In the hunt for the elusive origins of Alzheimer’s disease, researchers may have to dig deeper than ever before. Deeper into the brain, that is. According to a November 4 report in Nature Communications, the basal forebrain, a subcortical structure that houses cholinergic neurons, falls prey to neurodegeneration before the entorhinal cortex—the region commonly considered the disease’s first victim. Researchers led by Nathan Spreng of Cornell University in Ithaca, New York, tapped into longitudinal imaging, biomarker, and cognitive data from the Alzheimer’s Disease Neuroimaging Initiative to track the pathological chain of events leading to AD. They report that the forebrain had already started shriveling in people who had abnormal levels of Aβ42 in their cerebrospinal fluid but were still cognitively normal. In contrast, the entorhinal cortex only began to shrink once a person started having memory problems as well.

The prevailing view in the field is that the entorhinal cortex—a region nestled next to the hippocampus—degenerates first (see Braak et al., 2006). Previous studies have suggested that following diffuse Aβ deposition broadly in the neocortex, incipient tau tangles in the EC somehow transform into neuronal killers there and in the hippocampus, and then ultimately move out into the neocortex as well (see Mar 2016 newsAug 2016 news). 

However, a different line of research has for many years focused on early degeneration in the basal forebrain, specifically in a hub of cholinergic neurons called the nucleus basalis of Meynert (NbM) (see Mesulam et al., 2004; Sassin et al., 2000; Grothe et al., 2010; and Kilimann et al., 2014). 

Axons from these NbM-dwelling cholinergic neurons spread far and wide to form myriad connections with neurons throughout the cortex. These neurons are the main source of acetylcholine (ACh) in the brain. NbM neurons receive input from the nearby EC, and researchers think the bond between these two regions allows people to focus on and remember new or important events, while relegating familiar or irrelevant events to the background (see Egorov et al., 2002). 

A previous study using cross-sectional ADNI data had linked NbM degeneration to Aβ accumulation (as assessed by amyloid PET) in preclinical AD (see Grothe et al., 2014), but the longitudinal data necessary to pin down when this degeneration occurs was lacking. Along with Spreng, first author Taylor Schmitz of the University of Cambridge in England now took advantage of a larger body of ADNI data to time basal forebrain degeneration in the span of AD progression.

The cohort analyzed consisted of 434 older adults who underwent brain imaging and cognitive tests at baseline, and again after one and two years. Of these participants, 150 were healthy, 103 had mild cognitive impairment but never progressed to AD, 84 had MCI and did progress to AD, and 97 had AD throughout the two-year time course.

Using voxel-based morphometry analysis of MRI data to measure the size of brain regions, the researchers first determined that both the NbM and the EC shrank with clinical progression of AD. But which region took the first hit? To find out, the researchers tracked how the size of each affected the other over time. They found that a small NbM at baseline—indicative of prior degeneration—correlated strongly with a shrinking EC one and two years later. The opposite was not true: Smaller baseline EC volumes did not portend a shrinking NbM at later time points. Furthermore, small baseline NbM volumes did not correlate with degeneration in other regions of the brain, indicating that the relationship between NbM and EC degeneration was unique.

To place this neurodegenerative relationship within the context of AD pathology, the researchers incorporated CSF Aβ42 measurements that were available for 216 participants. Based on CSF, they designated each participant as Aβ-positive or -negative. Among cognitively normal people, who were categorized as “controls” at enrollment, the researchers found significantly more NbM degeneration in people who were Aβ-positive than in those who were Aβ-negative, but they had no degeneration in the EC. This implied that in the presymptomatic stage of AD, degeneration already occurs in the NbM but not yet in the EC. EC degeneration became apparent once cognitive symptoms surfaced: People with MCI and low CSF Aβ who had not progressed to AD had modest EC degeneration, while people with progressive MCI or with AD lost as much EC tissue.

To define the relationship between degeneration in the NbM and EC, the researchers did mediation analyses, which uses multiple regression to test how one variable depends on another. A temporal cascade emerged: In the presence of Aβ pathology, NbM degeneration led to EC degeneration, which triggered memory problems.

