Cholesterol Depletion Regulates Axonal Growth and Enhances Central and Peripheral Nerve Regeneration.

Cristina Roselló-Busquets,Marta Pascual,Ashraf Muhaisen,Ramón Martínez-Mármol,Natalia De La Oliva,Eduardo Soriano,Jaume Del Valle,Xavier Navarro,Marc Hernaiz-Llorens

doi:10.3389/fncel.2019.00040

Abstract

Axonal growth during normal development and axonal regeneration rely on the action of many receptor signaling systems and complexes, most of them located in specialized raft membrane microdomains with a precise lipid composition. Cholesterol is a component of membrane rafts and the integrity of these structures depends on the concentrations present of this compound. Here we explored the effect of cholesterol depletion in both developing neurons and regenerating axons. First, we show that cholesterol depletion in vitro in developing neurons from the central and peripheral nervous systems increases the size of growth cones, the density of filopodium-like structures and the number of neurite branching points. Next, we demonstrate that cholesterol depletion enhances axonal regeneration after axotomy in vitro both in a microfluidic system using dissociated hippocampal neurons and in a slice-coculture organotypic model of axotomy and regeneration. Finally, using axotomy experiments in the sciatic nerve, we also show that cholesterol depletion favors axonal regeneration in vivo. Importantly, the enhanced regeneration observed in peripheral axons also correlated with earlier electrophysiological responses, thereby indicating functional recovery following the regeneration. Taken together, our results suggest that cholesterol depletion per se is able to promote axonal growth in developing axons and to increase axonal regeneration in vitro and in vivo both in the central and peripheral nervous systems.

Highlights

Axonal guidance during the development of the nervous system is thought to be highly regulated through interactions of transmembrane receptors with attractive, repulsive, and trophic cues
Our results show that cholesterol depletion increases the growth cone area in hippocampal and external granular layer (EGL) neurons in the Central Nervous System (CNS)
These contradictions can be explained by the fact that, in addition to a variety of proteins and molecules that promote axonal regeneration, such as receptors of neurotrophic factors, lipid rafts contain proteins and molecules that exert an inhibitory role, such as receptors of myelin-associated proteins (Kappagantula et al, 2014)

Summary

Introduction

Axonal guidance during the development of the nervous system is thought to be highly regulated through interactions of transmembrane receptors with attractive, repulsive, and trophic cues. The transected axon undergoes morphological changes to form the growth cone, a highly dynamic structure that senses the environment and leads the regenerative growth (Allodi et al, 2012). Membrane receptors localized in the growth cone have an important role in axonal signaling (Guirland et al, 2004; Kamiguchi, 2006). The regenerative shift of axotomized neurons is promoted by injury-induced signals, which stimulate the transcription of various trophic factors, Cholesterol and Axonal Regeneration adhesion molecules, growth-associated proteins and structural components needed for axonal regrowth and cell survival (Rishal and Fainzilber, 2014). Due to the importance of these signals during axonal degeneration and regeneration after peripheral nerve injury, microdomains in the membrane that cluster a range of proteins and molecules related to cellular signaling may play a key role in the regulation of these pathways. Lipid rafts have been described as cholesterol-enriched cell membrane microdomains that compartmentalize lipids and protein to form signaling platforms (Golub et al, 2004)

Methods

Results

Conclusion