Effect Of Resveratrol On The Hypoxic Responses Of Cortical Pyramidal Neurons

Hypoxic brain damage occurred in a variety of diseases such as neonatal hypoxic ischemia encephalopathy, cerebrovascular disease, brain trauma, altitude stress and so on may cause temporary or permanent brain dysfunction, even to death. Therefore, to investigate the mechanism of hypoxic brain damage and protective treatments to improve the patients quality of life and saving lives ha ve important clinical significance.Cerebral tissue is the most sensitive to hypoxic brain damage of the mammalian brain areas, which is the important part of the hypoxic brain damage in mammalian CNS. Hypoxia may lead to increasing of the internal flow of Na~+ which causes neuronal depolarization, leads directly to the neurons swelling, and causes the excess intracellular Ca2+ and presynaptic glutamate. As a result, the increasing of internal flow of Na~+ lead to a serie of traumatic changes in neurons, which is the important factor of hypoxic brain injury research.The hypoxic change of neuronal membrane potential and excitability is an important mechanism of mammalian hypoxic brain damage. The depolarization and increased excitability in early hypoxia are considered important trigger signal of neuron damage, which may cause injury or death of neurons, if not recovered in a few minutes, and hence as the important factor of hypoxic brain injury researc h.Resveratrol is the main active ingredient of traditional Chinese medicine Polygonum cuspidatum. It is confirmed that resveratrol possesses neuroprotective effect, and has the ability to pass the blood brain barrier quickly, thus it may be developed into an ideal neuroprotective medicine used for clinical purposes. However, the effect of resveratrol on the hypoxic responses of neurons is uncertain. This research adopts the whole cell patch clamp technique to observe the effect of resveratrol on the hypoxic responses of Na~+ currents of human cortical pyramidal neurons and effect of resveratrol on the hypoxic responses of membrane potential and excitability of rat cortical pyramidal neurons, and then provides experimental basis for its neuroprotective effect.1. Effect of Resveratrol on the hypoxic responses of Na~+ currents of human cortical pyramidal neurons Objective:To study the hypoxic responses of Na~+ currents of human cortical pyramidal neurons and the effect of resveratrol on it. Methods:Whole-cell patch-clamp technique was used to record the TTX-sensitive voltage-dependent Na~+ currents of pyramidal neurons in human cortical slices, in order to investigate the effects of acute hypoxia and resveratrol on Na~+ current amplitude and activation properties. Results:Acute hypoxia induced Na~+ current of human cortical pyramidal neurons a brief and slightly increase followed by a long- lasting inhibition in amplitude(P < 0.05). I-V curve and the activation curve of Na~+ currents were shifted to left(in the negative direction). AMPA receptor antagonist NBQX blocked the hypoxic increase of Na~+ currents(P < 0.01), and enhanced the posthypoxic inhibition of Na~+ currents(P < 0.01); GABAA receptor antagonist Bicuculline had no significant effect on the hypoxic alterations of Na~+ currents in human cortical pyramidal neurons(P > 0.05). Both of NBQX and Bicuculline did not affect the shifting of I-V curve and activation curve. 50?M resveratrol blocked the hypoxic increase of Na~+ currents of human cortical neurons(P < 0.01), and aggravated the posthypoxic inhibition of Na~+ currents(P < 0.05); 100?M resveratrol significantly delayed the responses of Na~+ currents to acute hypoxia(P < 0.05), and completely blocked the hypoxic increase of Na~+ currents and decreased the posthypoxic inhibition(P > 0.01). The negative shifting of I-V curve and the activation curve of Na~+ currents were shift to right( close to the curves in control group) by either 50?M or 100?M. Conclusion:Acute hypoxia mainly caused long-lasting inhibition in Na~+ currents of human cortical pyramidal neurons. AMPA receptor may effecte the responses of Na~+ channel to acute hypoxia. The effects of resveratrol upon hypoxic responses of Na~+ currents of human cortical neurons may be dose-related: small dose of resveratrol can mimic the effectiveness of N BQX to the hypoxic alteration of Na~+ current, and big dose of resveratrol depress the sensitivity of Na~+ channel to hypoxia.2. Effect of Resveratrol on the hypoxic responses of membrane potential of rat cortical pyramidal neurons Objective:To study the hypoxic responses of membrane potential of rat cortical pyramidal neurons and the effect of resveratrol on it. Methods:Whole-cell patch-clamp technique was used to record the membrane potential of pyramidal neurons in rat cerebral slices, in order to investigate the effect of resveratrol on the hypoxic responses of neuronal membrane potential. Results:Acute hypoxia induced an early depolarization, followed by a long- lasting posthypoxic hyperpolarization which lasted for more than 30 min in membrane potential of rat cortical pyramidal neurons;50?M resveratrol suppressed early depolarization and posthypoxic hyperpolarization(P<0.05); 100?M resveratrol completely blocked early depolarization and suppressed posthypoxic hyperpolarization(P<0.05). Conclusion:Acute hypoxia mainly caused an early depolarization and a long- lasting posthypoxic hyperpolarization in membrane potential of rat cortical pyramidal neurons. Resveratrol inhibited hypoxic changes in membrane potential of rat cortical pyramidal neurons, and may be dose-related.3. Effect of Resveratrol on the hypoxic responses of excitability of rat cortical pyramidal neurons Objective:To study the hypoxic responses of excitability of rat cortical pyramidal neurons and the effect of resveratrol on it. Methods:Whole-cell patch-clamp technique was used to evoke the action potential by injecting depolarized current into rat cortical pyramidal neurons under current-clamp mode and record the firing rate of cortical pyramidal neurons, in order to investigate the effect of resveratrol on the hypoxic responses of neuronal excitability. Results:Acute hypoxia induced of increase threshold value and decrease of the frequency of action potentials of rat cortical pyramidal neurons; Both 50?M and 100?M resveratrol suppressed the hypoxic increase of threshold value and hypoxic decrease of the frequency of action potentials of rat cortical pyramidal neurons(P<0.05), and the effect of 50?M resveratrol is more markedly than 100?M resveratrol(P<0.05). Conclusion:Acute hypoxia depressed the excitability of rat cortical pyramidal neurons. Resveratrol inhibited hypoxic changes in excitability of rat cortical pyramidal neurons, and may be dose-related.