Ca²⁺‐Independent, Ca²⁺‐Dependent, and Carbachol‐ Mediated Arachidonic Acid Release from Rat Brain Cortex Membrane

Abstract

Synaptoneurosomes obtained from the cortex of rat brain prelabeled with [14C]arachidonic acid [( 14C]AA) were used as a source of substrate and enzyme in studies on the regulation of AA release. A significant amount of AA is liberated in the presence of 2 mM EGTA, independently of Ca2+, primarily from phosphatidic acid and polyphosphoinositides (poly-PI). Quinacrine, an inhibitor of phospholipase A2 (PLA2), suppressed AA release by about 60% and neomycin, a putative inhibitor of phospholipase C (PLC), reduced AA release by about 30%. An additive effect was exhibited when both inhibitors were given together. Ca2+ activated AA release. The level of Ca2+ present in the synaptoneurosomal preparation (endogenous level) and 5 microM CaCl2 enhance AA liberation by approximately 25%, whereas 2 mM CaCl2 resulted in a 50% increase in AA release relative to EGTA. The source for Ca(2+)-dependent AA release is predominantly phosphatidylinositol (PI); however, a small pool may also be liberated from neutral lipids. Carbachol, an agonist of the cholinergic receptor, stimulated Ca(2+)-dependent AA release by about 17%. Bradykinin enhanced the effect of carbachol by about 10-15%. This agonist-mediated AA release occurs specifically from phosphoinositides (PI + poly-PI). Quinacrine almost completely suppresses calcium-and carbachol-mediated AA release. Neomycin inhibits this process by about 30% and totally suppresses the effect of bradykinin. Our results indicate that both phospholipases PLA2 and PLC with subsequent action of DAG lipase are responsible for Ca(2+)-independent AA release. Ca(2+)-dependent and carbachol-mediated AA liberation occurs mainly as the result of PLA2 action. A small pool of AA is probably also released by PLC, which seems to be exclusively responsible for the effect of bradykinin.

In vitro activation of cyclo‐oxygenase in the rabbit carotid body: effect of its blockade on [3H]catecholamine release.

“…It is conceivable, however, that the intense hypercapnic-acidic stimulus and high K+ activate additional intraglomic structures to produce PGE2. The notion that chemoreceptor cells are the origin of stimulus-induced PGE2 is also strengthened by the fact that all the stimuli tested in the present study increase [Ca21]i in chemoreceptor cells (Biscoe & Duchen, 1990;Obeso, Rocher, Fidone & Gonzalez, 1992;Gonzalez et al 1992;Sato, Yoshizaki & Koyamo, 1993), this ion being an activator of phospholipases (Irvine, 1982;Berridge, 1987;Strosznajder & Samochocki, 1991), the enzymes that provide arachidonate to cyclo-oxygenase. Taking this into account, it seems plausible that the increase in PGE2 synthesis that takes place in the CB during stimulation is, at least in part, secondary to stimulus-induced elevation of [Ca2+]i in chemoreceptor cells, although other possibilities exist.…”

“…[Ca21]i in chemoreceptor cells (Biscoe & Duchen, 1990;Obeso, Rocher, Fidone & Gonzalez, 1992;Gonzalez et al 1992;Sato, Yoshizaki & Koyamo, 1993), this ion being an activator of phospholipases (Irvine, 1982;Berridge, 1987;Strosznajder & Samochocki, 1991), the enzymes that provide arachidonate to cyclo-oxygenase. Taking this into account, it seems plausible that the increase in PGE2 synthesis that takes place in the CB during stimulation is, at least in part, secondary to stimulus-induced elevation of [Ca2+]i in chemoreceptor cells, although other possibilities exist.…”

Role of arachidonic acid in the regulation of the NMDA-evoked release of acetylcholine in striatal compartments

“…This interpretation is supported by several observations: 1) As shown on striatal neuronal cultures, in the absence of magnesium NMDA stimulates in a concentration-dependent manner the release of arachidonic acid (Dumuis et al, 1988). Similarly, by acting through M 1 muscarinic receptors ACh also stimulates the release of arachidonic acid (Strosznajder and Samochocki, 1991;Tencé et al, 1994) and, in addition, important synergistic effects on arachidonic acid formation are observed under the combined application of NMDA and ACh (or carbachol) (Tencé et al, 1995). 2) As shown in our laboratory, in the rat striatum arachidonic acid inhibits the reuptake of GABA and induced a marked facilitation of its release process through a mechanism which is calcium-dependent and partially results from the activation of a protein kinase C. In addition, the latter effects are not reproduced by other fatty acids and persist following blockade of the main metabolic pathways of arachidonic acid (Chéramy et al, 1996).…”

Aging modulates calcium-dependent phosphatidylinositol degradation by cerebral cortex synaptic plasma membrane phospholipases

“…The pellet was resuspended in 5 mL of fresh buffer and gently filtered through three layers of nylon mesh (pore size, 100 µm) using a plastic syringe (20-mL capacity), and the resulting filtrate was centrifuged at 1100g for 15 min to obtain a synaptoneurosomal pellet. This fraction was examined by electron microscopy as described previously by Strosznajder and Samochocki (1991).…”