Drug-induced changes in connectivity to midbrain dopamine cells revealed by rabies monosynaptic tracing
- Program in Mathematical, Computational, and Systems Biology, University of California, Irvine, United States
- Department of Physiology and Biophysics, University of California, Irvine, United States
- Department of Biomedical Engineering, University of California, Irvine, United States
- Department of Neurobiology and Behavior, University of California, Irvine, United States
- Department of Pharmaceutical Sciences, University of California, Irvine, United States
Figures
Figure 1 with 1 supplement
PCA-based dimensionality reduction of RABV input tracing to three different VTA cell types.
(A) Schematic and timeline of viral injections into mice. (B) Representative images of the BNST, GPe, and DCN of DAT-Cre, GAD2-Cre, and vGlut2-Cre mice. Green indicates RABV-labeled cells. Scale, 200 μm. (C) Bar plot showing percent of RABV-labeled cells in each input region for DAT-Cre, GAD2-Cre, and vGlut2-Cre mice. (D) PCA of input labeling from brains from DAT-Cre, GAD2-Cre, and vGlut2-Cre mice. For this and other figures, ellipsoids were centered at the average coordinate of a condition and stretched one standard deviation along the primary and secondary axes. (E) Box plot comparisons of PC1. One-way ANOVA p = 0.010, pairwise t-tests DAT-Cre vs. GAD2-Cre multiple comparisons adjusted p = 0.0087, DAT-Cre vs. vGluT2-Cre p = 0.46, GAD2-Cre vs. vGluT2-Cre p = 0.19. n = 8 for DAT-Cre, n = 13 for GAD2-Cre, and n = 7 for vGluT2-Cre. (F) Box plot comparisons of PC2. One-way ANOVA p = 0.016, pairwise t-tests DAT-Cre vs. GAD2-Cre multiple comparisons adjusted p = 0.65, DAT-Cre vs. vGluT2-Cre p = 0.016, GAD2-Cre vs. vGluT2-Cre p = 0.051. (G) Heatmap of the contributions of each brain region, or feature, in the data to PCs 1–3. For this and all figures, error bars = ± 1 SEM, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.
Figure 1—figure supplement 1
Representative images showing anatomical boundaries for the 22 defined input regions.
Each input region is denoted where it is present, and each region is denoted on at least one section.
Figure 2
Projection portraits and UMAP of RABV inputs to VTA cells.
(A) Schematic of how projection portraits are constructed. A representative slice of the VTA is considered for experiments from the Allen Mouse Brain Connectivity Atlas with injections into relevant brain regions that provide input to the VTA. Projection density values in this slice are averaged and visualized to obtain the projection portrait in the VTA for that group of regions. (B) Projection portrait of the VTA based on inputs from regions with a high positive contribution to PC1: BNST, EP, GPe, CeA, and ZI. (C) Projection portrait of the VTA based on inputs from regions with a large negative contribution to PC1: anterior cortex, NAcMed, VP, PO, septum, LHb, MHb, and LH. (D) Projection portrait of the VTA based on inputs from regions with a high positive contribution to PC2: LHb, MHb, EP, ZI, DCN, and anterior cortex. (E) Projection portrait of the VTA based on inputs from regions with a large negative contribution to PC2: NAcMed, NAcLat, NAcCore, DStr, and GPe. (F) UMAP embedding of input labeling of brains from DAT-Cre, GAD2-Cre, and vGlut2-Cre mice. (G) Correlogram generated using the average Euclidean distance of data points from each other from 20 UMAP embeddings.
Figure 3
PCA can deconvolve drug-induced effects from experimental variation.
