No abstract

In our in vitro model, rasagiline a selective irreversible monoamine oxidase-B (MAO-B) inhibitor, protected nerve growth factor (NGF)-differentiated PC12 cells from cell death under oxygen and glucose deprivation (OGD). The severity of the OGD insult, as expressed by cell death, was time-dependent. Exposure of the cells to OGD for 3 hr followed by 18 hr of reoxygenation caused about 30-40% cell death. Under these conditions, the neuroprotective effect of rasagiline was dose-dependent: rasagiline reducing OGD-induced cell death by 68% and 80% at 100 nM and 1 microM, respectively. The neuroprotective effect of rasagiline was also observed when added after the OGD insult (55% reduction in cell death). Under rasagiline treatment, there was a lesser decrease in ATP content in cultures exposed to OGD compared with that in untreated cultures. OGD followed by reoxygenation resulted in a several fold increase in PGE(2) release into the extracellular medium. Rasagiline (100 nM-1 microM) markedly inhibited OGD-induced PGE(2) release. Clorgyline, a monoamine oxidase-A (MAO-A) inhibitor, did not protect NGF-differentiated PC12 cells against OGD-induced cell death. As NGF-differentiated PC12 cells contain exclusively MAO type A, these data suggest that the neuroprotective effect of rasagiline under OGD conditions is independent of MAO inhibition.

In our in vitro model, rasagiline a selective irreversible monoamine oxidase-B (MAO-B) inhibitor, protected nerve growth factor (NGF)-differentiated PC12 cells from cell death under oxygen and glucose deprivation (OGD). The severity of the OGD insult, as expressed by cell death, was time-dependent. Exposure of the cells to OGD for 3 hr followed by 18 hr of reoxygenation caused about 30-40% cell death. Under these conditions, the neuroprotective effect of rasagiline was dose-dependent: rasagiline reducing OGD-induced cell death by 68% and 80% at 100 nM and 1 microM, respectively. The neuroprotective effect of rasagiline was also observed when added after the OGD insult (55% reduction in cell death). Under rasagiline treatment, there was a lesser decrease in ATP content in cultures exposed to OGD compared with that in untreated cultures. OGD followed by reoxygenation resulted in a several fold increase in PGE(2) release into the extracellular medium. Rasagiline (100 nM-1 microM) markedly inhibited OGD-induced PGE(2) release. Clorgyline, a monoamine oxidase-A (MAO-A) inhibitor, did not protect NGF-differentiated PC12 cells against OGD-induced cell death. As NGF-differentiated PC12 cells contain exclusively MAO type A, these data suggest that the neuroprotective effect of rasagiline under OGD conditions is independent of MAO inhibition.

We report the isolation of a novel pardaxin isoform from the toxic secretion of the Red Sea Moses sole (Pardachirus marmoratus). Mass spectrometrical analysis of the newly purified peptide revealed a different primary structure compared to the previously known pardaxin isoforms. Sequence analysis disclosed an aspartic acid residue instead of glycine at position 31 of the new isoform. According to the novel sequence, a synthetic Asp-31-peptide was compared with the native compound as well as with synthetic Gly-31-pardaxin. The isolated Asp-31-pardaxin isoform and its synthetic analog exhibited identical elution properties during reverse-phase HPLC, as well as similar dosedependent lytic effects on human erythrocytes at a concentration of 10 3T to 10 3S M. The hemolytic activity of Asp-31-pardaxins was lower than that of Gly-31-pardaxin and no synergistic effect between these peptides was found. The additional negative charge introduced by Asp-31 is likely to affect the selectivity of pardaxin pores towards a variety of ions.z 1998 Federation of European Biochemical Societies.

Pardaxin (PX) is a voltage-dependent ionophore that stimulates catecholamine exocytosis from PC-12 pheochromocytoma cells both in the presence and absence of extracellular calcium. Using a battery of phospholipase A(2) inhibitors we show that PX stimulation of phospholipase A(2) (PLA(2)) enzymes is coupled with induction of exocytosis. We investigated the relationship between PX-induced PLA(2) activity and neurotransmitter release by measuring the levels of arachidonic acid (AA), prostaglandin E(2) (PGE(2)), and dopamine release. In the presence of extracellular calcium, the cytosolic PLA(2) inhibitor arachidonyl trifluoromethyl ketone (AACOCF(3)) inhibited by 100, 70, and 73%, respectively, the release of AA, PGE(2), and dopamine induced by PX. The mitogen-activated protein kinase/extracellular signal-regulated kinase inhibitor 2'-amino-3'-methoxyflavone (PD98059) reduced by 100 and 82%, respectively, the release of AA and PGE(2) induced by PX. In the absence of extracellular calcium, the calcium-independent PLA(2) (iPLA(2)) inhibitors methyl arachidonyl fluorophosphonate, AACOCF(3), and bromoenol lactone (BEL) inhibited by 80 to 90% PX stimulation of AA release, by 65 to 85% PX stimulation of PGE(2) release, and by 80 to 90% PX-induced dopamine release. Using vesicle fusion-based enzyme-linked immunosorbent assay we found similar levels of inhibition of PX-induced exocytosis by these inhibitors. Also, PX induced the formation of soluble N-ethylmaleimide-sensitive factor attachment protein receptor complexes, an effect that was augmented by N-methylmaleimide. This complex formation was completely inhibited by BEL. Botulinum toxins type C1 and F significantly inhibited the release of AA, PGE(2), and dopamine induced by PX. Our data suggest that PX stimulates exocytosis by activating cystolic PLA(2) and iPLA(2), leading to the generation of AA and eicosanoids, which, in turn, stimulate vesicle competence for fusion and neurotransmitter release.