Downregulation of β1 -adrenergic receptors in rat C6 glioblastoma cells by hyperforin and hyperoside from St John's wort.

Objectives: While the use of St John's wort extracts as treatment for mild to moderate depression is well established the mode of action is still under investigation. Individual constituents of St John's wort extract were tested for possible effects on the β1 AR density and a subsequent change in downstream signalling in rat C6 glioblastoma cells.

Methods: The effect of compounds from St John's wort extract on the downregulation of β1 -adrenergic receptor-GFP fusion proteins (β1 AR-green fluorescent protein (GFP)) of transfected rat C6 gliobastoma cells (C6-β1 AR-GFP) was investigated by means of confocal laser scanning microscopy (LSM). The influence on the lateral mobility of β1 AR-GFP in C6-β1 AR-GFP was investigated by fluorescence correlation spectroscopy. The formation of second messenger was determined by c-AMP-assay.

Key Findings: Confocal LSM revealed that pretreatment of cells with 1 μm of hyperforin and hyperoside for 6 days, respectively, led to an internalization of β1 AR-GFP under non-stimulating conditions. Observation by fluorescence correlation spectroscopy showed two diffusion time constants for control cells, with τdiff1  = 0.78 ± 0.18 ms and τdiff2  = 122.53 ± 69.41 ms, similarly distributed. Pretreatment with 1 μm hyperforin or 1 μm hyperoside for 3 days did not alter the τdiff values but decreased the fraction of τdiff1 whereas the fraction of τdiff2 increased significantly. An elevated level of β1 AR-GFP with hindered lateral mobility was in line with β1 AR-GFP internalization induced by hyperforin and hyperoside, respectively. A reduced β1 -adrenergic responsiveness was assumed for C6 gliobastoma cells after pretreatment for 6 days with 1 μm of both hyperforin and hyperoside, which was confirmed by decreased cAMP formation of about 10% and 5% under non-stimulating conditions. Decrease in cAMP formation by 23% for hyperforin and 15% for hyperoside was more pronounced after stimulation with 10 μm dobutamine for 30 min.

Conclusions: The treatment of C6 gliobastoma cells with hyperforin and hyperoside results in a reduced β1 AR density in the plasma membrane and a subsequent reduced downstream signalling.