Absolute configuration at C45 in 45-hydroxyyessotoxin, a marine polyether toxin isolated from shellfish. (9/156)

The marine polyether toxin, 45-hydroxyyessotoxin, was isolated together with yessotoxin from the scallop, Patinopecten yessoensis. The 45-hydroxy group in the side chain was esterified with (S)- and (R)-alpha-methoxyalpha-trifluoromethylphenylacetic acids (MTPA). A detailed analysis of the 2D NMR spectra of the two esters established the R configuration at C45.  (+info)

Insulinotropic toxins as molecular probes for analysis of glucagon-likepeptide-1 receptor-mediated signal transduction in pancreatic beta-cells. (10/156)

Cholera toxin, pertussis toxin, mastoparan, maitotoxin, and alpha-latrotoxin are complex protein or polyether-based toxins of bacterial, insect, or phytoplankton origin that act with high potency at the endocrine pancreas to stimulate secretion of insulin from beta-cells located in the islets of Langerhans. The remarkable insulinotropic properties of these toxins have attracted considerable attention by virtue of their use as selective molecular probes for analyses of beta-cell stimulus-secretion coupling. Targets of the toxins include heptahelical cell surface receptors, GTP-binding proteins, ion channels, Ca(2+) stores, and the exocytotic secretory apparatus. Here we review the value of insulinotropic toxins from the perspective of their established use in the study of signal transduction pathways activated by the blood glucose-lowering hormone glucagon-like peptide-1 (GLP-1). Our analysis of one insulinotropic toxin (alpha-latrotoxin) leads us to conclude that there exists a process of molecular mimicry whereby the 'lock and key'analogy inherent to hormone-receptor interactions is reproduced by a toxin related in structure to GLP-1.  (+info)

Maitotoxin-induced membrane blebbing and cell death in bovine aortic endothelial cells. (11/156)

BACKGROUND: Maitotoxin, a potent cytolytic agent, causes an increase in cytosolic free Ca2+ concentration ([Ca2+]i) via activation of Ca2+-permeable, non-selective cation channels (CaNSC). Channel activation is followed by formation of large endogenous pores that allow ethidium and propidium-based vital dyes to enter the cell. Although activation of these cytolytic/oncotic pores, or COP, precedes release of lactate dehydrogenase, an indication of oncotic cell death, the relationship between CaNSC, COP, membrane lysis, and the associated changes in cell morphology has not been clearly defined. In the present study, the effect maitotoxin on [Ca2+]i, vital dye uptake, lactate dehydrogenase release, and membrane blebbing was examined in bovine aortic endothelial cells. RESULTS: Maitotoxin produced a concentration-dependent increase in [Ca2+]i followed by a biphasic uptake of ethidium. Comparison of ethidium (Mw 314 Da), YO-PRO-1 (Mw 375 Da), and POPO-3 (Mw 715 Da) showed that the rate of dye uptake during the first phase was inversely proportional to molecular weight, whereas the second phase appeared to be all-or-nothing. The second phase of dye uptake correlated in time with the release of lactate dehydrogenase. Uptake of vital dyes at the single cell level, determined by time-lapse videomicroscopy, was also biphasic. The first phase was associated with formation of small membrane blebs, whereas the second phase was associated with dramatic bleb dilation. CONCLUSIONS: These results suggest that maitotoxin-induced Ca2+ influx in bovine aortic endothelial cells is followed by activation of COP. COP formation is associated with controlled membrane blebbing which ultimately gives rise to uncontrolled bleb dilation, lactate dehydrogenase release, and oncotic cell death.  (+info)

The mechanism of maitotoxin-induced elevation of the cytosolic free calcium level in rat cerebrocortical synaptosomes. (12/156)

