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Growth cone and dendrite dynamics in zebrafish embryos: early events in synaptogenesis imaged in vivo


We used time-lapse fluorescence microscopy to observe the growth of Mauthner cell axons and their postsynaptic targets, the primary motor neurons, in spinal cords of developing zebrafish embryos. Upon reaching successive motor neurons, the Mauthner growth cone paused briefly before continuing along its path. Varicosities formed at regular intervals and were preferentially associated with the target regions of the primary motor neurons. In addition, the postsynaptic motor neurons showed highly dynamic filopodia, which transiently interacted with both the growth cone and the axon. Both Mauthner cell and motor neurons were highly active, each showing motility sufficient to initiate synaptogenesis.

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Figure 1: Illustration of the Mauthner cell–primary motor neuron system.
Figure 2: Time-lapse imaging of the Mauthner cell growth cone.
Figure 3: Time-lapse imaging of primary motor neurons and filopodia dynamics.
Figure 4: Simultaneous two-photon imaging of the Mauthner growth cone and primary motor neurons.
Figure 5: Projections of the Mauthner axon and sets of primary motor neurons.
Figure 6: Formation of an axonal varicosity.
Figure 7: Interaction of dendritic filopodia with the Mauthner axon.


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Susan Pike conducted initial work on this project. We thank S. Pike and faculty members of the MBL Neural Development and Genetics of the Zebrafish course (Woods Hole, Massachusetts) for help and advice. J.D.J. is a fellow of the Helen Hay Whitney Foundation. This work was supported by NIH grants to S.J.S.

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Correspondence to Stephen J. Smith.

Supplementary information

Video clips corresponding to Figs. 3, 4, 6 and 7

Supplementary Figure 3 (MPEG 503 KB)

Time-lapse imaging of primary motor neurons and filopodia dynamics. Video clips show brief time-lapse sequences of two RoP motor neurons.

Supplementary Figure 4 (MPEG 899 KB)

Simultaneous two-photon imaging of the Mauthner growth cone and primary motor neurons. The video sequence shows maximum-intensity projections from a representative time-lapse sequence during which an M-cell growth cone migrated past a CaP motor neuron. The growth cone was highly active and transiently interacted with the CaP cell, one of its synaptic partners. The growth cone moved past the CaP cell without collapsing, stalling or significantly altering its morphology.

Supplementary Figure 6 (MPEG 899 KB)

Formation of an axonal varicosity. This two-photon time-lapse sequence shows the formation of an axonal varicosity in close association with a CaP motor neuron. The images are maximum-intensity projections of image stacks displayed at five-minute intervals. Initially, the axon swelled, forming a large, elongated varicosity close to the CaP cell. Over time, the varicosity shrank and became more spherical. Throughout the time-lapse, there was extensive filopodia activity on the CaP cell near the developing varicosity.

Supplementary Figure 7 (MPEG 589 KB)

Interaction of dendritic filopodia with the Mauthner axon. This two-photon time-lapse sequence illustrates the interaction of a Mauthner growth cone with a RoP motor neuron as these two cells first came into contact. At the start of this time-lapse sequence, the Mauthner growth cone first reached the RoP ventral dendrite. Each image is a maximum-intensity projection of 14 sections collected at 1-mm steps. Numerous dendritic filopodia extended and retracted from the RoP ventral dendrite, many of which interacted transiently with the Mauthner axon. During this experiment, no stable cell-cell contacts were observed. Because a synapse will ultimately form between the Mauthner axon and the RoP ventral dendrite, it is possible that a filopodium may be responsible for initiating synaptogenic contact.

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Jontes, J., Buchanan, J. & Smith, S. Growth cone and dendrite dynamics in zebrafish embryos: early events in synaptogenesis imaged in vivo . Nat Neurosci 3, 231–237 (2000).

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