Association Of The Phosphorylation Status Of Dynamin 1 With Neuronal Excitability

Objective:This study aimes to investigate the expression of the total and phospho-dynamin 1 as well as the level of activity-dependent bulk endocytosis(ADBE)in neurons incubated in 0-Mg2+ media for 3 hours and in control neurons.The effect of dynamin 1 inhibitor(dynasore)and agonist(PIP2)on these parameters were also determined.Methods: A sombati epileptic neuron model was made by incubating in0-Mg2+ media for 3 hours.Western blotting was used to investigate the expression of total and phospho-dynamin 1 in epileptic neurons,control neurons and epileptic neurons incubated by dynamin 1 inhibitor(dynasore)and agonist(PIP2).Q-PCR was used to determine the m RNA of dynamin 1expression level.Patch clamp techniques was used to determine the effect of dynasore on epileptiform discharges of neurons.Live cell fluorescence imaging was used to investigate the uptake of dextran by epileptic neurons,control neurons and epileptic neurons incubated by dynasore and PIP2.Results: Compared with normal neurons,the frequency of action potential in epileptic neurons was significantly higher,the level of dephospho-dynamin 1 was significantly increased(the phospho-dynamin 1level decreased while the total dynamin 1 level did not change)and that ADBE was enhanced in epileptic neurons.The dynamin 1 inhibitor(dynasore)reversed the increased expression of dephospho-dynamin 1 and the enhanced ADBE.Patch clamp studies revealed dynasore's inhibitory effect on neuronal excitability.PIP2 increased the level of dephospho-dynamin 1 in epileptic neurons,but it had no effect on the uptake of dextran.Conclusion: Our study demonstrated that the level of dephospho-dynamin 1 was significantly increased and that ADBE was enhanced in epileptic neurons.Dynasore reversed the increased expression of dephospho-dynamin 1 and the enhanced ADBE.What's more,dynasore inhibited the excitability of epileptic neurons.These results indicate that the expression of dephospho-dynamin 1 is closely related to neuronal hyperexcitability through its role in ADBE-mediated SV retrieval,which helps neurons meet the demands of abnormally enhanced neurotransmission.