Advantages And Limitations Of The Zebrafish Model

The application of our larval zebrafish model to epilepsy research is probably limited only by one's imagination. As discussed in the previous section, a simple application of this model would be to evaluate drugs useful in generalized epilepsy syndromes. One might also consider forward- and

FIGURE 4 Response to antiepileptic drugs. a: Representative tectal field recordings obtained from a zebrafish bathed in normal Ringer's medium plus 15 mM pentylenetetrazol (PTZ) (baseline) and 45 minutes after application of 100 ||M diazepam. b: Representative tectal field recordings obtained from a zebrafish bathed in normal Ringer's medium plus 15 mM PTZ (baseline) and 45 minutes after application of 5mM valproic acid.

FIGURE 4 Response to antiepileptic drugs. a: Representative tectal field recordings obtained from a zebrafish bathed in normal Ringer's medium plus 15 mM pentylenetetrazol (PTZ) (baseline) and 45 minutes after application of 100 ||M diazepam. b: Representative tectal field recordings obtained from a zebrafish bathed in normal Ringer's medium plus 15 mM PTZ (baseline) and 45 minutes after application of 5mM valproic acid.

chemical-genetic screening, commonly used methods to identify gene mutations related to a specific behavior (or phenotype) in an unbiased manner (Malicki, 2000; Patton and Zon, 2001; Specht and Shokat, 2002). Such screening methods provide a strategy to test available AEDs on muta-genized zebrafish larvae, and perhaps will lead to a better understanding of why some patients are refractory to drug treatment. Pharmacogenic strategies (Weinshilboum and Wang, 2004) designed to identify gene mutations that underlie inter-individual variations in AED responsiveness are also possible. When designing zebrafish pharmacologic studies, it is important to consider an empirical testing of drug concentrations; as mentioned above, drug availability and distribution data are not available. Of course this model need not be limited to AED discovery; and not unlike many of the animal models described in this volume, it can be used to study directly how an epileptic brain functions. Epilepsy, particularly pediatric forms of epilepsy, is a neurologic disease that is defined by abnormal electrical discharge, and insights into the basis of this activity often follow from experimental animal model research. In particular the now widespread use of extracellular and intracellular recording techniques in rodent seizure models has provided evidence for intrinsic cellular and synaptic dysfunction that underlie generation of interictal and ictal epileptic seizures. Such research has led, for example, to identification of fuctional changes in H-channels in a febrile seizure model (Bender et al., 2004), and insight into how M-type potassium channel mutations lead to benign familial neonatal convulsions (BFNC) in mutant mice (Watanabe et al., 2000) have benefited from electrophysiology research. Extracellular recording techniques were demonstrated to be feasible (and relatively easy to obtain) in our zebrafish model, and it is not difficult to imagine that intracellular approaches are also possible. Because larval zebrafish possess far fewer neurons and synaptic circuits than immature rodents (but exhibit similar types of complex epileptiform discharge; see Figure 2), analysis of cellular mechanisms in this model could be a fruitful experimental strategy to identify the basic elements required for generation of epileptic seizures. Indeed, owing to the high degree of homology between zebrafish and human genomes, it is likely that identification of genes involved in the basic processes of seizure genesis, seizure-induced neurogenesis, and epileptogenesis will lead directly to insights into the human condition. If one combines this model with morpholino antisense oligonucleotides (Corey and Abrams, 2001) or transgenic methods (Stuart et al., 1990) to alter gene expression in developing zebrafish, a still wider set of experimental questions can be addressed. Perhaps the most significant application of our model, and one that our laboratory currently employs, is to use the zebrafish model to examine the genetic basis of epilepsy. Several types of applications can be envisioned, and we have chosen a forward-genetic screening strategy to identify

"seizure resistant" zebrafish mutants. This research program is based on a fundamental question of how to identify genes that prevent, or protect, an individual from developing epilepsy. Because larval zebrafish are a pediatric epilepsy model, perhaps these types of studies could lead to new therapies for the large number of children suffering with medically intractable forms of epilepsy. Although it is too early to speculate on how data from zebrafish will be extrapolated to human pediatric epilepsy disorders, especially considering the vast complexity of the human brain in relation to the relative simplicity of the zebrafish CNS, it is safe to predict that the process of discovery will lead to novel insights.

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