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Laser surgery of zebrafish (Danio rerio) embryos using femtosecond laser pulses: optimal parameters for exogenous material delivery, and the laser's effect on short- and long-term development.

Kohli V, Elezzabi AY - BMC Biotechnol. (2008)

Bottom Line: An exogenous fluorescent probe, fluorescein isothiocyanate (FITC), was successfully introduced into blastomere cells and found to diffuse throughout all developing cells.In our study, no significant differences in hatching rates and developmental morphologies were observed in laser-manipulated samples relative to controls.This tool represents an effective non-destructive technique for potential medical and biological applications.

View Article: PubMed Central - HTML - PubMed

Affiliation: 9107-116 St, Ultrafast Photonics and Nano-Optics Laboratory, Department of Electrical and Computer Engineering, University of Alberta, Edmonton, T6G2V4, Canada. vkohli@ece.ualberta.ca

ABSTRACT

Background: Femtosecond (fs) laser pulses have recently received wide interest as an alternative tool for manipulating living biological systems. In various model organisms the excision of cellular components and the intracellular delivery of foreign exogenous materials have been reported. However, the effect of the applied fs laser pulses on cell viability and development has yet to be determined. Using the zebrafish (Danio rerio) as our animal model system, we address both the short- and long-term developmental changes following laser surgery on zebrafish embryonic cells.

Results: An exogenous fluorescent probe, fluorescein isothiocyanate (FITC), was successfully introduced into blastomere cells and found to diffuse throughout all developing cells. Using the reported manipulation tool, we addressed whether the applied fs laser pulses induced any short- or long-term developmental effects in embryos reared to 2 and 7 days post-fertilization (dpf). Using light microscopy and scanning electron microscopy we compared key developmental features of laser-manipulated and control samples, including the olfactory pit, dorsal, ventral and pectoral fins, notochord, pectoral fin buds, otic capsule, otic vesicle, neuromast patterning, and kinocilia of the olfactory pit rim and cristae of the lateral wall of the ear.

Conclusion: In our study, no significant differences in hatching rates and developmental morphologies were observed in laser-manipulated samples relative to controls. This tool represents an effective non-destructive technique for potential medical and biological applications.

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Related in: MedlinePlus

High magnification SEM images of the olfactory pit rim and kinocilia projecting from cristae on the lateral wall into the lumen. (a) Olfactory pit rim in a laser-manipulated and (b) a control larva showing kinocilia covering the inside wall of the olfactory pit. Kinocilia projecting from lateral cristae in (c) a laser-manipulated and (d) a control larva. Scale bars for (a, b, c, d) represent 1 μm.
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Figure 13: High magnification SEM images of the olfactory pit rim and kinocilia projecting from cristae on the lateral wall into the lumen. (a) Olfactory pit rim in a laser-manipulated and (b) a control larva showing kinocilia covering the inside wall of the olfactory pit. Kinocilia projecting from lateral cristae in (c) a laser-manipulated and (d) a control larva. Scale bars for (a, b, c, d) represent 1 μm.

Mentions: The zebrafish olfactory organ is responsible for detecting and discriminating thousands of odorants [42]. Courtship depends upon the chemosensitivity of the organ, where specific pheromones act as attractants [42]. Zebrafish rendered anosmic are unable to reproduce [42], indicating that this organ is functionally essential. It has been hypothesized that the organ forms from subepidermal cells, some of which form the olfactory placode [43]. The placode cells differentiate into receptors, supporting cells and ciliated cells [43], which eventually give rise to the olfactory organ. As the olfactory organ develops, the epidermal cells spread, revealing the olfactory pit located on either side of the head posterior to the eye. Figures 11(a) and 11(d) depict the olfactory pits of a laser-manipulated and a control larva reared to 7 dpf, respectively. In both SEM images, epidermal cells surrounded the olfactory pit. Inside the pits, ciliated receptor cells and kinocilia were observed in both laser-manipulated and control larvae. Figures 13(a) and 13(b) show high magnification SEM images of the olfactory pit rim. We noticed that in both larvae, the pit rims were covered by long kinocilia densely covering the inside walls. Together, Figures 11(a,d) and 13(a,b) indicate that the olfactory pits of the laser-manipulated larvae appear to develop similarly to the controls.


