Mechanism of Inhibition of TRPC Cation Channels by 2-Aminoethoxydiphenylborane

Abstract

We investigated the actions of the organoborane, 2-aminoethoxydiphenylborane (2APB), on Ca 2ϩ signaling in wild-type human embryonic kidney (HEK) 293 cells and in HEK293 cells stably expressing canonical transient receptor potential (TRPC) channels. Previous reports have suggested that 2APB inhibits agonist activation of TRPC channels because of its ability to act as a membrane-permeant inhibitor of inositol 1,4,5-trisphosphate (IP 3 ) receptors. 2APB was specifically said to inhibit TRPC3 channels when activated through a phospholipase Clinked receptor but not when activated more directly by a synthetic diacylglycerol, oleyl-acetyl-glycerol (OAG) [Science (Wash DC) 287:1647-1651, 2000]. However, we subsequently reported that IP 3 does not activate TRPC3; rather the mechanism of activation by phospholipase C-linked receptors seemed to result from diacylglycerol [J Biol Chem 278: 16244 -16252, 2003]. Thus, the current study was carried out to address the mechanism of action of 2APB in inhibiting TRPC channels. We found that, although the release of Ca 2ϩ by a muscarinic agonist was reduced by high concentrations of 2APB, this effect was indistinguishable from that seen when stores were discharged by thapsigargin, which does not involve IP 3 receptors. This indicates that 2APB is incapable of significant inhibition of IP 3 receptors when applied to intact cells. We found that 2APB partially inhibits divalent cation entry in cells expressing TRPC3, TRPC6, or TRPC7 and that this partial inhibition was observed whether the channels were activated by a muscarinic agonist or by OAG. Thus, as concluded for store-operated channels, 2APB seems to inhibit TRPC channels by a direct mechanism not involving IP 3 receptors.

TRPM7 regulates quiescent/proliferative metabolic transitions in lymphocytes

“…Each cell type was activated with M4, an IgM monoclonal antibody that activates the BCR or thapsigargin, an inhibitor of the SERCA (sarco endoplasmic reticulum Ca 2+ ATPase) pump responsible for Ca 2+ transport into the ER. 23,24 Surface expression of IgM was confirmed with a specific anti-IgM antibody (Fig. 3A, top middle part) and release of calcium from intracellular stores were prominently higher than those observed in TRPM7-KO and WT cells treated with rapamycin, an inhibitor of mTOR that induces G 1 arrest.…”

Regulation of transient receptor potential canonical channel 1 (TRPC1) by sphingosine 1-phosphate in C2C12 myoblasts and its relevance for a role of mechanotransduction in skeletal muscle differentiation

“…As shown in Table 2, the treatment with 50 μM GdCl3 or 100 μM 2-APB, an inositol-3-phosphate receptor-dependent SOC inhibitor (Lievremont et al, 2005), reduced G m /C m by approximately 27% and 40% in unstimulated cells, respectively, and by approximately 60% and 65% in S1P-stimulated cells. Moreover, the ion current was completely abolished when the stimulated cells were silenced for TRPC1 expression.…”

Ca2+Influx through Mechanosensitive Channels Inhibits Neurite Outgrowth in Opposition to Other Influx Pathways and Release from Intracellular Stores

“…6c). The inhibitory effects of 2APB were particularly potent, which may be attributable to the nonspecific inhibition of TRPC channels (Lievremont et al, 2005) and activation of TRPV channels (Hu et al, 2004) in addition to blocking IP 3 receptors. These data suggest that Ca 2ϩ release from intracellular stores supports neurite outgrowth and is required for maximum growth rates observed when inhibitory influx through SACs is blocked.…”