In recent years, scientists have characterized a hidden set of lymphatic vessels in the meninges, the membranes that line the skull and envelope the brain (Oct 2017 news). These meningeal lymphatic vessels carry waste from the cerebrospinal and interstitial fluids to the deep cervical lymph nodes for disposal. Might these vessels weaken with age and play a role in disease? Yes, suggest scientists led by Jonathan Kipnis, University of Virginia, Charlottesville, in the July 25 Nature. They manipulated drainage through these vessels in young, aging, and transgenic mouse models and found cerebrospinal fluid flow slowed, animals had poorer memories, and they accumulated more Aβ. The results suggest the lymph is an important route of waste clearance from the brain, and contributes to both aging and disease if damaged.

  • Lymphatic vessels in the meninges clear waste from the brain.
  • These vessels drain less effectively in aged mice.
  • Interfering with their function exacerbates Aβ deposition.

“Collectively, these experiments suggest that drainage of brain ISF [interstitial fluid] and CSF [cerebrospinal fluid] by the meningeal lymphatics is necessary for proper cognitive function,” wrote Berislav Zlokovic and Melanie Sweeney, University of Southern California, Los Angeles, in an accompanying News and Views (see also comment below). “The finding has implications for normal aging and disorders such as Alzheimer’s disease.”

First author Sandro Da Mesquita and colleagues used several techniques to hinder drainage through the meningeal lymphatic vessels. They genetically inhibited lymphatic development in wild-type mice, gave three-month-old WT animals a drug that damaged the lymphatic vessels around the brain, or surgically tied vessels off so they couldn’t drain to the deep cervical lymph nodes. In all cases, the scientists found that perfusion of CSF through the brain slowed, as did drainage of macromolecules from ISF. CSF normally flows through the brain via the glymphatic system that runs alongside blood vessels in the brain, and scientists think the fluid eventually reaches the lymph system (Aug 2012 news). “These lymphatic vessels in the meninges are not physically connected to the brain perivascular routes, yet are able to regulate a process that takes place alongside the brain vasculature,” wrote Da Mesquita to Alzforum. “We show that there is a meningeal lymphatic-glymphatic connection, which seems to play important roles in rodent models of aging and Alzheimer’s disease.”

From Bad to Worse.

Aβ plaques (red) accumulated more readily in transgenic mice lacking functioning lymph vessels around the brain (right) than in those with intact vessels (left). Neurons are labeled cyan. [Courtesy of Da Mesquita et al., 2018. Nature.]

After a few weeks of impaired drainage, young mice had trouble remembering a sound associated with a slight foot shock. They took longer than intact animals to find the submerged platform in the Morris water maze, either when it was in a familiar location or had been moved, and they spent much less time searching the correct location when the platform was removed altogether.

Would aging similarly impair drainage? In 20-month-old animals, vessel diameter and vessel coverage in the meninges paled compared with that in young mice, and the CSF filtered more slowly through the brain tissue and into the lymph nodes. The authors were able to correct some of this by giving vascular endothelial growth factor C (VEGF-C) to the older mice. This increased vessel diameter, restored CSF flow, and improved their ability to find the platform in the water maze.

Would VEGF-C help remove Aβ from the brain? Da Mesquita and colleagues treated six- to seven-month-old J20 mice with the growth factor to no avail. VEGF-C had no effect on Aβ levels in the CSF or the amyloid burden in the hippocampus, and was not able to calm hyperactivity in these animals. In fact, by following a CSF tracer in the J20s and in three- to four-month-old 5xFAD mice, the researchers concluded that the morphology and expanse of the lymphatic vessels in these animals was no different from that in controls.

What about in older animals? To mimic aging of the vasculature, Da Mesquita and colleagues wiped out meningeal vessels in two-month-old 5xFAD mice and in six- to seven-month-old J20s. During the next six to 12 weeks, both strains accumulated more plaques in the hippocampus than did controls with intact lymph vessels (see image above). In addition, lymph ablation caused Aβ to build up in the meninges of the mice. When the authors examined postmortem brain tissue from nine AD patients, they found Aβ deposited in their cortical meninges, as well.

“As the authors speculate, it will be extremely interesting to see whether aged animals of less aggressive AD models show deposition of Aβ in the lymphatic vessels or meninges, as postmortem human AD brains in this study,” wrote Tarja Malm and Heikki Tanila from the A.I. Virtanen Institute for Molecular Sciences, University of Eastern Finland, Kuopio (see full comment below).

