Department of Pharmacology, School of Pharmaceutical Sciences, University of Shizuoka, Shizuoka City, Japan.
Am J Physiol Cell Physiol. 2019 Mar 1;316(3):C434-C443. doi: 10.1152/ajpcell.00210.2017. Epub 2019 Jan 16.
β-Cell swelling induces membrane depolarization, which has been suggested to be caused at least partly by the activation of cation channels. Here, we show the identification of the cation channels. In isolated mouse pancreatic β-cells, the exposure to 30% hypotonic solution elicited an increase in cytosolic Ca concentration ([Ca]). The [Ca] elevation was partially inhibited by ruthenium red, a blocker of several Ca-permeable channels including transient receptor potential vanilloid receptors [transient receptor potential cation channel subfamily V (TRPV)], and by nicardipine, but not by the depletion of intracellular Ca stores with thapsigargin and caffeine. The hypotonic stimulation also increased insulin secretion from isolated mouse islets, which was significantly suppressed by ruthenium red. Expression of TRPV2 and TRPV4 was confirmed in mouse pancreatic islets and the MIN6 β-cell line by RT-PCR, Western blot, and immunohistochemical analyses. However, neither 4α-phorbol 12,13-didecanoate nor GSK1016790A, TRPV4 activators, showed any apparent effect on [Ca] in isolated mouse β-cells or in MIN6 cells. In contrast, probenecid, a TRPV2 activator, induced an increase in [Ca] in MIN6 cells, which was attenuated by ruthenium red. Moreover, the [Ca] elevation induced by 30% hypotonic stimulation was significantly reduced by knockdown of TRPV2 with siRNA and by tranilast, a TRPV2 inhibitor. The knockdown of TRPV2 also decreased insulin secretion induced by the hypotonic stimulation. In addition, glucose-stimulated insulin secretion was also significantly reduced in the TRPV2-knockdown MIN6 cells. These results suggest that osmotic cell swelling activates TRPV2 in mouse β-cells, thereby causing membrane depolarization and subsequent activation of voltage-dependent Ca channels and insulin secretion.
β 细胞肿胀会引起膜去极化，这至少部分归因于阳离子通道的激活。在这里，我们鉴定了这些阳离子通道。在分离的小鼠胰腺 β 细胞中，暴露于 30%的低渗溶液会引起细胞内 Ca 浓度 ([Ca]) 的增加。钙敏感受体红（ruthenium red），一种包括瞬时受体电位香草酸受体（transient receptor potential cation channel subfamily V，TRPV）在内的几种 Ca 通透性通道的阻滞剂，以及尼卡地平（nicardipine）部分抑制了[Ca]的升高，但细胞内 Ca 储存的耗竭（用 thapsigargin 和 caffeine）则没有。低渗刺激也增加了分离的小鼠胰岛的胰岛素分泌，而钙敏感受体红则显著抑制了这种分泌。通过 RT-PCR、Western blot 和免疫组织化学分析，在小鼠胰腺胰岛和 MIN6 β 细胞系中证实了 TRPV2 和 TRPV4 的表达。然而，4α-佛波醇 12,13-二癸酸酯（4α-phorbol 12,13-didecanoate）或 TRPV4 激活剂 GSK1016790A 对分离的小鼠 β 细胞或 MIN6 细胞中的[Ca]均没有明显影响。相比之下，TRPV2 激活剂丙磺舒（probenecid）在 MIN6 细胞中诱导[Ca]增加，而钙敏感受体红则减弱了这种增加。此外，用 siRNA 敲低 TRPV2 和用 TRPV2 抑制剂曲尼司特（tranilast）处理可显著降低由 30%低渗刺激引起的[Ca]升高。TRPV2 的敲低也降低了低渗刺激引起的胰岛素分泌。此外，TRPV2 敲低的 MIN6 细胞的葡萄糖刺激的胰岛素分泌也显著降低。这些结果表明，渗透压细胞肿胀激活了小鼠 β 细胞中的 TRPV2，从而导致膜去极化和随后电压依赖性 Ca 通道的激活以及胰岛素的分泌。
