Investigating glymphatic system and AQP4 water channels with novel drugs and MRI techniques

Alghanimy, Alaa (2025) Investigating glymphatic system and AQP4 water channels with novel drugs and MRI techniques. PhD thesis, University of Glasgow.

Abstract

The glymphatic system serves as a vital low resistance pathway for the efficient removal of toxic waste products from the brain and its malfunction is implicated in numerous neuropathological conditions. Aquaporin-4 (AQP4) water channels are membrane-tied and highly expressed at the end-feet of astrocytic cells in the brain. They are thought to be crucial to the glymphatic clearance system, water circulation, and homeostasis of the brain. Pharmacologically targeting AQP4 presents a promising therapeutic strategy for various neurological diseases. In 2009, Huber et al. developed TGN-020, a potent AQP4 inhibitor that significantly reduced cerebral oedema in stroke models. In 2018, they introduced TGN-073, a novel AQP4 facilitator that enhanced fluid turnover and interstitial fluid clearance. This thesis investigates the effects of these AQP4 modulators, with a particular focus on TGN-073, using advanced MRI techniques and immunofluorescence staining in rat models to elucidate their potential therapeutic benefits.

The initial objective was to employ an H2 17O tracer to evaluate the effect of the novel AQP4 facilitator TGN-073 on glymphatic transport. Despite extensive optimization efforts, the tracer signal remained low and unreliable, precluding its use in conducting our studies. Consequently, we assessed the impact of TGN-073 on glymphatic transport using dynamic contrast-enhanced MRI. This involved catheterizing the cisterna magna to infuse the MRI contrast agent Gd-DTPA into the cerebrospinal fluid. Our findings indicated that rats treated with TGN-073 exhibited a more extensive distribution and higher parenchymal uptake of GdDTPA compared to the vehicle group, suggesting TGN-073's potential in enhancing glymphatic function. Following this, I developed and established an immunohistochemistry protocol for AQP4 staining using immunofluorescence, a first in our department. The aim was to optimize this technique to its fullest potential, ensuring precision and reliability for the following experiments.

Given the invasive nature of the method used to investigate the impact of TGN073 on glymphatic transport, which requires cisterna magna cannulation, noninvasive alternatives were explored. Therefore, the impact of both AQP4 modulators, TGN-020 and TGN-073, was assessed without the necessity of exogenous contrast agents. These evaluations utilized T2 mapping and stimulated echo diffusion-weighted echo planar imaging (STE-DW-EPI), followed by immunofluorescence labelling of AQP4. No significant changes in the diffusion coefficient were observed across all observation times in any animal group, indicating no substantial alterations in brain microstructure. However, T2 values significantly decreased following the administration of TGN-073, suggesting enhanced water exchange. In contrast, T2 values significantly increased following the administration of TGN-020, while remaining unchanged in the vehicle group. These findings underscore the role of AQP4 in modulating water exchange between tissue compartments. Immunofluorescence staining revealed significantly higher AQP4 expression in the brains treated with TGN-073, contrasting with a significant decrease in AQP4 expression in the brains treated with TGN-020, compared to the vehicle-treated group.

To advance our understanding of the positive effects of AQP4 facilitators on glymphatic function, we investigated the impact of TGN-073 in a rat model of vascular cognitive impairment, specifically the bilateral common carotid artery stenosis (BCAS) model. This model, known for inducing chronic cerebral hypoperfusion and vascular dementia, represents a novel application within our institution. To our knowledge, this is the first study to evaluate glymphatic transport in BCAS rat models and to assess the impact of an AQP4 facilitator in this context. We successfully established cerebral hypoperfusion in the BCAS model, as evidenced by a significant reduction in cerebral blood flow (CBF). Our findings demonstrated glymphatic dysfunction and altered AQP4 expression associated with BCAS. Importantly, TGN-073, effectively mitigated these effects by restoring AQP4 expression, enhancing glymphatic function, and alleviating CBF reduction. This study highlights the potential of AQP4 facilitators in ameliorating the adverse effects of cerebral hypoperfusion and associated glymphatic dysfunction. TGN-073 shows promise for preventing the progression of neurodegenerative diseases and improving the quality of life for affected individuals.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Subjects: R Medicine > R Medicine (General); R Medicine > RC Internal medicine > RC0321 Neuroscience. Biological psychiatry. Neuropsychiatry
• Colleges/Schools: College of Medical Veterinary and Life Sciences > School of Psychology & Neuroscience
• Supervisor's Name: Holmes, Dr. William and Work, Dr. Lorraine
• Date of Award: 2025
• Depositing User: Theses Team
• Unique ID: glathesis:2025-85218
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 19 Jun 2025 13:15
• Last Modified: 19 Jun 2025 13:18
• Thesis DOI: 10.5525/gla.thesis.85218
• URI: https://theses.gla.ac.uk/id/eprint/85218

Related URLs

• Article DOI
• Article DOI
• Article DOI