Decreases in plasma membrane Ca²⁺-ATPase in brain synaptic membrane rafts from aged rats.
Precise regulation of free intracellular Ca(2+)  concentrations [Ca(2+) ](i)  is critical for normal neuronal function, and alterations in Ca(2+)  homeostasis are associated with brain aging and neurodegenerative diseases. One of the most important proteins controlling [Ca(2+) ](i)  is the plasma membrane Ca(2+) -ATPase (PMCA), the high-affinity transporter that fine tunes the cytosolic nanomolar levels of Ca(2+) . We previously found that PMCA protein in synaptic plasma membranes (SPMs) is decreased with advancing age and the decrease in enzyme activity is much greater than that in protein levels. In this study, we isolated raft and non-raft fractions from rat brain SPMs and used quantitative mass spectrometry to show that the specialized lipid microdomains in SPMs, the rafts, contain 60% of total PMCA, comprised all four isoforms. The raft PMCA pool had the highest specific activity and this decreased progressively with age. The reduction in PMCA protein could not account for the dramatic activity loss. Addition of excess calmodulin to the assay did not restore PMCA activity to that in young brains. Analysis of the major raft lipids revealed a slight age-related increase in cholesterol levels and such increases might enhance membrane lipid order and prevent further loss of PMCA activity.
Department of Pharmacology and Toxicology and Higuchi Biosciences Center, The University of Kansas, Lawrence, KS, USA.
Lei
L
Jiang
Misty D
MD
Bechtel
Nadezhda A
NA
Galeva
Todd D
TD
Williams
Elias K
EK
Michaelis
Mary L
ML
Michaelis
D000375:Aging / Q000378:metabolism*; D000818:Animals; D001921:Brain / Q000201:enzymology / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000032:analysis / Q000378:metabolism*; D002853:Chromatography, Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D015151:Immunoblotting; D008297:Male; D013058:Mass Spectrometry; D021962:Membrane Microdomains / Q000737:chemistry / Q000201:enzymology* / Q000378:metabolism; D051381:Rats; D011916:Rats, Inbred F344; D013570:Synaptic Membranes / Q000737:chemistry / Q000201:enzymology* / Q000378:metabolism
D000252:Calcium-Transporting ATPases