(A) Bar graph representation of inputs from DAT-Cre mice anesthetized with isoflurane and treated with a single injection of saline or fluoxetine. n = 4 for saline, n = 5 for fluoxetine. Two-way ANOVA interaction term p = 0.9365. (B) PCA plot showing PC1 and PC2 of brains from saline or fluoxetine-treated mice. (C) PCA plot showing PC2 and PC3 of brains from saline or fluoxetine-treated mice. (D) Box plot of PC1, p = 0.75. n = 4 for saline, n = 5 for fluoxetine. (E) Box plot of PC2, p = 0.78. (F) Box plot of PC3, p = 0.81. (G) Bar graph representation of inputs from DAT-Cre mice anesthetized with isoflurane and treated with a single injection of saline or fluoxetine (controls), or one of the addictive drugs cocaine, methamphetamine, amphetamine, nicotine, or morphine. n = 9 for control, n = 24 for combined addictive drugs. Two-way ANOVA, interaction term p = 0.0001; unpaired t-tests with Sidak corrections for multiple comparisons DStr p = 0.0051, GPe p = 0.0036. (H) PCA plot showing PC1 and PC2 of brains from control or drug-treated mice. (I) PCA plot showing PC2 and PC3 of brains from control or drug-treated mice. (J) Box plot of PC1, p = 0.78. n = 9 for control, n = 24 for drugs. (K) Box plot of PC2, p = 0.0009. (L) Box plot of PC3, p = 0.024. (M) Heatmap of the contributions of each brain region to PC1–PC3 for data shown in panels H and I.
Figure 4 with 1 supplement
Differences in starter cell location do not explain differences in RABV input labeling patterns between addictive drug and control groups.
(A) Distribution of the starter cell center of mass along the medial–lateral axis for each experimental condition. Saline n = 4, fluoxetine n = 5, cocaine n = 5, methamphetamine n = 4, amphetamine n = 5, nicotine n = 5, morphine n = 5. (B) PC1 vs. PC2 for control vs. drug conditions as shown in Figure 3H, with each point colored according to the medial–lateral coordinate of the center of mass of starter cells for each brain. (C) PC2 vs. PC3 for control vs. drug conditions as shown in Figure 3I, with each point colored according to the medial–lateral coordinate of the center of mass of starter cells for each brain. (D) Linear regression between the x-coordinate of the center of mass of starter cells for each brain vs. PC1. r2 = 0.770, p = 1.62e−7. (E) Linear regression between the x-coordinate of the center of mass of starter cells for each brain vs. PC2. r2 = 0.308, p = 0.082. (F) Linear regression between the x-coordinate of the center of mass of starter cells for each brain vs. PC3, r2 = 0.085, p = 0.638.
Figure 4—figure supplement 1
Example images of starter cell distribution from mice in different treatment groups.
Red reflects expression of TVA-mCherry; green reflects expression of GFP from RABV; yellow reflects overlap. Related to Figure 4A.
Figure 5 with 1 supplement
Dimensionality reduction analysis of brains from animals treated with a single drug exposure.
(A) PCA plot showing PC1 and PC2 of brains from DAT-Cre and GAD2-Cre mice. DAT-Cre mice were anesthetized with K/X and treated with saline or anesthetized with isoflurane and treated with saline or one of the drugs cocaine, methamphetamine, amphetamine, nicotine, morphine, or fluoxetine. GAD2-Cre mice were anesthetized with isoflurane and treated with saline or cocaine. (B) Box plot of PC1, one-way ANOVA p < 0.0001, pairwise t-tests DAT-Cre saline vs. DAT-Cre fluoxetine multiple comparisons adjusted p = 0.99, DAT-Cre saline vs. DAT-Cre K/X p = 1.0, DAT-Cre saline vs. DAT-Cre drugs p = 0.23, DAT-Cre saline vs. GAD2-Cre p < 0.0001, DAT-Cre fluoxetine vs. DAT-Cre K/X p = 1.0, DAT-Cre fluoxetine vs. DAT-Cre drugs p = 0.46, DAT-Cre fluoxetine vs. GAD2-Cre p < 0.0001, DAT-Cre K/X vs. DAT-Cre drugs p = 0.31, DAT-Cre K/X vs. GAD2-Cre p < 0.0001, DAT-Cre drugs vs. GAD2-Cre p < 0.0001. n = 4 for DAT-Cre saline and DAT-Cre K/X, n = 5 for DAT-Cre fluoxetine, n = 24 for DAT-Cre drugs, and n = 17 for GAD2-Cre. (C) Heatmap of the contributions of each brain region to PC1–PC3 for data shown in panel A. (D) Representative images of brain slices from DAT-Cre mice showing, from left to right, the NAc, BNST, GPe, and PBN. Scale, 500 μm. (E) Bar plot showing percentage of RABV-labeled cells in each input region for the DAT-Cre mice shown in panel A. (F) PCA plot showing PC1 and PC2 of brains from DAT-Cre mice anesthetized with isoflurane and treated with cocaine, methamphetamine, amphetamine, nicotine, morphine, fluoxetine, or saline, or anesthetized with K/X and treated with saline. (G) PC2 and PC3 of the same group of brains as shown in panel F. (H) Box plot of PC1, one-way ANOVA p = 0.81. (I) Box plot of PC2, one-way ANOVA p = 0.0093, pairwise t-tests DAT-Cre saline vs. DAT-Cre fluoxetine multiple comparisons adjusted p = 0.94, DAT-Cre saline vs. DAT-Cre K/X p = 0.87, DAT-Cre saline vs. DAT-Cre drugs p = 0.034, DAT-Cre fluoxetine vs. DAT-Cre K/X p = 0.99, DAT-Cre fluoxetine vs. DAT-Cre drugs p = 0.10, DAT-Cre K/X vs. DAT-Cre drugs p = 0.27. (J) Box plot of PC3, one-way ANOVA p = 0.086. (K) Heatmap of the contributions of each brain region to PC1–PC3 for data shown in panels F and G.
