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GEN News Highlights : Mar 7, 2012

Antibodies against Dkk1 Prevent Synaptic Loss Associated with Early Alzheimer Disease

Elevated levels of Dkk1 trigger synapse disassembly prior to evidence of cell death.

Investigators suggest that antibodies targeting the Wnt antagonist Dickkopf-1 (Dkk1) could represent a new approach to slowing synaptic loss in Alzheimer disease (AD). A team at University College London’s department of cell and developmental biology built on previous work indicating that Dkk1 is increased in brains of AD patients and showed that acute exposure to amyloid- β (Aβ) induces Dkk1 expression and loss of synaptic sites.

Their studies in mice further demonstrated that administration of Dkk1-neutralizing antibodies suppress this synaptic loss in mouse brain slices. Silvia A. Purro, Ph.D., Ellen M. Dickins, Ph.D., and Patricia C. Salinas, Ph.D., report their work in the Journal of Neuroscience in a paper titled “The secreted Wnt antagonist Dickkopf-1 is required for amyloid β-mediated synaptic loss.”

AD is associated with synaptic dysfunction, early synaptic loss, and accumulation of Aβ, but soluble forms of Aβ decrease the number and strength of synapses before there is evidence of neuronal cell death, the researchers explain. Numerous lines of evidence have also implicated Wnt signaling deficits in AD, and prior research identified elevated levels of the secreted Wnt antagonist Dkk1 in the post-mortem brains of AD patients and in those from transgenic mouse models of AD.

The team first evaluated whether Aβ directly regulates Dkk1 expression in acute brain slices, using Aβ-derived diffusible ligands. Short-term administration of the ligands led to rapid loss of synapses and increased the level of Dkk1 mRNA. Administering a neutralizing antibody against Dkk1 completely removed the effect of Aβ on synapse loss. Interestingly, blocking Dkk1 had no effect on the number of synapses under basal conditions, which the team says supports the notion that Dkk1 is normally expressed at very low levels in the hippocampus.

They then used hippocampal dissociated neurons to look more closely at the effect of Dkk1 on synapses. Dkk1 was found to decrease the number of presynaptic and postsynaptic markers by up to 50% as well as the number of release sites but did not affect neuronal viability. Notably, this effect was reversible, as removal of Dkk1 resulted in the recovery of synaptic puncta numbers to control levels.

Interestingly, Western blot analyses suggested that Dkk1 doesn’t affect levels of synaptic proteins, which suggested that the Wnt inhibitor acts to decrease the number of synaptic sites by triggering disassembly of synapses. Indeed they found that Dkk1 decreased by about 30% the number of co-localized vGlut1 (presynaptic marker) and PSD-95 (postsynaptic marker) puncta, “indicating a decrease in the apposition of presynaptic and postsynaptic components,” the researchers state. Similarly, in mature hippocampal neurons, elevated Dkk1 reduced by 20% co-localization of the synaptic vesicle protein VAMP2 protein with the active zone protein Bassoon.

The evidence pointing to Dkk1’s role in synaptic disassembly prompted the team to measure the size of synaptic vesicle clusters. As expected, Dkk1 induced a 17% increase in the number of small puncta, and a parallel 40% decrease in the number of large clusters, further supporting the notion that “blockade of Wnt signaling results in the shrinkage of synapses,” they write.

Time-lapse recordings of hippocampal dissociated neurons added more depth to the findings and showed that in the presence of Dkk1, 70% of stable puncta are lost within 60 minutes, with the number of clusters decreasing just after 25 minutes. Moreover, Dkk1 induced a 78% reduction in the average area of stable clusters and reduced the size of both small and large puncta. 

Effectively, Dkk1 triggers synapse loss by inducing the dispersal of synaptic components, the team concludes. “Given the rapid increase of Dkk1 expression by Aβ and that cell death is a late event in AD resulting from synaptic dysfunction, our studies suggest that Dkk1 modulates synaptic loss without cell death in the early stages of AD ... As Dkk1 functions by blocking Wnt signaling through binding to the LRP6 receptor, these studies demonstrate a role for canonical Wnt signalling in the maintenance of central synapses.”