Notes: Abstract: The effects of maitotoxin (MTX) on endogenous amino acid release were tested on highly purified striatal neurons differentiated in primary culture. MTX induced a large and concentration-dependent release of γ-aminobutyric acid (GABA). This effect was abolished when experiments were performed in the absence of external Ca2+, and restored when Ca2+ ions were added after removing the MTX-containing Ca2+-free solution. MTX-induced amino acid release was not affected by 1 μM nifedipine and only slightly inhibited by 1 mM Co2+. MTX also induced a massive accumulation of 45Ca2+ in the neurons which, in contrast to the MTX-evoked GABA release, was totally blocked in the presence of 1 mM Co2+. Whereas 500 nM tetrodotoxin was without significant effect, MTX-evoked GABA release was dependent on the presence of external Na+ and sensitive to nipecotic acid, a GABA uptake inhibitor. It is concluded that, on striatal neurons, MTX induced Na+ influx only in the presence of external Ca2+. The increase in cytoplasmic Na+ ions then triggers the release of GABA.

Notes: Abstract: Maitotoxin (MTX) stimulated inositol phosphate (IP) formation in primary cultures of rat cerebellar granule cells. MTX-induced IP production was dependent on extracellular Ca2+ but independent of extracellular Na+. The stimulation of IP formation elicited by MTX was unaffected by pretreatment of cells with phorbol dibutyrate. pertussis toxin, and a variety of Ca2+ entry blockers, such as nimodipine, nisoldipine, Co2+, and Mn2+. The presence of MTX markedly attenuated IP production induced by carbachol and glutamate, with no apparent effect on the responses to norepinephrine (NE), histamine, 5-hydroxytryptamine (5-HT), and endothelin-1. The inhibition of the carbachol- and glutamate-induced responses by MTX was dose dependent with IC50 values of 1.2 and 0.5 ng/ml, respectively. Pretreatment of cells with a lower concentration of MTX (0.3 ng/ml) also attenuated carbachol- and glutamate-induced IP formation, in a time-dependent manner, with a decrease observed after 30 min prestimulation, but failed to affect NE-, histamine-, 5-HT-, endothelin-1, and sarafotoxin S6b-induced responses. Thus, MTX elicited a marked Ca2+-dependent phosphoinositide (PI) turnover in cerebellar granule cells and selectively inhibited carbachol- and glutamate-induced PI hydrolysis. Possible mechanisms underlying these selective modulations are discussed.

Notes: Abstract: The biological activities of maitotoxin are strictly dependent on the extracellular calcium concentration and are always associated with an increase of the free cytosolic calcium level. We tested the effects of voltage-sensitive calcium channel blockers (nicardipine and ω-conotoxin) on maitotoxin-induced intracellular calcium increase, membrane depolarization, and inositol phosphate production in PC 12 cells. Maitotoxin dose dependently increased the cytosolic calcium level, as measured by the fluorescent probe fura 2. This effect disappeared in a calcium-free medium; it was still observed in the absence of extracellular sodium and was enhanced by the dihydropyridine calcium agonist Bay K 8644. Nicardipine inhibited the effect of maitotoxin on intracellular calcium concentration in a dose-dependent manner. The maitotoxin-induced calcium rise was also reduced by pretreating cells with w-conotoxin. Pretreatment of cells with maitotoxin did not modify 125,I-ω-conotoxin and [3H]PN 200-110 binding to PC 12 membranes. Nicardipine and ω-conotoxin inhibition of maitotoxin-evoked calcium increase was reduced by pertussis toxin pretreatment. Maitotoxin caused a substantial membrane depolarization of PC 12 cells as assessed by the fluorescent dye bisoxonol. This effect was reduced by pretreating the cells with either nicardipine or ω-conotoxin and was almost completely abolished by the simultaneous pretreatment with both calcium antagonists. Maitotoxin stimulated inositol phosphate production in a dose-dependent manner. This effect was reduced by pretreating the cells with 1 μM nicardipine and was completely abolished in a calcium-free EGTA-containing medium. The findings on maitotoxin-induced cytosolic calcium rise and membrane depolarization suggest that maitotoxin exerts its action primarily through the activation of voltage-sensitive calcium channels, the increase of inositol phosphate production likely being an effect dependent on calcium influx. The ability of nicardipine and ω-conotoxin to inhibit the effect of maitotoxin on both calcium homeostasis and membrane potential suggests that l-and N-type calcium channel activation is responsible for the influx of calcium following exposure to maitotoxin. and not that a depolarization of unknown nature causes the opening of calcium channels.