Marsel Mesulam of Northwestern University in Chicago thought the study may reinvigorate the field’s interest in the cholinergic system, which has faded from view in recent years. As with any study that leans on statistics to infer causality from correlation, Mesulam urged caution in interpreting the results. He said future studies will need to more carefully tie this NbM degeneration to cholinergic neurons, because structural MRI makes it difficult to exclude the involvement of nearby noncholinergic neurons. This task could be accomplished by PET to single out cholinergic neurons, along with tau PET as a proxy for their neurodegeneration, he wrote.

Where does Aβ fit into the picture? After all, the researchers only observed NbM degeneration in people with abnormal CSF Aβ levels. Michel Grothe of the German Center for Neurodegenerative Diseases in Rostock pointed out that NbM cholinergic neurons—dubbed the “drain of the brain”—may take on Aβ from their numerous cortical contacts and that could be toxic for the cells (see Ovsepian and Herms, 2013). 

However, Grothe noted another, albeit controversial, chain of events, namely that cholinergic degeneration sets off Aβ pathology in cortical neurons. Previous reports indicated that cholinergic lesions in rodents triggered Aβ accumulation in connected cortical regions (see Roher et al., 2000). In people with AD, researchers uncovered more amyloid plaques in regions with cholinergic connections, and observed cholinergic dysfunction in preclinical AD (see Arendt et al., 1985; and Potter et al., 2011). In theory, degeneration in the NbM could initiate Aβ pathology in the neocortex, Grothe speculated.

Researchers proposed that the NbM could be responsible for the spread of tau pathology as well. Years ago, Mesulam reported that tau tangles arose in these neurons prior to memory problems, and accumulated there as AD progressed (see Mesulam et al., 2004). Schmitz and others acknowledged the possibility that tau aggregates might spread transynaptically from the NbM to the EC, just as previous studies have indicated they can spread from the EC into the hippocampus and beyond (see Nov 2010 news; Feb 2012 news). Elliott Mufson of the Barrow Neurological Institute in Phoenix added that one possible trigger of tauopathy in cholinergic neurons could be misregulation of nerve growth factor signaling, which reportedly causes tangles to form there.

While “tau-first” theories are enticing, Grothe acknowledged that unlike a cholinergic lesion in a rodent, the process of AD in humans is a complex chain of interwoven events. Once Aβ pathology begins, for example, it could amplify degeneration in the NbM and subsequently, the EC.

Still other groups have proposed that degeneration starts even deeper in the brain—in the locus coeruleus of the brain stem, which has extensive connections with the NbM (see Dec 2010 webinar). Carefully tracking cholinergic degeneration and amyloid accumulation via PET imaging would be a way to unravel this relationship in future studies, Grothe said.—Jessica Shugart 


  1. The cholinergic hypothesis of AD has been a staple of the field for many years. This hypothesis was the result of the classic studies showing a loss of cholinergic neurons in the basal forebrain and a decrease of cortical cholinergic activity in AD (see review in Mufson et al., 2016). These findings lead to investigations of the connectivity of the basal forebrain cholinergic neurons, which were demonstrated to provide the major cholinergic innervation to the entire cortical mantle (Mesulam et al., 1983). By contrast, most cortical regions do not display reciprocal connections with the cholinergic basal forebrain (Ch4), with a few exceptions including the entorhinal cortex (Mesulam et al., 1984), a component of the tri-synaptic medial temporal lobe memory circuit that degenerates early during the onset of AD. Since cholinergic pharmacotherapy has not yet resulted in consistent improvements in cognition, the cholinergic hypothesis has fallen out of favor in the AD community. Recent findings in molecular biology suggest that AD pathology spreads via a trans-synaptic mechanism between anatomically and functionally interconnected neuron populations (Clavaguera et al., 2009; de Calignon et al., 2012). However, how the interactive temporal sequence of disease pathology associates with well-defined afferent and efferent connectional pathways during initial stages of the disease remains unclear. Although this concept has been studied in rodent models of AD, the field is just beginning to apply imaging techniques to evaluate related phenomena during the onset of AD in humans. The subcortical Ch4 to entorhinal cortex (EC) projection system provides a human-based model to test such hypotheses. In their elegant article, Schmitz and colleagues demonstrate in a large cohort of age-matched older adults with a clinical diagnosis ranging from cognitively normal to MCI to AD, that Ch4 basal forebrain volume predicted longitudinal decreases in EC cortex degeneration but not vice versa. Data also provide evidence linking the Ch4 to EC degenerative sequence to memory dysfunction and showing that this process is dependent upon increased CSF amyloid neuropathology late in the disease process. The authors argue that Ch4 degeneration occurs early in non-demented elder people but it is not until EC volume reduction accompanies basal forebrain pathology that cognitive impairment appears.

