Preferential accumulation of amyloid-beta in presynaptic glutamatergic terminals (VGluT1 and VGluT2) in Alzheimer's disease cortex

Abstract

Amyloid-beta (Aβ) is thought to play a central role in synaptic dysfunction (e.g. neurotransmitter release) and synapse loss. Glutamatergic dysfunction is involved in the pathology of Alzheimer’s disease (AD) and perhaps plays a central role in age-related cognitive impairment. Yet, it is largely unknown whether Aβ accumulates in excitatory boutons. To assess the possibility that glutamatergic terminals are lost in AD patients, control and AD synaptosomes were immunolabeled for the most abundant vesicular glutamate transporters (VGluT1 and VGluT2) and quantified by flow cytometry and immunoblot methods. In post-mortem parietal cortex from aged control subjects, glutamatergic boutons are fairly abundant as approximately 40% were immunoreactive for VGluT1 (37%) and VGluT2 (39%). However, the levels of these specific markers of glutamatergic synapses were not significantly different among control and AD cases. To test the hypothesis that Aβ is associated with excitatory terminals, AD synaptosomes were double-labeled for Aβ and for VGluT1 and VGluT2, and analyzed by flow cytometry and confocal microscopy. Our study demonstrated that Aβ immunoreactivity (IR) was present in glutamatergic terminals of AD patients. Quantification of Aβ and VGluT1 in a large population of glutamatergic nerve terminals was performed by flow cytometry, showing that 42% of VGluT1 synaptosomes were immunoreactive for Aβ compared to 9% of VGluT1 synaptosomes lacking Aβ-IR. Percentage of VGluT2 synaptosomes immunoreactive for Aβ (21%) was significantly higher than VGluT2 synaptosomes lacking Aβ-IR (9%). Moreover, Aβ preferentially affects VGluT1 (42% positive) compared to VGluT2 terminals (21%). These data represent the first evidence of high levels of Aβ in excitatory boutons in AD cortex and support the hypothesis that Aβ may play a role in modulating glutamate transmission in AD terminals.

Predominant loss of glutamatergic terminal markers in a β-amyloid peptide model of Alzheimer's disease

“…Indeed, a reduction of vGluT1 density in cortical regions was also observed in post-mortem studies in AD patients Kirvell et al, 2007 Q2 ; Kashani et al, 2008), as well as in animal models of AD Minkeviciene et al, 2008). This might be related to the tight interplay between APP-derived formation of Ab and the activity of glutamatergic synapses (Cirrito et al, 2008) and to the accumulation of Ab in glutamatergic nerve terminals in hippocampal regions at early stages in AD models (Lacor et al, 2004;Sokolow et al, 2012). Furthermore, several studies in different animal models of AD have converged in the identification of early dysfunctions of synaptic plasticity at cortical and hippocampal glutamatergic synapses (Hsia et al, 1999;Oddo et al, 2003;Trinchese et al, 2004;Jacobsen et al, 2006;Liu et al, 2008;Auffret et al, 2009), which result from a combined alteration of the set-up of AMPA and NMDA receptors in synapses (Almeida et al, 2005;Hsieh et al, 2006;D'Amelio et al, 2011), and from a decreased density and efficiency of astrocytic glutamate transporters Matos et al, 2012) which is also observed in the afflicted brain regions of AD patients (Hardy et al, 1987a;Westphalen et al, 2003).…”

Glutamate system, amyloid β peptides and tau protein: functional interrelationships and relevance to Alzheimer disease pathology

“…56,57 More recent studies using synaptosome-enriched fractions also showed a trend for reduced levels of VGLUT1 and 2 in the parietal cortex of individuals with Alzheimer disease compared with controls, but they did not reach significant levels. 58 Whether this could be due to the region of the brain examined or the relatively small number of samples used in the study remains to be established. Nevertheless, this study showed that amyloid β peptide accumulates more in VGLUT1/2-containing terminals than in non-VGLUT terminals, thus suggesting that these peptides may preferentially affect presynaptic glutamatergic terminals in the cortex of individuals with Alzheimer disease.…”

“…Nevertheless, this study showed that amyloid β peptide accumulates more in VGLUT1/2-containing terminals than in non-VGLUT terminals, thus suggesting that these peptides may preferentially affect presynaptic glutamatergic terminals in the cortex of individuals with Alzheimer disease. 58 The gene expression and protein level of EAAT1 and EAAT2, on the other hand, have also been shown to be reduced in the cortex and hippocampus of individuals with Alzheimer disease, 59,60,71 suggesting that glutamate clearance from the synaptic cleft is attenuated. In one study, it has been demonstrated that reduced levels of EAAT2, produced mainly by astrocytes, inversely correlates with APP751/770 mRNA levels, thus indicating that abnormal functioning and/or processing of APP may be involved in regulating EAAT2 levels/functions.…”

“…Reduced protein levels before cell loss and onset of pathology [56][57][58] GLAST/EAAT1 Reduced protein at early clinical stages 59,60 GLT-1/EAAT2 Reduced protein levels at early clinical stages 59,60 Glutamine synthetase Reduced protein levels…”

Glutamate transporter EAAT2: regulation, function, and potential as a therapeutic target for neurological and psychiatric disease

“…The cognitive deficits are more significantly correlated with reduced glutamatergic pre-synaptic-bouton density than with neurofibrillary tangles or amyloid-b burden [180]. In addition, deficiencies in the glutamatergic system (e.g., reduced glutamate uptake) have been observed in AD and are correlated with cognitive decline [181][182][183][184][185][186]. Reduced glutamate uptake function may result in increased extracellular glutamate levels which, in turn, potentially increase amyloid-b production over time [187][188][189].…”