Figure 5—figure supplement 1
Brains from animals treated with a single exposure to a drug (Figure 5A), but with the brain identities scrambled.
(A) PCA plot showing the data with scrambled identities along PC1 and PC2. (B) The same scrambled data along PC1 and PC3. (C) UMAP of the scrambled data. (D) Correlogram of 20 UMAP embeddings of the scrambled data.
Figure 6
Projection portraits and UMAP plot of VTADA cells following a single drug exposure.
(A) Projection portrait of the VTA based on inputs from regions with a large negative contribution to PC1, based on data from Figure 5K: NAcCore, NAcLat, DStr, and GPe. (B) Projection portrait based on inputs from regions with a high positive contribution to PC1, based on data from Figure 5K: LHb, DR, VP, PO, EAM, and LH. (C) Projection portrait based on inputs from regions with a high positive contribution to PC2, based on data from Figure 5K: EAM, PBN, DR, EP, LH, GPe, and ZI. (D) Projection portrait based on inputs from regions with a large negative contribution to PC2, based on data from Figure 5K: NAcMed, NAcCore, DStr, Septum, and MHb. (E) UMAP embedding of data from the brains of DAT-Cre mice shown in Figure 5E. (F) Correlogram showing average Euclidean distance of data points over 20 UMAP embeddings of the data presented in panel E.
Figure 7 with 3 supplements
Dimensionality reduction analysis of brains from animals anesthetized with isoflurane and treated with either cocaine or saline, or anesthetized with ketamine/xylazine and treated with saline.
(A) PCA plot of input data from VTADA cells from mice anesthetized either with isoflurane or K/X and given saline. Box plot comparisons of PC1 and PC2 are shown to the right. PC1, p = 0.086, PC2, p = 0.037, unpaired t-tests. n = 4 for both. (B) Bar plot showing percentage of RABV-labeled cells in each input region from mice anesthetized either with isoflurane or K/X and given saline. (C) Heatmap of the contributions from each brain region to PC1–PC3 for data shown in panel A. (D) Projection portrait of regions with high PC1 contributions in all VTADA cells: VP, PO, EAM, PVH, LH, and PBN. (E) Projection portrait of regions with high PC2 contributions in VTADA cells: NAcLat, DStr, GPe, and PVH. (F) UMAP plot of input data from VTADA cells from mice anesthetized either with isoflurane or K/X and given saline. (G) PCA plot of RABV input mapping experiments to VTADA → NAcLat cells in mice anesthetized with isoflurane and given saline or cocaine, or anesthetized with K/X and given saline. Box plot comparisons of PC1 and PC2 are shown to the right. PC1, one-way ANOVA p = 0.0003, pairwise t-tests saline vs. cocaine multiple comparisons adjusted p = 0.011, saline vs. K/X p = 0.071, K/X vs. cocaine p = 0.0002. PC2, one-way ANOVA p = 0.16. n = 4 for saline and K/X, n = 5 for cocaine. (H) Bar plot showing percent of RABV-labeled cells in each input region for cTRIO experiments from VTADA → NAcLat cells. (I) Heatmap of the contributions from each brain region to PC1–PC3 for data shown in panel G. (J) Projection portrait of regions with low PC1 contributions in VTADA → NAcLat cells: cortex, NAcMed, NAcLat, NAcCore, septum, PVH, and MHb. (K) Projection portrait of regions with high PC2 contributions in VTADA → NAcLat cells: VP, PO, EAM, LHb, and LH. (L) UMAP plot of input mapping experiments to VTADA → NAcLat cells. (M) PCA plot of input mapping experiments to VTADA → Amygdala cells. Box plot comparisons of PC1 and PC2 are shown to the right. PC1, one-way ANOVA p = 0.48. PC2, one-way ANOVA p = 0.30. n = 4 for saline, n = 5 for K/X and cocaine. (N) Bar plot showing percent of RABV-labeled cells in each input region for cTRIO experiments from VTADA → Amygdala cells. (O) Heatmap of the contributions of each brain region to PC1–PC3 for data shown in panel M. (P) UMAP plot of input mapping experiments to VTADA → Amygdala cells.
