The Spider Toxin GsMTx-4 Blocks TRPV4 Cation Channels Expressed in HEK-293 Cells

TRPV4型 机械敏感通道 瞬时受体电位通道 化学 HEK 293细胞 生物物理学 转染 细胞内 激活剂(遗传学) 呋喃-2 荧光 离子通道 受体 生物化学 生物 胞浆 物理 量子力学 基因
作者
Christian S.J. Kesselring,Mirjam Krautwald,Yaxin Zhang,Heinrich Brinkmeier
出处
期刊:Biophysical Journal [Elsevier]
卷期号:108 (2): 126a-126a 被引量:1
标识
DOI:10.1016/j.bpj.2014.11.702
摘要

The transient receptor potential channel TRPV4 is a calcium conducting, osmosensitive cation channel and expressed in motoneurons and skeletal muscles. In mouse muscle fibers TRPV4 forms or contributes to a mechanosensitive cation channel. The molecular architecture of this channel is still unclear. Since mechanosensitive cation channels can be blocked by the peptide toxin GsMTx-4, we studied whether TRPV4 itself is sensitive to GsMTx-4. To this end a TRPV4-YFP construct was functionally expressed in HEK-293 cells. Intracellular Ca2+ concentrations were monitored with the fluorescent Ca2+ indicator Fura-2 and the data presented as Fura-2 fluorescence ratios after alternate excitation at 340 and 380 nm. Non-transfected cells (n=47) had an average resting ratio of 0.39 (range 0.38 and 0.42), while TRPV4 transfected cells (n=524) showed on average an increased fluorescence ratio of 1.11 ranging from 0.44 to 2.71. Application of 4a-PDD (5 µM), a known TRPV4 activator, increased the fluorescence ratios in the HEK cells to 1.72, 1.73 and 1.77 with a peak or plateau at 1-3 min (n=13-19 cells, each experiment). The effect of 4a-PDD could be reversed within about 10-30 min by application of the TRPV4 blocker HC-067047 (1 µM) and GsMTx-4 (5 µM). The final resting ratios were 0.77 for HC-067047 and 1.16 for GsMTx-4. At a concentration of 1 µM the toxin was almost ineffective. The results indicate that TRPV4 activity can be inhibited by GsMTx-4. However, the required concentration of the toxin is higher than those required for inhibition of TRPC6, TRPC1 and voltage-gated sodium channels. The results support the view that TRPV4 may be part of mechanosensitive cation channels that are sensitive to the channel blocker GsMTx-4. The work was supported by the Deutsche Duchenne Stiftung der aktion benni & co e. V.
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