Activation of muscarinic acetylcholine receptors induces Ca(2+) mobilization in FRT cells.
The effect of the muscarinic receptors agonist carbachol (Cch) on intracellular calcium concentration ([Ca(2+)](i)) and cAMP level was studied in polarized Fischer rat thyroid (FRT) epithelial cells. Cch provoked a transient increase in [Ca(2+)](i), followed by a lower sustained phase. Thapsigargin, a specific microsomal Ca(2+)-ATPase inhibitor, caused a rapid rise in [Ca(2+)](i) and subsequent addition of Cch was without effect. Removal of extracellular Ca(2+) reduced the initial transient response and completely abolished the plateau phase. Ryanodine, an agent that depletes intracellular Ca(2+) stores through stimulation of ryanodine receptors (RyRs), had no effect on [Ca(2+)](i). However, the transitory activation of [Ca(2+)](i) was dose-dependently attenuated in cells pretreated with U73122, a specific inhibitor of phospholipase C (PLC). These data suggest that the Cch-stimulated increment of [Ca(2+)](i) required IP(3) formation and binding to its specific receptors in Ca(2+) stores. Further studies were performed to investigate whether the effect of Cch on Ca(2+) entry into FRT cells was via L-type voltage-dependent Ca(2+) channels (L-VDCCs). Nicardipine, a nonspecific L-type Ca(2+) channel blocker, decreased Cch-induced increase on [Ca(2+)](i), while Bay K-8644, an L-type Ca(2+) channel agonist, slightly increased [Ca(2+)](i) in FRT cells. These data indicate that Ca(2+) entry into these nondifferentiated thyroid cells occurs through an L-VDCC, and probably through another mechanism such as a capacitative pathway. Cch did not affect the intracellular cAMP levels, but its effects on [Ca(2+)](i) were significantly reduced when cells were pretreated with forskolin, suggesting the existence of an intracellular cross-talk between PLC and cAMP mechanisms in the regulation of intracellular Ca(2+) mobilization in neoplastic FRT cells.
Departamento de Bioquímica y Biología Molecular, Facultad de Medicina, Universidad de Málaga, Spain.
M
M
Montiel
J
J
Pavía
S
S
Marsigliante
E
E
Jiménez
D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002118:Calcium / Q000378:metabolism*; D020746:Calcium Channels, L-Type / Q000187:drug effects / Q000378:metabolism; D002217:Carbachol / Q000378:metabolism / Q000494:pharmacology*; D002460:Cell Line; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D018721:Muscarinic Agonists / Q000494:pharmacology*; D018727:Muscarinic Antagonists / Q000494:pharmacology*; D051381:Rats; D011916:Rats, Inbred F344; D011976:Receptors, Muscarinic / Q000378:metabolism*; D012433:Ryanodine / Q000494:pharmacology; D015398:Signal Transduction; D013961:Thyroid Gland; D013997:Time Factors
D020746:Calcium Channels, L-Type; D004791:Enzyme Inhibitors; D018721:Muscarinic Agonists; D018727:Muscarinic Antagonists; D011976:Receptors, Muscarinic; D012433:Ryanodine; D005576:Colforsin; D002217:Carbachol; D000242:Cyclic AMP; D002118:Calcium