The Mechanism And Intervention Of Cocaine-Induced Seizure

Cocaine-induced seizure(CIS),characterized by abnormal brain activity and repeated or rhythmic jerking muscle movements,has always been considered one of the major effects of cocaine poisoning.Although CIS has become an insidious and global public health problem for years,the unstanding about mechanism of CIS is still at an early stage,and there are few studies on it and the underlying mechanism of it.The most effective medication used for treating seizures in clinics is benzodiazepine accompanied by the nonnegligible psychoactive or sedative side effects.Although there are more and more tips for treating seizures in clinics:Cannabinoids has increasingly been used as an antiepileptic medication in clinics,and Epidiolex,a purification reagent of cannabidiol(CBD),has recently been approved by the U.S.Food and Drug Administration(FDA)and proved to be effective in treating patients over 2 years old with seizures related to Dravet syndrome and LennoxGastaut syndrome.There is also evidence indicating the therapeutic effects of CBD on cocaine-induced seizures in a few animal studies,although the cellular and molecular mechanisms remain unsolved.In this study,we first successfully established a mouse model of CIS.Next,consistent with the current clinical medication for the treatment of seizures,tetrahydrocannabinol(THC,the psychoactive components of cannabinoids)and cannabidiol(CBD,the non-psychoactive component of cannabinoids)were respectively used to evaluate their therapeutic effects on CIS.According to the target of cannabinoids,specific antagonists were used to initially explore the molecular mechanism of CIS,we found that the specific antagonists against cannabinoid receptors were both incapable of interrupting the effects of cannabinoids on CIS,which indicates that the restoring effects of cannabinoids on CIS may involve other targets.Then,based on the molecular targets of cocaine and cannabinoids,single-cell patch-clamp electrophysiology technique was used and we found that high dose of cocaine suppressed 50%drop in glycine activated currents in HEK293 cells transfected with glycine receptors and cultured primary cortical neurons,which means that cocaine significantly inhibited glycine receptors function.And cocaine-induced dysfunction of the glycine receptor was restored by cannabinoids.Furthermore,molecular dynamic simulation(MDS)and molecular docking studies and single-cell patch-clamp electrophysiology were performed to determine that DH-CBD can reduce the hydrogen-bonding interaction between cocaine and threonines at positions 264 and 265 of the glycine receptor,thereby alleviating the inhibitory effect of cocaine on glycine receptor function.Since cocaine has a good blood-brain barrier permeability,common mass spectrometry and acid-enhanced desorption electrospray ionization-MS(aDESIMS)imaging were used to exclude the effect of DH-CBD on the concentration and distribution of cocaine entering the brain during CIS.Therefore,the early gene c-fos immunofluorescence staining experiment was used to explore the effects of cocaine on brain neuronal excitability,we found that the number of neurons triggered by cocaine increased 2～3 times in the prefrontal cortex(PFC),CA1(cornu ammonis 1)and dentate gyrus(DG)in the hippocampus,while cannabinoids significantly suppressed cocaine-induced neuronal hyperactivity in these brain regions.Finally,in vitro patchclamp electrophysiological techniques,localized drug administration in local brain regions,and in vivo electrophysiological techniques were used to clarify that DH-CBD restored cocaine-induced dysfunction of the glycine receptors in prefrontal cortex and hippocampus CA1 and DG by interacting with extrasynaptic glycine receptors in these brain regions to treat cocaine-induced seizures.In conclusion,this research has innovatively proposed and proved that cannabinoids can restore cocaine-induced seizures through glycine receptors.To explore the underlying mechanism of CIS and cannabinoids treatment,we utilized a a variety of techniques and found that cocaine triggers the hyperexcitability of neurons in the prefrontal cortex and hippocampus CA1 and DG by inhibiting the function of glycine receptors in these brain regions,while cannabinoids can reduce the hydrogenbonding interaction between cocaine and glycine receptors by interacting with extrasynaptic glycine receptors in these brain regions,thereby restoring cocaineinduced dysfunction of the glycine receptors in these brain regions,and then treating CIS.At the same time,DH-CBD,a synthetic cannabinoid modified from tetrahydrocannabinol by our previous studies,has been clarified its therapeutic effect on cocaine-induced seizures,and broadened the therapeutic effect of cannabinoids and the role of glycine receptors in the central nervous system.