Fluoride, which is used commonly as a pharmacological tool to activate
phosphoinositide-phospholipase C coupled to the heterotrymeric Gq/11
proteins, inhibited the phosphorylation of
phosphatidylinositol (
PtdIns) to
polyphosphoinositides (
PtdIns4P and PtdIns4,5P2) in membranes from rat brain cortex.
Fluoride enhanced basal production of 3H-inositol
phosphates in membranes prepared from brain cortical slices that had been prelabeled with [3H]
inositol, but inhibited the stimulation elicited by
carbachol in the presence of
GTPgammaS. However in both cases
fluoride depleted [3H]
PtdIns4P content by 95%. The inhibitory effects of
fluoride on the release of 3H-inositol
phosphates in slices were not apparent in a pulse [3H]
inositol-labeling strategy, but became dramatic in a continuous labeling protocol, particularly at long incubation times. Prelabeling slices with [3H]
inositol in the presence of
fluoride precluded
polyphosphoinositide labeling, and eliminated
phospholipase C responsiveness to
carbachol under normal or depolarizing conditions, and to the
calcium ionophore ionomycin. The lack of response of 3H-polyphosphoinositide-depleted slices to
phospholipase C stimuli was not due to
fluoride poisoning, unaccessibility of the [3H]
inositol label to
phospholipase C or desensitization of Gq/11, as the effect of
carbachol and
GTPgammaS was restored, in the presence of
ATP, in membranes prepared from slices that had been labeled in the presence of
fluoride. In conclusion, our data show that
fluoride, at a concentration similar to that used to stimulate directly Gq/11-coupled
phospholipase C, effectively blocks the synthesis of
phospholipase C substrates from
PtdIns.