Introduction: Brain aging involves a complex interplay of cellular and molecular changes, including metabolic alterations and the accumulation of senescent cells. These changes frequently manifest as dysregulation in glucose metabolism and mitochondrial function, leading to reduced energy production, increased oxidative stress, and mitochondrial dysfunction—key contributors to age-related neurodegenerative diseases.Methods: We conducted experiments on two models: young (3–4 months) and aged (over 18 months) mice, as well as cultures of senescent and control mouse astrocytes. Mitochondrial content and biogenesis were analyzed in astrocytes and neurons from aged and young animals. Cultured senescent astrocytes were examined for mitochondrial membrane potential and fragmentation. Quantitative PCR (qPCR) and immunocytochemistry were used to measure fusion- and fission-related protein levels. Additionally, transmission electron microscopy provided morphological data on mitochondria.Results: Astrocytes and neurons from aged animals showed a significant reduction in mitochondrial content and a decrease in mitochondrial biogenesis. Senescent astrocytes in culture exhibited lower mitochondrial membrane potential and increased mitochondrial fragmentation. qPCR and immunocytochemistry analyses revealed a 68% increase in fusion-related proteins (mitofusin 1 and 2) and a 10-fold rise in DRP1, a key regulator of mitochondrial fission. Transmission electron microscopy showed reduced perimeter, area, and length-to-diameter ratio of mitochondria in astrocytes from aged mice, supported by elevated DRP1 phosphorylation in astrocytes of the cerebral cortex.Discussion: Our findings provide novel evidence of increased mitochondrial fragmentation in astrocytes from aged animals. This study sheds light on mechanisms of astrocytic metabolic dysfunction and mitochondrial dysregulation in brain aging, highlighting mitochondrial fragmentation as a potential target for therapeutic interventions in age-related neurodegenerative diseases.

Microglia, the resident immune cells of the central nervous system (CNS), play essential roles in maintaining brain homeostasis. While transient activation is protective, chronic microglial reactivity contributes to neuroinflammatory damage and neurodegeneration. The mitochondrial mechanisms underlying this shift remain poorly understood. Here, we investigated whether lipopolysaccharide (LPS) induces coordinated mitochondrial and metabolic alterations in BV-2 microglial cells. LPS stimulation (100 ng/mL, 24 h) induced a reactive phenotype, with increased Iba1 (+82%), F4/80 (+132%), and Cd68 (+44%), alongside elevated hydrogen peroxide (~6-fold) and nitrite (~45-fold). Cytotoxicity increased by 40% (LDH assay), and cell viability dropped to ~80% of the control (MTT). Extracellular lactate increased, indicating glycolytic reprogramming. However, LPS-primed cells showed greater ATP depletion under antimycin A challenge, reflecting impaired metabolic flexibility. Hoechst staining revealed a ~4-fold increase in pyknotic nuclei, indicating apoptosis. Mitochondrial dysfunction was confirmed by a 30–40% reduction in membrane potential (TMRE, JC-1), a ~30% loss of Tomm20, and changes in dynamics: phospho-Drp1 increased (+23%), while Mfn1/2 decreased (33%). Despite a ~70% rise in Lamp2 signal, Tomm20–Lamp2 colocalization decreased, suggesting impaired mitophagy. High-resolution respirometry revealed decreased basal (−22%), ATP-linked (24%), and spare respiratory capacity (41%), with increased non-mitochondrial oxygen consumption. These findings demonstrate that LPS induces mitochondrial dysfunction, loss of metabolic adaptability, and increased apoptotic susceptibility in microglia. Mitochondrial quality control and energy flexibility emerge as relevant targets to better understand and potentially modulate microglial responses in neuroinflammatory and neurodegenerative conditions.

Astrocytes are critical for maintaining brain homeostasis through metabolic support, neurotransmitter regulation, and blood–brain barrier integrity. In the aging brain and neurodegenerative conditions, astrocytes undergo functional and morphological changes that culminate in a state of cellular senescence. Astrocytic senescence—characterized by irreversible cell-cycle arrest and a pro-inflammatory senescence-associated secretory phenotype (SASP)—is emerging as a key contributor of chronic neuroinflammation and synaptic dysfunction in aging. This review examines the molecular mechanisms underlying astrocyte senescence, highlighting how persistent DNA damage responses (DDR), oxidative stress, and mitochondrial dysfunction disrupt essential astrocytic functions (e.g., glutamate uptake, K⁺ buffering, and metabolic coupling with neurons). These senescent changes in astrocytes lead to impaired synaptic plasticity and contribute to age-related cognitive decline. Collectively, astrocytic senescence represents a pivotal and targetable mechanism in age-related neurodegeneration, and therapeutic strategies aimed at eliminating senescent cells or modulating the SASP hold promise for restoring synaptic function and promoting healthy brain aging.

Aging is a complex biological process that significantly impacts the central nervous system (CNS). Astrocytes, critical support cells in the brain, undergo senescence with age, contributing to neurodegenerative diseases. While previous studies have utilized murine models to investigate astrocyte senescence, human astrocytes offer a more physiologically relevant system to study age‐related neurodegenerative changes. This study presents a novel protocol for inducing senescence in both human primary and murine astrocytes using a combination of cellular stressors, such as lactacystin, H₂O₂, rotenone, and doxorubicin. Our results demonstrate that doxorubicin treatment effectively induces a robust senescent phenotype in both human and murine astrocytes, characterized by increased expression of senescence markers such as p21 and β‐galactosidase, along with activation of the DNA damage response (γ‐H2AX and 53BP1). Doxorubicin treatment increased nuclear size and induced cell cycle arrest in astrocytes, as revealed by reduced BrdU incorporation and decreased cell density, without inducing cytotoxic effects. This phenotype is accompanied by a pronounced pro‐inflammatory profile, with elevated expression of cytokines including MMP3, IL‐6, and IL‐1β, indicative of a strong senescence‐associated secretory phenotype (SASP). These findings provide a novel in vitro model for murine and human astrocyte senescence induced by doxorubicin, highlighting its relevance for studying mechanisms underlying age‐related neuroinflammation and neurodegeneration. By establishing a robust model of human astrocyte senescence, this study provides a valuable tool for exploring the molecular and cellular mechanisms driving age‐related neurodegenerative processes, serving as an alternative approach to traditional murine models. image