Store depletion and store-operated Ca2+ current in human prostate cancer LNCaP cells: involvement in apoptosis.
1. In the present study, we investigated the mechanisms involved in the induction of apoptosis by the Ca2+-ATPase inhibitor thapsigargin (TG), in androgen-sensitive human prostate cancer LNCaP cells. 2. Exposure of fura-2-loaded LNCaP cells to TG in the presence of extracellular calcium produced an increase in intracellular Ca2+, the first phase of which was associated with depletion of intracellular stores and the second one with consecutive extracellular Ca2+ entry through plasma membrane, store-operated Ca2+ channels (SOCs). 3. For the first time we have identified and characterized the SOC-mediated membrane current (Istore) in prostate cells using whole-cell, cell-attached, and perforated patch-clamp techniques, combined with fura-2 microspectrofluorimetric and Ca2+-imaging measurements. 4. Istore in LNCaP cells lacked voltage-dependent gating and displayed an inwardly rectifying current-voltage relationship. The unitary conductance of SOCs with 80 mM Ca2+ as a charge carrier was estimated at 3.2 +/- 0.4 pS. The channel has a high selectivity for Ca2+ over monovalent cations and is inhibited by Ni2+ (0.5-3 mM) and La3+ (1 microM). 5. Treatment of LNCaP cells with TG (0.1 microM) induced apoptosis as judged from morphological changes. Decreasing extracellular free Ca2+ to 200 nM or adding 0.5 mM Ni2+ enhanced TG-induced apoptosis. 6. The ability of TG to induce apoptosis was not reduced by loading the cells with intracellular Ca2+ chelator (BAPTA-AM). 7. These results indicate that in androgen-sensitive prostate cancer cells the depletion of intracellular Ca2+ stores may trigger apoptosis but that there is no requirement for the activation of store-activated Ca2+ current and sustained Ca2+ entry in induction and development of programmed cell death.
Skryma
R
R
Laboratoire de Physiologie Cellulaire, INSERM EPI-9938, Villeneuve d'Ascq, France.
Mariot
P
P
Bourhis
X L
XL
Coppenolle
F V
FV
Shuba
Y
Y
Vanden Abeele
F
F
Legrand
G
G
Humez
S
S
Boilly
B
B
Prevarskaya
N
N
D000728:Androgens / Q000494:pharmacology; D017209:Apoptosis; D002118:Calcium / Q000378:metabolism / Q000502:physiology*; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D004553:Electric Conductivity; D006801:Humans; D007811:Lanthanum / Q000494:pharmacology; D008297:Male; D008856:Microscopy, Fluorescence; D009532:Nickel / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D011471:Prostatic Neoplasms / Q000378:metabolism / Q000503:physiopathology*; D019284:Thapsigargin / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D000728:Androgens; D015220:Calcium Channels; D019284:Thapsigargin; D007811:Lanthanum; D009532:Nickel; D000252:Calcium-Transporting ATPases; D002118:Calcium