    Of interest is the observation that abnormal Ch4 degeneration is clinically silent or not being detected by current neuropsychological tests in the early stages of AD. Together the data suggest that loss of basal forebrain afferent innervation to the EC induces pathology in this paralimbic cortical region. The factors that drive this Ch4 to EC putative trans-synaptic degenerative sequence during the progression of human AD remains unknown. Since Ch4 neurons display tau-positive neurofibrillary tangles (NFTs), it is possible that a toxic form of tau is transported to the EC, which activates a disease process resulting in cellular dysfunction as the disease progresses. Such a temporal course would require an extended preclinical period.

    Since basal forebrain cellular degeneration occurs prior to EC dysfunction, it is imperative to determine what triggers basal forebrain degeneration early in the disease. Given that the survival of Ch4 neurons depends upon the neurotrophin nerve growth factor (NGF) and its high (trkA) and low affinity (p75NTR) receptors, and that NGF dysregulation induces NFTs, impaired trophic factor support may initiate NFT formation in Ch4 neurons (Mufson et al., 2016). Once tau aggregation occurs in Ch4 neurons, it can then be transported via anterograde transport to anatomically connected brain regions such as the EC. The transport of aggregated tau allows this putative toxic moiety to be incorporated into EC neurons and then be transported to the hippocampus and onward over years of disease, resulting in more severe cognitive decline. On the other hand, Ch4 and EC neurons may be selectively vulnerable to a degenerative intraneuronal process(es), which is activated at different stages of the disease process and is not trans-synaptic dependent. Since CFS amyloid levels seem to play a role late in AD, amyloid may not be necessary or sufficient for the initiation of cellular dysfunction during the early phases of the disease. Neuropathological evidence suggests that other subcortical pathways (e.g., the brainstem locus coeruleus noradrenergic forebrain projection system) are affected either before or after Ch4 degeneration. Whether subcortical pathological spread is a common feature in AD remains an intriguing question. The work of Schmitz and co-workers adds more fuel to the fire that the trans-synaptic transport of a toxic prion-like protein plays a pivotal role in the spread of AD pathology across a wide range of subcortical anatomically and functionally linked brain regions. Finally, these data will reinvigorate interest in the cholinergic hypothesis of AD and suggest a polypharmaceutical approach for the treatment of the disease, which follows a temporal time course.


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    . Molecular and cellular pathophysiology of preclinical Alzheimer's disease. Behav Brain Res. 2016 Sep 15;311:54-69. Epub 2016 May 13 PubMed.

  2. The exact temporal sequence of events underlying sporadic AD is still hotly contested. This is an important question because it holds the key to understanding the mechanism of pathogenesis and for identifying critical intervention targets. It is generally accepted that amyloid deposition precedes neurofibrillary tangle (NFT) formation, that cortical NFTs initially emerge in the entorhino-hippocampal complex, and that the resultant neurodegeneration of the medial temporal lobe is responsible for the memory loss. It is also accepted that the Ch4 (i.e., cholinergic) neurons of the nucleus basalis become subjected to early NFT formation. The resultant neurodegeneration of Ch4 causes the cortical cholinergic denervation that has become the target of the first effective pharmacologic intervention in AD. However, the causal and temporal relationship between the cholinergic lesion and the other two ingredients of AD pathology remain unclear. Many interactions have been proposed but none have been convincing.

    In their elegant paper based on ADNI data, Schmitz and Spreng use longitudinal structural MRI, amyloid imaging, and cognitive assessment to construct a model of sequential interactions among molecular markers of AD, sites of atrophy, and the onset of memory impairment. They conclude that atrophy of the nucleus basalis precedes and presumably drives the atrophy of the entorhinal cortex. They also conclude that the memory loss does not emerge until neurodegeneration encompasses both the nucleus basalis and entorhinal cortex (EC). To quote the authors, “…our findings strongly suggest that a subcortical-cortical pathologic spread from Ch4 to EC defines the earliest link in the predictive pathological staging of AD.”

