcomes of prior studies lay a physiological and molecular foundation for ultrastructural analyses. The purpose of this pilot study was to establish methods that will permit analysis of the node of Ranvier and internodal lengths, thus indicating the location of Schwann cells found throughout neuronal fiber bundles in the sciatic nerve of Xenopus. Light microscopy was implemented to evaluate sciatic nerve sections using a modified protocol of the myelin stain, Luxol fast blue, and the nuclear stain, hematoxylin. These regions were then compared against confocal images from adjacent serial sections that detailed the length of Schwann cells between nodes of Ranvier. Schwann cells were detected by greater localization of actin-containing structures (AlexaFluor 488 phalloidin) near the nodes of Ranvier, and by the presence of Schwann cell nuclei (Hoechst 33342). Images were overlapped to examine myelinated regions stained in Luxol fast blue with Schwann cell regions detected in confocal microscopy.  Future directions of this project include implementing transgenic animals to study the myelination of Xenopus sciatic nerve and the use of TEM to capture details of myelin ultrastructure. Data from internodal regions measured in this study can provide a basis for analysis of Schwann cell development and myelin production and degeneration, using similar imaging protocols. Research support: NIH P50GM068762; NSF DBI 0959817.