Lifespan Imaging Suggests Midlife as a Strategic Window to Protect Microvascular Mitochondria, Blood-Brain Barrier Integrity and Neurovascular Coupling

Mitochondria are emerging as master regulators of brain endothelial function, fueling blood-brain barrier (BBB) integrity, sustaining neurovascular coupling (NVC), and shaping the resilience of the aging cerebrovasculature, yet how microvascular endothelial mitochondria change across the lifespan remains poorly understood. Here, we introduce the first lifespan-spanning, high-resolution atlas of endothelial mitochondrial aging in the living brain using longitudinal in vivo two-photon imaging to track microvascular endothelial mitochondrial architecture and dynamics in VECAD-mito-Dendra2 mice from early adulthood to old age. After implantation of a TPX cranial window at 5 months, monthly imaging from 5 to 24 months quantified mitochondrial number, vascular coverage, and fusion/fission behavior. Our analysis uncovered a previously unrecognized biphasic mitochondrial aging program, with mitochondrial number and vascular coverage progressively increasing and reaching their highest levels around 12 months, before declining. This inflection point identifies a critical window in which endothelial bioenergetic vulnerability emerges. To test whether this age-related deterioration could be prevented, a subset of mice received the mitochondrial-targeted therapies SS31 or the 5-HT1F receptor agonist Lasmiditan beginning at 12 months, coinciding with the onset of decline. Together, this work defines the natural longitudinal progression of endothelial mitochondrial aging and establishes a framework to test whether maintaining endothelial mitochondrial integrity during midlife could help sustain BBB stability and NVC function into aging. Funding: NIH R03AG070479, K01AG073614, R21AG080775, AHA (CDA941290 and 24TPA1299954, 25IPA1456700, 25POST1377493), Hevolution/AFAR. 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.