Nearly four years ago, Steven Paul and coworkers published a provocative paper that showed that a single administration of an anti-Aβ antibody rapidly improved memory in APP-overexpressing mice, without budging a bit of the plaque in their brains (see ARF related news story). The work supported a dawning realization at that time that Aβ (perhaps soluble, or as small multimers) could have acute and reversible effects on nerve and synapse function.

Paul and colleagues at Eli Lilly and Co., Indianapolis, now report a possible explanation for that surprising result. In a paper published online February 23 in the Journal of Clinical Investigation, the researchers show that the PDAPP mice display an Aβ-dependent decrease in hippocampal acetylcholine (ACh) release, which manifests as a learning deficit. Administration of the m266 anti-Aβ antibody restored cholinergic tone and reversed the learning deficit.

Just as this week’s other news shows that stimulation of the cholinergic system can rein in Aβ production, Paul’s new data suggests that Aβ can be hazardous to ACh levels. The results support the idea that passive immunization of people early in the course of AD might be sufficient to elicit some functional improvement.

To measure cholinergic function in vivo, first author Kelly Bales and colleagues performed in vivo microdialysis measurements of acetylcholine in the hippocampus of free-running PDAPP and control wild-type mice. Under basal conditions, the PDAPP mice showed lower acetylcholine levels than did age-matched controls. When the mice were stimulated, they showed a profound dysregulation in evoked ACh release. Putting the mice into a new cage (environmental stimulation) caused a much higher and prolonged evoked ACh release in the PDAPP mice. In contrast, pharmacological treatment with the muscarinic receptor antagonist scopolamine resulted in much lower ACh release in the transgenic compared to wild-type.

While searching for a reason for the reduction in basal ACh in PDAPP mice, the researchers found an increase in extracellular choline. This led them to examine the effect of Aβ on the choline transporter ChT-1. Coimmunopreciptation experiments indicated that Aβ42 physically associates with ChT-1 in hippocampus. Aβ affected choline transport, but not in the direction expected. In synaptosomes from hippocampus, Aβ treatment caused an increase in choline uptake. The investigators hypothesize that the increase may reflect a compensatory mechanism in response to failed cholinergic signaling.

Whatever the details of the odd brain chemistry of PDAPP mice, all was brought nearly back to normal by administration of the anti-Aβ antibody m266. Whether in microdialysis measures or synaptosome uptake, a shot of m266 antibody was sufficient to restore basal and evoked ACh release, and normalize choline transport.

Normalizing cholinergic signals also affected the mice’s performance in one behavioral test. Mice placed in a new environment run around for a while and eventually settle down, in a process of habituation. Previously, the same workers had shown that PDAPP failed to habituate like wild-type mice, demonstrating a higher locomotor activity throughout the hour-long test (Dodart et al., 1999). Mice treated with the m266 antibody habituated normally, suggesting that returning ACh to normal resulted in restoration of this learning response.

The results uphold the idea that cholinergic defects, thought to underlie memory loss in humans, could result at least in part from a direct “cholinotoxic” effect of Abeta. Previously, Abeta has been shown to bind the alpha7 nicotinic ACh receptor and there have been suggestions that this interaction promotes tau phosphorylation (see ARF related news story). Just how Abeta association with alpha7 or the choline transporter might be linked to the observed ACh dysregulation will be a subject for further study.

The reversal of ACh perturbations by the antibody m266 is promising, but it is important to keep in mind that for all their plaques, these mice do not undergo neurodegeneration, a process that sets in quite early in people with AD. Understanding the limits of reversibility in humans versus mice will be critical to developing anti-Abeta treatments—Pat McCaffrey.

Reference:
Bales KR, Tzavara ET, Wu S, Wade MR, Bymaster FP, Paul SM, Nomikos GG. Cholinergic dysfunction in a mouse model of Alzheimer disease is reversed by an anti-Abeta antibody. J Clin Invest. 2006 Feb 23; [Epub ahead of print] Abstract

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  1. In short, Bales et al. provide data suggesting that Aβ association with the choline transporter, ChT-1, leads to augmentation of enzyme activity. In the PDAPP mouse hippocampus, this apparently leads to dysregulation of ACh synthesis and release. Treatment of these animals with a monoclonal antibody that selectively interacts with soluble Aβ completely reverses the ACh phenotype. The fact that a conformationally selective antibody, m266, reverses impaired baseline ACh efflux as well as dysregulation of pharmacologically or behaviorally induced ACh efflux suggests that the binding of Aβ to ChT-1 is conformation-dependent. This also indirectly demonstrates that m266 recognizes a toxic form of Aβ in that administration of the antibody, and not other anti-Aβ antibodies, to PDAPP mice selectively reversed the ACh phenotype.