The present study was conducted to elucidate the mechanism of the maitotoxin (MTX)-induced increase in intrasynaptosomal free calcium level ([Ca2+]i). The MTX (1 ng/ml)-induced increase in [Ca2+]i was partially inhibited by the omission of extracellular Ca2+ (Ca2+e) or the addition of verapamil, but not by adding nifedipine, omega-agatoxin IVA, omega-conotoxin GVIA and omega-conotoxin MVIIC. An increase in [Ca2+]i in the absence of Ca2+e was sensitive to procaine, TMB-8, genistein and verapamil, but not to ryanodine and U-73122. These results may suggest that MTX increases [Ca2+]i by stimulating Ca2+ entry through voltage-independent nonselective cation channels and Ca2+ release from stores through a phospholipase C-gamma1-mediated pathway in rat cerebrocortical synaptosomes.  (+info)

Heterogeneous increases of cytoplasmic calcium: distinct effects on down-regulation of cell surface sodium channels and sodium channel subunit mRNA levels. (13/156)

1. Long-term (> or = 12 h) treatment of cultured bovine adrenal chromaffin cells with A23187 (a Ca(2+) ionophore) or thapsigargin (TG) [an inhibitor of sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA)] caused a time- and concentration-dependent reduction of cell surface [(3)H]-saxitoxin (STX) binding capacity, but did not change the K:(D:) value. In A23187- or TG-treated cells, veratridine-induced (22)Na(+) influx was reduced (with no change in veratridine EC(50) value) while it was enhanced by alpha-scorpion venom, beta-scorpion venom, or Ptychodiscus brevis toxin-3, like in nontreated cells. 2. The A23187- or TG-induced decrease of [(3)H]-STX binding was diminished by BAPTA-AM. EGTA also inhibited the decreasing effect of A23187. A23187 caused a rapid, monophasic and persistent increase in intracellular concentration of Ca(2+) ([Ca(2+)](i)) to a greater extent than that observed with TG. 2,5-Di-(t-butyl)-1,4-benzohydroquinone (DBHQ) (an inhibitor of SERCA) produced only a rapid monophasic increase in [Ca(2+)](i), without any effect on [(3)H]-STX binding. 3. Reduction in [(3)H]-STX binding capacity induced by A23187 or TG was attenuated by Go6976 (an inhibitor of conventional protein kinase C) or calpastatin peptide (an inhibitor of calpain). When the internalization rate of cell surface Na(+) channels was measured in the presence of brefeldin A (an inhibitor of vesicular exit from the trans-Golgi network), A23187 or TG accelerated the reduction of [(3)H]-STX binding capacity. 4. Six hours treatment with A23187 lowered Na(+) channel alpha- and beta(1)-subunit mRNA levels, whereas TG had no effect. 5. These results suggest that elevation of [Ca(2+)](i) caused by A23187, TG or DBHQ exerted differential effects on down-regulation of cell surface functional Na(+) channels and Na(+) channel subunit mRNA levels.  (+info)

Up-regulation of cell surface sodium channels by cyclosporin A, FK506, and rapamycin in adrenal chromaffin cells. (14/156)

Treatment of cultured bovine adrenal chromaffin cells with cyclosporin A (CsA) increased cell surface [(3)H]saxitoxin ([(3)H]STX) binding by 56% in a time (t(1/2) = 15.2 h)- and concentration (EC(50) = 2.9 microM)-dependent manner but did not change the K(d) value. In CsA-treated cells, veratridine-induced (22)Na(+) influx was augmented with no change in the EC(50) of veratridine; also, alpha- and beta-scorpion venom and Ptychodiscus brevis toxin-3 enhanced veratridine-induced (22)Na(+) influx in a more than additive manner, as in nontreated cells. CsA treatment for 1 to 24 h inhibited calcineurin activity, measured by the in vitro assay, with the IC(50) of 0.6 microM but did not alter cellular level of calcineurin. FK506 or rapamycin elevated [(3)H]STX binding by 36 or 25%, whereas GPI-1046, an immunophilin ligand incapable to inhibit calcineurin, or okadaic acid, an inhibitor of protein phosphatases 1 and 2A, had no increasing effect. The rise of [(3)H]STX binding by CsA was attenuated by the coincident treatment with brefeldin A (BFA), an inhibitor of vesicular exit from the trans-Golgi network. The internalization rate of cell surface Na(+) channels, as determined in the presence of BFA, was decreased in CsA (but not rapamycin)-treated cells (t(1/2) = 20.3 h), compared with nontreated cells (t(1/2) = 13.7 h). CsA treatment, however, did not elevate cellular levels of Na(+) channel alpha-subunit and Na(+) channel alpha- and beta(1)-subunit mRNAs. In CsA-treated cells, veratridine-induced (45)Ca(2+) influx via voltage-dependent Ca(2+) channels and catecholamine secretion were enhanced, whereas high K(+)-induced (45)Ca(+) influx was not. Thus, the inhibition of calcineurin or rapamycin-binding protein causes up-regulation of cell surface functional Na(+) channels via modulating externalization and internalization of Na(+) channels, thus enhancing Ca(2+) channel gating and catecholamine secretion.  (+info)