Figure 7—figure supplement 1
Dimensionality reduction analysis of brains from animals anesthetized with isoflurane and treated with either cocaine or saline, or anesthetized with ketamine/xylazine and treated with saline, focusing on PC3.
(A) PCA plot of input data from VTADA cells from mice anesthetized either with isoflurane or K/X and given saline, showing PC2 and PC3. (B) Box plot comparisons of PC3, p = 0.99, unpaired t-tests, n = 4 for both. (C) PCA plot of RABV input mapping experiments to VTADA → NAcLat cells in mice anesthetized with isoflurane and given saline or cocaine, or anesthetized with K/X and given saline, showing PC2 and PC3. (D) Box plot comparisons of PC3, one-way ANOVA p = 0.12. n = 4 for saline and K/X, n = 5 for cocaine. (E) PCA plot of input mapping experiments to VTADA → Amygdala cells, showing PC2 and PC3. (F) Box plot comparisons of PC3, PC1, one-way ANOVA p = 0.014, pairwise t-tests saline vs. K/X multiple comparisons adjusted p = 0.011, saline vs. cocaine p = 0.24, K/X vs. cocaine p = 0.16. n = 4 for saline, n = 5 for K/X and cocaine. (G) PCA plot of input mapping experiments to VTADA → NAcMed cells, showing PC2 and PC3. (H) Box plot comparisons of PC3, one-way ANOVA p = 0.64. PC2, one-way ANOVA p = 0.16, n = 4 for saline and K/X, n = 5 for cocaine. (I) PCA plot of input mapping experiments to VTADA → mPFC cells, showing PC2 and PC3. (J) Box plot comparisons of PC3, one-way ANOVA p = 0.67. PC2, one-way ANOVA p = 0.083. n = 4 for saline and cocaine, n = 5 for K/X.
Figure 7—figure supplement 2
Additional cTRIO data from mice anesthetized with K/X and given saline or isoflurane and given saline or cocaine.
(A) PCA plot of input mapping experiments to VTADA → NAcMed cells. Box plot comparisons of PC1 and PC2 are shown to the right. PC1, one-way ANOVA p = 0.64. PC2, one-way ANOVA p = 0.27. n = 4 for saline and K/X, n = 5 for cocaine. (B) Bar plot showing percent of RABV-labeled cells in each input region for cTRIO experiments from VTADA → NAcMed cells. (C) Heatmap of the contributions of each brain region to PC1–PC3 for data shown in panel A. (D) UMAP plot of input mapping experiments to VTADA → NAcMed cells. (E) PCA plot of input mapping experiments to VTADA → mPFC cells. Box plot comparisons of PC1 and PC2 are shown to the right. PC1, one-way ANOVA p = 0.67. PC2, one-way ANOVA p = 0.97. n = 4 for saline and cocaine, n = 5 for K/X. (F) Bar plot showing percent of RABV-labeled cells in each input region for cTRIO experiments from VTADA → mPFC cells. (G) Heatmap of the contributions of each brain region to PC1–PC3 for data shown in panel E. (H) UMAP plot of input mapping experiments to VTADA → mPFC cells.
Figure 7—figure supplement 3
Inputs to brains of mice anesthetized with isoflurane and receiving saline or cocaine, or K/X-saline (Figure 7), but with the brain identities scrambled.
