In this study, we showed that insulin induced translocation and insertion of TRPV2 into the plasma membrane. Consistent with these observations, calcium entry as monitored by fura-2 fluorescence was elevated in cells pretreated with insulin, which was inhibited by an inhibitor of TRPV2 and knockdown of TRPV2. Collectively, exogenous insulin induces translocation of TRPV2 to the plasma membrane and augments calcium entry. In our experimental condition, translocation of TRPV2 was observed to some extent under basal conditions. This is in sharp contrast to CHO cells and macrophages (12
), where the amount of TRPV2 in the plasma membrane is very low in an unstimulated condition. This difference may be explained by the basal secretion of insulin, since the basal level of translocation is reduced by loading BAPTA, which reduces basal secretion, and by anti-insulin antibody. These observations imply that translocation of TRPV2 induced by basal release of insulin may contribute to basal calcium entry. Furthermore, translocation of TRPV2 was induced by insulin secretagogues including a high concentration of glucose. Taken together, insulin released from β-cells further augments calcium entry by recruiting TRPV2 to the plasma membrane. Given that calcium is a critical regulator of insulin secretion, this is a feed-forward mechanism to accelerate insulin secretion. With regard to the functional significance of TRPV2, inhibition of the TRPV2 activity by tranilast or by knockdown of TRPV2 using shRNA reduces glucose-induced insulin secretion. Augmented calcium entry through TRPV2 may contribute partly to glucose-induced insulin secretion. Also, TRPV2 is involved in insulin secretion induced by other secretagogues. For example, potassium-induced secretion is also inhibited partly by inhibiting TRPV2. Presumably, insulin released by depolarization of β-cells in turn recruits TRPV2 to the plasma membrane, which leads to additional calcium entry through TRPV2 and thereby enhances insulin secretion. It is known that voltage-dependent calcium channels are downregulated (so-called inactivation) rather quickly after depolarization of the plasma membrane. In contrast, the TRPV2 channel is not downregulated quickly. Instead, it promotes calcium entry for a relatively long period (11
). Consequently, recruitment of TRPV2 by insulin may prolong calcium entry into β-cells. TRPV2 is a calcium-permeable channel and also permeates sodium as well (11
). Translocation of TRPV2 thus also increases sodium entry into β-cells. This leads to depolarization of the plasma membrane and subsequent activation of the voltage-dependent calcium channels. Recently, it was reported that TRPM2 and TRPM4 are involved in glucose-induced insulin secretion in pancreatic β-cells (22
). TRPM2 and TRPM4 permeate calcium and/or sodium, which leads to depolarization of the plasma membrane and activation of the voltage-gated calcium channel. In addition to the well-known role of the voltage-gated calcium channels, members of the TRP channel family may contribute to the stimulus-secretion coupling in β-cells.
The autocrine effect of secreted insulin on β-cell function has been a matter of debate. Both positive and negative feedback regulations have been postulated, and there have been many reports supporting these ideas (rev. in 24
). In this regard, a study using β-cell–specific knockout of the IR gene demonstrated that blocking of the insulin action in β-cells impairs glucose-induced insulin secretion and also reduces the β-cell mass in adults (1
). The present results suggest that TRPV2 is one of the target molecules of insulin and may participate at least partly in the trophic action of this hormone in β-cells. Inhibition of the TRPV2 reduces insulin secretory response and DNA synthesis in these cells. Given that stimulation of calcium entry is a prerequisite for cell-cycle progression induced by growth factors (25
), it is likely that TRPV2 plays a role in insulin-induced proliferation. Taken together, TRPV2 functions as one of the molecular targets of the insulin action and modulates calcium entry.
We studied the role of TRPV2 in MIN6 cells and dispersed β-cells. It is uncertain at present whether or not insulin secreted from β-cells actually controls translocation of TRPV2 in islet β-cells in vivo. It is possible that basal secretion of insulin induces translocation of TRPV2 and TRPV2 is always operative in the plasma membrane. It is also possible that growth factors in plasma, for example IGF-I, induces translocation and TRPV2 is always functional. If so, insulin secretagogues do not regulate TRPV2 translocation. Instead, changes in the expression of TRPV2, if any, would alter the basal rate of calcium entry. Further studies are necessary to assess the in vivo role of TRPV2 in β-cells.
The present results provide for the first time evidence that calcium-permeable channel TRPV2 is a signaling molecule involved in the insulin action. In this regard, insulin is shown to recruit various types of ion channels to the plasma membrane in neuronal cells (27
). TRPV2 is one of the channel molecules regulated by insulin. Many issues are still unsolved. For example, it is not known whether insulin directly modifies gating of the TRPV2 in addition to induction of translocation. Further studies are necessary to elucidate the role of TRPV2 in the insulin action.