GABAA receptors in human temporal lobe epilepsy

Abstract

Restricted Item. Print thesis available in the University of Auckland Library or may be available through Inter-Library Loan. Temporal lobe epilepsy, the most frequent cause of focal seizures, is associated with a characteristic histological pattern of damage in the hippocampus known as hippocampal sclerosis (HS). Understanding the chemical and receptor changes in the affected hippocampus could lead to the development of effective drugs to treat this condition and an increased understanding of epilepsy. γ-aminobutyric acid (GABA), as the principal inhibitory neurotransmitter in the central nervous system, is likely to play a key role in seizures, which are considered an uncontrolled electrical discharge due to excess excitation or insufficient inhibition. The location, quantity and composition of GABAA receptors, a pentameric structure located on many different cell types, was studied in the human hippocampus using immunohistochemistry and in situ hybridisation. The hippocampi for study were obtained from surgical resections to treat temporal lobe epilepsy and compared both with post mortem tissue from patients free of neurological disease and with surgical specimens from patients with seizures but without HS. Using immunohistochemistry, 5 of the 18 known GABAA receptor subunits were studied. Also studied were three GABA transporters (GAT-1, GAT-2, GAT-3), involved in GABA reuptake from the extracellular space, glial fibrillary acidic protein (GFAP), a marker of astrocytes, and glutamic acid decarboxylase (GAD), the enzyme responsible for the production of GABA were also studied. Using in situ hybridisation, 10 GABAA receptor subunits plus GAT-1 were studied. In hippocampal sclerosis there is a dramatic loss of principal cells, particularly in the CA1 region and to a lesser degree in the CA2, CA3 and hilar regions. In order to evaluate the GABAA receptor subunit changes in the face of this cell loss, cell counts were performed throughout the hippocampus in the control and epilepsy cases. Quantitative analysis was performed in immunohistochemistry and in situ hybridisation studies using computerised density measurements followed by detailed statistical analysis. Immunohistochemistry showed that the majority of GABAA receptors are located on the processes of the principal cells, but with regional variation. Receptors are located on interneurons throughout the hippocampus. Co-localisation of receptor subunits suggested a number of receptor compositions in the normal and epileptic hippocampus including α1β2,3γ2, α2β2,3γ2, α2βxγ2, α3βxγ2 and α3β2,3γ2 Cell counts demonstrated a drastic loss of principal cells ranging from 41% to 76% in the CA regions in HS consistent with previous reports. Overall, quantitative assessment of density in immunohistochemical studies of the 5 GABAA receptor subunits showed a remarkable preservation of density in all regions. This was confirmed by density studies using in situ hybridisation. This suggests massive upregulation of GABAA receptors on the remaining principal cells. Some subunit switching within receptors was proposed to account for some of the regional changes in individual subunits though this could not be confirmed due to the small numbers of cases studied. The δ subunit, studied with in situ hybridisation, showed a significant reduction in HS. This is of particular interest in view of its role in the modification of zinc sensitivity of inhibitory synaptic transmission as described by Saxena, 1994. δ containing receptors may play a role in preventing seizures by generation of tonic inhibition via long-lasting inhibitory post-synaptic potentials. GAT-1, studied with both techniques, was significantly reduced in HS. Immunohistochemistry showed the reduction to be in CA1, CA3 and hilus, being the regions with the most marked cell loss. GAT-2 was not present in the normal or epileptic hippocampus. GAT-3 was present on the soma of the principal cells and in astrocytes. Immunohistochemistry showed a marked increase in GAT-3 in the epileptic hippocampus. GFAP and GAD both showed a striking increase in the epileptic hippocampus compared to controls. In summary, GABAA receptors showed variation in regional location and individual subunit composition in the normal and epileptic hippocampus with the most striking finding being the dramatic up-regulation of total GABAA receptors in the epileptic hippocampus despite major cell loss in most regions.