(A) PCA plot of scrambled data from all VTADA cells including isoflurane-anesthetized and K/X-anesthetized mice. (B) UMAP plot of scrambled data from all VTADA cells including isoflurane-anesthetized and K/X-anesthetized mice. (C) PCA plot of scrambled data from input mapping experiments to VTADA → NAcLat cells. (D) UMAP plot of scrambled data from input mapping experiments to VTADA → NAcLat cells. (E) PCA plot of scrambled data from input mapping experiments to VTADA → Amygdala cells. (F) UMAP plot of scrambled data from input mapping experiments to VTADA → Amygdala cells. (G) PCA plot of scrambled data from input mapping experiments to VTADA → NAcMed cells. (H) UMAP plot of scrambled data from input mapping experiments to VTADA → NAcMed cells. (I) PCA plot of scrambled data from input mapping experiments to VTADA → mPFC cells. (J) UMAP plot of scrambled data from input mapping experiments to VTADA → mPFC cells.
Figure 8 with 3 supplements
GO analysis of top 50 genes within the Allen Gene Expression Atlas correlated with differences in RABV input labeling obtained from experimental vs. control mice.
(A, B) GO from an average of the addictive drug conditions vs. saline-treated controls, all anesthetized using isoflurane. (C, D) GO from K/X- vs. isoflurane-anesthetized saline-treated mice. Panels A and C show molecular function-related gene classes whose expression was positively correlated with RABV labeling differences, and panels B and D show GO analyses that reflect biological process-related gene classes whose expression was negatively correlated with RABV labeling differences.
Figure 8—figure supplement 1
GO analysis of top 50 expressed genes correlated to differences in RABV input labeling obtained from drug-treated vs. control mice.
GO from mice treated with (A) amphetamine, (B) cocaine, (C) methamphetamine, (D) morphine, or (E) nicotine-treated relative to saline-injected controls are shown. Each GO analysis reflects molecular function-related gene classes whose expression was negatively correlated with RABV labeling differences.
Figure 8—figure supplement 2
GO analysis of top 50 expressed genes correlated to differences in RABV input labeling obtained from drug-treated vs. control mice.
GO analyses from mice treated with (A) amphetamine, (B) cocaine, (C) methamphetamine, (D) morphine, or (E) nicotine-treated relative to saline-injected controls are shown. Each GO analysis reflects biological process-related gene classes whose expression was positively correlated with RABV labeling differences.
Figure 8—figure supplement 3
GO analysis of the top 50 genes correlated with differences in RABV input labeling obtained from fluoxetine vs.
saline-treated controls.
Panel (A) shows molecular function-related gene classes whose expression was positively correlated with RABV labeling differences, and panel (B) shows GO analyses that reflect biological process-related gene classes whose expression was negatively correlated with RABV labeling differences.
Figure 9 with 3 supplements
Relationship between RABV labeling ratio in addictive drug/saline-treated groups and mean basal gene expression of gene subgroups, subtypes of ion channels, and neurotransmitter receptors.
Linear regressions are shown for all five examined drugs against different gene classes expressed by brain region within the Allen Gene Expression Atlas, including (A) ion channel-related genes, (B) neurotransmitter-related genes, (C) synapse-related genes, (D) Ca2+ channels, (E) Cl− channels, (F) K+ channels, (G) Na+ channels, (H) glutamate receptors, (I) acetylcholine receptors, (J) GABA receptors, and (K) glycine receptors.
Figure 9—figure supplement 1
Relationship between RABV labeling ratio in addictive drug/saline-treated groups and mean basal gene expression of gene subgroups.
Linear regressions are shown for all five examined drugs against all genes and different gene classes expressed by brain region within the Allen Gene Expression Atlas, including (A) all genes, (B) substance dependence-related genes, (C) endocytosis- and exocytosis-related genes, (D) mitochondrial genes, (E) sugar transporter genes, (F) tumor-associated genes, (G) ligand-gated ion channel genes, (H) voltage-gated ion channel genes, and (I) other ion channel genes.
Figure 9—figure supplement 2
Relationship between RABV labeling ratio in K/X-saline vs. isoflurane-saline groups and mean basal gene expression of gene subgroups.
Linear regressions are shown for K/X-saline vs. isoflurane-saline groups against different gene classes, including (A) ion channel-related genes, (B) synapse-related genes, (C) neurotransmitter-related genes, (D) DEGs related to substance misuse, (E) voltage-gated ion channels, (F) ligand-gated ion channels, (G) other ion channels, (H) K+ channels, (I) Ca2+ channels, (J) Cl− channels, (K) Na+ channels, (L) glutamate receptors, (M) glycine receptors, (N) acetylcholine receptors, and (O) GABA receptor genes.
