NDLI: Role of TRPC3 channels in ATP-induced Ca2+ signaling in principal cells of the inner medullary collecting duct

Content Provider	PubMed Central
Author	Goel, Monu Schilling, William P.
Abstract	The transient receptor potential channel TRPC3 is exclusively expressed in the apical membrane of principal cells of the collecting duct (CD) both in vivo and in the mouse CD cell line IMCD-3. Previous studies revealed that ATP-induced apical-to-basolateral transepithelial Ca2+ flux across IMCD-3 monolayers is increased by overexpression of TRPC3 and attenuated by a dominant negative TRPC3 construct, suggesting that Ca2+ entry across the apical membrane occurs via TRPC3 channels. To test this hypothesis, we selectively measured the Ca2+ permeability of the apical membrane of fura-2-loaded IMCD-3 cells using the Mn2+ quench technique. Mn2+ influx across the apical membrane was increased 12- to 16-fold by apical ATP and was blocked by the pyrazole derivative BTP2, a known inhibitor of TRPC3 channels, with an IC50 value <100 nM. In contrast, Mn2+ influx was only increased ∼2-fold by basolateral ATP. Mn2+ influx was also activated by apical, but not basolateral, 1-stearoyl-2-acetyl-sn-glycerol (SAG), a known activator of TRPC3 channels. Apical ATP- and SAG-induced Mn2+ influx was increased by overexpression of TRPC3 and completely blocked by expression of the dominant negative TRPC3 construct. Mn2+ influx was also stimulated ∼2-fold by thapsigargin applied to either the apical or basolateral side. Thapsigargin-induced flux was blocked by BTP2 but was unaffected by overexpression of TRPC3 or by dominant negative TRPC3. Apical ATP, but not basolateral ATP, increased transepithelial 45Ca2+ flux. These results demonstrate that the apical membrane of IMCD-3 cells has two distinct Ca2+ influx pathways: 1) a store-operated channel activated by thapsigargin and basolateral ATP and 2) TRPC3 channels activated by apical ATP. Only activation of TRPC3 leads to net transepithelial apical-to-basolateral Ca2+ flux. Furthermore, these results demonstrate that native TRPC3 is not a store-operated channel in IMCD-3 cells.
Related Links	http://dx.doi.org/10.1152/ajprenal.00670.2009
Starting Page	225
File Format	PDF
ISSN	03636127
e-ISSN	15221466
Journal	American Journal of Physiology - Renal Physiology
Issue Number	1
Volume Number	299
Language	English
Publisher	American Physiological Society
Publisher Date	2010-07-01
Access Restriction	Open
Rights Holder	American Physiological Society
Subject Keyword	Physiology Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Physiology

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