Cerebrospinal Fluid (CSF) plays a crucial role in the central nervous system, providing mechanical protection, nourishment, and facilitating the clearance of neurotransmitters, metabolites, and protein aggregates such as amyloid-β and tau. These functions are essential for maintaining neurological health and preventing disorders such as Alzheimer’s disease. The CSF is continuously secreted by the choroid plexus, filling the cerebral ventricles and subarachnoid space, and it undergoes a turnover rate of 3 to 5 times per day.
Recent research has heightened the focus on the regulation of CSF production, circulation, and drainage. There is growing evidence that suggests a reduction in CSF secretion or clearance during ageing could contribute to the development of neurodegenerative conditions like Alzheimer’s disease. This correlation underscores the significance of understanding CSF dynamics in the context of neurological health.
One of the known drainage routes for CSF from the subarachnoid space involves the lymphatics in the cribriform plate, adjacent to where the olfactory nerves pass through the ethmoid bone, and in the perineurium of cranial nerves. Dura mater lymphatics also serve as a CSF drainage path. However, understanding the connections between the subarachnoid space and the extracranial lymphatics, which are crucial in CSF clearance, remains challenging.
Pioneering studies have shed light on this complex system. A landmark study discovered Richardson’s blue dye in the lymphatics of the nasal and palatal mucosa following its injection into the subarachnoid space, indicating a pathway for CSF outflow to the nasopharynx. Other research using dyes or Microfil silicone rubber traced the flow from the subarachnoid space to nasal lymphatics and cranial nerve perineurium. MRI studies have further supported these findings, showing evidence of CSF outflow through the cribriform plate to the nasopharynx.
Innovative techniques have been employed to better understand these processes. For instance, light-sheet fluorescence microscopy and MRI with gadolinium-based contrast agents have been used to observe connections between intracranial and extracranial lymphatics. However, challenges persist, such as the limited resolution of MRI preventing the identification of connections to extracranial lymphatics and issues with tracer phagocytosis by macrophages.
To address these challenges, recent research has employed fluorescence microscopy imaging in anaesthetized Prox1-GFP reporter mice. This study revealed a distinct lymphatic plexus in the nasopharynx, functioning as a hub for CSF outflow from the cribriform plate and other intracranial regions to deep cervical lymph nodes.
Notably, this nasopharyngeal lymphatic plexus (NPLP) undergoes regression and transcriptomic changes with ageing, though deep cervical lymphatics, which transport CSF from the plexus to lymph nodes, do not exhibit these age-related changes. The contractile properties of these lymphatics are regulated by α-adrenergic and nitric oxide signalling, and it was found that pharmacological activation can increase CSF transport in aged mice.
These findings are crucial in understanding the complexities of CSF dynamics and their implications for ageing and neurological diseases. The research not only advances our comprehension of the anatomical and functional aspects of CSF circulation but also opens new avenues for therapeutic interventions in age-related neurodegenerative disorders. By elucidating the pathways and mechanisms underlying CSF clearance, these studies offer hope for developing strategies to maintain or restore healthy CSF dynamics, potentially preventing or mitigating the effects of debilitating neurological conditions.
Discussion
Our comprehensive study sheds light on the nasopharyngeal lymphatic plexus (NPLP), pinpointing its pivotal role as a hub for cerebrospinal fluid (CSF) outflow. This lymphatic plexus, characterized by its unique valves, short lymphangions, and absence of smooth-muscle coverage, serves as a primary drainage route for CSF from crucial areas of the subarachnoid space, including the pituitary gland, cavernous sinus, and cribriform plate. This CSF then flows to the deep cervical lymph nodes (dcLNs), with the medial cervical lymphatics identified as the predominant pathway for this transfer.
The contrast between the structure of the lymphatics in the plexus and the medial cervical lymphatics is striking. The latter, equipped with semilunar valves and typical lymphangion smooth-muscle coverage, are adept at pumping CSF towards lymph nodes. Our findings reveal that CSF outflow via the medial route exceeds that of the lateral route from basolateral dural lymphatics by an average of 180%, highlighting the significance of the NPLP in CSF drainage.
A remarkable aspect of our study is the discovery that CSF outflow is actively modulated by adrenergic and nitric oxide (NO) signalling within the lymphatic smooth muscle. This modulation, preserved even in the context of ageing and atrophy of the nasopharyngeal plexus, opens new possibilities for pharmacological intervention to enhance CSF transport, particularly when clearance is impaired by ageing. Our findings demonstrate that CSF drainage to dcLNs can be effectively increased through the topical application of phenylephrine or sodium nitroprusside to deep cervical lymphatics. This proof of concept for pharmacological manipulation provides a foundation for future clinical applications, though it necessitates innovative drug targeting or delivery methods.
However, our study is not without limitations. The use of deep anaesthesia and surgical techniques to expose the nasopharynx and medial cervical lymphatics may influence the physiological dynamics of CSF drainage. Additionally, cerebral blood flow and vascular pulsation, which are known to contribute to CSF circulation, might be affected by these surgical interventions, thereby altering CSF outflow.
Intriguingly, the nasopharyngeal plexus, while showing a propensity for expansion in response to VEGF-C overexpression and natural regression with ageing, could not be selectively ablated in our experiments. This raises questions about the adaptability of other CSF clearance routes and underscores the need for further research into the mechanisms underlying these alterations and strategies for their prevention or reversal.
Our study also delves into the alterations in lymphatic endothelial cells (LECs) due to ageing, with a particular focus on apoptosis and regression in aged mice. The changes in LECs, including a decrease in valves, contribute to the reduction in CSF outflow, highlighting the importance of understanding the ageing process in lymphatic function.
Single-cell RNA sequencing analysis of LECs isolated from the nasopharyngeal mucosa revealed distinct subclusters, with notable differences in aged LECs, such as an enrichment in genes related to inflammation and type I interferon signalling. This finding aligns with the known hallmarks of vascular ageing and suggests a potential link between inflammation, immune surveillance, and lymphatic function.
Furthermore, the role of lymphatics in neurological diseases such as Alzheimer’s disease and their involvement in immune surveillance and central nervous system (CNS) immune cell turnover is increasingly evident. The diverse activities of lymphatics, from CSF clearance to CNS immune surveillance, underscore their multifaceted contribution to both normal and pathological processes.
In conclusion, our study not only underscores the crucial role of the NPLP in CSF outflow but also highlights the potential for pharmacologically enhancing CSF transport through medial cervical lymphatics under pathological conditions. This opens avenues for future research and potential therapeutic strategies targeting CSF dynamics, particularly in the context of ageing and neurological disorders.
reference link : https://www.nature.com/articles/s41586-023-06899-4#Sec12
