Cerebellar granule-cell-specific GABAAreceptors attenuate benzodiazepine-induced ataxia: evidence from α6-subunit-deficient mice
Benzodiazepine- and alcohol-induced ataxias in rodents have been proposed to be affected by the gamma-aminobutyric acid type A (GABAA) receptor alpha 6 subunit, which contributes to receptors specifically expressed in cerebellar granule cells. We have studied an alpha 6 -/- mouse line for motor performance and drug sensitivity. These mice, as a result of a specific genetic lesion, carry a precise impairment at their Golgi-granule cell synapses. On motor performance tests (rotarod, horizontal wire, pole descending, staircase and swimming tests) there were no robust baseline differences in motor function or motor learning between alpha 6 -/- and alpha 6 +/+ mice. On the rotarod test, however,…
Characterization of γ-aminobutyrate type A receptors with atypical coupling between agonist and convulsant binding sites in discrete brain regions
Abstract γ-Aminobutyric acid type A (GABA A ) receptor ionophore ligand t -[ 35 S]butylbicyclophosphorothionate ([ 35 S]TBPS) was used in an autoradiographic assay on brain cryostat sections to visualize and characterize atypical GABA-insensitive [ 35 S]TBPS binding previously described in certain recombinant GABA A receptors and the cerebellar granule cell layer. Picrotoxinin-sensitive but 1-mM GABA-insensitive [ 35 S]TBPS binding was present in the rat cerebellar granule cell layer, many thalamic nuclei, subiculum and the internal rim of the cerebral cortex, amounting in these regions up to 6% of the basal binding determined in the absence of exogenous GABA. Similar binding properties wer…
Quantification of GABA(A) receptor subunit mRNAs by non-radioisotopic competitive RT-PCR utilizing plate-based EIA methodology.
We developed a non-radioisotopic quantitative competitive RT-PCR method for the measurement of gamma-aminobutyric acid (GABA) type A receptor subunit mRNA levels. The specificity of the method was optimized by the use of four subunit-specific oligonucleotides in the sequential steps: reverse transcription, polymerase chain reaction (PCR), and detection. The biotinylated PCR products were bound on streptavidin-coated microtiter plates allowing detection of the products using dinitrophenyl (DNP)-labeled probes and anti-DNP alkaline phosphatase conjugate. The method was set up for the six major cerebellar GABA(A) receptor subunits: alpha1; alpha6; beta2; beta3; gamma2 and delta. The method is …
Coupling between agonist and chloride ionophore sites of the GABA(A) receptor: agonist/antagonist efficacy of 4-PIOL.
Eight gamma-aminobutyric acid (GABA) mimetics were tested on their ability to differentiate native GABA(A) receptor subtypes present in various rat brain regions. In rat brain cryostat sections, little regional variations by the agonistic actions of muscimol, thiomuscimol, 4,5,6,7-tetrahydroisoazolo(5,4-c)pyridin-3-ol, piperidine-4-sulphonic acid, taurine and beta-alanine on [35S]t-butylbicyclophosphorothionate ([35S]TBPS) binding to GABA(A) receptor channels were found. They were very similar to those found for GABA itself and indicated no direct correlation with single subunit distributions for any of these compounds. Only the low-efficacy GABA mimetic 5-(4-piperidyl)isoxazol-3-ol (4-PIOL…
Furosemide action on cerebellar GABA(A) receptors in alcohol-sensitive ANT rats.
Furosemide increases the basal tert-[35S]butylbicyclophosphorothionate ([35S]TBPS) binding and reverses the inhibition of the binding by gamma-aminobutyric acid (GABA) in the cerebellar GABA(A) receptors containing the alpha6 and beta2/beta3 subunits. These effects are less pronounced in the alcohol-sensitive (ANT) than in the alcohol-insensitive (AT) rat line. The difference between the rat lines in the increase of basal [35S]TBPS binding was removed after a longer preincubation with ethylendiaminetetraacetic acid (EDTA) containing buffer, but long preincubation did not reduce the GABA content of the incubation fluid or remove the difference in GABA antagonism by furosemide. The GABA sensi…