Muscarinic receptor-induced phosphoinositide hydrolysis at resting cytosolic Ca2+ concentration in PC12 cells.
In PC12 cells, cultured in the presence of nerve growth factor to increase their complement of muscarinic receptors, treatment with carbachol induces muscarinic receptor-dependent rises in free cytosolic Ca2+ as well as hydrolysis of membrane phosphoinositides. Experiments were carried out to clarify the relationship between these two receptor-triggered events. In particular, since inositol-1,4,5-trisphosphate (the hydrophilic metabolite produced by the hydrolysis of phosphatidylinositol-4,5-bisphosphate) is believed to mediate intracellularly the release of Ca2+ from nonmitochondrial store(s), it was important to establish whether it can be generated at resting cytoplasmic concentration of Ca2+ (approximately 0.1 microM). Cells incubated in Ca2+-free medium were depleted of their cytoplasmic Ca2+ stores by pretreatment with ionomycin. When these cells were then treated with carbachol, their cytosolic concentration of Ca2+ remained at the resting level, whereas inositol-1,4,5-trisphosphate generation was still markedly stimulated. Our results demonstrate that an increase in the concentration of cytosolic Ca2+ is not a necessary intermediate between receptor activation and phosphoinositide hydrolysis, and therefore support the second-messenger role of inositol-1,4,5-trisphosphate.
Vicentini
L M
LM
Ambrosini
A
A
Di Virgilio
F
F
Pozzan
T
T
Meldolesi
J
J
D000818:Animals; D002118:Calcium / Q000502:physiology*; D002460:Cell Line; D004533:Egtazic Acid / Q000494:pharmacology; D006868:Hydrolysis; D007295:Inositol Phosphates / Q000378:metabolism; D009414:Nerve Growth Factors / Q000494:pharmacology; D009490:Neurosecretory Systems; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols / Q000378:metabolism*; D011976:Receptors, Muscarinic / Q000502:physiology*
D007295:Inositol Phosphates; D009414:Nerve Growth Factors; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols; D011976:Receptors, Muscarinic; D004533:Egtazic Acid; D002118:Calcium