Intercellular crosstalk mediated by tunneling nanotubes between central nervous system cells. What we need to advance

Abstract

Long-range intercellular communication between Central Nervous System (CNS) cells is an essential process for preserving CNS homeostasis. Paracrine signaling, extracellular vesicles, neurotransmitters and synapses are well-known mechanisms involved. A new form of intercellular crosstalk mechanism based on Tunneling Nanotubes (TNTs), suggests a new way to understand how neural cells interact with each other in controlling CNS functions. TNTs are long intercellular bridges that allow the intercellular transfer of cargoes and signals from one cell to another contributing to the control of tissue functionality. CNS cells communicate with each other via TNTs, through which ions, organelles and other signals are exchanged. Unfortunately, almost all these results were obtained through 2D in-vitro models, and fundamental mechanisms underlying TNTs-formation still remain elusive. Consequently, many questions remain open, and TNTs role in CNS remains largely unknown. In this review, we briefly discuss the state of the art regarding TNTs identification and function. We highlight the gaps in the knowledge of TNTs and discuss what is needed to accelerate TNTs-research in CNS-physiology. To this end, it is necessary to: 1) Develop an ad-hoc TNTs-imaging and software-assisted processing tool to improve TNTs-identification and quantification, 2) Identify specific molecular pathways involved into TNTs-formation, 3) Use in-vitro 3D-CNS and animal models to investigate TNTs-role in a more physiological context pushing the limit of live-microscopy techniques. Although there are still many steps to be taken, we believe that the study of TNTs is a new and fascinating frontier that could significantly contribute to deciphering CNS physiology.

Keywords: central nervous system; in-vitro 3D model; intercellular communication; super resolution live-cell microscopy; tunneling nanotubes.

FIGURE 1

Ideal workflow and imaging techniques to deciphering tunneling nanotubes mediated crosstalk between central nervous system cells. (A) To make progress in understanding the triggering factors and functions of TNTs in the central nervous system, we require an integrated system including: 1) specific in vitro and in vivo models, 2) super-resolution live microscopy and, 3) software-assisted data analysis. In vitro and in vivo models are valuable for investigating the intercellular transfer of cargoes such as ions, RNAs, proteins, and organelles. High-resolution live-cell confocal microscopy techniques, including spinning disk and structured illumination super-resolution microscopy, are used to analyze these models. The data produced by 3D reconstruction are then analyzed using artificial intelligence (AI) approaches to obtain unbiased TNT quantification. For in vitro data, the AI-based software must be capable of discriminating TNT as F-actin positive, straight structures detached from the substrate that are capable of transferring cargo from one cell to another. With the use of these tools, we can quickly and objectively test candidate stimuli that trigger or destroy TNTs. Ultimately, this approach could speed up the identification of specific molecular pathways involved in TNT formation and the functions of TNTs in the CNS. (B) Sample type and imaging techniques for efficient TNTs analysis in fixed and live samples.