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Mechanisms underlying the physiological role of amyloid precursor protein glutamatergic synapses
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N-methyl-D-aspartate receptors (NMDARs) are glutamatergic ionotropic receptors essential for synaptic maturation during development and synaptic plasticity in adult stages, whose properties are known to change depending on the life stage.
In particular, the subunit composition of synaptic NMDARs has important implications for NMDAR function. In the hippocampus, NMDARs are mainly composed by GluN2A or GluN2B subunits, with GluN2B-NMDARs being associated with slower kinetics, more calcium charge transfer and higher mobility.
GluN2B-NMDARs are predominant in immature synapses during development, contributing to the synaptic maturation process, which involves a GluN2B to GluN2A shift. Thus, GluN2A-NMDARs are the most abundant subtype in adult stages, when most synapses are in the mature state. Much less is known about the subunit contribution in aged synapses. However, previous reports showed age-related alterations in NMDAR properties, such as slower responses, lower current amplitudes and a negative correlation between GluN2B levels and memory performance.
These alterations in NMDAR properties, from development to aging, might be caused by different regulation mechanisms. The amyloid precursor protein (APP), which is mainly known to be involved in Alzheimer’s Disease, has emerged as a putative regulator of NMDARs. Although the physiological role of APP is not fully understood, it is known to regulate synaptogenesis and synaptic plasticity and might have different effects when acting through the full-length protein or its derived fragments. Additionally, APP has shown to interact and regulate NMDAR surface levels and currents but the functional relevance of this interaction at different life stages, as well as the underlying mechanisms of regulation have not been explored so far.
Thus, we hypothesized that APP regulates NMDARs in an age-dependent manner and defined as the main aims of this work to study APP-NMDAR regulation mechanisms in immature, mature and aged synapses. To address these questions in physiological conditions, we used as our experimental models the hippocampus of wild-type C57Bl/6 mice at different life stages (infant (7-10 days), adults (10-16 weeks) and aged (18 – 20 months), as well as postmortem brain tissue from human subjects with different ages (18-89 years old) and rodent hippocampal primary neuronal cultures.
By combining patch-clamp electrophysiology and molecular approaches, we have unraveled a dual mechanism by which APP controls GluN2B-NMDARs, depending on the life stage. In the present study, we show that APP is highly abundant at the post synapse in infant mice, where it interacts with GluN2B-NMDARs, controlling its mediated currents. Moreover, APP knockdown in primary neuronal cultures caused a reduction in GluN2B-NMDAR synaptic content, suggesting that APP might be important to stabilize the receptors at the synapse. Considering the crucial role of GluN2B-NMDAR in synapse maturation, this mechanism might potentially be important to achieve functional, mature synapses during development.
Although this interaction is maintained in adult/aged synapses, NMDAR-mediated currents showed to be unaltered when interfering with the APP C-terminal during a short period at these ages, contrary to the results obtained in infant mice. Thus, we concluded that the APP-NMDAR regulatory mechanisms are different in adult/aged mice when compared to infants.
We hypothesize that alterations in the APP-NMDAR regulation could be the underlying mechanism for age-related alterations in NMDAR properties. Accordingly, we found that aged mice exhibit an increase in GluN2B-NMDAR relative currents, which does not correlate with alterations in subunit levels. Moreover, we found an increase in APP processing into intracellular fragments upon aging. Importantly, when we inhibited APP processing or interfered with APP intracellular signaling in aged mice, we were able to normalize GluN2B-NMDAR synaptic contribution to adult-like levels. Thus, we propose that signaling pathways mediated by APP intracellular fragments induce an increase in GluN2B-NMDAR relative currents upon aging. Additionally, we show that APP processing into intracellular fragments also tends to increase in aged humans, suggesting that a similar mechanism might occur in mice and humans. Considering the impact of NMDAR on synaptic plasticity, this increase in GluN2B-NMDAR relative currents can potentially contribute to age-related synaptic and memory impairments.
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PPA (proteína precursora de amiloide) recetor NMDA envelhecimento desenvolvimento pós-natal sinapse glutamatérgica APP (amyloid precursor protein) NMDA receptor aging postnatal development glutamatergic synapse