Laser surgery of zebrafish (Danio rerio) embryos using femtosecond laser pulses: optimal parameters for exogenous material delivery, and the laser's effect on short- and long-term development.

Kohli V, Elezzabi AY - BMC Biotechnol. (2008)

High magnification SEM images of the olfactory pit rim and kinocilia projecting from cristae on the lateral wall into the lumen. (a) Olfactory pit rim in a laser-manipulated and (b) a control larva showing kinocilia covering the inside wall of the olfactory pit. Kinocilia projecting from lateral cristae in (c) a laser-manipulated and (d) a control larva. Scale bars for (a, b, c, d) represent 1 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2270268&req=5

Figure 13: High magnification SEM images of the olfactory pit rim and kinocilia projecting from cristae on the lateral wall into the lumen. (a) Olfactory pit rim in a laser-manipulated and (b) a control larva showing kinocilia covering the inside wall of the olfactory pit. Kinocilia projecting from lateral cristae in (c) a laser-manipulated and (d) a control larva. Scale bars for (a, b, c, d) represent 1 μm.
Mentions: The zebrafish olfactory organ is responsible for detecting and discriminating thousands of odorants [42]. Courtship depends upon the chemosensitivity of the organ, where specific pheromones act as attractants [42]. Zebrafish rendered anosmic are unable to reproduce [42], indicating that this organ is functionally essential. It has been hypothesized that the organ forms from subepidermal cells, some of which form the olfactory placode [43]. The placode cells differentiate into receptors, supporting cells and ciliated cells [43], which eventually give rise to the olfactory organ. As the olfactory organ develops, the epidermal cells spread, revealing the olfactory pit located on either side of the head posterior to the eye. Figures 11(a) and 11(d) depict the olfactory pits of a laser-manipulated and a control larva reared to 7 dpf, respectively. In both SEM images, epidermal cells surrounded the olfactory pit. Inside the pits, ciliated receptor cells and kinocilia were observed in both laser-manipulated and control larvae. Figures 13(a) and 13(b) show high magnification SEM images of the olfactory pit rim. We noticed that in both larvae, the pit rims were covered by long kinocilia densely covering the inside walls. Together, Figures 11(a,d) and 13(a,b) indicate that the olfactory pits of the laser-manipulated larvae appear to develop similarly to the controls.

Bottom Line: An exogenous fluorescent probe, fluorescein isothiocyanate (FITC), was successfully introduced into blastomere cells and found to diffuse throughout all developing cells.In our study, no significant differences in hatching rates and developmental morphologies were observed in laser-manipulated samples relative to controls.This tool represents an effective non-destructive technique for potential medical and biological applications.

View Article: PubMed Central - HTML - PubMed

Affiliation: 9107-116 St, Ultrafast Photonics and Nano-Optics Laboratory, Department of Electrical and Computer Engineering, University of Alberta, Edmonton, T6G2V4, Canada. vkohli@ece.ualberta.ca

ABSTRACT

Background: Femtosecond (fs) laser pulses have recently received wide interest as an alternative tool for manipulating living biological systems. In various model organisms the excision of cellular components and the intracellular delivery of foreign exogenous materials have been reported. However, the effect of the applied fs laser pulses on cell viability and development has yet to be determined. Using the zebrafish (Danio rerio) as our animal model system, we address both the short- and long-term developmental changes following laser surgery on zebrafish embryonic cells.

Results: An exogenous fluorescent probe, fluorescein isothiocyanate (FITC), was successfully introduced into blastomere cells and found to diffuse throughout all developing cells. Using the reported manipulation tool, we addressed whether the applied fs laser pulses induced any short- or long-term developmental effects in embryos reared to 2 and 7 days post-fertilization (dpf). Using light microscopy and scanning electron microscopy we compared key developmental features of laser-manipulated and control samples, including the olfactory pit, dorsal, ventral and pectoral fins, notochord, pectoral fin buds, otic capsule, otic vesicle, neuromast patterning, and kinocilia of the olfactory pit rim and cristae of the lateral wall of the ear.

Conclusion: In our study, no significant differences in hatching rates and developmental morphologies were observed in laser-manipulated samples relative to controls. This tool represents an effective non-destructive technique for potential medical and biological applications.

Show MeSH
Related in: MedlinePlus