Directing growth cones of optic axons growing with laser scissors and laser tweezers

We have combined a laser scissors and a laser tweezers to study, (1) the response of nerve fiber growth cones to laserinduced damage on single axons, and (2) localized microfluidic flow generated by laser-driven spinning birefringent particles. In the laser scissors study, sub-axotomy damage elicits a growth cone response whether damage is on the same or an adjacent axon. In laser tweezers study, the axon growth cones turn in response to the optically driven microfluidic flow. In summary, both the laser scissors and the laser tweezers studies elicit growth cone turning responses

Intraocular BDNF promotes ectopic branching, alters motility and stimulates abnormal collaterals in regenerating optic fibers

A great deal of effort has been invested in using trophic factors and other bioactive molecules to promote cell survival and axonal regeneration in the adult central nervous system. Far less attention has been paid to investigating potential effects that trophic factors may have that might interfere with recovery. In the visual system, BDNF has been previously reported to prevent regeneration. To test if BDNF is inherently incompatible with regeneration, BDNF was given intraocularly during optic nerve regeneration in the adult goldfish. In vivo imaging and anatomical analysis of selectively labeled axons were used as a sensitive assay for effects on regeneration within the tectum. BDNF had no detectable inhibitory effect on the ability of axons to regenerate. Normal numbers of axons regenerated into the tectum, exhibited dynamic growth and retractions similar to controls, and were able to navigate to their correct target zone in the tectum. However, BDNF was found to have additional effects that adversely affected the quality of regeneration. It promoted premature branching at ectopic locations, diminished the growth rate of axons through the tectum, and resulted in the formation of ectopic collaterals. Thus, although BDNF has robust effects on axonal behavior, it is, nevertheless, compatible with axonal regeneration, axon navigation and the formation of terminal arbors

Neuronal growth cones respond to laser-induced axonal damage.

Although it is well known that damage to neurons results in release of substances that inhibit axonal growth, release of chemical signals from damaged axons that attract axon growth cones has not been observed. In this study, a 532 nm 12 ns laser was focused to a diffraction-limited spot to produce site-specific damage to single goldfish axons in vitro. The axons underwent a localized decrease in thickness ('thinning') within seconds. Analysis by fluorescence and transmission electron microscopy indicated that there was no gross rupture of the cell membrane. Mitochondrial transport along the axonal cytoskeleton immediately stopped at the damage site, but recovered over several minutes. Within seconds of damage nearby growth cones extended filopodia towards the injury and were often observed to contact the damaged site. Turning of the growth cone towards the injured axon also was observed. Repair of the laser-induced damage was evidenced by recovery of the axon thickness as well as restoration of mitochondrial movement. We describe a new process of growth cone response to damaged axons. This has been possible through the interface of optics (laser subcellular surgery), fluorescence and electron microscopy, and a goldfish retinal ganglion cell culture model

Intraocular BDNF promotes ectopic branching, alters motility and stimulates abnormal collaterals in regenerating optic fibers

A great deal of effort has been invested in using trophic factors and other bioactive molecules to promote cell survival and axonal regeneration in the adult central nervous system. Far less attention has been paid to investigating potential effects that trophic factors may have that might interfere with recovery. In the visual system, BDNF has been previously reported to prevent regeneration. To test if BDNF is inherently incompatible with regeneration, BDNF was given intraocularly during optic nerve regeneration in the adult goldfish. In vivo imaging and anatomical analysis of selectively labeled axons were used as a sensitive assay for effects on regeneration within the tectum. BDNF had no detectable inhibitory effect on the ability of axons to regenerate. Normal numbers of axons regenerated into the tectum, exhibited dynamic growth and retractions similar to controls, and were able to navigate to their correct target zone in the tectum. However, BDNF was found to have additional effects that adversely affected the quality of regeneration. It promoted premature branching at ectopic locations, diminished the growth rate of axons through the tectum, and resulted in the formation of ectopic collaterals. Thus, although BDNF has robust effects on axonal behavior, it is, nevertheless, compatible with axonal regeneration, axon navigation and the formation of terminal arbors

A photon-driven micromotor can direct nerve fibre growth

Axonal path-finding is important in the development of the nervous system, nerve repair and nerve regeneration. The behaviour of the growth cone at the tip of the growing axon determines the direction of axonal growth and migration. We have developed an optical-based system to control the direction of growth of individual axons (nerve fibres) using laser-driven spinning birefringent spheres. One or two optical traps position birefringent beads adjacent to growth cones of cultured goldfish retinal ganglion cell axons. Circularly polarized light with angular momentum causes the trapped bead to spin. This creates a localized microfluidic flow generating an estimated 0.17 pN shear force against the growth cone that turns in response to the shear. The direction of axonal growth can be precisely manipulated by changing the rotation direction and position of this optically driven micromotor. A physical model estimating the shear force density on the axon is described