Vascular endothelial growth factor (VEGF) is an angiogenic cytokine that stimulates angiogenesis, vasodilatation and increased vascular permeability. All these effects depend on a VEGF-dependent increase in endothelial intracellular Ca²⁺ concentration ([Ca²⁺]_i) (Bates, et al. 1999). However it is unclear how VEGF induces the increase of [Ca²⁺]_i. The aim of this study was to determine whether VEGF could stimulate cation entry through a transient receptor potential (TRP) channel.

Chinese hamster ovary (CHO) cells were co-transfected with VEGF receptor 2 (VEGF-R2) and TRPC3 and were used for whole cell recording and analysis within 48-72 h. The bath solution contained (in mM) 140 NaCl, 5 CsCl, 2 CaCl₂, 1 MgCl₂, 10 glucose, and 10 Hepes (pH 7.4 with NaOH). The pipette solution contained (in mM) 135 CsCl, 2 MgCl₂, 3.62 CaCl₂, 10 EGTA, 30 Hepes (pH 7.2 with CsOH) with a calculated free [Ca²⁺] of 100 nM. Cells were held at a potential of -60 mV, and current-voltage (I-V) relations were obtained every 5 s from voltage ramps between 100 and +100 mV with a duration of 400 ms (Jung et al. 2002).

VEGF (1 nM) increased net inward and outward current during applied voltage ramps, with a time to peak-response of about 2 min. VEGF-activated current was blocked by gadolinium chloride (100 µM). A similar current was activated in TRPC3-transfected CHO cells, by application of 1-oleoyl-2-acetyl-sn-glycerol (OAG) (100 µM). No VEGF-activated currents could be recorded from control cells transfected with the pcDNA3 vector. VEGF-activated currents were recorded from 3 of 10 cells from VEGF-R2 and TRPC3 doubly-transfected cultures. Of the 7 VEGF-unresponsive cells, 1 responded to OAG whilst 6 responded to neither VEGF nor OAG. Transient transfection efficiency was determined from results of X-gal staining of LacZ-transfected cells, which showed 60-70 % cells were transfected. This is consistent with 30-40 % efficacy for double-transfection of cells and the incidence of VEGF-activated current in our recordings. The amplitude of currents we recorded was similar with that reported by Hofmann (Hofmann, et al. 1999) who used TRPC6-microinjected CHO-K1 cells.

Collectively, these findings suggest that VEGF is able to stimulate cation entry through VEGF-R2 mediated activation of TRPC3 channels.This work was funded by the British Heart Foundation.