Figure 9—figure supplement 3
Relationship between RABV ratio labeling in fluoxetine vs. saline-treated groups, both anesthetized with isoflurane, and mean basal gene expression of gene subgroups.
Linear regressions are shown for fluoxetine/saline-treated groups against different gene classes, including (A) ion channel-related genes, (B) synapse-related genes, (C) neurotransmitter-related genes, (D) DEGs related to substance misuse, (E) voltage-gated ion channels, (F) ligand-gated ion channels, (G) other ion channels, (H) K+ channels, (I) Ca2+ channels, (J) Cl− channels, (K) Na+ channels, (L) glutamate receptors, (M) glycine receptors, (N) acetylcholine receptors, and (O) GABA receptor genes.
Figure 10 with 1 supplement
Reduction of Cacna1e expression in the NAcLat lowers the number of RABV-labeled inputs from the NAcLat onto VTADA cells.
(A) Schematic of viral injections performed. (B) Schematic of connectivity in relevant brain regions and explanation of the normalized input metric. (C) Representative image of fluorescence in starter cells in the VTA. Main image scale, 1 mm; inset scale, 100 μm. (D) Representative image of fluorescence in the NAcLat after either administration of no guide RNA (gRNA) (left) or gRNA targeting Cacna1e (right). Main image scale, 1 mm; inset scale, 100 μm. (E) qPCR results showing slight reduction of Cacna1e in the NAcLat. (F) Difference in RABV-labeled inputs after CRISPRi-mediated knockdown of Cacna1e compared to controls (Beier et al., 2017), and no gRNA. The control conditions are based on previously published data where no AAVs were injected into NAcLat (Beier et al., 2015).
Figure 10—figure supplement 1
AGEA data showing expression of Cacna1e.
Shown are both coronal (A–C) and parasagittal (D–F) sections. The NAc is highlighted in panels B and E. The colors in panels C and F reflect the expression energy of Cacna1e; the color bar is shown to the right of panel F. Expression of Cacna1e is relatively uniform and high throughout the striatum, including the NAc. All panels are reproduced from the Allen Mouse Brain Atlas, under the tab for ISH Data (see here; Lein et al., 2007). (A, B) are showing ISH signal from experiment 74988669 (see here and here), and also use the Allen’s Mouse P56 coronal reference atlas. (C) is showing a quantification of expression from experiment 74988669. (D, E) are showing ISH signal from experiment 69236897 (see here and here), and also use the Allen’s Mouse P56 sagittal reference atlas. (F) is showing a quantification of expression from experiment 69236897.
© 2008, Allen Institute for Brain Science. Images are from the Allen Mouse Brain Atlas, 2008 (https://mouse.brain-map.org/), and are not available under the terms of a Creative Commons Attribution License. Further reproductions of these images should adhere to the Allen Institute’s Citation policy (https://alleninstitute.org/legal/citation-policy/).
