Methods for Assessing Neurotoxicity in Zebrafish - Zebrafish: Methods for Assessing Drug Safety and Toxicity - page 124

such as differentiation of specific cell type, and proliferation and migration of nascent
neurons (Barone et al., 2000). In addition, the developing brain is more exposed to
blood-borne toxins prior to formation of the BBB. Therefore, critical assessment of
developmental neurotoxicity requires use of an embryonic model. The U.S. Envi-
ronmental Protection Agency (U.S. EPA) and the Organization for Economic
Cooperation and Development (OECD) guidelines for developmental toxicity testing
contain protocols using a rat embryonic model (U.S. EPA, 1998; Mileson and
Ferenc, 2001; OECD, 2003). This model is well developed and includes several
developmental, histological, and behavioral end points. Developmental landmarks
include size, time of vaginal opening, and time of preputial separation. Histological
methods include assessment of brain weight, histopathology, and qualitative and
semiquantitative morphometric analyses. Behavioral end points include measure-
ment of motor activity, auditory startle, and learning and memory. The rat model
allows comprehensive and predictive testing; however, these studies require a
minimum of 60 days per test and are experimentally complex, which hinders testing.
In a recent study, seven well-characterized compounds were selected to validate
zebrafish as a developmental neuorotoxicity model (Table 10.1) (Ton et al., 2006).
Embryos were exposed by semistatic immersion from 6hpf to 96 or 120hpf;
teratogenicity was assessed using a modified method previously reported
(Ton et al., 2006). Atrazine, dichlorodiphenyltrichloroethane (DDT), and 2,3,7,8-
tetrachlorodibenzo- p -dioxin (TCDD) were found to be primarily teratogenic and not
specifically neurotoxic. 2,4-Dichlorophenoxyacetic acid (2,4-D), dieldrin, and non-
ylphenol showed specific neurotoxicity; dieldrin and nonylphenol were specifically
toxic to catecholaminergic neurons. Exposure to 100 Mdieldrin also caused reduced

axon tracts in the brain (Fig. 10.8). Although not teratogenic, malathion showed some
nonspecific toxicity. Induction of brain apoptosis or necrosis was confirmed as an
indicator of developmental neurotoxicity and an effect on motor neurons in the caudal
third of the embryo was shown to correlate with expected defects in motility. In
another developmental neurotoxicity project funded by the U.S. National Science
Foundation, 200 environmental contaminants were screened for effects on brain
apoptosis, axon tract and motor neuron formation, and catecholaminergic neurons in
developing zebrafish and several compounds that showed effects on one or more of the
neurotoxicity parameters were identified and some compounds showed broad toxicity

m

Table 10.1 Comparison of Teratogenicity in Mammals and Zebrafish

Compound Mammals Zebrafish (96hpf)

2,4-D No (rats, rabbits), slight (mouse) Slight

Atrazine No (mouse) Yes

DDT Yes (mouse) Yes

Dieldrin No No

Malathion No (rat, rabbit) No

Nonylphenol No (rat) No

TCDD High (rat, mouse, hamster) High