Many brain malformations and abnormalities are caused by genetic and environmental factors. Several animal models of CD have been previously described: genetic, fetal insults, and neonatal lesion models (Sarkisian, 2001).
Genetic models of CD
Although models that clearly mimic human malformations are still limited, some genetic models seem to reproduce some rare dysplastic pathologies. The telencephalic internal structural heterotopia (TISH) model describes a mutant rat exhibiting a forebrain anomaly resembling the rare human neuronal migration disorder of double cortex. The bilateral heterotopia is prominent below the frontal and parietal neocortices, but is rarely observed in temporal neocortex. The cells in the heterotopia exhibit a “rim-to-core” neurogenetic pattern rather than the characteristic “inside-out” pattern observed in normotopic neocortex. Studies have shown that some of the animals with the TISH phenotype exhibit spontaneous recurrent electrographic and behavioral seizures (Lee et al., 1997).
Difficulties encountered in the studying of such models are the determination of how mutations in diverse sets of genes ultimately lead to alterations in brain excitability (Sarkisian, 2001). Moreover, spontaneous seizures were observed only in genetic models that showed bilateral or diffuse lesions (Lee et al., 1997).
Fetal insult models of CD
These models include those induced by administration of methylazoxymethanol acetate (MAM) and fetal irradiation of pregnant rats. Both these methods induce multifocal CD in newborn rats (Roper et al., 1995, 1997; Roper, 1998; Baraban et al., 2000; Castro et al., 2001; Kondo et al., 2001; Calcagnotto et al., 2002; Castro et al., 2002; Marin-Padilla et al., 2003; Kellinghaus et al., 2004).
In rats, prenatal exposure to methylazoxymethanol (MAM) consistently results in offspring with multifocal brain malformations, microcephaly, cortical thinning, loss of lamination and clusters of displaced neurons in the hippocampus, i.e., hippocampal heterotopia (Colacitti et al., 1999; Baraban et al., 2000). These animals have an increased sensitivity to various proconvulsant agents (de Feo et al., 1995; Baraban and Schwartzkroin, 1996; Germano et al., 1998; Chevassus-au-Louis et al., 1999). Isolated mini-slices containing heterotopia are capable of independent burst generation in vitro (Baraban et al., 2000) and seizure activity induced in hippocampal slices from MAM-exposed rats is resistant to commonly available antiepileptic drugs (Smyth et al., 2002). Although these animals exhibit salient features of the clinical condition, and have played an important role in determining how a malformed brain can generate abnormal electrical discharge, our overall understanding of hippocampal heterotopia remains limited. As for most animal models of CD, there is a lack of definite data on the spontaneous epileptogenicity in MAM-induced CD, although recent data indicate that a small percentage of these animals (<20%) exhibit spontaneous seizures arising from the hippocampus or the neocortex (Harrington et al., in press).
The in utero irradiation rat model of CD: The in utero irradiation of pregnant rats leads to the development of various degrees of cortical malformations and architectural abnormalities in the neocortical areas that are similar to those seen in some forms of CD in humans (Roper et al., 1995; Kondo et al., 2001). Hippocampal formation abnormalities also involve cell dispersion in the CA1 and to a lesser extent in CA3 (Roper et al., 1995; Kondo et al., 2001). The significance of these hippocampal abnormalities remains unclear as a similar pattern in human is yet to be described. Dyslamination and lack of columnar organization are seen in multiple areas of the neocortex as well as clustering of neurons in the molecular layer. Studies also showed that the cortical malformations inflicted in these rats would cause interictal epileptic discharges in a large number of the animals and spontaneous seizures in a much smaller percentage of these rats as compared to normal controls that is estimated around 10–20% (Kondo et al., 2001; Kellinghaus et al., 2004). Moreover, higher doses of in utero radiation leads to more severe pathological changes (microcephaly, absence of corpus callosum, and more diffuse and severe cortical dysplastic abnormalities), but these rats exhibit no spontaneous seizures. In vivo epileptogenicity increased with mild to moderate radiation doses, but occurred less frequently in the high-dose radiation group. There is a positive correlation between the radiation doses the rats received in utero and the severity of the cortical and hippocampal disorganization (Jensh, 1987; Fukui, 1991; Fushiki, 1996; Miki, 1999; Kellinghaus et al., 2004), suggesting a nearly linear dose–response curve regarding the extent of histological abnormalities seen in the brain postnatally. It has been shown that migratory cells are the most radiosensitive (Altman, 1968). In the rat, most of the pyramidal cells destined for layer V are generated at E16 and E17 (Ignacio, 1995) and are migrating to the cortical plate at the time of radiation (Ferrer, 1993b), whereas layer II/III neurons are generated later and are likely to be trapped near their periventricular origin due to radial glia damage. Therefore, it is not surprising that disorganization of both hippocampus and cortex (including subcortical heterotopias) increases with increasing radiation doses. It is interesting to note the dissociation between not only the severity of pathology and the expression of spontaneous epileptogenicity but also between in vitro and in vivo electrophysiological results: extracellular recordings from neocortical slices of rats treated with 225cGy on E17 showed evidence of hyperexcitability (Roper, 1997). Single-cell recordings from the dysplastic neocortex (Roper, 1997) showed reduced inhibition in that model that was correlated with a reduction of parvalbumin- and calbindin-reactive (i.e., probably inhibitory) neurons in the dysplastic normotopic neocortex (Roper, 1999). Therefore, changes in the inhibitory mechanisms may significantly contribute to the epileptogenicity. However, rats exposed to 175cGy on E17 with histological findings resemble most closely the model used for the in vitro studies (225cGy on E17) showed only rare epileptiform discharges and no spontaneous seizures.
