A Spinal Neural Circuitry For Converting Touch To Itch Sensation Induced By Tac2 Neurons

Mechanical itch is a kind of itch causing scratching sensation induced by light touch stimulation,which could be inhibited by neuropeptide Y neuron(NPY neuron).How innocuous touch is converted into itch,however,remains elusive.In order to figure it out,we regulated the spinal interneurons expressing Tachykinin 2-Cre(Tac2Cre)by chemogenetic and optogenetic approaches and abolished gastrin-releasing peptide receptor(GRPR)neurons,itch specific related neurons,by intrathecal injection of bombesin saporin(BB-sap),to study their roles in mechanical itch.In the meanwhile,we also worked on the communication among NPY,Tac2 and GRPR neurons to clarify the mechanism of inhibiting mechanical itch by chemogenetic,pharmacology and histomorphology methods.We'd like to explore the transmission pathway of mechanical itch in spinal cord according to our experiments.Furthermore,by applying our findings in clinical,we could provide a new therapeutic target in alloknesis symptom of chronic itch patients to improve their life quality.The followings are the main findings of this project:1.Tac2 and GRPR neurons in spinal cord could transmit the mechanical itch.(1).According to the results of immunofluorescence,the c-Fos induced by mechanical itch showed co-expression with Tac2 neurons in spinal cord,while chemical itch not.(2).According to the results of in vitro electrophysiology,Tac2 neurons in spinal cord could receive the input of A? fiber which transmitting the touch information.(3).According to the results of behavior tests,activation of Tac2 neurons in cervical spinal cord by optogenetic stimulation could induce scratching behavior,while chemogenetic inhibition of Tac2 neurons or GRPR neurons could both inhibit the mechanical itch behavior.Moreover,ablation of GRPR neurons in spinal cord could nearly abolish mechanical itch behavior.2.The NPY-NPY1R pathway could participate in inhibiting the mechanical and chemical itch.(1).According to the results of behavior tests,both intrathecal injection of NPY1R agonist(LP-NPY)and chemogenetic inhibition of NPY neurons in spinal cord could inhibit mechanical and chemical itch behaviors.On the contrary,intrathecal injection of Npy1r siRNA could increase that.(2).According to the results of RNAscope,Npylr has colocalization with Grpr in RNA level,whereas not with Tac2 RNA.(3).According to the results of in vitro electrophysiology,LP-NPY could increase the mIPSCs of GRPR neurons,while have no influence on Tac2 neurons.3.Tac2 and NPY neurons in spinal cord could play different roles in alloknesis.In dry skin model,intrathecal injection of LP-NPY could inhibit its spontaneous itch scratching behavior and mechanical itch behavior.Moreover,both chemogenetic activation of NPY neurons and inhibition of Tac2 neurons in cervical spinal cord could also inhibit the scratching behavior.Together,we conclude that Tac2 neurons acted as the first relay station to convert innocuous touch to itch through GRPR neurons,and NPY neurons could inhibit mechanical itch behavior by NPY1R on GRPR neurons.An imbalance between Tac2,GRPR and NPY neurons contributed to alloknesis associated with pathological itch.