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Gene (Mus musculus, males and females) | Cacna1e | NCBI | NM_009782.3 | |
| Strain, strain background (Mus musculus, males and females) | C57BL/6J | The Jackson Laboratory | Strain #: 000664, RRID:IMSR_JAX:000664 | |
| Strain, strain background (Mus musculus, males and females) | vGluT2-Cre | The Jackson Laboratory | Strain #: 016963, RRID:IMSR_JAX:016963 | |
| Strain, strain background (Mus musculus, males and females) | DAT-Cre | The Jackson Laboratory | Strain #: 006660, RRID:IMSR_JAX:006660 | |
| Strain, strain background (Mus musculus, males and females) | GAD2-Cre | The Jackson Laboratory | Strain #: 028867, RRID:IMSR_JAX:028867 | |
| Cell line (Homo sapiens) | HEK293T | ATCC | CRL-3216 RRID:CVCL_0063 | |
| Strain, strain background (adeno-associated virus) | AAV5-CAG-FLExloxP-TC | University of North Carolina, vector core | Titer: 2.4 × 1012 genome copies (gc)/ml | |
| Strain, strain background (adeno-associated virus) | AAV8-CAG-FLExloxP-G | University of North Carolina, vector core | Titer: 1.0 × 1012 gc/ml | |
| Strain, strain background (adeno-associated virus) | AAV5-CAG-FLExFRT-TC | University of North Carolina, vector core | Titer: 2.6 × 1012 gc/ml | |
| strain, strain background (adeno-associated virus) | AAV8-CAG-FLExFRT-G | University of North Carolina, vector core | Titer: 1.3 × 1012 gc/ml | |
| Strain, strain background (canine adenovirus) | CAV-FLExloxP-Flp | Plateforme de Vectorologie de Montpellier, France | Titer: 5.0 × 1012 gc/ml | |
| Strain, strain background (Lyssavirus rabies) | RABVΔG GFP | Made in lab | Titer: 5.0 × 108 colony forming units (cfu)/ml | |
| Strain, strain background (adeno-associated virus) | AAV9-CamKII-0.4-Cre-SV40 | Addgene | 05558-AAV9 | Titer: 1.5 × 1013 gc/ml |
| Recombinant DNA reagent | pJEP317 pAAV-U6SaCas9gRNA(SapI)-EFS-GFP-KASH-pA | Addgene | Plasmid #113694, RRID:Addgene_113694 | |
| Recombinant DNA reagent | pJEP317-pAAV-U6-Cacna1eAC-EFS-GFP-KASH-pA | Made in lab | ||
| Recombinant DNA reagent | pHelper | A gift from Matthew Banghart at the University of California, San Diego | ||
| Recombinant DNA reagent | pAAV2-8 | Addgene | Plasmid #112864, RRID:Addgene_112864 | |
| Recombinant DNA reagent | pAAV2-5 | Addgene | Plasmid #232922, RRID:Addgene_232922 | |
| Commercial assay or kit | Q5 Site-directed mutagenesis kit | New England Biolabs | E0554S | |
| Commercial assay or kit | Luna Universal One-Step RT-qPCR Kit | New England Biolabs | E3005L | |
| Chemical compound, drug | PEIMAX | Polysciences | 24765-100 | |
| Chemical compound, drug | PEG 8000 | Fisher Scientific | AA4344336 | |
| Chemical compound, drug | DAPI | Thermo Fisher Scientific | D1306 | |
| Chemical compound, drug | Isoflurane | Patterson Veterinary | 78938441 | |
| Chemical compound, drug | Ketamine | Patterson Veterinary | 78935790 | |
| Chemical compound, drug | Xylazine | Patterson Veterinary | 78945244 | |
| Chemical compound, drug | Vetameg | Patterson Veterinary | 78924375 | |
| Chemical compound, drug | Vetbond | Patterson Veterinary | 78055031 | |
| Chemical compound, drug | Fluoxetine | Tocris Bioscience | 0927 | |
| Chemical compound, drug | Cocaine | Sigma | C5776-1G | |
| Chemical compound, drug | Morphine | Patterson Veterinary | 78924699 | |
| Chemical compound, drug | Methamphetamine | Cayman Chemical company | 13997 | |
| Chemical compound, drug | Amphetamine | Sigma | A5880-1G | |
| Chemical compound, drug | Nicotine | Tocris Bioscience | 3546 | |
| Software, algorithm | Python | Version 3.11 | ||
| Other | Superfrost Plus microscope slides | Fisher Scientific | 1255015 | |
| Other | Coverslips | Thermo Fisher Scientific | 152460 | |
| Other | Fluoromount-G | SouthernBiotech | 0100-01 | |
| Other | TRIzol | Thermo Fisher Scientific | 15596026 |
Additional files
-
MDAR checklist
- https://cdn.elifesciences.org/articles/93664/elife-93664-mdarchecklist1-v1.pdf
-
Supplementary file 1
List of experimental conditions included in this manuscript for brain-wide RABV tracing analysis.
- https://cdn.elifesciences.org/articles/93664/elife-93664-supp1-v1.xlsx
-
Supplementary file 2
Table of statistical comparisons performed in this manuscript.
- https://cdn.elifesciences.org/articles/93664/elife-93664-supp2-v1.xlsx
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Drug-induced changes in connectivity to midbrain dopamine cells revealed by rabies monosynaptic tracing
eLife 13:RP93664.
https://doi.org/10.7554/eLife.93664.3
