Pecze László, Blum Walter, Schwaller Beat
Department of Medicine, University of Fribourg, Fribourg, Switzerland.
Biochim Biophys Acta. 2013 Jul;1833(7):1680-91. doi: 10.1016/j.bbamcr.2012.08.018. Epub 2012 Sep 5.
Transient receptor potential vanilloid subtype 1 (TRPV1) receptor is a pain-sensing, ligand-gated, non-selective cation channel expressed in peripheral sensory neurons. Prolonged activation of TRPV1 by capsaicin leads to cell swelling and formation of membrane blebs in rat dorsal root ganglion (DRG) neurons. Similar results were obtained in NIH3T3 fibroblast cells stably expressing TRPV1. Here, we assessed the contribution of Ca(2+) and Na(+) ions to TRPV1-mediated changes. Cell swelling was caused by a substantial influx of extracellular Na(+) via TRPV1 channels, causing concomitant transport of water. In the absence of extracellular Na(+), the membrane blebbing was completely inhibited, but Ca(2+) influx did not change under these conditions. Na(+) influx was modulated by the intracellular Ca(2+) concentration ([Ca(2+)]i). Elevation of [Ca(2+)]i by ionomycin sensitized/activated TRPV1 channels causing cell swelling in TRPV1-positive cells. In the absence of extracellular Ca(2+), capsaicin caused only little increase in [Ca(2+)]i indicating that the increase in [Ca(2+)]i observed after capsaicin application is derived essentially from extracellular Ca(2+) and not from internal Ca(2+) stores. In the absence of extracellular Ca(2+) also the process of cell swelling was considerably slower. Calretinin is a Ca(2+) buffer protein, which is expressed in a subset of TRPV1-positive neurons. Calretinin decreased the amplitude, but slowed down the decay of Ca(2+) signals evoked by ionomycin. Cells co-expressing TRPV1 and calretinin were less sensitive to TRPV1-mediated, capsaicin-induced volume increases. In TRPV1-expressing NIH3T3 cells, calretinin decreased the capsaicin-induced Ca(2+) and Na(+) influx. Swelling and formation of membrane blebs resulted in impaired plasma membrane integrity finally leading to cell death. Our results hint towards a mechanistic explanation for the apoptosis-independent capsaicin-evoked neuronal loss and additionally reveal a protective effect of calretinin; we propose that the Ca(2+)-buffering capacity of calretinin reduces the susceptibility of calretinin-expressing DRG neurons against cell swelling/death caused by overstimulation of TRPV1 channels. This article is part of a Special Issue entitled:12th European Symposium on Calcium.
瞬时受体电位香草酸亚型1（TRPV1）受体是一种在周围感觉神经元中表达的疼痛感知、配体门控、非选择性阳离子通道。辣椒素对TRPV1的长期激活会导致大鼠背根神经节（DRG）神经元细胞肿胀和膜泡形成。在稳定表达TRPV1的NIH3T3成纤维细胞中也获得了类似结果。在此，我们评估了Ca(2+)和Na(+)离子对TRPV1介导变化的作用。细胞肿胀是由细胞外Na(+)通过TRPV1通道大量内流引起的，并伴随水的转运。在没有细胞外Na(+)的情况下，膜泡形成被完全抑制，但在此条件下Ca(2+)内流没有变化。Na(+)内流受细胞内Ca(2+)浓度（[Ca(2+)]i）调节。离子霉素使[Ca(2+)]i升高，从而使TRPV1通道敏感化/激活，导致TRPV1阳性细胞肿胀。在没有细胞外Ca(2+)的情况下，辣椒素仅使[Ca(2+)]i略有增加，这表明辣椒素作用后观察到的[Ca(2+)]i增加主要源于细胞外Ca(2+)，而非细胞内Ca(2+)储存。在没有细胞外Ca(2+)的情况下，细胞肿胀过程也明显减慢。钙视网膜蛋白是一种Ca(2+)缓冲蛋白，在一部分TRPV1阳性神经元中表达。钙视网膜蛋白降低了离子霉素诱发的Ca(2+)信号的幅度，但减缓了其衰减。共表达TRPV1和钙视网膜蛋白的细胞对TRPV1介导的、辣椒素诱导的体积增加不太敏感。在表达TRPV1的NIH3T3细胞中，钙视网膜蛋白减少了辣椒素诱导的Ca(2+)和Na(+)内流。肿胀和膜泡形成导致质膜完整性受损，最终导致细胞死亡。我们的结果为与凋亡无关的辣椒素诱发的神经元损失提供了一种机制解释，此外还揭示了钙视网膜蛋白的保护作用；我们提出，钙视网膜蛋白的Ca(2+)缓冲能力降低了表达钙视网膜蛋白的DRG神经元对TRPV1通道过度刺激引起的细胞肿胀/死亡的易感性。本文是名为：第12届欧洲钙研讨会的特刊的一部分。