    As in all other attempts to map the temporal sequence of events in AD, these results need to be interpreted cautiously. For one, although the region of the nucleus basalis shown in Figure 1a was rigorously defined according to existing probabilistic maps of Ch4, it is likely to include not only cholinergic cell bodies (i.e., the Ch4) but also components of the ansa peduncularis, the ansa lenticularis and several non-cholinergic cell populations. Is the Schmitz and Spreng temporal (and presumably causal) sequence specifically mediated by a cholinergic mechanism? This question can be addressed through the use of cholinergic ligands and tau PET imaging. The former would determine whether the atrophy in the region of the nucleus basalis is also associated with cortical cholinergic denervation whereas the latter would help to determine whether the atrophy is associated with NFT within Ch4 neurons. The second caveat relates to the obvious limitation of using correlation to infer causality, even when the statistical analyses are rigorous and innovative. This is where animal models would be essential. Does a lesion of Ch4 lead to entorhinal neurodegeneration?

    Once considered the prime mover of the pathologic cascade, the cholinergic component of AD has gradually moved away from the limelight. The Schmitz and Spreng paper may reverse this trend and increase the enthusiasm for developing newer and more effective cholinomimetic treatments.

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News Citations

  1. Tau PET Aligns Spread of Pathology with Alzheimer’s Staging
  2. Brain Imaging Suggests Aβ Unleashes the Deadly Side of Tau
  3. Insidious Spread of Aβ: More Support for Synaptic Transmission
  4. Mice Tell Tale of Tau Transmission, Alzheimer’s Progression

Webinar Citations

  1. Focus on the Locus! (Ceruleus, That Is, in Alzheimer’s Disease)

Paper Citations

  1. . Staging of Alzheimer disease-associated neurofibrillary pathology using paraffin sections and immunocytochemistry. Acta Neuropathol. 2006 Oct;112(4):389-404. PubMed.
  2. . Cholinergic nucleus basalis tauopathy emerges early in the aging-MCI-AD continuum. Ann Neurol. 2004 Jun;55(6):815-28. PubMed.
  3. . Evolution of Alzheimer's disease-related cytoskeletal changes in the basal nucleus of Meynert. Acta Neuropathol. 2000 Sep;100(3):259-69. PubMed.
  4. . Reduction of basal forebrain cholinergic system parallels cognitive impairment in patients at high risk of developing Alzheimer's disease. Cereb Cortex. 2010 Jul;20(7):1685-95. PubMed.
  5. . Subregional basal forebrain atrophy in Alzheimer's disease: a multicenter study. J Alzheimers Dis. 2014;40(3):687-700. PubMed.
  6. . Graded persistent activity in entorhinal cortex neurons. Nature. 2002 Nov 14;420(6912):173-8. PubMed.
  7. . Basal forebrain atrophy and cortical amyloid deposition in nondemented elderly subjects. Alzheimers Dement. 2014 Oct;10(5 Suppl):S344-53. Epub 2014 Jan 10 PubMed.
  8. . Drain of the brain: low-affinity p75 neurotrophin receptor affords a molecular sink for clearance of cortical amyloid β by the cholinergic modulator system. Neurobiol Aging. 2013 Nov;34(11):2517-24. PubMed.
  9. . Cortical cholinergic denervation elicits vascular A beta deposition. Ann N Y Acad Sci. 2000 Apr;903:366-73. PubMed.
  10. . Neuronal loss in different parts of the nucleus basalis is related to neuritic plaque formation in cortical target areas in Alzheimer's disease. Neuroscience. 1985 Jan;14(1):1-14. PubMed.
  11. . Pre- and post-synaptic cortical cholinergic deficits are proportional to amyloid plaque presence and density at preclinical stages of Alzheimer's disease. Acta Neuropathol. 2011 Jul;122(1):49-60. PubMed.

Further Reading

Primary Papers

  1. . Basal forebrain degeneration precedes and predicts the cortical spread of Alzheimer's pathology. Nat Commun. 2016 Nov 4;7:13249. PubMed.