Associative Learning and CA3–CA1 Synaptic Plasticity Are Impaired in D₁R Null,Drd1a^−/−Mice and in Hippocampal siRNA SilencedDrd1aMice

Abstract

Associative learning depends on multiple cortical and subcortical structures, including striatum, hippocampus, and amygdala. Both glutamatergic and dopaminergic neurotransmitter systems have been implicated in learning and memory consolidation. While the role of glutamate is well established, the role of dopamine and its receptors in these processes is less clear. In this study, we used two models of dopamine D 1 receptor (D 1 R, Drd1a) loss, D 1 R knock-out mice (Drd1a

Impairment of acquisition of intravenous cocaine self-administration by RNA-interference of dopamine D1-receptors in the nucleus accumbens shell

“…The construction of Drd1a-siRNA expressing lentiviruses has been described previously in detail (Bahi and Dreyer, 2012;Ortiz et al, 2010). In brief, three 19-nucleotide Drd1a-siRNA sequences were added to the U6 promoter by PCR.…”

“…Vectors were resuspended in PBS-BSA and stored at À80 C until use (Bahi, 2013;Bahi and Dreyer, 2013). The three distinct Drd1a-siRNA sequences were successfully used to knock-down the expression Drd1a mRNA in the NAc (Bahi and Dreyer, 2012) and proteins in the hippocampus (Ortiz et al, 2010).…”

“…After 15 days of recovery from surgical procedures, which is sufficient to reach a maximal degree of D1aR silencing in vivo (Ortiz et al, 2010), rats were trained to self-administer cocaine intravenously (unit doses, 0.25 mg/kg in 24 ml) in single daily 1-h sessions for 5 days/week, under fixed ratio schedule 1 (FR-1, 1 nose-poke for 1 infusion), which was then increased to FR-5 (5 nose-pokes for 1 infusion) starting from the 15th session in each experiment. At this point, in at least one experimental group, rats reached the criterion for acquisition (set at 85% of responses on the active hole), keeping a stable pattern over three sessions.…”

“…RNAi is triggered in eukaryotic cells by double-stranded RNAs having 19e21 nucleotides (short interfering RNA, siRNA; Fire et al, 1998), which, when recognized as aberrant, induce a polyenzymatic process leading to degradation of homologous mRNAs (Hannon, 2002). For in vivo application, viral mediated expression of siRNAs, which takes place a few days after transduction (Bahi and Dreyer, 2012;Ortiz et al, 2010), allows stable long term RNAi, thus silencing the expression of the relative genes (Bahi et al, 2004(Bahi et al, , 2005Bahi and Dreyer, 2012;Ortiz et al, 2010;Scherr et al, 2003;Van den Haute et al, 2003). Viral mediated silencing of targeted proteins offers higher temporal (RNAi starts right after transduction of targeted cells) and spatial control (inoculation of defined volumes and concentrations of viral vectors restricts silencing to targeted areas) of manipulations, compared to conventional knockout or pharmacological strategies.…”

Lentiviral Delivery of a Vesicular Glutamate Transporter 1 (VGLUT1)-Targeting Short Hairpin RNA Vector Into the Mouse Hippocampus Impairs Cognition

“…The skull surface was exposed and holes drilled at the injection sites. Mice received bilateral injections of nontarget control shRNA or VGLUT1 shRNA 5 encoding lentiviral vectors (2 ml per side; 9.68 Â 10 7 viral particles/ml, a similar titer to previous studies; Ortiz et al, 2010) into the dorsal hippocampus (anterioposterior À 2.0, mediolateral ± 1.5, dorsoventral À 2.5 mm from Bregma) using a 33-gauge microinjector connected to a Hamilton syringe. Injections occurred over 2 min and the injector remained in place for a further 2 min.…”