Early in the course of Alzheimer disease, blockages in axonal traffic lead to sick axons swollen with the jumbled pile-up of traffic components. The blockages precede overt amyloid pathology in AD mouse models by a year and mark the brains of people who died at early stages of AD. Axonal traffic blockage should be studied as a potential cause for this devastating disease and might open new research avenues toward early diagnosis. These are the provocative claims of a paper by Larry Goldstein’s group at the University of California, San Diego, which will appear tomorrow in Science magazine. In collaboration with Eliezer Masliah and others at UCSD and Peter Davies at Albert Einstein College of Medicine in New York, the Goldstein group extends to mice and humans a line of investigation they originally established in fruit flies (see ARF related news story and ARF news story).
The axonal transport hypothesis is appealing to some scientists because its underlying mechanism engages both major pathologies of Alzheimer disease—neurofibrillary tangles and amyloid plaques. Tau protein, the major component of tangles, is well-established as regulating axonal traffic through its microtubule-binding function (see, for example, ARF related news story and ARF news story). Based on his earlier work, Goldstein had suggested that APP does so, too, possibly by way of serving as a binding partner for the anterograde motor protein kinesin. A slowdown of axonal traffic, so the hypothesis goes, may cause aberrant generation of the Aβ peptide en route to nerve terminals, leading to synaptic damage and, ultimately, plaques. This research has created a buzz but also ruffled feathers in the AD field. Some researchers have said that the findings are hard to reproduce, and many have eagerly awaited follow-up data from mouse and humans to put the hypothesis to rigorous scrutiny.
In the present paper, first author Gorazd Stokin and colleagues looked for axonal defects that represent transport deficits. In doing so, they harked back to early observations about axonal pathology by AD research pioneer Bob Terry (Terry et al., 1964) and others. The UCSD researchers first examined various fiber tracts in two different transgenic APP mouse models. As had earlier investigators before them, they, too, found axonal swellings distended up to 3 micrometers in diameter and filled with axonal transport components. These occurred in axons of the nucleus basalis of Meynert (NBM), an area that provides cholinergic input to the cerebral cortex and atrophies in AD, as well as in cortex and hippocampus of transgenic, but not wild-type mice. At four months of age, long before amyloid deposits have formed, and prior to the established loss of fibers in the NBM of these mouse models, these axonal swellings were as abundant as at 20 months, according to the paper. The sample sizes in all groups were 4 to 5 animals each. The scientists also assessed fiber changes in the NBM of a small human sample of three people each who had died at Braak AD stages 0, I-III, or IV-VI. Similarly, nearly half the cholinergic fibers at stages I-III had axonal swellings, well prior to when amyloid deposition becomes detectable.
What’s in those swellings? Electron microscopy from the young mice indicated that they brim with mitochondria, vacuoles, and vesicles. Dense bodies, dense axoplasm, and debris gave some swellings the appearance of early-stage axonal degeneration. Light microscopy indicated accumulation of kinesin, hinting at a transport deficit.
To explore the role of transport and kinesin further, Stokin and colleagues recapitulated in mice their earlier genetic experiment in flies, in which they bred strains to cut the dosage of kinesin light chain by half. As before in the flies, tightening the kinesin supply in this way almost doubled the formation of axonal swellings in the APP-transgenic mice. In one test of how the amount of kinesin affects APP transport, the scientists cultured hippocampal neurons from kinesin wild-type and kinesin-reduced mice and then transfected the cells with APP linked to yellow fluorescent protein. In the neurons from the kinesin-reduced mice, anterograde transport of APP-containing particles decreased, and retrograde transport increased. Beyond that, the paper does not quantify how axonal transport rates change in the APP-kinesin knockout crosses. Even so, this data indicates that if kinesin is in short supply, APP transport changes and swellings form, the authors state. To date, one small genetic association study has linked kinesin polymorphisms to AD, but it has not yet been independently confirmed (see Alzgene entry).
Furthermore, the paper suggests that slowing down traffic to axon terminals by limiting the kinesin supply leads to a selective increase in the ratio of Aβ42 to Aβ40 peptides, and also to the peptide’s intraneuronal accumulation, in APP-transgenic mice. The topic of intraneuronal Aβ accumulation has garnered growing interest in the research community in recent years, though some scientists question its relevance to the primary disease process (see ARF related conference story). In the present model, kinesin reduction not only boosts Aβ42 generation prior to amyloid deposition, it also accelerates amyloid deposition. The brains of old APP-transgenic mice with reduced kinesin contained more plaques, larger plaques, and more diffuse extracellular deposits than did APP-transgenics with normal kinesin levels. The brains of middle-aged APP-transgenic mice with reduced kinesin had as much amyloid deposition as did old APP-transgenics with both copies of kinesin. This, and further analysis, suggests “that swellings precede, and participate in the formation of, amyloid plaques,” the authors write.
The authors also write that their data might warrant a reconsideration of dystrophic neurites, which are widely assumed to form because of the presence of plaques. By contrast, axonal swellings do not form in response to plaques, the authors contend, suggesting that they may instead be precursors to some dystrophic neurites.
Reports implicating axonal transport to AD are not new (for a recent review on axonal transport deficits in the neurodegeneration literature, see Roy et al. 2005). What is new is that Stokin and colleagues have placed them prior to other aspects of AD pathogenesis in vivo and staked out a claim for a possible cause for AD. The study does not prove a cause-effect relationship. It also does not address how axonal swellings might relate to other factors that are in contention as early-stage insults, including oligomeric forms of Aβ or tau, oxidative stress, or risk factors such as the apoE4 allele. The study also does not go so far as to compare behavioral differences between the APP-transgenics and their kinesin-reduced brethren.
In summary, the paper puts forth this scenario for testing by the research community: Impaired axonal transport could lead to axonal swellings, which promote aberrant Aβ generation locally at the sites of blockage. This local Aβ increase leads to Aβ secretion or lysis of the swellings, which could trigger amyloid deposition in these spots. Additional transport vesicles coming upon a blockage would get diverted back to the cell body and dendrites, where aberrant Aβ generation, accumulation, and deposition might then occur. People with genetic reductions of kinesin would be especially prone to this latter process. The authors connect these processes into a hypothetical autocatalytic spiral, in which blockages and APP processing would keep stimulating each other and lead to synaptic loss. Finally, differences in how fast axonal transport slows down with age might underlie some cases of sporadic AD, the researchers speculate.—Gabrielle Strobel