Na +/Ca 2+ exchange activity is increased in Alzheimer's disease brain tissues

Abstract

These studies were performed to determine the changes that occur in Na +/Ca 2+ exchange activity in Alzheimer's disease (AD) brain tissues. Cerebral plasma membrane vesicles were purified by sucrose density gradient centrifugation from frozen postmortem hippocampla/temporal cortex tissue slices derived from age matched brains of normal, AD and non-Alzheimer dementia (NAD) origin (autopsy confirmed). Membrane marker assays (Na/K ATPase, muscarinic receptor, cytochromec oxidase) revealed no change in membrane purity across different preparations. Thin-section electron microscopy revealed predominantly intact unilamellar vesicles. Vesicles were preincubated for 15 min (37°C) in buffer containing 132 mM NaCl, 5 mM KCl, 1.3 mM MgCl 2, 10 mM glucose and 10 mM HEPES (pH 7.4). Ca 2+ uptake was initiated by diluting vesicles 20-fold with buffer containing either 132 mM NaCl or 132 mM choline chloride and 45CaCl 2 then terminated by addition of 200 μM LaCl 3 and rapid filtration. Ca 2+ content increased rapidly at first and then maintained a steady plateau for up to 5 min. When the Ca 2+ ionophore A23187 (10 μM) with 100 μM EGTA was added after 4 min, Ca 2+ content was reduced to 10% of its original value. Ruthenium red (10 μM) had no effect on Ca 2+ content. Na +-dependent Ca 2+ uptake (Ca 2+ content measured in choline chloride minus that measured in NaCl) was increased in AD brains as evidenced by both an increased in the initial rise in Ca 2+ content and in elevated values of peak plateau Ca 2+ content. TheK mandV max for Na +/Ca + exchange was estimated from Lineweaver-Burk analysis of the effect of increasing extravesicular concentrations of Ca 2+ on Na +-dependent Ca 2+ uptake after 30 s. The values obtained for theK m (μM) andV max (nmol/mg/min) were (respectively): normal(n = 6) 57.9 ± 28.1and4.45 ± 0.58;AD(n = 6) 71.2 ± 29.3and6.68 ± 1.58;NAD(n = 4) 66.1 ± 3.3and3.76 ± 1.50 (mean±S.D.). TheV max for Na +/Ca 2+ exchange in AD brain tissues was significantly (P < 0.05) elevated when compared with normal or NAD brains. There was no correlation evident in either normal, AD, NAD or the total sample population between Na +/Ca 2+ exchangeV max orK m and autolysis time or age at death. The results suggest that in brain regions suffering the greatest degeneration due to AD the surviving neurons have increased Na +/Ca 2+ exchange activity. This increase in Na +/Ca 2+ exchange activity may provide clues to a better understanding of the pathogenesis of nerve cell degeneration in AD.