The red tide toxin, brevetoxin, induces embryo toxicity and developmental abnormalities. (15/156)

Brevetoxins are lipophilic polyether toxins produced by the red tide dinoflagellate Gymnodinium breve, and their neurotoxic effects on adult animals have been documented. In this study, we characterized adverse developmental effects of brevetoxin-1 (PbTx-1) using an exposure paradigm that parallels the maternal oocyte transfer of toxin. Medaka fish (Oryzias latipes) embryos were exposed to PbTx-1 via microinjection of toxin reconstituted in a triolein oil droplet. Embryos microinjected with doses of 0.1-8.0 ng/egg (ppm) of brevetoxin-1 exhibited pronounced muscular activity (hyperkinesis) after embryonic day 4. Upon hatching, morphologic abnormalities were commonly found in embryos at the following lowest adverse effect levels: 1.0-3.0 ppm, lateral curvature of the spinal column; 3.1-3.4 ppm, herniation of brain meninges through defects in the skull; and 3.4-4.0 ppm, malpositioned eye. Hatching abnormalities were also commonly observed at brevetoxin doses of 2.0 ppm and higher with head-first, as opposed to the normal tail-first, hatching, and doses > 4.1 ng/egg produced embryos that developed but failed to hatch. Given the similarity of developmental processes found between higher and lower vertebrates, teratogenic effects of brevetoxins have the potential to occur among different phylogenetic classes. The observation of developmental abnormalities after PbTx-1 exposure identifies a new spectrum of adverse effects that may be expected to occur following exposure to G. breve red tide events.  (+info)

Biomonitoring brevetoxin exposure in mammals using blood collection cards. (16/156)

A method has been tested in laboratory mice to monitor for the presence of brevetoxins in blood after exposure. The use of blood collection cards is an adaptation of a method employed for routine diagnostic and genetic testing of newborns. Blood is collected and applied to a 0.5-inch diameter circle on a specially prepared blood collection card and allowed to dry. The blood spots are then extracted and the presence of toxin activity is first screened using a high throughput receptor binding assay. Positive samples are then examined for specific brevetoxin congeners by liquid chromatography-tandem mass spectrometry. Preliminary experiments tested the efficiency and linearity of toxin extraction from blood spiked with brevetoxin-3 (PbTx-3). Blood from treated mice was tested for the presence of brevetoxin at different times following exposure to a sublethal dose (180 microg/kg PbTx-3). Brevetoxin activity determined by receptor assay increased to 25 +/- 7.4 nM PbTx-3 equivalents within 4 hr after exposure and was still detectable in three of four animals 24 hr after exposure. Tandem mass spectrometry provided confirmation of PbTx-3, which also increased for the time points between 0.5 and 4.0 hr exposure. However, PbTx-3 was not detected at 24 hr, which suggested the formation of a biologically active metabolite. We anticipate that this approach will provide a method to biomonitor brevetoxins in living marine resources (e.g., finfish), protected species, and humans.  (+info)