Roles of thapsigargin-sensitive Ca2+ stores in the survival of developing cultured neurons.
The roles of the intracellular calcium pool involved in regulating the Ca2+ profile and the neuronal survival rate during development were studied by using thapsigargin (TG), a specific inhibitor of endoplasmic reticulum (ER) Ca2+-ATPase in cultured cerebellar granule neurons. Measuring the neuronal [Ca2+]i directly in the culture medium, we found a bell-shaped curve for [Ca2+]i versus cultured days in cerebellar granule neurons maintained in medium containing serum and 25 mM K+. The progressive increase in [Ca2+]i of the immature granule neurons (1-4 days in vitro) was abolished by TG, which resulted in massive neuronal apoptosis. When the [K+] was lowered from 25 to 5 mM, neither the progressively increasing [Ca2+]i nor the survival of immature granule neurons was significantly changed over 24-h incubation. Similarly, TG caused a dramatic decrease in the [Ca2+]i and survival rate of these immature neurons when switched to 5 mM K+ medium. Following maturation, the granule neurons became less sensitive to TG for both [Ca2+]i and neuronal survival. However, TG can protect mature granule neurons from the detrimental effect of switching to a 5 mM K+ serum-free medium by decreasing [Ca2+]i to an even lower level than in the respective TG-free group. Based on these findings, we propose that during the immature stage, TG-sensitive ER Ca2+-ATPase plays a pivotal role in the progressive increase of [Ca2+]i, which is essential for the growth and maturation of cultured granule neurons.
Yao
C J
CJ
Institutes of Toxicology, College of Medicine, National Taiwan University, Taipei, ROC.
Lin
C W
CW
Lin-Shiau
S Y
SY
D000818:Animals; D001798:Blood Proteins / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D016922:Cellular Senescence / Q000187:drug effects; D002531:Cerebellum / Q000166:cytology; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004721:Endoplasmic Reticulum / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D009474:Neurons / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology*; D018696:Neuroprotective Agents / Q000494:pharmacology; D011188:Potassium / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D019284:Thapsigargin / Q000494:pharmacology*
D001798:Blood Proteins; D016895:Culture Media, Serum-Free; D004791:Enzyme Inhibitors; D018696:Neuroprotective Agents; D019284:Thapsigargin; D000252:Calcium-Transporting ATPases; D011188:Potassium; D002118:Calcium