Fast rearrangement of the neuronal growth cone’s actin cytoskeleton following VEGF stimulation

Abstract

The neuronal growth cone plays a crucial role in the development of the nervous system. This highly motile structure leads the axon to its final destination by translating guidance cues into cytoskeletal rearrangements. Recently, vascular endothelial growth factor (VEGF), which is essential for angiogenesis and vascular sprouting, has been found to exert a trophic activity also on neurons, leading to an increased axonal outgrowth, similar to the well-known nerve growth factor (NGF). The neurotrophic properties of VEGF are likely to be promoted via the VEGF receptor 2 (VEGFR-2) and neuropilin-1 (NRP-1). In the long term, VEGF attracts and influences the growth cone velocity and leads to growth cone enlargement. The present study focuses on immediate VEGF effects using RFP-actin and GFP-NF-M microinjected chicken dorsal root ganglia for live cell imaging of the neuronal growth cone. We analyzed actin and neurofilament dynamics following VEGF and NGF treatment and compared the effects. Furthermore, key signaling pathways of VEGF were investigated by specific blocking of VEGFR-2 or NRP-1. With the aid of confocal laser scanning microscopy and stimulated emission depletion microscopy, we show for the first time that VEGF has a quick effect on the actin-cytoskeleton, since actin rearrangements were identifiable within a few minutes, leading to a dramatically increased motion. Moreover, these effects were strongly enhanced by adding both VEGF and NGF. Most notably, the effects were inhibited by blocking VEGFR-2, therefore we propose that the immediate effects of VEGF on the actin-cytoskeleton are mediated through VEGFR-2.

Rapid Impact of Progesterone on the Neuronal Growth Cone

“…Controls were obtained by incubating samples in nutrient medium, lacking any supple- mented growth factors but horse serum. In concordance with other studies (18,25) limited neuritic outgrowth was detectable, whereas treatment with 50 ng/mL NGF ( Figure 1B), 10nM progesterone ( Figure 1C), and progesterone plus NGF ( Figure 1D) augmented the outspread, extending several hundreds of micrometers with ramified neuronal cell processes. Neurons exposed to NGF showed many neurites extending into the periphery and forming dense networks ( Figure 1B).…”

“…Nevertheless, it has been described that the application of chemoattractants leads to an increased protrusion, an accumulation of F-actin, and an augmented barbed-end density within the growth cone near the locus of application (38). Such observations have been reported for instance for vascular endothelial growth factor (18,25,58). Furthermore, studies by Argiro and colleagues (59) showed a direct correlation between growth cone shape and motility.…”

“…NGF was shown to be an attractive cue for neuronal growth cones in numerous studies, favoring actin polymerizations and stabilizations by interaction with actin binding proteins like actin depolymerizing factor/cofilin (63)(64)(65). Thus, an up-regulation of the amount of NGF receptor would increase the growth cone´s sensitivity for NGF and in turn increase the growth cone's area and circumference (18,53,57,(62)(63)(64)(65)(66). Furthermore, ventral midline antigen/VEM1, a certain isoform of PGRMC1, was discussed as being involved in axonal pathfinding by the regulation of surface availability of guidance receptors (12,67).…”

ALS as a distal axonopathy: molecular mechanisms affecting neuromuscular junction stability in the presymptomatic stages of the disease

“…During development, the neuronal growth cone directs the developing axon by translating environmental guidance cues into cytoskeletal rearrangements. New evidence has revealed that upon VEGF stimulation, the actin cytoskeleton in the growth cone undergoes immediate and fast rearrangement that enables increased motion (Olbrich et al, 2013). One could speculate that a similar event is occurring at the NMJ in ALS mice upon trophic factor administration; the cytoskeleton of the motor neuron terminal is modified to a highly active state by the presence of the trophic factor at the NMJ enabling it to remain stable in an otherwise destructive environment.…”

Sensor potency of the moonlighting enzyme-decorated cytoskeleton: the cytoskeleton as a metabolic sensor

“…IQGAPs are actin-binding proteins that transmit extracellular signals to the actin network so as to influence mitogenic, morphological and migratory cell behavior [59,60]. In growth cones, ezrin/radixin/moesin (ERM) proteins tether actin filaments to the cell membrane and transmit Nerve growth factor and neurotrophin-3 signals to the actin network, which leads to its remodeling and to the redistribution of adhesion receptors [61]; similarly, the vascular endothelial growth factor receptor 2 transduces the VEGF signal into major actin rearrangements [62]. In A .…”