At this point there has been no clear explanation of the dissociation between pathology and epileptogenicity in some animal models of CD. Therefore, the question remains: would these animals become epileptic under the right circumstance? If so, what are the conditions that may transform a potentially epileptogenic pathology to an epileptic phenotype? Data from human studies suggest that some patients with congenital/perinatal dysplastic lesions do not express epilepsy till later in life and if they do, epilepsy appears after some type of a trigger. In order to address these questions we recently studied the in vivo epileptogenicity in rats exposed to in utero radiation (Oghlakian, 2006): The majority of the radiated rats did not display spontaneous epileptic activities, but a second hit (a single treatment with a low dose of the proconvulsant agent pentylenetetrazole, PTZ) in rats that did not develop spontaneous ictal patterns rendered the majority of dysplastic rats (but not age matched control rats) into epileptic ones as these rats started to exhibit spontaneous epileptic seizures after a single injection of PTZ (Oghlakian, 2006). These results are interesting as they mirror the natural history in a significant number of patients with CD (thought to be due to prenatal/congenital or perinatal insults) in whom the epileptic phenotype does not develop till an otherwise nonepileptic stressor such as trauma, acute infection, stress, sleep deprivation … leads to the transformation of a nonepileptic pathology into an epileptic phenotype. As the mechanisms of this transformation from a dormant pathology into an epileptic phenotype are unknown further studies are needed.
As the pathological abnormalities in the in utero injury models are diffuse it is difficult to use these animals for studies that aim to directly correlate intrinsic epileptogenicity with histopathological/cellular, and/or molecular changes but these models are of interest for the studying of the dissociation between pathology and epileptogenicity and for the studying of the mechanisms underlying the transformation of nonepileptic dysplastic lesions into epileptic ones.
Neonatal lesion models
The hypothesis of an association between neonatal injuries and development of CD has been supported by extensive experimental data. Cowen et al. (1970) first reported that a closed head injury in rat pups could result in focal microdysgenesis. Thereafter, several studies reported on the occurrence of various types of malformations in the cortex of neonatal rats (up to 3–4 days postnatal) as a result of different types of injuries that include freeze lesions, laser and ionizing radiations, electrocoagulation, electroshock, focal injection of ibotenate, focal aspiration, and punctures (Dvorak and Feit, 1977; Ferrer and Catala, 1991; Humphreys et al., 1991; Suzuki and Choi, 1991; Ferrer, 1993a; Ferrer et al., 1993; Marret et al., 1995). The description of the early models focused mainly on the pathological characterization of the lesions with no attempt for electrophysiological studies till the second half of the last decade. Therefore, the functional significance (from an epilepsy perspective) of most of these elegant pathological models remains unknown.
One of the models of focal CD in the rat that was extensively studied is the neonatal freeze lesion model (Prince et al., 1997): The application of a deep-freeze probe to the skull of newborn rats results in a dysplastic lesion similar to polymicrogyria with spontaneous in vitro epileptogenicity in the immediate vicinity of the lesion (Jacobs et al., 1996; Prince et al., 1997; Jacobs et al., 1999). Although neocortical freeze lesions show histological characteristics of some forms of CD, and brain slices show in vitro hyperexcitability (Jacobs et al., 1996, 1999; DeFazio and Hablitz, 2000; Hablitz and DeFazio, 2000), recent studies showed a lack of epileptogenicity in vivo: Holmes et al. (1999) reported no significant differences in seizure susceptibility, as measured by after discharge threshold and kindling rate, between animals subjected to neocortical freeze lesions (1 or 3 lesions) at birth and age matched control animals. Long-term EEG monitoring has failed to find in vivo epileptiform discharges or behavioral seizures in this model (Kellinghaus, in press).
This illustrates some of the main problems with some animal models of epileptic pathologies where there is dissociation between pathology and electrophysiology.
Despite extensive studies, it is safe to say that most of the animal models using various species (that ranged from drosophila to primates) to this day have failed to meet the validation criteria as defined above. Further research is needed to determine the effect of a “second hit” on the potential transformation of various models of “dormant” pathology into an active (spontaneous) epileptic pathologies. These studies would reproduce some of the features that are characteristic of the natural history of some forms of human pathology and would ultimately lead to the identification of some of the cellular mechanisms of epileptogenesis and the future design of “preventative” interventions.