Together, the results hint at another way for waste to find its way out of the brain. “In sporadic AD, researchers think that the main problem is getting rid of amyloid,” explained Costantino Iadecola, Weill Cornell Medical College, New York, who was not involved in the study. “Inasmuch as this pathway is used in clearing the amyloid, coupled with the aging effect on the lymphatics that they’ve shown, one can presume there may be a link between dysfunction of this system and the accumulation of Aβ in the brain.”—Gwyneth Dickey Zakaib

Comments

  1. Previous work has shown that the cerebral blood vessel system and the blood-brain barrier play a major role in regulating the composition of brain interstitial fluid and the distribution of molecules within the CNS, which is required for proper neuronal and synaptic functioning. Da Mesquita et al. importantly show that the meningeal lymphatic system takes a share in clearance of molecules from brain ISF and CSF. Their data indicate that lymphatic vessels interact with the blood vessels to regulate brain proteostasis, which is important for maintaining cognitive functions and could play a role in the pathogenesis of Alzheimer’s disease. It remains to be determined whether treatments directed at meningeal lymphatics can also improve the impaired function of blood vessels with age, and whether enhancing clearance at the BBB can improve lymphatic drainage. Whether the meningeal lymphatic system can also influence immune responses to modulate AD pathology remains to be seen.

    This study was not primarily concerned with blood vessels and the BBB, as the authors selectively targeted and damaged lymphatic vessels, leaving blood vessels functional with an intact BBB, as their data show. Therefore in their experiments, when they injected tracers in brain ISF, transport of tracers occurred along blood vessels to reach CSF.

    And vice versa, when they injected high concentrations of tracers into CSF, tracers were diffusing back into the brain along blood vessels. But the changes in blood vessels in terms of expression of different receptors, transporters, and other proteins have not been studied, leaving this an important topic for future studies, as we have emphasized in News and Views.

  2. The extensive and elegant study by Kipnis et al. sheds light on the impact of aging and brain amyloidosis in ISF-CSF circulation. This work extends earlier findings of peripheral lymphatic vessel dysfunction in aging to that of dural lymphatic vessels.

    An exciting finding of the study is the impact of aging on the lymphatic cell phenotype and functions in mice, and that the morphological changes were accompanied by changes in the gene expression level. Moreover, the authors show that viral transduction into cisterna magna, and local meningeal application of VEGF-C into aged mice, increased the diameter of the meningeal lymphatic vessels and partially corrected the transport of the dye into the deep cervical lymph nodes. These are encouraging findings demonstrating that age-related degeneration in brain lymphatic vessels can be potentially pharmacologically targeted.

    The authors also tested the effect of VEGF-C treatment on two AD mouse models, APPswe(Ind)J20 (six to seven months of age) and 5xFAD (three to four months of age). The treatment failed to have any observable effects on meningeal lymphatic vessels, CSF Aβ levels, amyloid deposition in the hippocampus, or cognitive deficits. On the other hand, neither one of the employed AD models showed significant deficits in the lymphatics compared to wild-types. These findings further emphasize the important contribution of aging on Alzheimer phenotype also in mouse models.

    As the authors speculate, it will be extremely interesting to see whether aged animals of less aggressive AD models show deposition of Aβ in the lymphatic vessels or meninges, as postmortem human AD brains in this study.

    In addition, it would be interesting to see the impact of VEGF-C in aged mouse models of amyloidosis. Interestingly, no amyloid deposition was detected in the meninges of the 5xFAD mice, whereas mice with ablated meningeal lymphatics with photoconversion showed amyloid deposits. It remains to be investigated whether photoconversion induces a local inflammatory reaction, which in turn promotes amyloid deposition.

    Finally, it will be extremely interesting to see the interplay between the various CSF-ISF drainage pathways, dural lymphatics, glymphatics, nasal lymphatics, intramural periarterial drainage pathway, as well as the drainage via perineural routes, and whether these are impaired upon aging.

  3. I am impressed with the variety of tools used in this study, which highlight that the dural lymphatics are affected by age and related functionally to drainage of both CSF and ISF. In examining the drainage of interstitial fluid, the authors focused on quantifying the tracers that remain in the brain one hour after injection. Clearance of the tracers was reduced at this timepoint in the absence of dural lymphatic function.

    This opens up an avenue for future questions that investigate the exact anatomical pathways between the capillary/arterial basement membranes, which are the sites of intramural periarterial drainage (IPAD), and dural lymphatics. Multiple mechanisms of clearance are working within one hour, including LRP1, IPAD, perivascular macrophages, microglia, so it will be interesting to define how exactly each mechanism is affected by the lack of dural lymphatic drainage.

