Perivascular macrophages (PVMs) are increasingly recognized as key players in maintaining brain homeostasis, yet their role in maintaining neurovascular–metabolic homeostasis has not been fully explored. We hypothesized that PVM depletion compromises cerebrospinal fluid-interstitial fluid exchange through glymphatic system (GS) dysfunction, thereby exacerbating cortical hyperexcitability manifested as increased epilepsy susceptibility and seizure intensity. Using clodronate liposomes (CLOs), we achieved >85% PVM depletion in mice. Following pentylenetetrazole (PTZ) challenge, PVM-depleted mice exhibited anxiety-like behaviors (reduced center time, p < 0.05), impaired working memory (decreased spontaneous alternation, p < 0.05), and increased cortical hyperexcitability, including shorter seizure latency and elevated EEG total power (p < 0.05). Mechanistically, PVM loss led to dysregulation of extracellular matrix components (increased laminin and collagen IV), impairing perivascular space integrity and GS function (reduced CSF tracer clearance, p < 0.05). AQP4 inhibition with TGN-020 further exacerbated PTZ-induced EEG abnormalities (increased total power, p < 0.05). Analysis of human epileptic tissue confirmed elevated collagen IV deposition in the seizure focus (p < 0.05) and a trend toward increased PVM density (p = 0.0638). These results highlight PVMs as essential modulators of the glymphatic–metabolic axis, linking vascular health to brain excitability. Targeting the PVM–GS interface offers therapeutic potential for disorders involving vascular dysfunction and neuronal hyperexcitability.
RasmussenMKMestreHNedergaardM.Fluid transport in the brain. Physiol Rev2022; 102: 1025–1151.
2.
Jiang-XieLFDrieuABhasiinK, et al. Neuronal dynamics direct cerebrospinal fluid perfusion and brain clearance. Nature2024; 627(8002): 157–164.
3.
Holstein-RønsboSGanYGiannettoMJ, et al. Glymphatic influx and clearance are accelerated by neurovascular coupling. Nat Neurosci2023; 26: 1042–1053.
4.
WenWChengJTangY.Brain perivascular macrophages: current understanding and future prospects. Brain2024; 147: 39–55.
5.
UekawaKHattoriYAhnSJ, et al. Border-associated macrophages promote cerebral amyloid angiopathy and cognitive impairment through vascular oxidative stress. Mol Neurodegener2023; 18: 73.
6.
SchaefferSIadecolaC.Revisiting the neurovascular unit. Nat Neurosci2021; 24: 1198–1209.
7.
Da MesquitaSRuaR. Brain border-associated macrophages: common denominators in infection, aging, and Alzheimer’s disease?Trends Immunol2024; 45: 346–357.
8.
DrieuADuSStorckSE, et al. Parenchymal border macrophages regulate the flow dynamics of the cerebrospinal fluid. Nature2022; 611: 585–593.
9.
GodderyENFainCELipovskyCG, et al. Microglia and perivascular macrophages act as antigen presenting cells to promote CD8 T cell infiltration of the brain. Front Immunol2021; 12: 726421.
10.
TaylorXClarkIMFitzgeraldGJ, et al. Amyloid-β (Aβ) immunotherapy induced microhemorrhages are associated with activated perivascular macrophages and peripheral monocyte recruitment in Alzheimer’s disease mice. Mol Neurodegener2023; 18: 59.
11.
LiCLiTHuM, et al. Novel perivascular macrophage mechanism to promote glymphatic Aβ clearance after stroke. Stroke2025; 56(9): 2695–2706.
12.
MestreHVermaNGreeneTD, et al. Periarteriolar spaces modulate cerebrospinal fluid transport into brain and demonstrate altered morphology in aging and Alzheimer’s disease. Nat Commun2022; 13: 3897.
13.
WardlawJMBenvenisteHNedergaardM, et al. Perivascular spaces in the brain: anatomy, physiology and pathology. Nat Rev Neurol2020; 16: 137–153.
14.
PercieduSertNHurstVAhluwaliaA, et al. The ARRIVE guidelines 2.0: updated guidelines for reporting animal research. J Cereb Blood Flow Metab2020; 40: 1769–1777.
15.
ParkLUekawaKGarcia-BonillaL, et al. Brain perivascular macrophages initiate the neurovascular dysfunction of alzheimer Aβ peptides. Circ Res2017; 121: 258–269.
16.
LinXKhalinIHarapanBN, et al. Perivascular macrophages mediate microvasospasms after experimental subarachnoid hemorrhage. Stroke2023; 54: 2126–2134.
17.
HouCLiuQZhangH, et al. Nimodipine attenuates early brain injury by protecting the glymphatic system after subarachnoid hemorrhage in mice. Neurochem Res2022; 47: 701–712.
18.
HuangWZhangYZhouY, et al. Glymphatic dysfunction in migraine mice model. Neuroscience2023; 528: 64–74.
19.
HarrisonIFIsmailOMachhadaA, et al. Impaired glymphatic function and clearance of tau in an Alzheimer’s disease model. Brain2020; 143: 2576–2593.
20.
KeezerMRSisodiyaSMSanderJW.Comorbidities of epilepsy: current concepts and future perspectives. Lancet Neurol2016; 15: 106–115.
