Sarco-endoplasmic reticulum Ca2+ ATPase (SERCA) inhibitors identify a novel calcium pool in the central nervous system.
Ca2+ uptake into the endoplasmic reticulum (ER) is mediated by Ca2+ ATPase isoforms, which are all selectively inhibited by nanomolar concentrations of thapsigargin. Using ATP/Mg2+-dependent 45Ca2+ transport in rat brain microsomes, tissue sections, and permeabilized cells, as well as Ca2+ imaging in living cells we distinguish two ER Ca2+ pools in the rat CNS. Nanomolar levels of thapsigargin blocked one component of brain microsomal 45Ca2+ transport, which we designate as the thapsigargin-sensitive pool (TG-S). The remaining component was only inhibited by micromolar thapsigargin, and thus designated as thapsigargin resistant (TG-R). Ca2+ ATPase and [32P]phosphoenzyme assays also distinguished activities with differential sensitivities to thapsigargin. The TG-R Ca2+ uptake displayed unique anion permeabilities, was inhibited by vanadate, but was unaffected by sulfhydryl reduction. Ca2+ sequestered into the TG-R pool could not be released by inositol-1,4,5-trisphosphate, caffeine, or cyclic ADP-ribose. The TG-R Ca2+ pool had a unique anatomical distribution in the brain, with selective enrichment in brainstem and spinal cord structures. Cell lines that expressed high levels of the TG-R pool required micromolar concentrations of thapsigargin to effectively raise cytoplasmic Ca2+ levels. TG-R Ca2+ accumulation represents a distinct Ca2+ buffering pool in specific CNS regions with unique pharmacological sensitivities and anatomical distributions.
Watson
William D
WD
Department of Neurology, Uniformed Services University of the Health Sciences, Bethesda, Maryland, USA.
Facchina
Stephen L
SL
Grimaldi
Maurizio
M
Verma
Ajay
A
D000818:Animals; D001253:Astrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D001923:Brain Chemistry; D002118:Calcium / Q000378:metabolism* / Q000493:pharmacokinetics; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors*; D002451:Cell Compartmentation / Q000502:physiology; D002478:Cells, Cultured; D002490:Central Nervous System / Q000166:cytology / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D066298:In Vitro Techniques; D008274:Magnesium / Q000378:metabolism; D008297:Male; D008861:Microsomes / Q000737:chemistry / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D009928:Organ Specificity; D051381:Rats; D017207:Rats, Sprague-Dawley; D017208:Rats, Wistar; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D019284:Thapsigargin / Q000494:pharmacology
D004791:Enzyme Inhibitors; D019284:Thapsigargin; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000252:Calcium-Transporting ATPases; D008274:Magnesium; D002118:Calcium