RESUMEN
Cholinergic interneurons, which provide the main source of acetylcholine (ACh) in the striatum, control the striatal local circuits and deeply involve in the pathogenesis of neurodegenerative diseases. Glycogen synthase kinase-3 (GSK-3) is a crucial kinase with diverse fundamental functions and accepted that deregulation of GSK-3 activity also plays important roles in diverse neurodegenerative diseases. However, up to now, there is no direct proof indicating whether GSK-3 activation is responsible for cholinergic dysfunction. In the present study, with combined intracerebroventricular injection of Wortmannin and GF-109203X, we activated GSK-3 and demonstrated the increased phosphorylation level of microtubule-associated protein tau and neurofilaments (NFs) in the rat striatum. The activated GSK-3 consequently decreased ACh level in the striatum as a result of the reduction of choline acetyltransferase (ChAT) activity. The alteration of ChAT activity was due to impaired ChAT distribution rather than its expression. Furthermore, we proved that cellular ChAT distribution was dependent on low phosphorylation level of NFs. Nevertheless, the cholinergic dysfunction in the striatum failed to induce significant neuronal number reduction. In summary, our data demonstrates the link between GSK-3 activation and cholinergic dysfunction in the striatum and provided beneficial evidence for the pathogenesis study of relevant neurodegenerative diseases.
Asunto(s)
Acetilcolina/metabolismo , Colina O-Acetiltransferasa/metabolismo , Cuerpo Estriado/metabolismo , Glucógeno Sintasa Quinasa 3/metabolismo , Interneuronas/metabolismo , Animales , Western Blotting , Activación Enzimática , Inmunohistoquímica , Masculino , Microscopía Confocal , Enfermedades Neurodegenerativas/metabolismo , Fosforilación , Ratas , Ratas WistarRESUMEN
Berberine was used to determine inhibition of arylamine N-acetyltransferase (NAT) activity in human bladder tumour cells. The NAT activity was measured by HPLC assaying for the amounts of N-acetyl-2-aminofluorene (AAF) and N-acetyl-p-aminobenzoic acid (N-Ac-PABA) and remaining 2-aminofluorene (AF) and p-aminobenzoic acid (PABA). Two assay systems were performed, one with cellular cytosols, the other with intact bladder tumour cell suspensions. The NAT activity in human bladder tumour cells was inhibited by berberine in a dose-dependent manner, that is, the higher the concentration of berberine, the higher the inhibition of NAT activity. The values of apparent Km and Vmax calculated from cytosol NAT and intact cells were also decreased by berberine. This report is the first demonstration to show berberine did affect human bladder tumour cell NAT activity.