VEGF and ATP act by different mechanisms to increase microvascular permeability and endothelial [Ca²⁺]_i

Abstract

Vascular endothelial growth factor (VEGF) increases hydraulic conductivity (L(p)) by stimulating Ca(2+) influx into endothelial cells. To determine whether VEGF-mediated Ca(2+) influx is stimulated by release of Ca(2+) from intracellular stores, we measured the effect of Ca(2+) store depletion on VEGF-mediated increased L(p) and endothelial intracellular Ca(2+) concentration ([Ca(2+)](i)) of frog mesenteric microvessels. Inhibition of Ca(2+) influx by perfusion with NiCl(2) significantly attenuated VEGF-mediated increased [Ca(2+)](i). Depletion of Ca(2+) stores by perfusion of vessels with thapsigargin did not affect the VEGF-mediated increased [Ca(2+)](i) or the increase in L(p). In contrast, ATP-mediated increases in both [Ca(2+)](i) and L(p) were inhibited by thapsigargin perfusion, demonstrating that ATP stimulated store-mediated Ca(2+) influx. VEGF also increased Mn(2+) influx after perfusion with thapsigargin, whereas ATP did not. These data showed that VEGF increased [Ca(2+)](i) and L(p) even when Ca(2+) stores were depleted and under conditions that prevented ATP-mediated increases in [Ca(2+)](i) and L(p). This suggests that VEGF acts through a Ca(2+) store-independent mechanism, whereas ATP acts through Ca(2+) store-mediated Ca(2+) influx.

Vascular endothelial growth factor-mediated activation of p38 is dependent upon Src and RAFTK/Pyk2

“…VEGF can induce an influx of extracellular calcium into the cell through membrane calcium channels (Criscuolo et al, 1989;Pocock et al, 2000;Garnier-Raveaud et al, 2001). Chelating extracellular calcium, by a 5 s pretreatment with EGTA, abolished the VEGF-induced activation of Figure 2 RAFTK/Pyk2, but not FAK, is required for VEGF stimulation of p38 MAPK.…”

Role of endothelial Ca2+ stores in the regulation of hydraulic conductivity of Rana microvessels in vivo

“…To determine whether Ca 2ϩ store release was sufficient to increase L p in the absence of Ca 2ϩ influx, the irreversible SERCA inhibitor 100 nM TG was perfused with the nonspecific cation channel blocker 5 mM NiCl 2 (Ni 2ϩ ). TG (100 nM) has previously been shown in this laboratory to transiently increase L p in the presence of Ca 2ϩ influx (25). Vessels were perfused with 1% BSA in FR and rat erythrocytes for a variable time period until the baseline L p stabilized and then perfused with TG and Ni 2ϩ for 20 min.…”

“…We have assumed that this is due to a delayed increase in [Ca 2ϩ ] i in response to these agonists, because previous work (25), where [Ca 2ϩ ] i and permeability were measured in the same model (although in different vessels), showed that there was a distinct time lag between the application of TG to the cells and the increase in both [Ca 2ϩ ] i and permeability. This is in contrast to many studies of endothelial cells in culture where TG results in an immediate transient increase in intracellular Ca 2ϩ , followed by a sustained increased [Ca 2ϩ ] i (8).…”

“…Bates and Curry (5) showed that the addition of the cation channel blocker Ni 2ϩ (5 mM) prevents Ca 2ϩ influx and completely blocks the Ca 2ϩ responses to VEGF. However, VEGF increases L p through a Ca 2ϩ store-independent mechanism, as demonstrated by the inability of the sarco(endo)plasmic reticulum (ER) Ca 2ϩ -ATPase (SERCA) inhibitor thapsigargin (TG) to block the response by first emptying stores (25).…”

Papel do fator de crescimento vascular endotelial na angiogênese e na retinopatia diabética

“…A ligação do VEGF a esses receptores causa influxo de cálcio citoplasmático, aumentando sua concentração em até quatro vezes, mudança da forma, divisão e migração celular (17,18,(31)(32)(33). Esse aumento da permeabilidade das vênulas às macromoléculas permite que proteínas plasmáticas extravasem para o espaço extravascular, levando a coagulação do fibrinogênio e deposição de gel de fibrina que serve como matriz provisória para o crescimento de novos vasos sanguíneos (15,17,18,23).…”