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Rammelkamp Center for Education and Research and Department of Physiology and Biophysics, Case Western Reserve University School of Medicine, Cleveland, Ohio 44109
The effects of
maitotoxin (MTX) on plasmalemma permeability are similar to those
caused by stimulation of P2Z/P2X7
ionotropic receptors, suggesting that
1) MTX directly activates
P2Z/P2X7 receptors or
2) MTX and
P2Z/P2X7 receptor stimulation
activate a common cytolytic pore. To distinguish between these two
possibilities, the effect of MTX was examined in
1) THP-1 monocytic cells before and
after treatment with lipopolysaccharide and interferon-
, a maneuver
known to upregulate P2Z/P2X7
receptor, 2) wild-type HEK cells and
HEK cells stably expressing the
P2Z/P2X7 receptor, and
3) BW5147.3 lymphoma cells, a cell
line that expresses functional P2Z/P2X7 channels that are poorly
linked to pore formation. In control THP-1 monocytes, addition of MTX
produced a biphasic increase in the cytosolic free
Ca2+ concentration
([Ca2+]i);
the initial increase reflects MTX-induced
Ca2+ influx, whereas the second
phase correlates in time with the appearance of large pores and the
uptake of ethidium. MTX produced comparable increases in
[Ca2+]i
and ethidium uptake in THP-1 monocytes overexpressing the
P2Z/P2X7 receptor. In both
wild-type HEK and HEK cells stably expressing the
P2Z/P2X7 receptor, MTX-induced
increases in
[Ca2+]i
and ethidium uptake were virtually identical. The response of BW5147.3
cells to concentrations of MTX that produced large increases in
[Ca2+]i
had no effect on ethidium uptake. In both THP-1 and HEK cells, MTX- and
Bz-ATP-induced pores activate with similar kinetics and exhibit similar
size exclusion. Last, MTX-induced pore formation, but not channel
activation, is greatly attenuated by reducing the temperature to
22°C, a characteristic shared by the
P2Z/P2X7-induced pore. Together,
the results demonstrate that, although MTX activates channels that are
distinct from those activated by
P2Z/P2X7 receptor stimulation, the
cytolytic/oncotic pores activated by MTX- and Bz-ATP are indistinguishable.
THP-1 monocytes; HEK cells; heterologous expression; oncosis; vital dyes
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