Calcium channels in PDGF-stimulated A172 cells open after intracellular calcium release and are not voltage-dependent.
Using laser image cytometry and Indo-1 fluorescence, we investigated the intracellular free Ca2+ concentration ([Ca2+]i) of confluent A172 human glioblastoma cells stimulated by the BB homodimer of platelet-derived growth factor (PDGF-BB). The shape of the calcium transients and the delay time between stimulation and the beginning of the transient varied considerably. The percentage of responsive cells, the peak [Ca2+]i and the duration of the response were directly related to PDGF-BB dose, while the delay time was inversely related; the maximal response occurred at a PDGF-BB concentration of 20 ng/ml. Studies with EGTA and inorganic calcium-channel blockers (Ni2+, La3+) showed that the increase of [Ca2+]i resulted from initial release of intracellular stores and subsequent calcium influx across the plasma membrane. Opening of calcium channels in the plasma membrane, monitored directly by studying Mn2+ quenching of Indo-1 fluorescence, was stimulated by PDGF-BB and blocked by La3+; the opening occurred 55 +/- 10 s after the initial increase in [Ca2+]i. Therefore, in these tumor cells, intracellular release always occurs before channel opening in the plasma membrane. Depolarization of cells with high extracellular [K+] did not generally induce calcium transients but did decrease calcium influx. L-type calcium-channel blockers (verapamil, nifedipine, and diltiazem) had little or no effect on the calcium influx induced by PDGF-BB. These results indicate that PDGF-BB induces calcium influx by a mechanism independent of voltage-sensitive calcium channels in A172 human glioblastoma cells.
Szöllösi
J
J
Department of Neurological Surgery, School of Medicine, University of California, San Francisco.
Feuerstein
B G
BG
Vereb
G
G
Pershadsingh
H A
HA
Marton
L J
LJ
D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000378:metabolism*; D004533:Egtazic Acid / Q000494:pharmacology; D004594:Electrophysiology; D005456:Fluorescent Dyes; D006801:Humans; D007091:Image Processing, Computer-Assisted; D007211:Indoles; D007811:Lanthanum / Q000494:pharmacology; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology*; D015398:Signal Transduction; D014407:Tumor Cells, Cultured
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D005456:Fluorescent Dyes; D007211:Indoles; D010982:Platelet-Derived Growth Factor; D004533:Egtazic Acid; D007811:Lanthanum; C048960:indo-1; D002118:Calcium