    The PDAPP mice have a rather complex phenotype regarding evoked versus baseline ACh efflux. In comparison to wild-type or m266-treated PDAPP, PDAPP hippocampus baseline ACh is reduced, extracellular choline is elevated, behaviorally induced ACh efflux is elevated, and blockade of muscarinic acetylcholine receptors with scopolamine results in reduced ACh release. The authors interpret this as possibly due to enrichment of ChT-1 to a synaptic vesicular pool (thus explaining the enhanced choline transport in synaptosomes) with subsequent inhibition of choline transport or inappropriate distribution of the transporter elsewhere. Impaired ACh biosynthesis, reduced ACh levels overall, lower ACh basal release, excess ACh efflux with behavioral activity, and impaired scopolamine-induced ACh release are attributed to the latter. Further studies should elucidate if ChT-1 distribution is altered under conditions of elevated Aβ.

    Based on the work of Cirrito et al., 2005, one might propose an additional mechanism in which synaptic activity stimulates Aβ production, which then interacts with ChT-1 to increase choline uptake and stimulate new ACh synthesis. This, in turn, leads to enhanced ACh release under behaviorally evoked conditions. In vitro experiments similar to the choline uptake studies performed in this work might shed light on the likelihood of this mechanism if ACh levels increase in synaptosomes treated with Aβ1-42.

    While it is presumed that the Aβ species used in the choline uptake studies is soluble, it is hoped that future studies will have a more extensive characterization of which conformation(s) bind to the ChT-1 to affect its function and a more direct demonstration that m266 interacts with and neutralizes the same.

    If the observations described for PDAPP mice in Bales et al. truly represent AD, it certainly has implications for the timing of AChE treatment. If in early stages of the disease, basal ACh synthesis is impaired and on-demand ACh synthesis is excessive, anti-cholinesterase treatment might not be the most efficacious strategy. It would be interesting to know if cholinesterase therapy in PDAPP mice improves or exacerbates memory performance at young ages.

    References:

    . Synaptic activity regulates interstitial fluid amyloid-beta levels in vivo. Neuron. 2005 Dec 22;48(6):913-22. PubMed.

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References

News Citations

  1. One-Shot Deal? Mice Regain Memory Day After Vaccination, Plaques Stay Put
  2. Cholinergic Transmission and Aβ: Boosting M1 Receptors Treats Model
  3. Nicotine and β Amyloid—Smoking Guns?

Paper Citations

  1. . Behavioral deficits in APP(V717F) transgenic mice deficient for the apolipoprotein E gene. Neuroreport. 2000 Feb 28;11(3):603-7. PubMed.
  2. . Cholinergic dysfunction in a mouse model of Alzheimer disease is reversed by an anti-A beta antibody. J Clin Invest. 2006 Mar;116(3):825-32. PubMed.

Further Reading

Papers

  1. . Cholinergic dysfunction in a mouse model of Alzheimer disease is reversed by an anti-A beta antibody. J Clin Invest. 2006 Mar;116(3):825-32. PubMed.
  2. . Targeting amyloid-beta peptide (Abeta) oligomers by passive immunization with a conformation-selective monoclonal antibody improves learning and memory in Abeta precursor protein (APP) transgenic mice. J Biol Chem. 2006 Feb 17;281(7):4292-9. PubMed.
  3. . Antibodies against beta-amyloid reduce Abeta oligomers, glycogen synthase kinase-3beta activation and tau phosphorylation in vivo and in vitro. J Neurosci Res. 2006 Feb 15;83(3):374-84. PubMed.
  4. . Vessel ultrastructure in APP23 transgenic mice after passive anti-Abeta immunotherapy and subsequent intracerebral hemorrhage. Neurobiol Aging. 2007 Feb;28(2):202-12. PubMed.

News

  1. “Natural” Aβ Oligomers Cause Transitory Cognitive Disruptions
  2. Passive Aggressive—Must Antibody Therapy Start Early to Be Effective?

Primary Papers

  1. . Cholinergic dysfunction in a mouse model of Alzheimer disease is reversed by an anti-A beta antibody. J Clin Invest. 2006 Mar;116(3):825-32. PubMed.