Background: Aging is a highly complex process that affects various tissues and systems in the body. Senescent changes are relatively more prevalent and severe in the postmitotic cells. Mitochondria play an important role in the aging process. Recently, cell cultures have been widely used as an in vitro model to study aging. The present study was designed to investigate mitochondrial dysfunction associated with aging in a long-term cell culture system.
Material/Methods: Rat hippocampal neurons were maintained in culture in serum-free medium for 30 days in vitro (DIV). The morphology and development of hippocampal neurons was observed by phase contrast microscope. The levels of cellular senescence were evaluated by cytochemical staining of senescence-associated beta-galactosidase (SA-beta-Gal) at DIV 5, 10, 15, 20, 25 and 30. In addition, we investigated the changes in mitochondrial membrane potential (Δψm) and intracellular reactive oxygen species (ROS) generation of hippocampal neurons by flow cytometry at different ages.
Results: The proportion of the senescent cells steadily increased with age in neuron cultures. Δψm decreased gradually with age in long-term culture, while ROS generation increased.
Conclusions: This study indicates an age-related decrease in mitochondrial function in long-term hippocampal neuronal culture and suggests that DIV 25 neurons could possibly serve as a platform for the future study of anti-aging from the perspective of mitochondrial function.

Keywords: Rhodamine 123 - metabolism, Reactive Oxygen Species - metabolism, Neurons - pathology, Mitochondria - pathology, Membrane Potential, Mitochondrial, Intracellular Space - metabolism, Hippocampus - cytology, Cell Aging, Staining and Labeling, Time Factors, beta-Galactosidase - metabolism