ABNORMAL EXPRESSION OF NMDA RECEPTOR SUBUNITS IN DISSOCIATED CELLS FROM HUMAN PEDIATRIC CORTICAL DYSPLASIA USING SINGLE CELL RT-PCR
Abstract number :
3.054
Submission category :
Year :
2002
Submission ID :
858
Source :
www.aesnet.org
Presentation date :
12/7/2002 12:00:00 AM
Published date :
Dec 1, 2002, 06:00 AM
Authors :
Veronique M. Andre, Mary Kay Lobo, Cepeda Carlos, Michael S. Levine, Gary W. Mathern. Mental Retardation Research Center, University of California, Los Angeles, Los Angeles, CA; Neurosurgery, University of California, Los Angeles, Los Angeles, CA
RATIONALE: Cortical dysplasia (CD) is the most common substrate leading to seizure surgery in children. Previous electrophysiological studies reported that abnormal cytomegalic cells have altered NMDA currents. To determine if electrophysiological changes were related to alterations in gene expression, we studied NMDA receptor subunits in normal and dysmorphic neurons from CD and non-CD patients.
METHODS: Neocortical samples from 9 patients (6CD and 3 non-CD) were collected and slices were prepared. Their ages ranged from 5 months to 8 years and were not significantly different (CD = 0.4-7.8 yrs; non-CD = 1.2-8 yrs; P = 0.18). For each dissociated cell, rt-PCR was performed using human primers to amplify the NMDA subunits NR1, NR2A, NR2B and one housekeeping gene. Expression of amplified cDNA was visualized on ethidium-bromide gels. Depending on the expression pattern of the 3 NMDA subunits, cells were classified into 4 groups (presence of NR1 + NR2A + NR2B; presence of NR1 + 1 NR2, presence of NR1 only; presence of NR2(s) without NR1).
RESULTS: In the CD group, 51 cells were processed for PCR among which 12 (23%) were classified as abnormal-looking while 100% of the cells in the non-CD group (n = 23) were classified as normal-looking cells (p = 0.01). The distribution of NMDA subunits was significantly different comparing CD and non-CD cells (P = 0.009). In the non-CD group, 100% of the cells showed co-localization of NR1, NR2A and NR2B mRNAs whether the cell shape was pyramidal or not. In the CD group, only 63% of the cells showed the presence of the 3 subunits while in 15%, NR1 and only one NR2 were expressed, in 10%, NR1 only was expressed and 12% of the cells showed no NR1 but NR2(s). Moreover, in CD, abnormal shaped cells showed lack of NMDA subunit(s) compared with normal shaped pyramidal neurons (P = 0.01).
CONCLUSIONS: In this study, we demonstrated that the mRNA expression for NMDA subunits NR1, NR2A and NR2B differed in CD and non-CD cells from pediatric surgical patients. In CD, some of the NMDA subunits are not expressed in all the cells. Since NMDA currents depend on subunit composition, the lack of NR2A/NR2B could induce different responses to glutamate and could be responsible for hyperexcitability and seizures. The lack of NR1, NR2A and NR2B could also be indicative for abnormally developped or immature cells in CD.
[Supported by: NIH RO1 NS 38992]