Role Of Epac In The Regulation Of ECB-mediated Inhibitory Synaptic Plasticity Of VTA Dopamine Neurons

Aims:The activation of phospholipase C?PLC?-?and PLC?has been linked to the facilitation of depolarization-induced suppression of inhibition?DSI?and long-term depression of inhibitory postsynaptic currents?IPSCs??I-LTD?in the ventral tegmental area?VTA?.So,PLC?might contribute to the facilitation of DSI and I-LTD in the VTA.Exchange protein directly activated by cAMP?Epac?activates the PLC?pathway.Thus,we hypothesized that the activation of Epac-PLC?pathway might play an important role in the facilitation of DSI and I-LTD in the VTA.This study investigated whether Epac was involved in the endocannabinoid dependent inhibitory synaptic plasticity in the VTA,with a focus on the role of Epac in the cocaine addiction and cocaine related conditioned place preference?CPP?performance.Methods:?1?Animals:Epac^+/+,Epac^-/-,Epac1^-/-,Epac2^-/-,PLC?^+/+?C57BL/6?and PLC?^-/-mice were used in this study.?2?Brain slice electrophysiology:Midbrain slices containing VTA were made.Whole-cell patch clamp recordings of IPSCs and sIPSCs were made in VTA dopamine neurons.DSI and I-LTD were induced using an internal solution with or without 8-?4-chlorophenylthio?adenosine 3',5'-cyclic monophosphate?8-CPT?.?3?In vivo single-unit extracellular recordings from VTA dopamine neurons were made to explore the role of Epac in the cocaine-induced increase in action potential?AP?firing.To confirm cell type,neurons were labeled with neurobiotin and brain slices were used for immunostaining of tyrosine hydroxylase?TH?after recordings.?4?Epac2^-/-,PLC?^-/-and wild type?WT?control mice were used in the cocaine CPP test to investigate whether Epac-PLC?pathway is required in cocaine-induced CPP.?5?The role of resveratrol pretreatment in improving cocaine CPP and cocaine addiction induced reduction of GABAergic inhibition in VTA dopamine neurons was investigated.Results:?1?8-CPT facilitates endocannabinoid-mediated DSI and I-LTD.?2?Epac2 knock-out blocks 8-CPT induced facilitation of DSI and I-LTD in the VTA.?3?PLC?knock-out blocks 8-CPT induced facilitation of DSI and I-LTD in the VTA.?4?Diacylglycerol lipase?DAGL?inhibitor DO34 blocks 8-CPT induced facilitation of DSI and I-LTD in the VTA.?5?Epac2 knock-out or PLC?knock-out blocks cocaine CPP.?6?Epac2 knock-out or PLC?knock-out blocks cocaine induced reduction of GABAergic inhibition in VTA dopamine neurons.?7?Epac2 knock-out blocks cocaine induced increase of VTA dopamine neurons excitability.?8?Resveratrol pretreatment improves cocaine CPP and cocaine addiction induced reduction of GABAergic inhibition in VTA dopamine neurons.Conclusions:Epac plays an important role in the regulation of?-aminobutyric acid?GABAergic?synaptic plasticity of VTA dopamine neurons.Epac facilitats endocannabinoid-mediatedDSIandI-LTDbyactivatingthe Epac2?PLC??DAGL?2-AG pathway.?2?The activation of Epac might be one of the molecular mechanisms of cocaine CPP.?3?Resveratrol pretreatment restores cocaine CPP by improving the reduction of GABAergic inhibition in VTA dopamine neurons.