Abstracts

The most common neurological symptom of human neocortical dysplasias is early-life epilepsy, including infantile spasms. Frequently, these patients develop pharmaco-resistant seizure disorders, which require surgical excision. The exact mechanisms of epileptogenesis and high incidence of pharmaco-resistance to conventional antiepileptic drugs (AED) in cortical dysplasias are poorly understood. GABA receptors (GABARs), which are normally hyperpolarizing, can become depolarizing under certain circumstances and actually increase neuronal excitation. This depends on the intracellular chloride concentration, regulated by the chloride-importer NKCC1 and the chloride-exporter KCC2. Increased NKCC1 and decreased KCC2 expression in hippocampal subiculum from temporal lobe epilepsy (TLE) patients renders GABARs depolarizing (PNAS 103:8465;2006). We hypothesized that similar to TLE, neurons from neocortical dysplastic tissue would demonstrate higher NKCC1 expression and lower KCC2 levels relative to control neurons, a profile consistent with the presence of depolarizing GABARs., Cortical tissue from 6 patients ages 1-6 years were obtained during brain surgery for drug-resistant epilepsy. In all cases, malformations of cortical development were confirmed by histopathological examination (tuberous sclerosis, n=3; focal cortical dysplasia, n=2; hemimegalencephaly, n=1). Age-matched autopsy cortical samples from cases with normal neurologic history were used as controls (n=4). Tissue was collected and handled in accordance with the Clinical Research Committee at Children[apos]s Hospital, Boston. 4% paraformaldehyde-fixed tissue blocks were cut at 50 microns and immunohistochemically double labeled with cell markers (NeuN, non-phosphorylated neurofilament, vimentin, GFAP) in combination with transporter antibodies NKCC1 (1:100, Chemicon) and KCC2 (1:500, Upstate)., Staining of dysplastic tissue with neuronal and glial markers demonstrated loss of cortical lamination, the presence of disoriented, misshaped neurons, an increase in number of white matter neurons and a variable degree of gliosis. In addition, in cortical tubers and hemimegalencephaly tissue, cytomegalic dysplastic neurons could be easily identified. In all cases, increased NKCC1 expression associated with a moderate decrease in KCC2 levels was observed in most normal-size and cytomegalic dysplastic neurons, compared to controls. Notably, strong NKCC1 immunoreactivity was observed in glial cell population as well., Our results demonstrate an imbalance in neuronal NKCC1 and KCC2 expression (increased NKCC1: KCC2 ratio) in human neocortical dysplasias that may render GABARs depolarizing. This mechanism may play a critical role in increased seizure susceptibility and relative refractoriness to GABAR agonists in these patients., (Supported by TS Alliance, Boston Neurosurgical Fdn, NIH NS31718.)