Insulin Normalizes Ionic Homeostasis and the State of Mitochondria after a Mechanical Damage to the Culture of Brain Neurons

Abstract

Mechanical damage to the primary neuronal culture serves as a convenient in vitro model for studying the molecular and cellular mechanisms involved in the spread of the lesion in mechanical brain trauma. In this study, changes in the intracellular concentrations of Ca²⁺ ([Ca²⁺]_i) and Na⁺ ([Na⁺]_i), as well as mitochondrial potential (ΔΨ_m) in response to mechanical damage to the primary culture of rat cortical neurons were studied. Rapid (within 1–2 s) damage to the neuronal network in the form of an approximately 3 × 0.1-mm scratch caused an abrupt increase in [Ca²⁺]_i and [Na⁺]_i as well as a sharp drop in ΔΨ_m. There was a gradual recovery of these parameters to the basal level in ~78% of the cells that responded to the culture damage. The second phase of the [Ca²⁺]_i rise (delayed calcium deregulation, DCD), synchronous with a marked drop in ΔΨ_m, occurred in 22% of the cells. In such cells, [Na⁺]_i remained at the level of a high plateau. The addition of insulin (100 nM) 5 min before the mechanical damage reduced the proportion of neurons that had DCD and a sustained increase in [Na⁺]_i. Thus, the presence of insulin contributed to the normalization of Ca²⁺ and Na⁺ homeostasis and the functioning of mitochondria, which were impaired under the in vitro simulation of mechanical brain trauma.