Abstract
Background: Patients with Alzheimer's disease (AD) develop blood–brain barrier dysfunction to varying degrees. How aging impacts Aβ pathology, blood–brain barrier function, and cognitive decline in AD remains largely unknown. In this study, we used 5xFAD mice to investigate changes in Aβ levels, barrier function, and cognitive decline over time. Methods: 5xFAD and wild-type (WT) mice were aged between 9.5 and 15.5 months and tested for spatial learning and reference memory with the Morris Water Maze (MWM). After behavior testing, mice were implanted with acute cranial windows and intravenously injected with fluorescent-labeled dextrans to assess their in vivo distribution in the brain by two-photon microscopy. Images were processed and segmented to obtain intravascular intensity, extravascular intensity, and vessel diameters as a measure of barrier integrity. Mice were sacrificed after in vivo imaging to isolate brain and plasma for measuring Aβ levels. The effect of age and genotype were evaluated for each assay using generalized or cumulative-linked logistic mixed-level modeling and model selection by Akaike Information Criterion (AICc). Pairwise comparisons were used to identify outcome differences between the two groups. Results: 5xFAD mice displayed spatial memory deficits compared to age-matched WT mice in the MWM assay, which worsened with age. Memory impairment was evident in 5xFAD mice by 2–threefold higher escape latencies, twofold greater cumulative distances until they reach the platform, and twice as frequent use of repetitive search strategies in the pool when compared with age-matched WT mice. Presence of the rd1 allele worsened MWM performance in 5xFAD mice at all ages but did not alter the rate of learning or probe trial outcomes. 9.5-month-old 15.5-month-old 5xFAD mice had twofold higher brain Aβ40 and Aβ42 levels (p < 0.001) and 2.5-fold higher (p = 0.007) plasma Aβ40 levels compared to 9.5-month-old 5xFAD mice. Image analysis showed that vessel diameters and intra- and extravascular dextran intensities were not significantly different in 9.5- and 15.5-month-old 5xFAD mice compared to age-matched WT mice. Conclusion: 5xFAD mice continue to develop spatial memory deficits and increased Aβ brain levels while aging. Given in vivo MP imaging limitations, further investigation with smaller molecular weight markers combined with advanced imaging techniques would be needed to reliably assess subtle differences in barrier integrity in aged mice.
| Original language | English |
|---|---|
| Article number | 29 |
| Journal | Fluids and Barriers of the CNS |
| Volume | 21 |
| Issue number | 1 |
| DOIs | |
| State | Published - Dec 2024 |
Bibliographical note
Publisher Copyright:© The Author(s) 2024.
Funding
The authors acknowledge current and former members of the Bauer and Hartz Laboratories for proofreading the manuscript. The content is solely the authors' responsibility and does not necessarily represent the official views of the National Institutes of Health. The funding for this study was provided by the National Institute of Health (Grant 2R01AG039621, PI: Hartz; R01AG075583, MPI: Hartz, Bauer; R01NS133250, PI: Hartz), Science Achievement Scholarship of Thailand (Yubolphan, 2019) and the Royal Jubilee Ph.D. Program Fellowship (Promsan, 2022). Animal experiments were approved by the Institutional Animal Care and Use Committee at the University of Kentucky (#2014–1233 and #2020–3455; PI: Hartz) and were in accordance with AAALAC regulations, the Guide of the Care and Use of Laboratory Animals, and the US Department of Agriculture Animal Welfare Act. Male transgenic 5xFAD mice (Tg6799; B6SJL-Tg (APPSwFlLonPSEN1*M146L*L286V)6799Vas/Mmjax; MMRRC Strain: 034840-JAX) and corresponding non-carrier littermates (WT mice) were obtained from Mutant Mouse Resource & Research Center (MMRRC) supported by the NIH at Jackson Laboratory (Bar Harbor, ME, USA). Animals were received at 4–8 weeks of age and group-housed in an AAALAC-accredited temperature-and-humidity-controlled vivarium with standard chow feed (Teklad 2918; Inotiv, West Lafayette, IN, USA) and water ad libitum under a 12 h light/dark cycle (temperature: 25 ± 1 °C; humidity: 50 ± 5%). Mice were genotyped for APPswe transgene and native Pde6b allele by polymerase chain reaction analysis (Transnetyx, Cordova, TN, USA) of ear and brain tissue samples and were aged 9–10 months and 15–16 months before experimentation. Pde6b status was assessed because this allele as a homozygote causes severe retinal deterioration and heterozygotes have visual deficiency []. 5xFAD and WT mice with heterozygous Pde6b allele are referred to as 5xFAD and WT mice in this study. 5xFAD and WT mice without Pde6b allele are referred to as 5xFAD and WT mice in this study. 133 mice were allocated to this study (n = 50 for 15.5-month-old 5xFAD mice, n = 33 for 15.5-month-old WT mice, n = 25 for 9.5-month-old 5xFAD mice, n = 25 for 9.5-month-old WT mice). Of these, 71 mice survived till endpoint (n = 18 for 15.5-month-old 5xFAD mice, n = 13 for 15.5-month-old WT mice, n = 21 for 9.5-month-old 5xFAD mice, n = 19 for 9.5-month-old WT mice). Assessment of rd1 allele revealed uneven distribution across ages and genotypes (Table ; χ = 32.875; p = 0.492).
| Funders | Funder number |
|---|---|
| Bauer and Hartz Laboratories | |
| Science Achievement Scholarship of Thailand | |
| MMRRC | |
| National Institutes of Health (NIH) | R01NS133250, R01AG075583, 2R01AG039621 |
| National Institutes of Health (NIH) |
Keywords
- 5xFAD
- Aβ
- Blood–Brain Barrier
- Pde6b
ASJC Scopus subject areas
- Neurology
- Developmental Neuroscience
- Cellular and Molecular Neuroscience