Maitotoxin-elevated cytosolic free calcium in GH4C1 rat pituitary cells nimodipine-sensitive and -insensitive mechanisms.
Maitotoxin includes an extracellular Ca2+-dependent membrane depolarization predominantly via activation of L-type voltage-dependent Ca2+ channels (L-VDCC) in GH4C1 rat pituitary cells. In contract to studies employing intracellular dyes, electrophysiological studies have indicated that maitotoxin activates voltage-independent conductances. In the present study, we used fura-2 calcium digital analysis to investigate the actions of very low concentrations of maitotoxin on cytosolic free calcium ([Ca2+]i) in GH4C1 cells in an effort to distinguish different calcium entry mechanisms. Maitotoxin at concentrations as low as 0.01 ng/mL elevated [Ca2+]i 35 +/- 3% and induced membrane depolarization. The concentration dependency for maitotoxin-elevated [Ca2+]i was biphasic with the first phase maximal at 0.05 to 0.5 ng/mL and the minimum EC50 of the second phase about 2.0 ng/mL. Nimodipine (100 nM), a dihydropyridine antagonist of L-VDCC, prevented the [Ca+2]i increase and depolarization induced by up to 0.1 ng/mL maitotoxin, but not at higher concentration (0.5 ng/mL) of maitotoxin. This indicates that lower concentrations (0.1 ng/mL) of maitotoxin require L-VDCC, whereas higher concentrations (>-0.5 ng/mL) of maitotoxin may require additional ionic mechanisms. Maitotoxin (0.5 ng/mL) induced 45Ca2+ uptake and depolarization in Ltk-cells which lack VDCC. Reducing extracellular Cl- from 123 to 5.8 microM increased the magnitude of membrane depolarization by maitotoxin (0.5 ng/mL), which suggests that a Cl- conductance participated in depolarization induced by higher maitotoxin concentrations. Taken together, our results indicate that maitotoxin activates at least two ionic mechanisms. At lower concentrations of maitotoxin, the primary ionic mechanism requires the activation of L-VDCC; however, at higher maitotoxin concentrations, additional ionic mechanisms are involve in the entry of extracellular Ca2+. This latter mechanism may represent the voltage-independent pathway evident under voltage clamp conditions.
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Marine Biotoxins Program of the U.S. Marine Fisheries Services, Charleston, SC 29412, USA
Kurtz
D T
DT
Ramsdell
J S
JS
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000187:drug effects; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D005110:Extracellular Space / Q000378:metabolism; D016257:Fura-2; D007477:Ions; D007700:Kinetics; D008387:Marine Toxins / Q000494:pharmacology*; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009553:Nimodipine / Q000494:pharmacology*; D035964:Oxocins; D010902:Pituitary Gland / Q000187:drug effects* / Q000378:metabolism*; D011188:Potassium / Q000494:pharmacology; D051381:Rats; D012680:Sensitivity and Specificity; D000095702:Polyether Toxins
D002118:Calcium; D015220:Calcium Channels; D016257:Fura-2; D007477:Ions; D008387:Marine Toxins; D009553:Nimodipine; D035964:Oxocins; D011188:Potassium; C020394:maitotoxin; D000095702:Polyether Toxins