RESUMO
The aim of this study was to investigate the mRNA expression of the two GABA(B1) receptor isoforms and the GABA(B2) subunit, in human postmortem control hippocampal sections and in sections resected from epilepsy patients using quantitative in situ hybridisation autoradiography. Utilising human control hippocampal sections it was shown that the oligonucleotides employed were specific to the receptor. Hippocampal slices from surgical specimens obtained from patients with hippocampal sclerosis and temporal lobe epilepsy were compared with neurologically normal postmortem control subjects for neuropathology and GABA(B) mRNA expression. Neuronal loss was observed in most of the hippocampal subregions, but in the subiculum no significant difference was detected. The localisation of GABA(B1a) and GABA(B1b) isoform mRNAs in human control hippocampal sections supported and extended earlier studies using the GABA(B1) pan probe, which does not distinguish between the two GABA(B1) isoforms. Moreover, the GABA(B2) mRNA location confirmed the heterodimerisation of the receptor. Thus, although there was an apparent correlation between GABA(B1b) and GABA(B2), GABA(B1a) exhibited no such relationship. GABA(B1b) and GABA(B2) showed a similar intensity of expression whilst GABA(B1a) displayed a lower hybridisation signal. Comparison of the expression of the three mRNAs between control and epileptic subjects showed significant decreases or increases in different hippocampal subregions.GABA(B) isoforms and subunit mRNA expression per remaining neuron was significantly increased in the hilus and dentate gyrus. These results demonstrate that altered GABA(B) receptor mRNA expression occurs in human TLE; possibly the observed changes may also serve to counteract ongoing hyperexcitability.
Assuntos
Epilepsia do Lobo Temporal/metabolismo , Variação Genética/fisiologia , Hipocampo/metabolismo , Receptores de GABA-B/biossíntese , Receptores de GABA/biossíntese , Adulto , Idoso , Idoso de 80 Anos ou mais , Epilepsia do Lobo Temporal/genética , Epilepsia do Lobo Temporal/patologia , Feminino , Regulação da Expressão Gênica/fisiologia , Hipocampo/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Isoformas de Proteínas/biossíntese , Isoformas de Proteínas/genética , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Receptores de GABA/genética , Receptores de GABA-B/genéticaRESUMO
1 The aim of this study was to investigate the binding of a novel GABA(B) receptor radioligand, [(3)H]-CGP62349, to human post-mortem control and epileptic hippocampal sections using quantitative receptor autoradiography. Utilizing human control hippocampal sections it was shown that [(3)H]-CGP62349 bound with high affinity (K(D) 0.5 nM) to this tissue. 2 Hippocampal slices from surgical specimens obtained from patients with hippocampal sclerosis (HS) and temporal lobe epilepsy (TLE) were compared with neurologically normal post-mortem control subjects for neuropathology and GABA(B) receptor density and affinity. Neuronal loss was observed in most of the hippocampal subregions, but in the subiculum no significant difference was detected. 3 The localization of GABA(B) receptors with the antagonist [(3)H]-CGP62349 in human control hippocampal sections supported and extended earlier studies using the agonist ligand [(3)H]-GABA. 4 The kinetics of binding to the GABA(B) receptor in human hippocampus using this novel compound was comparable to previous data obtained in rat hippocampal membranes. 5 GABA(B) receptor density (B(max)) was significantly reduced in CA3, hilus, and dentate gyrus (DG); the affinity was increased exclusively in DG. The trend is identical in all the hippocampal subregions with the agonist and the antagonist, although significant differences with the antagonist where recorded in CA3 and hilus, whereas with the agonist a significant reduction was reported in all of the hippocampal subfields. 6 GABA(B) receptor expression per remaining neuron appeared significantly increased in CA3 and hilus. These results suggest altered GABA(B) receptor function may occur in human TLE, possibly as a result of synaptic reorganization, and may contribute to epileptogenesis.
Assuntos
Benzoatos/farmacologia , Epilepsia do Lobo Temporal/metabolismo , Hipocampo/metabolismo , Compostos Organofosforados/farmacologia , Receptores de GABA-B/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Autorradiografia , Contagem de Células , Epilepsia do Lobo Temporal/patologia , Feminino , Hipocampo/patologia , Humanos , Técnicas In Vitro , Cinética , Masculino , Pessoa de Meia-Idade , Neurônios/patologia , Ensaio RadioliganteRESUMO
The distribution of GABA(B) receptor subunits GABA(B(1a)), GABA(B(1b)) and GABA(B2), has been examined in the cerebral cortex and thalamus of adult rats using an immunocytochemical technique. GABA(B(1a)) and GABA(B(1b)) subunits co-localized with GABA(B2) in the cortex, where afferent thalamic GABAergic axons project to pyramidal neurones. The expression patterns of GABA(B(1a)), GABA(B(1b)) and GABA(B2) were similar throughout the thalamus. The data suggest that the GABA(B(1b)) subunit might be the presynaptic isoform in the thalamo-cortical pathway with the GABA(B(1a)) subunit possibly present at postsynaptic sites on cell bodies. This contrasts with our previous data, obtained in cerebellum and spinal cord which indicate opposite locations. Thus, it seems unlikely that functional role along with cellular location can be assigned in a general manner to specific GABA(B) receptor subunit splice variants.