  4. This study is important for a couple of reasons. First, somewhat unexpectedly, the authors observed that impairment of lymphatic drainage impaired glymphatic exchange of CSF and interstitial solutes. This suggests that the processes governing exchange between the CSF and interstitial compartments, and between the CSF and lymphatic drainage, are in some way functionally linked. It will be important to figure out what the basis for this functional connection is. It is possible that disruption of lymphatic function promotes reactive gliosis, which could account for the changes in glymphatic function, although no differences in blood-brain barrier permeability were noted. This needs to be evaluated. It could also be that disruption of lymphatic drainage pathways alters cisternal CSF flow pathways or pressure gradients that support CSF-ISF exchange.

    It is also important that the authors observe that impaired lymphatic drainage results in neurocognitive deficits while amyloid deposition within parenchyma is exacerbated in mouse models of amyloidosis. This suggests that meningeal lymphatic function can modulate interstitial amyloid dynamics.

    However, when meningeal lymphatic function was increased in two different rodent models of amyloidosis, no effect on amyloid deposition was observed. This raises the question of whether age- or disease-related impairment of lymphatic function contribute to the development of amyloid plaques. While it is possible that these fast-developing rodent models may not be the appropriate model to evaluate this question, it will be important to evaluate this for its relevance to human Alzheimer’s disease to be fully understood.

  5. It’s exciting that several of the methods that were used in this paper to show the effects of diminishing or enhancing the function of meningeal lymphatic vessels can be translated directly to humans. These include measuring the diameter of the vessels themselves, which our group demonstrated is possible through a variety of MRI techniques, as well as tracking the flow of MRI contrast dye injected into the cerebrospinal fluid. These methods might therefore provide proof-of-concept outcome measures for early phase testing of drugs that modulate the function of dural lymphatics, accelerating the pathway toward discovery of new therapies for dementia. 

  6. We share the authors' enthusiasm regarding the possible role of CNS lymphatic clearance in Alzheimer’s disease. With novel pharmaceutical approaches or lifestyle changes, one might even be able to stem the decline in lymphatic function that occurs with aging and, perhaps, during the disease itself. This may improve the homeostasis of the brain tissue to help prevent aggregation of amyloid plaques, even if lymphatic vessels may not themselves be responsible for the clearance of the Aβ.

    That said, we must object to the authors’ concept of the recently rediscovered meningeal lymphatic vessels draining a significant proportion of cerebrospinal fluid (CSF). In our recently published paper, we clearly demonstrated using fluorescence imaging that the major routes of CSF outflow in mice were through the cribriform plate and along cranial nerves to reach lymphatic vessels outside the skull, not via the meningeal lymphatic vessels (Ma et al., 2017). This finding was consistent with dozens of other reports in many species going back 150 years (Schwalbe et al., 1869; Bradbury and Cserr, 1985; Koh et al, 2005). The authors have appeared to ignore any evidence that argues against the importance of the meningeal lymphatic vessels, to the point of not citing relevant literature in their present report.

    It is evident that more work is needed to clarify the exact roles of the lymphatic system in CSF clearance and in the pathology of Alzheimer’s disease.

    References:

    . Outflow of cerebrospinal fluid is predominantly through lymphatic vessels and is reduced in aged mice. Nat Commun. 2017 Nov 10;8(1):1434. PubMed.

    . Die Arachnoidalraum ein Lymphraum und sein Zusammenhang mit den Perichorioidalraum. [The arachnoidal space as a lymphatic space with connection to the perichoroidal compartment.]. . Zbl. Med. Wiss. 1869; 7, 465–467.

    . Experimental Biology of the Lymphatic Circulation. Elsevier, Amsterdam, 1985

    . Integration of the subarachnoid space and lymphatics: is it time to embrace a new concept of cerebrospinal fluid absorption?. Cerebrospinal Fluid Res. 2005 Sep 20;2:6. PubMed.

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References

News Citations

  1. Lymphatic Vessels Found in Human Brain
  2. Brain Drain—“Glymphatic” Pathway Clears Aβ, Requires Water Channel

Research Models Citations

  1. J20 (PDGF-APPSw,Ind)
  2. 5xFAD

Further Reading

Primary Papers

  1. . Functional aspects of meningeal lymphatics in ageing and Alzheimer's disease. Nature. 2018 Aug;560(7717):185-191. Epub 2018 Jul 25 PubMed.