On the role of agonist-evoked Ca2+ mobilization in sustaining the ongoing phosphoinositide hydrolysis. A study on intact SK-N-BE(2) neuroblastoma cells subjected to muscarinic stimulation.
Stimulation of SK-N-BE(2) cells with 1 mM carbachol (Cch) elicited phosphoinositide (PPI) hydrolysis and a rapid elevation of cytosolic Ca2+ concentration ([Ca2+]i) from 115 nM to about 500 nM, followed by a plateau around 200 nM. In myo [3H]inositol-labelled cells, Cch-evoked accumulation of [3H]inositol phosphate (IPs) was not affected when [Ca2+]i was clamped at resting by cell loading with 10 microM BAPTA/AM; under these conditions, maximal 1,4,5-inositol trisphosphate accumulation was not reduced either. When [Ca2+]i was clamped around 700 nM by cell treatment with 600 nM ionomycin, Cch-evoked [3H]IPs accumulation was enhanced by less than 20%, but it was impaired by a 30% and a 55% after [Ca2+]i reduction to about 70 nM and 35-50 nM, by cell loading with 20 microM or 40 microM BAPTA/AM, respectively. These results show that, in SK-N-BE(2) cells, Cch-activated PPI-specific phospholipase C is sensitive to [Ca2+]i but it already operates under suboptimal conditions at resting [Ca2+]i.
Limatola
C
C
Department of Biology, University of Rome Tor Vergata, Italy.
Pacini
L
L
Candi
E
E
Spinedi
A
A
D002118:Calcium / Q000378:metabolism; D002120:Calcium Channel Agonists / Q000494:pharmacology*; D002217:Carbachol / Q000494:pharmacology; D002614:Chelating Agents / Q000494:pharmacology; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004594:Electrophysiology; D006801:Humans; D006868:Hydrolysis; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D008916:Miotics / Q000494:pharmacology; D009447:Neuroblastoma; D010716:Phosphatidylinositols / Q000378:metabolism*; D014407:Tumor Cells, Cultured / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism
D002120:Calcium Channel Agonists; D002614:Chelating Agents; D007476:Ionophores; D008916:Miotics; D010716:Phosphatidylinositols; D004533:Egtazic Acid; D015759:Ionomycin; D002217:Carbachol; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D002118:Calcium