An intact insect embryo for developmental neurotoxicity testing of directed axonal elongation

Abstract

proliferation, neurochemical differentiation, migration of neural precursor cells, and electrical activity in randomly formed neural networks (Frank et al., 2018). Most in vitro systems do not recapitulate many of the complex cell-cell and cell-matrix interactions or morphogen gradients in the intact organism that are necessary for normal brain formation and may be subject to significant influence by toxicants (Lein et al., 2005). To address possible adverse effects of chemicals on these complex mechanisms, non-mammalian models such as zebrafish embryos can be successfully employed (Dach et al., 2019). Alternatively, the applicability of invertebrate models such as Caenorhabditis (Avila et al., 2012), planarians (Hagstrom et al., 2019), or Drosophila (Rand, 2010) is recognized. In spite of the phylogenetical distance between vertebrates and invertebrates, mechanisms of neural development appear to be highly conserved (Sánchez-Soriano et al., 2007). Comparative DNT studies between a zebrafish and a

Scanning laser optical tomography resolves developmental neurotoxic effects on pioneer neurons

“…With respect to axonal outgrowth, intact locust embryos can provide an alternative to animal experiments in DNT testing 19 . There are several advantages of this system.…”

“…In a single range finding experiment, the test group was exposed to 10 µM MeHgCl for 18 hours. Using conventional fluorescence microscopy and the scoring scheme of Bergmann et al 19 , we detected reductions of axon extension of the pioneer neurons in a number of specimens. Figure 2A shows the immediately fixed start situation (n = 6) after dissecting the embryo out of the egg.…”

“…To quantify the developmental neurotoxic effect of MeHgCl on pioneer axon extension in our test system, we generated concentration-response curves using the established scoring scheme 19 and the resazurin assay (Fig. 4A).…”

“…Due to conservation of those guidance molecules between vertebrates and invertebrates, the pioneer neurons of the locust embryo may provide a useful model for examining blocking effects of neurotoxic chemicals on axonal outgrowth and navigation. We recently devised an intact embryo culture system in which defects in axonal elongation caused by exposure to chemicals are quantified in concentration-response curves 19 . The comparison of these curves to the biochemically measured general viability of the embryo allows for an evaluation of the DNT potential of chemicals, with regard to the endpoint axon elongation.…”

“…Using the quantification method of Bergmann et al 19 for conventional fluorescence microscopy images, we generate concentration-response curves of axon elongation and determine the acute toxicity both for methylmercury chloride and the alkaloid staurosporine. Our assay identifies not only methylmercury chloride, but also the pro-apoptotic compound staurosporine as developmental neurotoxicants.…”