Methods for Assessing Neurotoxicity in Zebrafish - Zebrafish: Methods for Assessing Drug Safety and Toxicity - page 122

compounds caused no significant effects, supporting the utility of zebrafish for iden-
tifying potential neuroprotectants (Parng, 2005; Parng et al., 2006).

10.3.6 Compound-Induced Effects on Motility

One of the greatest challenges in developing methods for assessing neurotoxicity is
associating neuromorphological, neurochemical, and neurophysiological alterations
with behavioral changes, frequently assessed as abnormal movement (NRC, 1992).
In zebrafish larvae, patterns of motility or locomotion are stage specific and
behavioral abnormalities can be readily distinguished (Granato and Nusslein-Vol-
hard, 1996). Embryonicmotor behavior develops sequentially and consists of an early
period of transient spontaneous coiling contractions, followed by the emergence of
twitching in response to touch, and later, by the ability to swim (Drapeau et al., 2002).
By 4dpf, embryos are free swimming and change direction spontaneously with
characteristic speed and distance intervals. Zebrafish also exhibit basic behavior
including memory, nonassociative learning, conditioned responses, and social be-
havior such as schooling (Burgess and Granato, 2007; Best et al., 2008).
Several studies have shown that zebrafish motility can be tracked using a
computer-driven motion detector (ViewPoint Life Sciences, Lyons, France) (Emran
et al., 2007; McGrath and Li, 2008; Winter et al., 2008). Using this automated system,
zebrafish movement is captured electronically with a high-resolution black and white
camera (Fig. 10.7a). The number of movements in a given time period, duration, and
distance traveled can be assessed (Fig. 10.7b). Investigators (Baraban et al., 2005;
Prober et al., 2006; Berghmans et al., 2007; McGrath and Li, 2008;Winter et al., 2008)
have demonstrated the usefulness of this assay format for assessing compound-
induced neurotoxicity, including seizures. Similar to results in mammals,
pentylenetetrazole (PTZ), a GABA antagonist known to induce convulsions in
humans, has been shown to induce seizures in zebrafish and the behavioral,
electrophysiological, and molecular changes in PTZ-treated zebrafish were compa-
rable to effects observed in a rodent seizure model. In recent experiments, using the
VideoTrack System, effect of treatment with PTZ on zebrafish swim movement was
assessed. Zebrafish were placed in 24-well microplates, one animal per well.
Acquisition software was programmed to separate swim movements into three
distinct color-coded patterns based on speed: inactivity/low-speed movement ( 4

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mm/s), normal movement (4-20 mm/s), and high-speed movement ( 20 mm/s).

Zebrafish exposed to PTZ exhibited a dramatic increase in high-speed swim move-
ment, compared to carrier control-treated zebrafish that primarily exhibited normal
movement during 60min of motility recording. Total distance traveled at high speed
by combining speed and duration of high-speed movement was analyzed at 1 min
intervals. The average distance traveled (mm) by 10 zebrafish in 1min intervals
increased dramatically and peaked

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7min after PTZ treatment; increased distance
traveled at high speed continued for the duration of the 60min recording period.
Although untreated zebrafish also exhibited high-speed swim movement, this re-
presented a small percentage of total swim movement and did not increase.