Red Fluorescent Labeling of Myelin by Membrane-Targeted tdTomato in Transgenic Mouse Lines

Myelin is a highly complex membranous structure wrapped around axons by oligodendrocytes or Schwann cells in the central and peripheral nervous system, respectively. Fluorescent labeling is widely used to study the structure and dynamics of myelin. Combining structural with functional imaging requires labeling of myelin with red fluorescence, as many functional sensors, including Ca²⁺ indicators and genetically encoded metabolite sensors, fluoresce in the green spectral range. However, in vivo tools enabling red fluorescent labeling of myelinating cells and their myelin sheaths remain limited. Here, we generated a set of seven transgenic mouse lines expressing a membrane-targeted variant of the red fluorescent protein tdTomato in myelinating oligodendrocytes and Schwann cells throughout the nervous system. The mouse lines provide a variety of expression patterns ranging from wide-spread labeling of myelin to a rather sparse expression, the latter enabling visualization of individual oligodendrocytes and their associated myelin sheaths. In the peripheral nervous system, the pattern of fluorescence in sciatic nerves indicates predominant localization of tdTomato to non-compact myelin compartments including the inner and outer tongues, paranodal loops and Schmidt-Lanterman incisures. In summary, our work provides a set of novel mouse lines with myelin labeled by red fluorescence, which are compatible with diverse imaging modalities in the green spectral range enabling integrated structural and functional imaging.