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. Author manuscript; available in PMC: 2019 Feb 8.
Published in final edited form as: Cell. 2018 Feb 1;172(4):706–718.e15. doi: 10.1016/j.cell.2018.01.008

Figure 5. RIM organizes release sites in dopamine neurons.

Figure 5

(A) Representative confocal images of VMAT2 and TH staining of the dorsal striatum in RIM control and RIM cKODA mice.

(B) Quantification of fluorescence intensity, size and density of VMAT2 and TH staining for images shown in (A). Each dot represents the mean of a single animal (average of 6–8 images). RIM cKODA values were normalized to their corresponding RIM control. For VMAT2, RIM control n = 3 mice, RIM cKODA, n = 3. For TH, RIM control n = 4, RIM cKODA n = 4.

(C) Quantification of tissue dopamine levels in the striatum measured by ELISA from mice after intraperitoneal injection of reserpine or DMSO two hours before the tissue harvest. Each dot is an average of two measurements from one animal. For DMSO, RIM control n = 4 mice, RIM cKODA n = 4. For reserpine, RIM control n = 5, RIM cKODA n = 3.

(D) Quantification of in vivo extracellular dopamine levels in the striatum measured by microdialysis in anesthetized mice before and during reverse dialysis of TTX. Dopamine levels are expressed normalized to the average in RIM control before reverse dialysis of TTX. RIM control n = 5 mice, RIM cKODA n = 5 (p < 0.001 for baseline levels, not significant for levels after TTX).

(E) Representative surface rendered images (5 × 5 × 2 μm3) of bassoon clusters within dopamine axons from RIM control and RIM cKODA mice.

(F) Quantification of density and size of bassoon clusters within dopamine axons in RIM control and RIM cKODA mice.

(G) Representative STED images of bassoon within dopamine synaptosomes from RIM control and RIM cKODA mice. Vesicles in dopaminergic neurons were labeled by crossing Cre-dependent synaptophysin-tdTomato mice (SYP-tdTomato) with DATIRES-Cre mice. For analysis of SYP-tdTomato expression in dopamine axons, and for analyses of Munc13 and Munc18 clusters, see Figure S5.

(H) Quantification of the bassoon area in (G). Each circle is the average of an image with 600–2100 synaptosomes. RIM control n = 14 images/3 mice, RIM cKODA n = 18/3.

All data are mean ± SEM. *** p < 0.001, **, p<0.01, ns, not significant; Mann-Whitney rank sum test for (B, F), Kruskal-Wallis analysis of variance with post hoc Dunn’s test for (C, D, H).