The Effect Of Lingo-1 On Axon With Mg²⁺-Free-induced Neuron Seizure-Like Discharge

Partâ…  the effect of axon and Lingo-1 with Mg2+-free-induced neuron seizure-like dischargeObjective To detect the changes of axon morphology and Lingo-1 expression after seizure-like discharge induced by Mg2+-free media.Methods Cortical neurons cultured for 10 d were exposed to normal media or Mg2+-free media for 3h, then divided into two groups: normal group and Mg2+-free group.The action potential frequency of two groups detected by patch clamp. Application of NF200 to test the purity of neuron and the changes of axons. Then the expression of Lingo-1 m RNA and protein were determined by Q-PCR and Western-blot at 6h,12 h,24h,48 h and 72 h after Mg2+-free or normal media treatment.Results 1) The discharge form of Mg2+-free group was myotonic discharge, and the action potential frequency was significantly higher than the normal group(P<0.05). 2) In normal group, neurons axon were thick and dense network connection.NF200 only expressed in the neurite. In Mg2+-free group, neurons axon were slender and sparse network connection. NF200 highly expressed in cell body. 3) From 6h to 72 h, compared with normal group,Lingo-1 expresion level increased at each point in time in Mg2+-free group(P<0.05).Conclusion The Mg2+-free treated can successfully establish the seizure-like discharge neuron model, there are axonal injury and Lingo-1 expression upregulation after Mg2+-free treatment.Part â…¡ the relationship between axonal injury and Lingo-1 expression upregulation after seizure-like discharge induced by Mg2+-free mediaObjective To study the relationship between axonal injury and Lingo-1 expression upregulation after seizure-like discharge induced by Mg2+-free media.Methods 1) Designed Lingo-1 sh RNA lentiviral inhibited Lingo-1 expression of neuron, and control sh RNA lentiviral as control, divided into normal group, control sh RNA group and Lingo-1 sh RNA group. Western-blot detected the Lingo-1 expression of the three groups.2) Cultured neuron were divided into four groups: normal group, Mg2+-free group, Mg2+-free + control sh RNA group and Mg2+-free + Lingo-1 sh RNA group. Observed the morphology changes of neuron axon and detected the neuron axon length with immunofluorescence.Results 1) The expression of Lingo-1: normal group and control sh RNA group had no statistically differences, while Lingo-1 sh RNA group significantly lower than control sh RNA group. 2) The length of axon: Mg2+-free group were less than normal group(P=0.000), Mg2+-free + control sh RNA group and Mg2+-free group had no statistically differences, Mg2+-free + Lingo-1 sh RNA group were greater than Mg2+-free + control sh RNA group(P=0.000).Concusion Lentiviral-mediated Lingo-1 sh RNA can successfully inhibit the expression of Lingo-1, and Lingo-1 expression downregulation can reduced the axonal injury. The study show Lingo-1 may be involved in axonal injury after Mg2+-free-induced neuron seizure-like discharge.