RESUMO
[3H]Propyl-6-azido-beta-carboline-3-carboxylate ([3H]ACCP) exhibited a high affinity for GABAA receptors affinity purified from the brains of adult rats, and binding of this compound could be inhibited by several ligands of the benzodiazepine binding site of GABAA receptors. On irradiation with UV light, [3H]ACCP, similarly to [3H]flunitrazepam, irreversibly labeled a protein with an apparent molecular weight of 51 kDa in affinity-purified GABAA receptors, and this labeling could be inhibited in the presence of diazepam. These data indicate that [3H]ACCP can be used as a photoaffinity label for GABAA receptors.
Assuntos
Marcadores de Afinidade/metabolismo , Carbolinas/metabolismo , Receptores de GABA-A/análise , Marcadores de Afinidade/farmacologia , Animais , Ligação Competitiva , Encéfalo/ultraestrutura , Química Encefálica , Carbolinas/farmacologia , Diazepam/metabolismo , Diazepam/farmacologia , Cinética , Ensaio Radioligante , Ratos , Receptores de GABA-A/isolamento & purificação , Receptores de GABA-A/metabolismo , TrítioRESUMO
Melatonin was extracted from serum by using Baker reversed-phase C-18 columns. More than 99% of the applied melatonin was retained by the columns, and more than 97% was eluted from the columns in 300 microL of methanol. We then determined melatonin in the serum extract by a modification of a standard radioimmunoassay, using filtration instead of centrifugation to collect the [3H]melatonin-antibody complex precipitated by saturated ammonium sulfate. These modifications allow more rapid, accurate, and reproducible determination of melatonin than do previously published procedures.