Real time CPP

Figure 4G- D1-cre
Figure 5H- retroAAV-cre
Figure S5D- retroAAV-cre control
Surgery: At 8-12 weeks of age mice were stereotactically injected for real-time CPP (rtCPP) experiments. The first experimental cohort consisted of D1-Cre male mice (n=8) stereotactically injected with 80nl of AAV9-CAGGS-FLEX-ChR2-TdTomato (purchased from the UPENN virus core) to the claustrum region (LM: 2.82, RC: 1, DV: -3.7). The second experimental cohort consisted of WT c57/bl6 mice (n=6) stereotactically injected with 200nl retroAAV-CKII-iCre to the ACC (LM: ±0.25, RC: 1.1, DV: -1.9) and OFC (LM: ±1, RC: 2.55, DV: -2.4) and 80nl of AAV9-CAGGS-FLEX-ChR2-TdTomato stereotactically injected to the claustrum region (LM: ±2.82, RC: 1, DV: -3.7). The control group consisted of 4 WT c57/bl6 mice stereotactically injected with 200nl retroAAV-CKII-iCre to the ACC (LM: ±0.25, RC: 1.1, DV: -1.9) and OFC (LM: ±1, RC: 2.55, DV: -2.4) and 80nl of AAV9-CAGGS-FLEX-ChR2-TdTomato stereotactically injected to the claustrum region (LM:±2.82 , RC: 1, DV: -3.7). Silica multimode fiber optic cannula (200-μm core; 240-μm outer diameter) mounted in a 1.25-mm zirconia ferrule (Doric lenses) were slowly lowered into the brain bilaterally following virus injection (LM:±2.82 , RC: 1, DV: -3.65), and cemented in place such that the fiber tip was located above the claustrum.
Optogenetics equipment: A blue laser (MBL-FN-437-200mW, CNI laser), was connected to 1x2 Intensity Division Fiberoptic Rotary Joint (FRJ_1x2i_SMA-2FC_0.22, Doric). Two 50cm Mono Fiberoptic Patchcord (MFP_200/230/900-0.37_0.5m_FC-ZF1.25(F), Doric) were connected to the implanted cannulae with a zirconia connection sleeve (1.25mm, Doric).

Real-time CPP: A 3-chamber compartment was used. The north chamber had a smooth floor and walls textured with black and white vertical stripes, while the south chamber had a grid floor and white walls with black dots. The central chamber lacked discerning features and was meant to increase the division between the lateral chambers. Either the north or south chambers were paired with laser stimulation of the claustrum, counterbalanced within each experimental group. The mouse’s position was calculated in real time with Ethovision XT 11.5 software. The laser (5mW) was illuminated in a cycle of 1second on / 3second off for the duration that the mouse remained in the laser-paired chamber (as in (Kravitz and Kreitzer, 2012)). All experimental sessions were 20min long. For each real-time CPP experiment repeated measurements were performed on the same group of mice. No experimental mice were excluded from analysis.

图4G- D1-cre
图5H- retroAAV-cre
图S5D- retroAAV-cre对照
手术：在8-12周龄的小鼠中，通过立体定向技术注入实时条件性帕利哌酮（rtCPP）进行实验。第一组实验小鼠为D1-Cre雄性小鼠（n=8），通过立体定向技术向纹状体区域（LM: 2.82, RC: 1, DV: -3.7）注入80nl的AAV9-CAGGS-FLEX-ChR2-TdTomato（购自UPENN病毒核心）。第二组实验小鼠为野生型c57/bl6小鼠（n=6），通过立体定向技术向前扣带回（ACC）（LM: ±0.25, RC: 1.1, DV: -1.9）和眶额叶皮质（OFC）（LM: ±1, RC: 2.55, DV: -2.4）注入200nl的retroAAV-CKII-iCre，并在纹状体区域（LM: ±2.82, RC: 1, DV: -3.7）注入80nl的AAV9-CAGGS-FLEX-ChR2-TdTomato。对照组由4只野生型c57/bl6小鼠组成，通过立体定向技术向ACC和OFC注入200nl的retroAAV-CKII-iCre，并在纹状体区域注入80nl的AAV9-CAGGS-FLEX-ChR2-TdTomato。在病毒注入后，采用硅基多模光纤导管（200-μm芯径；240-μm外径）和1.25-mm氧化锆 ferrule（Doric镜头）缓慢地双侧降低至大脑，并固定在适当位置，使光纤尖端位于纹状体之上。
光遗传学设备：连接至1x2强度分配光纤旋转关节（FRJ_1x2i_SMA-2FC_0.22，Doric）的蓝色激光（MBL-FN-437-200mW，CNI激光），通过50cm单光纤连接线（MFP_200/230/900-0.37_0.5m_FC-ZF1.25(F)，Doric）连接至植入的导管，导管通过氧化锆连接套筒（1.25mm，Doric）进行连接。
实时条件性帕利哌酮实验：使用三室隔间。北部隔间具有光滑的地板和黑白相间的垂直条纹纹理墙壁，而南部隔间具有网格地板和白色墙壁上的黑色圆点。中央隔间缺乏显著特征，旨在增加侧室之间的分隔。北或南隔间与纹状体的激光刺激配对，在每组实验中均进行平衡。使用Ethovision XT 11.5软件实时计算小鼠的位置。激光（5mW）以1秒开/3秒关的周期在激光配对的隔间中照亮，持续时间与小鼠在激光配对的隔间中停留的时间相同（如Kravitz和Kreitzer，2012年所述）。所有实验会话均为20分钟长。对于每个重复的实时条件性帕利哌酮实验，对同一组小鼠进行重复测量。没有实验小鼠被排除在分析之外。