Cholesterol depletion perturbs calcium handling by rat submandibular glands
Copyright policy )Cholesterol depletion perturbs calcium handling by rat submandibular glands
AbstractThe sensitivity to cholesterol depletion of calcium handling by rat submandibular glands was investigated. The glands were digested with collagenase. After homogenization, the lysate was extracted at 4°C with 0.5% Triton X‐100 and the extract was submitted to an ultracentrifugation in a sucrose discontinuous gradient. A population of detergent‐resistant membranes (DRM) was collected at the 5%–35% interface. The DRM had a higher content of cholesterol, saturated and long‐chain fatty acids. Caveolin‐1 and αq/11 were located in these membranes. They were more ordered than vesicles from total cellular lysate as determined by anisotropy measurement. They disappeared after cholesterol extraction with methyl‐β‐cyclodextrin (MCD). Exposure of the cellular suspension with MCD nearly abolished the response to carbachol, epinephrine, and substance P and inhibited the activation of phospholipase C (PLC) by these agonists and by sodium fluoride. MCD did not affect the mobilization of intracellular pools of calcium by thapsigargin. It increased the uptake of extracellular calcium or barium and did not inhibit the uptake of calcium after depletion of the intracellular stores of this ion. From these results, it is concluded that Triton X‐100 can extract a fraction of membrane resistant to detergents. Treatment of the cells with MCD disrupts these membranes. The coupling between the heterotrimeric GTP‐binding protein Gq/11 and poly‐phosphoinositide‐specific PLC is affected by disruption of these membrane fractions. At the opposite, the store‐operated calcium channel (SOCC) is not affected by DRM‐disruption. © 2004 Wiley‐Liss, Inc.
Intracellular Fluid, Male, Caveolin 1, Wistar, Intracellular Fluid -- drug effects, Calcium Signaling -- physiology, Extracellular Fluid -- metabolism, Calcium Signaling -- drug effects, Cell Membrane -- metabolism, Octoxynol -- pharmacology, Epithelial Cells -- metabolism, Down-Regulation -- drug effects, Down-Regulation -- physiology, Gq-G11 -- drug effects, Cell Membrane -- drug effects, Sciences bio-médicales et agricoles, Type C Phospholipases -- metabolism, GTP-Binding Protein alpha Subunits, Subcellular Fractions -- drug effects, Cholesterol, Gq-G11 -- metabolism, beta-Cyclodextrins -- pharmacology, Epithelial Cells -- drug effects, Subcellular Fractions, Calcium Channels -- drug effects, Octoxynol, Submandibular Gland, Down-Regulation, Caveolins, Cholesterol -- metabolism, Animals, Calcium Signaling, Rats, Wistar, Intracellular Fluid -- metabolism, Type C Phospholipases -- antagonists & inhibitors, Calcium Channels -- metabolism, Extracellular Fluid -- drug effects, Caveolins -- metabolism, Cell Membrane, Epithelial Cells, Extracellular Fluid, Submandibular Gland -- drug effects, Rats, Subcellular Fractions -- metabolism, Type C Phospholipases, Caveolins -- drug effects, Calcium -- metabolism, GTP-Binding Protein alpha Subunits, Gq-G11, Calcium, Submandibular Gland -- metabolism, Calcium Channels
Intracellular Fluid, Male, Caveolin 1, Wistar, Intracellular Fluid -- drug effects, Calcium Signaling -- physiology, Extracellular Fluid -- metabolism, Calcium Signaling -- drug effects, Cell Membrane -- metabolism, Octoxynol -- pharmacology, Epithelial Cells -- metabolism, Down-Regulation -- drug effects, Down-Regulation -- physiology, Gq-G11 -- drug effects, Cell Membrane -- drug effects, Sciences bio-médicales et agricoles, Type C Phospholipases -- metabolism, GTP-Binding Protein alpha Subunits, Subcellular Fractions -- drug effects, Cholesterol, Gq-G11 -- metabolism, beta-Cyclodextrins -- pharmacology, Epithelial Cells -- drug effects, Subcellular Fractions, Calcium Channels -- drug effects, Octoxynol, Submandibular Gland, Down-Regulation, Caveolins, Cholesterol -- metabolism, Animals, Calcium Signaling, Rats, Wistar, Intracellular Fluid -- metabolism, Type C Phospholipases -- antagonists & inhibitors, Calcium Channels -- metabolism, Extracellular Fluid -- drug effects, Caveolins -- metabolism, Cell Membrane, Epithelial Cells, Extracellular Fluid, Submandibular Gland -- drug effects, Rats, Subcellular Fractions -- metabolism, Type C Phospholipases, Caveolins -- drug effects, Calcium -- metabolism, GTP-Binding Protein alpha Subunits, Gq-G11, Calcium, Submandibular Gland -- metabolism, Calcium Channels
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