Decreases in plasma membrane Ca²⁺‐ATPase in brain synaptic membrane rafts from aged rats

Abstract

Precise regulation of free intracellular Ca2+ concentrations [Ca2+]i is critical for normal neuronal function, and alterations in Ca2+ homeostasis are associated with brain aging and neurodegenerative diseases. One of the most important proteins controlling [Ca2+]i is the plasma membrane Ca2+-ATPase (PMCA), the high affinity transporter that fine tunes the cytosolic nanomolar levels of Ca2+. 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 the present 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 of 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 CaM 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.

Presynaptic Control of Glycine Transporter 2 (GlyT2) by Physical and Functional Association with Plasma Membrane Ca2+-ATPase (PMCA) and Na+-Ca2+ Exchanger (NCX)

“…In this case, triple co-localization spots are highlighted by inverted arrows. Taken together, these biochemical and immunocytochemical results suggest that GlyT2 and PMCA pumps are associated in lipid raft subdomains, where both proteins have been shown to be optimally active (11,(32)(33)(34)(35).…”

“…The distribution of PMCA isoforms in isolated DRMs (or lipid rafts) is a point of controversy because in synaptic plasma membranes (SPMs) from pig cerebellum PMCA4 seems to be the only isoform located in raft domains (31), although in primary cortical neurons (32) or hippocampus membranes 3 all isoforms appear to be raft-associated. Recently, Jiang et al (33) have described by quantitative mass spectrometry that the four isoforms are associated with rafts in synaptic plasma membranes from whole rat brain, representing about 60% of the total PMCA. This DRM-associated PMCA pool has been shown to have higher specific activity (32)(33)(34)(35) according to the initial proposition that PMCAs are more active when included in these membrane subdomains (36).…”

“…Recently, Jiang et al (33) have described by quantitative mass spectrometry that the four isoforms are associated with rafts in synaptic plasma membranes from whole rat brain, representing about 60% of the total PMCA. This DRM-associated PMCA pool has been shown to have higher specific activity (32)(33)(34)(35) according to the initial proposition that PMCAs are more active when included in these membrane subdomains (36). Similarly, we have described that GlyT2 can be found in DRMs where it displays the highest transport activity (11) suggesting that GlyT2 presence/absence in lipid rafts could be a versatile regulatory mechanism for the transporter (9,10).…”

The Plasma Membrane Calcium ATPasesand Their Role as Major New Players in Human Disease

“…PMCA activity and abundance declines substantially with age in synaptic plasma membranes from rats, as does the extent of PMCA activation by aged calmodulin (CaM) due to the oxidation of CaM methionine residues (172,240,418). Likewise, a range of oxidative agents have been shown to induce proteolytic degradation and a reduction of PMCA activity in rat synaptic membranes and cortical neurons (417,419), as well as causing PMCA internalization and loss of expression in primary hippocampal neurons (184).…”

A switch in the mode of the sodium/calcium exchanger underlies an age-related increase in the slow afterhyperpolarization

“…Indeed, changes to plasma membrane and sarco-endoplasmic membrane ATPase activity (Jiang, et al, 2012, Michaelis, et al, 1992, voltage gated Ca 2+ channel currents (Campbell, et al, 1996, Frolkis, et al, 1984, Moyer and Disterhoft, 1994, Ca 2+ binding proteins (Verkhratsky and Toescu, 1998) and mitochondrial structure and function have been noted to change with age, some of which can lead to activation of KCa currents (Foster, 2007).…”