Aim: 

Oxygen deprivation induces loss of energy supply, oxidative stress and consequent cell injury. The goal of the present work was to establish the early electrophysiological changes induced by the reduction of oxygen in the recording solution.

Methods: 

We performed patch-clamp recordings to investigate the effects induced on the cortical neuron membrane potentials by perfusion of sodium dithionite (10 mM). Dithionite induced a significant rapid transient depolarization (20.8+ 4.9 mV; p<0.01) that recovered to the resting membrane potential, although the hypoxic insult persisted. This depolarization was mediated by Reactive Oxygen Species (ROS) since treatment of cortical neurons with the free radical scavenger Edaravone (100 mM), prevented the membrane potential changes induced by dithionite. To investigate the ionic currents that sustained this depolarization, we utilized an external solution in which Ca²⁺ was removed and Na⁺ was substituted with Tris(hyroxymethyl)aminomethane.

Results: 

In this condition, dithionite-induced depolarization was not observed, indicating that it was due to Na⁺- and Ca²⁺- channel activation. The repolarization, instead, was due to the opening of ATP-dependent potassium channels since bath application of tolbutamide (100 mM), a selective antagonist of these channels, significantly reduced the repolarization.

Conclusion: 

This hypoxic in vitro model, described here allows the study of the early events of hypoxia with the possibility to test potential therapeutic drugs.