Reciprocal Regulation of Axonal Filopodia and Outgrowth during Neuromuscular Junction Development

Abstract

BackgroundThe assembly of the vertebrate neuromuscular junction (NMJ) is initiated when nerve and muscle first contact each other by filopodial processes which are thought to enable close interactions between the synaptic partners and facilitate synaptogenesis. We recently reported that embryonic Xenopus spinal neurons preferentially extended filopodia towards cocultured muscle cells and that basic fibroblast growth factor (bFGF) produced by muscle activated neuronal FGF receptor 1 (FGFR1) to induce filopodia and favor synaptogenesis. Intriguingly, in an earlier study we found that neurotrophins (NTs), a different set of target-derived factors that act through Trk receptor tyrosine kinases, promoted neuronal growth but hindered presynaptic differentiation and NMJ formation. Thus, here we investigated how bFGF- and NT-signals in neurons jointly elicit presynaptic changes during the earliest stages of NMJ development.Methodology/Principal FindingsWhereas forced expression of wild-type TrkB in neurons reduced filopodial extension and triggered axonal outgrowth, expression of a mutant TrkB lacking the intracellular kinase domain enhanced filopodial growth and slowed axonal advance. Neurons overexpressing wild-type FGFR1 also displayed more filopodia than control neurons, in accord with our previous findings, and, notably, this elevation in filopodial density was suppressed when neurons were chronically treated from the beginning of the culture period with BDNF, the NT that specifically activates TrkB. Conversely, inhibition by BDNF of NMJ formation in nerve-muscle cocultures was partly reversed by the overexpression of bFGF in muscle.ConclusionsOur results suggest that the balance between neuronal FGFR1- and TrkB-dependent filopodial assembly and axonal outgrowth regulates the establishment of incipient NMJs.

The ketone body metabolite β‐hydroxybutyrate induces an antidepression‐associated ramification of microglia via HDACs inhibition‐triggered Akt‐small RhoGTPase activation

“…Rac1 and Cdc42, the most famous small RhoGTPases, are considered to mediate the formation of lamellipodia and filopodia, two structures that are essential for phagocytosis, cell mobility, and neuronal outgrowth (Giralt, Coura, & Girault, 2016;Li, Zhou, Meng, Madhavan, & Peng, 2012;Neubrand et al, 2014). We showed here that the proramification and pro-phagocytosis effect of BHB were dependent on Akt and small RhoGTPase activation.…”

“…The cell shape change is associated with the rearrangement of cellular actin filaments and microtubule cytoskeletons (Seiberlich et al, ), and this process is mediated by small RhoGTPases and their upstream kinases such as phosphatidylinositol‐3‐kinase (PI3K) and protein kinase B (Akt) (Huang et al, ; Neubrand et al, ). Rac1 and Cdc42, the most famous small RhoGTPases, are considered to mediate the formation of lamellipodia and filopodia, two structures that are essential for phagocytosis, cell mobility, and neuronal outgrowth (Giralt, Coura, & Girault, ; Li, Zhou, Meng, Madhavan, & Peng, ; Neubrand et al, ). We showed here that the pro‐ramification and pro‐phagocytosis effect of BHB were dependent on Akt and small RhoGTPase activation.…”

Heparanase 2, mutated in urofacial syndrome, mediates peripheral neural development in Xenopus

“…Given that (i) heparanase 2 inhibits the activity of heparanase 1 ( 17 ), (ii) heparanase 1 enhances FGF2 bioavailability and its signalling via pERK ( 14 – 16 ), (iii) FGF signalling is implicated in neuromuscular development ( 40 , 41 ) and (iv) transcripts encoding heparanase 1 and FGF2 were elevated in heparanase 2 morphants, we next analysed pERK levels in heparanase 2 morphants versus controls and compared tissue expression patterns of pERK and heparanase 2 in wild-type embryos. Protein was extracted from pools of whole embryos (three for each blot) harvested at different stages and both pERK and total ERK (tERK) immunoreactive proteins sought by western blotting (Fig.…”

From gene discovery to new biological mechanisms: heparanases and congenital urinary bladder disease

“…These emerging data place HPSE2 and LRIG2 within developing peripheral nerves. Growth factors, including several bound by HSPGs, are implicated in mediating key steps of motor neuron differentiation, including the specification of neuron precursor cells, axonal growth from these cells and synaptogenesis at neuromuscular junctions [ 48 , 51 , 52 ]. What is less well understood, however, is how these processes are regulated so that growth factor signalling is tuned to avoid over- or underactivity, either of which could fatally compromise the generation of functional neuromuscular units.…”