Neuronal nitric oxide synthase expression in resected epileptic dysplastic neocortex

Laboratory investigation

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  • 1 Departments of Neurosurgery,
  • | 2 Neurology, and
  • | 3 Pathology, Cleveland Clinic, Cleveland, Ohio
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Object

Nitric oxide has been associated with epileptogenesis. Previous studies have shown increased expression of N-methyl-d-aspartate (NMDA) subunit NR2B receptors in epileptic dysplastic human neocortex. The expression of neuronal nitric oxide synthase (nNOS), and its relation to this subunit NR2B in epileptic dysplastic tissue has never been addressed.

Methods

Ten patients with medically intractable epilepsy caused by focal cortical dysplasia (CD), and 2 patients with mesial temporal sclerosis (control group) underwent pre- and/or intraoperative invasive monitoring evaluations. Cortical samples from epileptogenic and nonepileptogenic areas were collected from each patient intraoperatively. Samples were processed for cresyl violet staining, immunocytochemical tests with nNOS, NeuN, and NR2B, and immunofluorescence analyses to evaluate colocalized immunoreactivity between nNOS and NR2B.

Results

. All samples obtained in the patients with epilepsy revealed CD in various degrees. In the nonepileptic sample group, cresyl violet staining revealed normal cortical architecture in 9 samples, but a mild degree of CD in 3. The density and intensity of nNOS-stained neurons was remarkably increased in the epileptic tissue compared with nonepileptic samples (p < 0.05). Two types of nNOS-stained neurons were identified: Type I, expressing strong nNOS immunoreactivity in larger neurons; and Type II, expressing weak nNOS immunoreactivity in slightly smaller neurons. Different from Type I neurons, Type II nNOS-stained neurons revealed immunoreactivity colocalized with NR2B antibody.

Conclusions

The overexpression of nNOS in the epileptic samples and the immunoreactivity colocalization between nNOS and NR2B may suggest a possible role of nNOS and NO in the pathophysiological mechanisms related to in situ epileptogenicity.

Abbreviations used in this paper:

CD = cortical dysplasia; NMDA = N-methyl-d-aspartate; NeuN = neuronal nuclei; nNOS = neuronal nitric oxide synthase; NR2B = NMDA receptor 2B; TBS = Trisbuffered saline.

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Contributor Notes

Address correspondence to: Jorge A. González-Martínez, M.D., Ph.D., Department of Neurosurgery, Cleveland Clinic, 9500 Euclid Avenue, Desk S80, Cleveland, Ohio 44195. email: gonzalj1@ccf.org.
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