The ontogenesis of the GABA-gated Cl- channel was investigated in the cerebral cortex of the rat by monitoring the binding parameters of [S-35]t-butylbicyclophosphorothionate ([S-35]TBPS) at intervals after birth (1-90 days). To investigate the influence of the developmental changes in the content of GABA on [S-35]TBPS-binding, the assays were carried out in unwashed membranes, in which the concentration of GABA was dependent on its content in vivo, and in repeatedly washed membranes in the presence of defined concentrations of exogenous GABA. At birth, the density (B-max) of [S-35]TBPS-binding sites in unwashed membranes was similar to that found in well-washed membranes. However, in unwashed membranes, the number of [S-35]TBPS-binding sites increased by two-fold within 10 days after birth whereas in washed membranes it increased by four-fold during the same period. The higher density of [S-35]TBPS-binding sites in washed membranes as compared with the unwashed counterparts persisted throughout development. In unwashed membranes, the apparent K-d for [S-35]TBPS-binding increased with age whereas in washed membranes the affinity of [S-35]TBPS for its binding sites remained constant throughout development. The binding of [S-35]TBPS to the GABA-gated Cl- channel is allosterically modulated by drugs acting on different sites of the GABA(A) receptor complex. Thus, GABA and diazepam decrease [S-35]TBPS-binding whereas the GABA(A) receptor antagonist, bicuculline, and the inverse agonist for benzodiazepine receptors, 6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylic acid methyl ester, increase it. The magnitude of the effects of positive and negative modulators of the GABA(A) receptor on [S-35]TBPS-binding did not change significantly from birth through adulthood, provided the concentration of GABA in the incubation medium was kept constant at 3 mu M. These results reflect the early development of the functional interactions between the different components of the GABA(A) receptor.
Allosteric modulation of [35S]TBPS-binding in the cerebral cortex of the rat during postnatal development
GIORGI, OSVALDO;LECCA, DANIELE;CORDA, MARIA GIUSEPPA
1994-01-01
Abstract
The ontogenesis of the GABA-gated Cl- channel was investigated in the cerebral cortex of the rat by monitoring the binding parameters of [S-35]t-butylbicyclophosphorothionate ([S-35]TBPS) at intervals after birth (1-90 days). To investigate the influence of the developmental changes in the content of GABA on [S-35]TBPS-binding, the assays were carried out in unwashed membranes, in which the concentration of GABA was dependent on its content in vivo, and in repeatedly washed membranes in the presence of defined concentrations of exogenous GABA. At birth, the density (B-max) of [S-35]TBPS-binding sites in unwashed membranes was similar to that found in well-washed membranes. However, in unwashed membranes, the number of [S-35]TBPS-binding sites increased by two-fold within 10 days after birth whereas in washed membranes it increased by four-fold during the same period. The higher density of [S-35]TBPS-binding sites in washed membranes as compared with the unwashed counterparts persisted throughout development. In unwashed membranes, the apparent K-d for [S-35]TBPS-binding increased with age whereas in washed membranes the affinity of [S-35]TBPS for its binding sites remained constant throughout development. The binding of [S-35]TBPS to the GABA-gated Cl- channel is allosterically modulated by drugs acting on different sites of the GABA(A) receptor complex. Thus, GABA and diazepam decrease [S-35]TBPS-binding whereas the GABA(A) receptor antagonist, bicuculline, and the inverse agonist for benzodiazepine receptors, 6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylic acid methyl ester, increase it. The magnitude of the effects of positive and negative modulators of the GABA(A) receptor on [S-35]TBPS-binding did not change significantly from birth through adulthood, provided the concentration of GABA in the incubation medium was kept constant at 3 mu M. These results reflect the early development of the functional interactions between the different components of the GABA(A) receptor.
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