Artikel
Non-selective transient receptor potential channel type 3 in spontaneously hypertensive rats
Transient receptor potential Kanal Typ 3 bei spontanhypertensiven Ratten
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Veröffentlicht: | 10. August 2005 |
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Gliederung
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Objective: Several investigators showed increased calcium influx in primary hypertension. Recently a new family of non-selective cation channels, so-called transient receptor potential (TRP) channels, have been described. The role of TRP channels has not been evaluated in primary hypertension yet.
Methods: Transient receptor channels type 3 (TRPC3) were investigated in monocytes from spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY) with in-cell Western blots using odyssey infrared imaging system (primary antibodies, rabbit anti-TRPC3-antibodies (1:1000); secondary antibodies, IRDye800CW-infrared fluorescent dye-conjugated goat anti-rabbit antibodies (1:1000); emission, 810nm; excitation, 780 nm). For ratiometric calcium imaging experiments monocytes were loaded with 2µmol/L fura2 and fluorescence was measured using a 96well-fluorescent plate reader at 510nm emission with excitation wavelengths of 340nm and 480nm.
Results: Using in-cell Western blotting and CD14 as internal reference, we identified significantly increased expression of TRPC3 channels in monocytes from SHR compared with normotensive WKY (TRPC3/CD14 ratio, 7.7±0.3 for SHR, and 2.9±0.5 for WKY; mean±SEM; each n=4; p<0.05). Calcium influx was significantly higher in monocytes from SHR compared with WKY (fluorescence ratio increase, 0.81±0.13 for SHR, n=9; and 0.34±0.10 for WKY, n=8; p<0.05). In the presence of the inhibitor, SKF-96365, the calcium increase was significantly reduced in monocytes from SHR to 0.19±0.11 (n=7; p<0.05 compared with control) and in monocytes from WKY it was reduced to 0.07±0.07 (n=8; p<0.05 compared with control). Calcium influx through TRP channels significantly increased reactive oxygen species in monocytes from rats as measured by dihydroethidium fluorescence.
Conclusion: Our study for the first time indicates increased TRPC3 channel expression in primary hypertension.