Regulation of the cell swelling-activated chloride conductance by cholesterol-rich membrane domains.
OBJECTIVE
The role of high cholesterol-containing microdomains in the signal transduction cascade leading to the activation of volume-regulated anion channels (VRACs) was studied.


METHODS
Osmotic cell swelling-induced efflux of 125I- was determined in human epithelial Intestine 407 cells and in skin fibroblasts obtained from healthy controls or Niemann-Pick type C (NPC) patients. Cellular cholesterol content was modulated by pre-incubation with 2-hydroxypropyl-beta-cyclodextrin in the presence of acceptor lipid vesicles.


RESULTS
Osmotic cell swelling of human Intestine 407 cells leads to the rapid activation of a compensatory anion conductance. Treatment of the cells with cyclodextrin enhanced the response to submaximal hypotonic stimulation by approx. twofold, but did not further increase the efflux elicited by a saturating stimulus. In contrast, the volume-sensitive anion efflux was markedly inhibited when cholesterol-loaded cyclodextrin was used. Potentiation of the response by cholesterol depletion was maintained in caveolin-1 deficient Caco-2 colonocytes as well as in sphingomyelinase-treated Intestine 407 cells, indicating that cholesterol-rich microdomains are not crucially involved. However, treatment of the cells with progesterone, an inhibitor of NPC1-dependent endosomal cholesterol trafficking, not only markedly reduced the hypotonicity-provoked anion efflux, but also prevented its potentiation by cyclodextrin. In addition, the volume-sensitive anion efflux from human NPC skin fibroblasts was significantly smaller when compared with control fibroblasts.


CONCLUSIONS
The results support a model of regulatory volume decrease involving recruitment of volume-sensitive anion channels from intracellular compartments to the plasma membrane.
Lim
C H
CH
Department of Biochemistry, Erasmus University Medical Center, Rotterdam, The Netherlands.
Schoonderwoerd
K
K
Kleijer
W J
WJ
de Jonge
H R
HR
Tilly
B C
BC
D000073738:2-Hydroxypropyl-beta-cyclodextrin; D001692:Biological Transport; D016022:Case-Control Studies; D051242:Caveolin 1 / Q000494:pharmacology; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D048429:Cell Size; D002478:Cells, Cultured; D018118:Chloride Channels / Q000378:metabolism*; D002784:Cholesterol / Q000378:metabolism*; D005079:Excipients / Q000494:pharmacology; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D007038:Hypotonic Solutions; D007413:Intestinal Mucosa / Q000378:metabolism*; D008051:Lipid Bilayers; D052556:Niemann-Pick Disease, Type C / Q000378:metabolism*; D011374:Progesterone / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D013108:Sphingomyelin Phosphodiesterase / Q000494:pharmacology; D047392:beta-Cyclodextrins / Q000494:pharmacology
D051242:Caveolin 1; D018118:Chloride Channels; D005079:Excipients; D007038:Hypotonic Solutions; D008051:Lipid Bilayers; D047392:beta-Cyclodextrins; D000073738:2-Hydroxypropyl-beta-cyclodextrin; D011374:Progesterone; D002784:Cholesterol; D013108:Sphingomyelin Phosphodiesterase