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.
Highlights
The development of the vertebrate neuromuscular junction (NMJ) is widely studied for understanding the cellular and molecular control of synaptogenesis [1]
Our results suggest that the balance between neuronal fibroblast growth factor (FGF) receptor 1 (FGFR1)- and tropomyosin-related kinase B (TrkB)-dependent filopodial assembly and axonal outgrowth regulates the establishment of incipient NMJs
We found that blocking filopodial assembly in either muscle or nerve hindered NMJ development in Xenopus nervemuscle cocultures [14,26], and that basic fibroblast growth factor (bFGF) produced by muscle activated FGFR1 in neurons to induce filopodia which facilitate NMJ establishment [14]
Summary
The development of the vertebrate neuromuscular junction (NMJ) is widely studied for understanding the cellular and molecular control of synaptogenesis [1]. During NMJ assembly the tip of a motor neuron – the axonal growth cone – approaches and contacts muscle and transforms into a presynaptic terminal specialized for secreting the neurotransmitter acetylcholine (ACh) The hallmark of this differentiation process is the accumulation of synaptic vesicles and mitochondria in the nerve terminal [1,2]. Supporting our BDNF results, relative to control GFP-neurons (Figure 5, A and A’) WTTrkB-expressing neurons developed fewer filopodia (Figure 5, B and B’) and TR-TrkB-expressing neurons grew more filopodia (Figure 5, C and C’) Comparison of these filopodial densities (in Figure 5D) with those from FGFR1 experiments above shows that the expression of WT-TrkB, like that of TR-FGFR1, reduced filopodial assembly, and that the introduction into neurons of ‘‘dominant-negative’’ TrkB enhanced filopodial growth, like WTFGFR1 overexpression. Here we investigated how bFGF- and NT-signals in neurons jointly elicit presynaptic changes during the earliest stages of NMJ development
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