Characterizing cytoskeleton changes during axonal degeneration

UNSAIN N ; BORDENAVE M; JALIL S; MARÍN JI; VON BILDERLING C; JOHNSTONE AD; BISBAL M; BARKER PA; STEFANI FD; CACERES AO

San Diego

Congreso; Society for Neuroscience Annual Meeting - Neuroscience 2016; 2016

Society for Neuroscience

Axonal fragmentation is a regulated process that uses a growing set of receptors, signaling molecules, proteases and other regulators to disintegrate the axonal compartment. Little is known about the changes (and possible role) of the axonal cytoskeleton during degeneration. In this study we aimed at describing cytoskeletal changes associated with axonal degeneration induced by of trophic factor (NGF) withdrawal (TFW) or injury in cultured DRG neurons. We first focused our attention in the actin-rich growth cone and found that growth cone collapse (GCC) is an early event observed in TFW and injured axons. Live-imaging shows that GCC is almost complete by 30 minutes after TFW, which is well before the 18-24 hours needed to observe axon fragmentation. Accompanying increased filopodia dynamics at the axonal tip suggest that GCC is caused by a sudden increase in F-actin dynamics. To our surprise, drugs (Nicotinamide Adenine Dinucleotide, EGTA, go6976, N-Acetyl Cysteine) or gene deletions (Caspase-3 -/-) known to prevent axonal fragmentation do not prevent GCC. Growth cone collapse does not represent a point of no return towards axonal fragmentation since re-adding NGF after GCC (6-12 hours) prevents axonal fragmentation and growth cones are re-form. Total F-actin staining decreases in every cell compartment during degeneration. On the other hand, axonal microtubules form tight bundles and we observed an early and marked microtubule de-bundling in degenerating axons and microtubules curves at the tip of axons where growth cones collapsed. These previously unnoticed early changes in the cytoskeleton suggest an instructive role for the cytoskeleton during axon fragmentation.