21.
KressBTIliffJJXiaM, et al. Impairment of paravascular clearance pathways in the aging brain. Ann Neurol2014; 76: 845–861.
22.
IshidaKYamadaKNishiyamaR, et al. Glymphatic system clears extracellular tau and protects from tau aggregation and neurodegeneration. J Exp Med2022; 219(3): e20211275.
23.
GomolkaRSHablitzLMMestreH, et al. Loss of aquaporin-4 results in glymphatic system dysfunction via brain-wide interstitial fluid stagnation. eLife2023; 12: e82232.
24.
WangSYuXChengL, et al. Dexmedetomidine improves the circulatory dysfunction of the glymphatic system induced by sevoflurane through the PI3K/AKT/ΔFosB/AQP4 pathway in young mice. Cell Death Dis2024; 15: 448.
25.
RasmussenMKMestreHNedergaardM.The glymphatic pathway in neurological disorders. Lancet Neurol2018; 17: 1016–1024.
26.
SimonMJMurchisonCIliffJJ.A transcriptome-based assessment of the astrocytic dystrophin-associated complex in the developing human brain. J Neurosci Res2018; 96: 180–193.
27.
SiXDaiSFangY, et al. Matrix metalloproteinase-9 inhibition prevents aquaporin-4 depolarization-mediated glymphatic dysfunction in Parkinson’s disease. J Adv Res2024; 56: 125–136.
28.
HouCLiJWangB, et al. Dynamic evolution of the glymphatic system at the early stages of subarachnoid hemorrhage. Front Neurol2022; 13: 924080.
29.
ZhangXWangYJiaoB, et al. Glymphatic system impairment in Alzheimer’s disease: associations with perivascular space volume and cognitive function. Eur Radiol2024; 34: 1314–1323.
30.
IliffJJWangMLiaoY, et al. A paravascular pathway facilitates CSF flow through the brain parenchyma and the clearance of interstitial solutes, including amyloid β. Sci Transl Med2012; 4: 147ra111.
31.
JessenNAMunkASLundgaardI, et al. The glymphatic system: a beginner’s guide. Neurochem Res2015; 40: 2583–2599.
32.
NedergaardMGoldmanSA.Glymphatic failure as a final common pathway to dementia. Science2020; 370: 50–56.
33.
LiWSunBZhangX, et al. Near-infrared-II imaging revealed hypothermia regulates neuroinflammation following brain injury by increasing the glymphatic influx. ACS Nano2024; 18: 13836–13848.
34.
ZouKDengQZhangH, et al. Glymphatic system: a gateway for neuroinflammation. Neural Regen Res2024; 19: 2661–2672.
35.
FultzNEBonmassarGSetsompopK, et al. Coupled electrophysiological, hemodynamic, and cerebrospinal fluid oscillations in human sleep. Science2019; 366: 628–631.
36.
HablitzLMVinitskyHSSunQ, et al. Increased glymphatic influx is correlated with high EEG delta power and low heart rate in mice under anesthesia. Sci Adv2019; 5: eaav5447.
37.
LiuKZhuJChangY, et al. Attenuation of cerebral edema facilitates recovery of glymphatic system function after status epilepticus. JCI Insight2021; 6(17): e151835.
38.
XieFZhouCJinH, et al. Bilateral glymphatic dysfunction and its association with disease duration in unilateral temporal lobe epilepsy patients with hippocampal sclerosis. Epilepsy Behav2024; 155: 109777.
39.
ZhangCXuKZhangH, et al. Recovery of glymphatic system function in patients with temporal lobe epilepsy after surgery. Eur Radiol2023; 33: 6116–6123.
40.
PrinzMMasudaTWheelerMA, et al. Microglia and central nervous system-associated macrophages—from origin to disease modulation. Annu Rev Immunol2021; 39: 251–277.
41.
HawkesCAMcLaurinJ.Selective targeting of perivascular macrophages for clearance of beta-amyloid in cerebral amyloid angiopathy. Proc Natl Acad Sci U S A2009; 106: 1261–1266.
42.
WanHBrathwaiteSAiJ, et al. Role of perivascular and meningeal macrophages in outcome following experimental subarachnoid hemorrhage. J Cereb Blood Flow Metab2021; 41: 1842–1857.
43.
MaloCSHugginsMAGodderyEN, et al. Non-equivalent antigen presenting capabilities of dendritic cells and macrophages in generating brain-infiltrating CD8 (+) T cell responses. Nat Commun2018; 9: 633.
44.
MestreHTithofJDuT, et al. Flow of cerebrospinal fluid is driven by arterial pulsations and is reduced in hypertension. Nat Commun2018; 9: 4878.
45.
XieLKangHXuQ, et al. Sleep drives metabolite clearance from the adult brain. Science2013; 342: 373–377.
46.
HlauschekGNicoloJPSinclairB, et al. Role of the glymphatic system and perivascular spaces as a potential biomarker for post-stroke epilepsy. Epilepsia Open2024; 9: 60–76.
47.
IliffJJWangMZeppenfeldDM, et al. Cerebral arterial pulsation drives paravascular CSF–interstitial fluid exchange in the murine brain. J Neurosci2013; 33: 18190–18199.
