Muscarinic receptors coupled to phosphoinositide hydrolysis and elevated cytosolic calcium in a human neuroblastoma cell line SK-N-SH.
1. The effects of the muscarinic agonist carbachol on phosphoinositide metabolism and its relationship to alteration of intracellular calcium were examined in SK-N-SH human neuroblastoma cells. Muscarinic receptors on these cells are coupled to phospholipase C and the myo [2-3H]-inositol phosphates resulting from receptor activation of cells labelled with [3H]-inositol accumulate rapidly. The breakdown of both inositol monophosphate (InsP1) and inositol bisphosphate (InsP2) is sensitive to lithium with inhibition of the latter only observed at higher concentrations of this ion. 2. Use of the calcium indicator dye Fura 2 revealed that carbachol stimulates a biphasic increase in intracellular calcium. 3. Carbachol was able to stimulate both [3H]-inositol phosphate production and intracellular calcium levels with respective EC50 values of 15.9 +/- 1.0 microM and 10.7 +/- 3.2 microM, indicating that no amplification occurs between these steps in the signal transduction pathway. 4. Inositol 1,4,5 trisphosphate (Ins(1,4,5)P3) released 45Ca2+ in a stereospecific and dose-related manner from intracellular stores of permeabilised cells. 5. These results suggest that this cell line may represent a useful model system to investigate receptor-mediated phosphoinositide metabolism and calcium homeostasis.
Baird
J G
JG
Department of Pharmacology and Therapeutics, Leicester.
Lambert
D G
DG
McBain
J
J
Nahorski
S R
SR
D001572:Benzofurans / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002217:Carbachol / Q000494:pharmacology; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D016257:Fura-2; D006801:Humans; D006868:Hydrolysis; D007293:Inosine Triphosphate / Q000494:pharmacology; D009447:Neuroblastoma / Q000378:metabolism*; D010716:Phosphatidylinositols / Q000096:biosynthesis / Q000378:metabolism*; D011976:Receptors, Muscarinic / Q000187:drug effects*; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism
D001572:Benzofurans; D010716:Phosphatidylinositols; D011976:Receptors, Muscarinic; D007293:Inosine Triphosphate; D002217:Carbachol; D002118:Calcium; D016257:Fura-2