Alamandine Restores Mitochondrial Functions in the Aging Brain Microvasculature

Background: Aging is a strong risk factor for the development of Alzheimer’s disease and dementia. Brain microvascular (BMV) dysfunctions are increasingly recognized as mediators of vascular cognitive impairment and dementia (VCID). Mitochondrial dysfunction in the endothelium directly contributes to the impaired barrier functions and neurovascular coupling. Alamandine (Ala) is a novel angiotensin heptapeptide, which was shown to be cardiovascular protective by acting on the Mas Related G Protein Coupled Receptor, member D (MrgD). This study tested the potential beneficial effects of Ala on the aging-associated cognitive decline and delineated the mitochondrial mechanisms. Hypothesis: This study tested the hypothesis that Ala restores cognitive functions in aging by restoring mitochondrial biogenesis, and mitophagy. Methods: Young (2 - 4 months) and Old (20 – 24 months) C57BL/6 mice were used in this study. Old mice were treated with Ala by using osmotic pumps (1 μg/kg/min, 28 days) subcutaneously. BMVs were collected from mice’ brains by ultracentrifugation. BMVs were processed for transmission electron microscopy (TEM), confocal microscopy or for western blotting. Expression of proteins PINK1, PARKIN, BAX, BCL-2, MFN-1, MFN-2 and DRP-1 were detected by western blotting. Oxidative phosphorylation (OxPhos) was evaluated by using the Agilent Seahorse bioanalyzer. Statistical significance was determined using Mann Whitney or Kruskal-Wallis followed by Dunn’s test. Results: Aging decreased the mitochondrial size and numbers in BMVs (P < 0.05, n = 3) which was reversed by Ala treatment (P < 0.0001, n = 3). Expression of MFN-1 (P < 0.05, n = 6), MFN-2 (p< 0.05, N=6), and DRP-1 (P < 0.001, n = 6) was reduced with aging and showed an increasing trend with treatment. Expression of mitophagy related proteins PINK1 (P < 0.05, n = 6) and PARKIN (P < 0.05, n = 6) were increased with aging that were decreased by Ala (P < 0.05 or 0.01, respectively). The ratio of pro- and anti-apoptotic proteins BAX and BCL-2 (P < 0.05, n = 6) was increased in the Old BMVs that was reversed by Ala. OxPhos was decreased in the Old BMVs as indicated by decreased basal, maximal and nonmitochondrial respiration (P < 0.05, n = 8) while ATP-linked respiration and spare respiratory capacity showed decreasing trends. These impairments were reversed by Ala (P < 0.05, n = 8). Conclusion: These findings show that the cognitive decline with aging is associated with impaired biogenesis, increased mitophagy and apoptosis and decreased OxPhos. Ala treatment reversed these dysfunctions and increased the number of mitochondria, which resulted in the increased cognitive functions. Funding Support: This study is supported by National Institute on Aging (NIA), AG056881. This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.