期刊
JOURNAL OF NEUROCHEMISTRY
卷 102, 期 2, 页码 454-465出版社
WILEY
DOI: 10.1111/j.1471-4159.2007.04592.x
关键词
brain aging Ca2+ dysregulation; plasma membrane Ca2+-ATPase; primary neurons; RNA-silencing
资金
- NCRR NIH HHS [P20RR017708] Funding Source: Medline
- NIA NIH HHS [P01AG12993] Funding Source: Medline
Intraneuronal calcium ([Ca2+](i)) regulation is altered in aging brain, possibly because of the changes in critical Ca2+ transporters. We previously reported that the levels of the plasma membrane Ca2+-ATPase (PMCA) and the V-max for enzyme activity are significantly reduced in synaptic membranes in aging rat brain. The goal of these studies was to use RNA(i) techniques to suppress expression of a major neuronal isoform, PMCA2, in neurons in culture to determine the potential functional consequences of a decrease in PMCA activity. Embryonic rat brain neurons and SH-SY5Y neuroblastoma cells were transfected with in vitro- transcribed short interfering RNA or a short hairpin RNA expressing vector, respectively, leading to 80% suppression of PMCA2 expression within 48 h. Fluorescence ratio imaging of free [Ca2+](i) revealed that primary neurons with reduced PMCA2 expression had higher basal [Ca2+](i), slower recovery from KCl-induced Ca2+ transients, and incomplete return to pre-stimulation Ca2+ levels. Primary neurons and SH-SY5Y cells with PMCA2 suppression both exhibited significantly greater vulnerability to the toxicity of various stresses. Our results indicate that a loss of PMCA such as occurs in aging brain likely leads to subtle disruptions in normal Ca2+ signaling and enhanced susceptibility to stresses that can alter the regulation of Ca2+ homeostasis.
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