Effect Of Lentivirus-mediated ShRNA Targeting Norepinephrine Transporter On Depression

Objective: To investigate the effect of lentivirus-mediated shRNA targeting norepinephrine transporter(NET)in locus coeruleus on depression-like behavior of rats,and the feasibility of experimental gene therapy for depression.Methods:(1)Recombinant lentiviral vector expressing shRNA against NET(ShRNA-1146,ShRNA-1222,ShRNA-330 and ShRNA-629)and nonsense shRNA as negative control(ShRNA-NC)were produced,and their efficiency in knocking down of NET in PC12 cells were assessed by Q-PCR and Western Blot analysis.(2)NET-shRNA were injected into the rat locus coeruleus bilaterally to investigate whether it can prevent the depressive-like behavior induced by chronic unpredictable mild stress(CUMS).3 weeks later,rats were tested for depressive-like behavior in the forced swimming test,the sucrose preference test and the open field test;the protein expressions of NET were measured by Western Blot;the levels of NE were measured by high performance liquid chromatography(HPLC).Results:(1)Q-PCR and Western Blot showed that the ShRNA-1146 has the best interference efficiency targeting on NET(P<0.01).(2)Compared with the depression model group,the immobility time in the forced swimming test decreased significantly(P<0.01),the sucrose preference in the sucrose preference test and the total scores in the open field test increased significantly(P<0.01)in the group treated with shRNA in locus coeruleus;compared with the depression model group,NET levels decreased significantly(P < 0.01)and NE levels increased significantly(P<0.05)in the group treated with shRNA in locus coeruleus.Conclusions: Lentivirus-mediated shRNA targeting NET in locus coeruleus downregulated NET both in vitro and in vivo,which also led to a significant reduction in depressive-like behavior of rats,opening a way for further translational studies.