Pharmacological Rescue of Long-Term Potentiation in Alzheimer Diseased Synapses

Abstract

Long-term potentiation (LTP) is an activity-dependent and persistent increase in synaptic transmission. Currently available techniques to measure LTP are time-intensive and require highly specialized expertise and equipment, and thus are not well suited for screening of multiple candidate treatments, even in animal models. To expand and facilitate the analysis of LTP, here we use a flow cytometry-based method to track chemically induced LTP by detecting surface AMPA receptors in isolated synaptosomes: fluorescence analysis of single-synapse long-term potentiation (FASS-LTP). First, we demonstrate that FASS-LTP is simple, sensitive, and models electrically induced LTP recorded in intact circuitries. Second, we conducted FASS-LTP analysis in two well-characterized Alzheimer's disease (AD) mouse models (3xTg and Tg2576) and, importantly, in cryopreserved human AD brain samples. By profiling hundreds of synaptosomes, our data provide the first direct evidence to support the idea that synapses from AD brain are intrinsically defective in LTP. Third, we used FASS-LTP for drug evaluation in human synaptosomes. Testing a panel of modulators of cAMP and cGMP signaling pathways, FASS-LTP identified vardenafil and Bay-73-6691 (phosphodiesterase-5 and -9 inhibitors, respectively) as potent enhancers of LTP in synaptosomes from AD cases. These results indicate that our approach could provide the basis for protocols to study LTP in both healthy and diseased human brains, a previously unattainable goal.

Cytokines and cytokine networks target neurons to modulate long-term potentiation

“…Indeed, synaptosomes have been widely used to study synaptic mechanisms by biochemical, structural and functional analysis (76–79). To focus on synapse-specific mechanisms, we have recently developed a novel flow-cytometry-based approach to study LTP in isolated synaptosomes, termed Fluorescence Analysis of Single-Synapse Long-Term Potentiation (FASS-LTP) (56, 80). FASS-LTP focuses on the insertion of AMPA receptors into the post-synaptic surface after cLTP.…”

“…Alternatively, surface levels of AMPA receptors after cLTP can be quantified by [ 3 H]-AMPA binding (81). Importantly, cLTP response in synaptosomes mechanistically parallels the facilitation of synaptic transmission following electrically-induced LTP in brain slices ( e.g., dependence on CaMKII and BDNF signaling) (80). Moreover, cLTP increases GluA1-PSD95 physical interaction in size-sorted synaptosomes (80).…”

“…Importantly, cLTP response in synaptosomes mechanistically parallels the facilitation of synaptic transmission following electrically-induced LTP in brain slices ( e.g., dependence on CaMKII and BDNF signaling) (80). Moreover, cLTP increases GluA1-PSD95 physical interaction in size-sorted synaptosomes (80). Importantly, FASS-LTP can test multiple samples in parallel (~40) using a minimal amount of tissue (milligrams) for each assay, thus opening the opportunity of studying multiple cytokines in parallel in brain samples from rats, mice and humans ( postmortem ) (80).…”

“…Moreover, cLTP increases GluA1-PSD95 physical interaction in size-sorted synaptosomes (80). Importantly, FASS-LTP can test multiple samples in parallel (~40) using a minimal amount of tissue (milligrams) for each assay, thus opening the opportunity of studying multiple cytokines in parallel in brain samples from rats, mice and humans ( postmortem ) (80). The use of synaptosomes from cryopreserved postmortem tissue provides a unique opportunity to study LTP modulation in the human brain.…”

“…Finally, flow cytometry identifies potentiated synapses by size and GluA1 labeling ( 4 ) (see details in (56)). A key advantage of this approach is the possibility of testing multiple agents in parallel (~ 40, (80)).…”