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Arachidonic acid (AA) exerts multiple physiological and pathophysiological effects in the brain. By continuously measuring the intracellular pH (pHi) and Ca2+ levels ([Ca2+]i) in primary cultured rat cerebellar granule cells, we have found, for the first time, that 20 min treatment with 10 μm AA resulted in marked increases in Ca2+ and H+ levels in both the cytosol and nucleus. A much higher concentration (40 mm) of another weak acid, propionic acid, was needed to induce a similar change in pHi. The [Ca2+]i increase was probably caused by AA‐induced activation of Ni2+‐sensitive cationic channels, but did not involve NMDA channels or the Na+‐Ca2+ exchanger. AA‐induced acidosis occurs by a different mechanism involving predominantly the passive diffusion of the un‐ionized form of AA, rather than a protein carrier, as proposed by Kamp & Hamilton for fatty acids (FAs) in artificial phospholipid bilayers (the ‘flip‐flop’ model). The following results, which are similar to those observed in lipid bilayers, support this conclusion: (1) FAs containing a ‐COOH group (AA, linoleic acid, α‐linolenic acid, and docosahexaenoic acid) induced intracellular acidosis, whereas a FA with a ‐COOCH3 group (AA methyl ester) had little effect on pHi, (2) a FA amine, tetradecylamine, induced intracellular alkalosis, and (3) the AA‐/FA‐induced pHi changes were reversed by bovine serum albumin. Further evidence in support of a passive diffusion model, rather than a membrane protein carrier, is that: (1) there was a linear relationship between the initial rate of acid flux and the concentration of AA (2‐100 μm), (2) acidosis was not inhibited by 4,4′‐diisothiocyanatostilbene‐2,2′‐disulphonic acid, a potent inhibitor of the plasma membrane FA carrier protein, and (3) the involvement of most known H+‐related membrane carriers and H+ conductance has been ruled out. Since AA can be released under both physiological and pathophysiological conditions, the possible significance of the AA‐evoked increases in H+ and Ca2+ in both the cytoplasm and nucleoplasm is discussed.

1.beta-Bungarotoxin, crotoxin and taipoxin, presynaptic neurotoxins of snake venom origin, have about the same phospholipid-splitting activities as a much less toxic cobra phospholipase A2 in the presence of Ca2+ and deoxycholate. 2. Sr2+ was a much less effective activator of the enzymes than is Ca2+, the activation by Sr2+ being only 3-6% for beta-bungarotoxin and crotoxin and 12% for taipoxin. 3. Sr2+ also inhibited the Ca2+ -activated enzymes by 80% in the cases of beta-bungarotoxin and crotoxin, but only 16% in the case of taipoxin. 4. Mg2" had no significant effect on beta-bungarotoxin or crotoxin, but activated taipoxin in the presence or absence of Ca2". 5. In Sr2+ -Tyrode lacking Ca2+ all three toxins exhibited the same immediate depression followed by facilitation in the rat and mouse diaphragms, but the final blocking activity was only 3-10% with beta-bungarotoxin and crotoxin and was 30% with taipoxin. 6. In Sr2+ -Tyrode, increasing in the rate of nerve stimulation had less accelerating effect on the development of neuromuscular block than in Ca2+ -Tyrode for any of the toxins. 7. Removal of Mg2+ from Sr2+ -Tyrode did not diminish the potency of taipoxin in blocking neuromuscular transmission, suggesting that enzyme activity at the outer surface of the axolemma does not contribute to the neuromuscular blocking action. 8. All of the results indicate that there are close correlations between the presynaptic activities of these toxins and their phospholipid-splitting activities in the cationic environment prevailing in the axoplasm. Apparently the final blocking effect of these toxins is due to phospholipase A action within the nerve terminal.

Arachidonic acid (AA) and other nonesterified fatty acids (FAs) have been shown to exert harmful effects during cardiac ischemia. By continuously measuring intracellular pH (pH(i)) changes in neonatal and adult cardiac myocytes, we have found, for the first time, that 10 micromol/L AA induces a substantial intracellular acidosis (0.3 to 0.4 pH units). We have ruled out the possibilities that the AA-induced acidosis is caused by (1) inhibition or stimulation of the pH(i) regulators, (2) protein kinase C activation or the generation of AA metabolites or free radicals, or (3) activation of NADPH oxidase or an inward H(+) current. The AA-induced acidosis fits to a simple diffusion mechanism, as proposed by Kamp and Hamilton (flip-flop model) for artificial phospholipid bilayers. The important properties found in the cardiac myocyte are that (1) the initial rate of acid flux (J(H)) increases with the AA concentration (2 to 50 micromol/L), (2) FAs with a (-)COOH group (eg, AA, oleic acid, and linoleic acid) induce intracellular acidification, but FAs with a (-)COOCH(3) group (eg, AA methyl ester) have little effect on the pH(i), (3) tetradecylamine (FA amine) induces intracellular alkalosis, and, most importantly, (4) both the AA- and tetradecylamine-induced pH(i) changes can be reversed by 0.3% BSA. Because a low concentration of AA (10 micromol/L) can induce a substantial acidosis, the possible involvement of the FA-evoked acidosis in the negative inotropic effect during cardiac ischemia is discussed. The full text of this article is available at http://www. circresaha.org.