• Home
  • Search Results
  • Sodium entry through endothelial store-operated calcium entry channels: regulation by Orai1.

Sodium entry through endothelial store-operated calcium entry channels: regulation by Orai1.

American journal of physiology. Cell physiology (2014-11-28)
Ningyong Xu, Donna L Cioffi, Mikhail Alexeyev, Thomas C Rich, Troy Stevens
ABSTRACT

Orai1 interacts with transient receptor potential protein of the canonical subfamily (TRPC4) and contributes to calcium selectivity of the endothelial cell store-operated calcium entry current (ISOC). Orai1 silencing increases sodium permeability and decreases membrane-associated calcium, although it is not known whether Orai1 is an important determinant of cytosolic sodium transitions. We test the hypothesis that, upon activation of store-operated calcium entry channels, Orai1 is a critical determinant of cytosolic sodium transitions. Activation of store-operated calcium entry channels transiently increased cytosolic calcium and sodium, characteristic of release from an intracellular store. The sodium response occurred more abruptly and returned to baseline more rapidly than did the transient calcium rise. Extracellular choline substitution for sodium did not inhibit the response, although 2-aminoethoxydiphenyl borate and YM-58483 reduced it by ∼50%. After this transient response, cytosolic sodium continued to increase due to influx through activated store-operated calcium entry channels. The magnitude of this sustained increase in cytosolic sodium was greater when experiments were conducted in low extracellular calcium and when Orai1 expression was silenced; these two interventions were not additive, suggesting a common mechanism. 2-Aminoethoxydiphenyl borate and YM-58483 inhibited the sustained increase in cytosolic sodium, only in the presence of Orai1. These studies demonstrate that sodium permeates activated store-operated calcium entry channels, resulting in an increase in cytosolic sodium; the magnitude of this response is determined by Orai1.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid, for molecular biology, ≥97.0%
Sigma-Aldrich
Thapsigargin, ≥98% (HPLC), solid film
Sigma-Aldrich
Ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid, ≥97.0%
Sigma-Aldrich
2-Aminoethyl diphenylborinate, 97%
Sigma-Aldrich
Ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid, BioUltra, for molecular biology, ≥99.0% (T)
Sigma-Aldrich
Krebs-Henseleit Buffer Modified, With 2000 mg/L glucose, without calcium chloride and sodium bicarbonate, powder, suitable for cell culture
Sigma-Aldrich
Fura 2-AM, ≥95% (HPLC)
Sigma-Aldrich
Ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid, BioXtra, ≥97 .0%
Sigma-Aldrich
Fura 2-AM, BioReagent, suitable for fluorescence, ≥95.0% (HPLC)
Sigma-Aldrich
YM-58483, ≥98% (HPLC)