Differential sensitivity of phosphoinositide metabolism to sodium fluoride and carbachol treatments in PC12 cells.
Exposure to sodium fluoride (NaF) resulted in an increased accumulation (up to 10-fold) of total [(3)H]inositol phosphates (T-InsP) in rat PC 12 cells. The magnitude of the NaF effect was comparable to that for muscarinic acetylcholine receptor-mediated stimulation of T-InsP accumulation in the presence of saturating concentrations of carbachol, but effects of NaF and muscarinic agonists were additive at subsaturating concentrations. The NaF effect was atropine insensitive; was not mimicked by effects of NaCl (10 mM), aluminum fluoride (1 to 100 muM), forskolin (up to 100 muM), or dibutyryl cyclic AMP (1 mM); and was not altered by treatment with pertussis or cholera toxins (1 mug/ml for 24 h). By contrast, the carbachol response was fully sensitive to atropine and partly sensitive to pertussis toxin. Chelation of extracellular calcium ion following 10 min of pretreatment with EDTA or EGTA (3 mM) inhibited carbachol-stimulated T-InsP accumulation by 50%, but resulted in an enhancement of NaF-stimulated T-InsP accumulation. By contrast, inhibition of the mobilization of intracellular calcium ion with 8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate inhibited NaF stimulation of T-InsP accumulation by more than 50% but inhibited carbachol-stimulated TInsP accumulation to a much lower extent. Enhanced calcium influx and cell depolarization stimulated by high extracellular concentrations of KCl markedly potentiated carbachol, but not NaF, stimulation of T-InsP accumulation. This differential sensitivity to muscarinic antagonists, cell depolarization, and manipulation of intra- and extracellular calcium ion indicates that different mechanisms underly NaF and carbachol stimulation of T-InsP accumulation. However, stimulation of T-InsP accumulation in the presence of carbachol alone, NaF alone, or carbachol plus NaF was inhibited to a similar extent in the presence of the phorbol ester, phorbol 12-myristate13-acetate. Taken together, these observations suggest that NaF and carbachol effects are mediated through distinct mechanisms but share a common target, perhaps a GTP-binding protein and/or phospholipase C, whose activity is known to be influenced by protein kinase C.
Bencherif
M
M
Division of Neurobiology, Barrow Neurological Institute, 350 West Thomas Road, Phoenix, Arizona 85013, USA.
Lukas
R J
RJ