Abstracts

The methylazoxymethanol (MAM) model of cortical dysplasia presents with neuronal heterotopia with unclear synaptic relationships to the surrounding brain. MAM-induced heterotopia in the hippocampal CA1 subfield, and the periventricular nodules (PN), have morphological features similar to heterotopia in human cortical dysplasia. In the present study, we have begun to investigate how abnormal connectivity of these cortical malformations may contribute to epileptogenesis. We hypothesize that aberrant synaptic connectivity between the heterotopia and hippocampus and/or neocortex contributes to an abnormal functional coupling between these brain regions.
Pregnant Spargue-Dawley rats (E15) were injected (25mg/kg, i.p.) with MAM. Rats (6 months old) exposed to MAM [italic]in utero[/italic] were injected with the anterograde tracer [italic]biotinylated dextran amine[/italic] (BDA) intrahippocampally. Injection were placed bilaterally; 1[micro]l per injection site was delivered over 5min. After a survival time of 8-10 days, rats were perfused with a solution consisting of 4% paraformaldehyde and 0.5% glutaraldehyde, followed by a histochemical procedure using a BDA Neuro Trace Kit (Molecular Probes, Eugene, OR). Electrophysiological studies were also carried out on brain slices from MAM-exposed rats(P21-P35). [/italic]Synaptic connectivity between the heterotopia and [ldquo]normal[rdquo] hippocampal pyramidal cells was studied by stimulating one region (heterotopia, CA1, Schaffer collaterals) and recording from cells in the other region (heterotopia, CA1) with sharp intracellular pipettes.
A single MAM injection at E15 produced morphological features of abnormal cortical development (i.e., microcephalic brains with reduce neocortical laminae thickness, heterotopia in the hippocampal CA1 subfield and periventricularly). BDA injections into the hippocampus labeled neuronal cell bodies, dendritic trees and axonal arbors both in normal cortical areas and heterotopic regions. Morphological features of BDA-labeled neurons in CA1 and PN heterotopia appeared similar to neocortical Lamina II/III pyramidal neurons and aspiny and sparsely-spiny non-pyramidal cells. Labeled axons that crossed the corpus callosum suggested bi-directional connectivity between CA1 heterotopia and neocortex. Hippocampus and PN heterotopia also exhibited extensive axonal connectivity, especially at the interface between stratum oriens and PNs. In preliminary electrophysiological analysis, excitatory postsynaptic potentials were recorded from heterotopic cells in response to stimulating Schaffer collaterals. Conversely, stimulation of the heterotopia evoked EPSPs in [ldquo]normal[rdquo] CA1 pyramidal neurons.
Preliminary studies of the MAM-exposed rat confirmed that there was extensive anatomical and electrophysiological connectivity between heterotopic cell clusters and hippocampus, and between heterotopia and neocortex. These results are consistent with the hypothesis that heterotopia serve as functional links between the hippocampus and neocortex.
[Supported by: NIH grant NS18895 (PAS)]