Characterization of the maitotoxin-activated cationic current from human skin fibroblasts.
The maitotoxin (MTX)-induced cationic current (I(mtx)) from human skin fibroblasts was characterized using the patch-clamp technique in whole-cell configuration. Under resting conditions (absence of MTX), the main current observed is produced by an outwardly rectifying K(+) channel which is inhibited by 1 mM TEA. The current reversal potential was -86 mV (n = 12). MTX (500 pM) activated a current with a linear current-voltage relationship and a reversal potential of -10 mV (n = 10). Replacing the extracellular Na(+) and K(+) with N-methyl-D-glucamine (NMDG) caused a shift of the reversal potential to a value below -100 mV, indicating that Na(+) and K(+), but not NMDG, carry I(mtx). Further ion selectivity experiments showed that Ca(2+) carries I(mtx) also. The resulting permeability sequence obtained with the Goldman-Hodgkin-Katz equation yielded Na(+) (1) approximately equal to K(+) (1) > Ca(2+) (0.87). The I(mtx) activation time course reflected the changes in intracellular Ca(2+) and Na(+) measured with the fluorescent indicators fura-2 and SBFI, respectively, suggesting that the activation of I(mtx) brings about an increment in intracellular Ca(2+) and Na(+). Reducing the extracellular Ca(2+) concentration below 1.8 mM prevented the activation of I(mtx) and the increment in intracellular Na(+) induced by MTX. Mn(2+) and Mg(2+) could not replace Ca(2+), but Ba(2+) could replace Ca(2+). MTX activation of current in 10 mM Ba(2+) was approximately 50 % of that induced in the presence of 1.8 mM Ca(2+). When 5 mM of the Ca(2+) chelator BAPTA was included in the patch pipette, MTX either failed to activate the current or induced a small current (less than 15 % of the control), indicating that intracellular Ca(2+) is also required for the activation of I(mtx). Intracellular Ba(2+) can replace Ca(2+) as an activator of I(mtx). However, in the presence of 10 mM Ba(2+) the activation by MTX of the current was 50 % less than the activation with nM concentrations of free intracellular Ca(2+).
Martínez-François
Juan Ramón
JR
Departamento de Biología Celular, Instituto de Fisiología Celular, UNAM, México D.F. 04510.
Morales-Tlalpan
Verónica
V
Vaca
Luis
L
D001464:Barium / Q000502:physiology; D002118:Calcium / Q000502:physiology; D002413:Cations, Divalent / Q000378:metabolism*; D002478:Cells, Cultured; D004553:Electric Conductivity; D005110:Extracellular Space / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects* / Q000502:physiology*; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism; D007473:Ion Channels / Q000502:physiology*; D008274:Magnesium / Q000502:physiology; D008387:Marine Toxins / Q000494:pharmacology*; D035964:Oxocins; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000502:physiology; D012867:Skin / Q000166:cytology; D012879:Skin Physiological Phenomena / Q000187:drug effects*; D012964:Sodium / Q000502:physiology; D000095702:Polyether Toxins
D001464:Barium; D002118:Calcium; D002413:Cations, Divalent; D007473:Ion Channels; D008274:Magnesium; D008387:Marine Toxins; D035964:Oxocins; D011188:Potassium; D012964:Sodium; C020394:maitotoxin; D000095702:Polyether Toxins