Large-scale 3D model of the dorsal striatum.

(a) Self-enveloped simulation space of 1003 µm3 with approximately 40.000 release sites from 150 neurons. Colours of individual release sites are not matched to neurons. (b) Simulation of a single release event after 5 ms and 10 ms. Colour-coded by DA concentration. (c) Comparison of analytical solution and simulation of diffusion after a single release event at three different time points. (d) Representative snapshot of steady state DA dynamics at 4 Hz tonic firing with parameters mirroring the dorsal striatum. (e) Cross-section of temporal dynamics for data shown in (d). (f) Histogram of DA concentrations ([DA]) across the entire space in (d). (g) DA release during three burst activity scenarios for all release sites in a 10×10×10 µm cube (black boxes) and spill-over into the surrounding space. Burst simulated as increase in firing rate on top of continued tonic firing of the surrounding space. Traces on top are average DA concentrations for the marked cubes, with bursts schematized by coloured lines below. First image row is at the end of the burst, second row another 100 ms after. Scale bars for traces are 200 ms and 500 nM. Scale bar for the images is 20 µm. (h) Peak DA concentration reached at different distances from area with phasic activity for the three firing scenarios. (i) Top: representative [DA] trace 1 µm away from a release site during pacemaker and burst activity. Bottom: Occupancy of D1Rs and D2Rs for same site. (j) Effect of complete pause in firing for 1 second on both average [DA] and D1R and D2R occupation.

List of variables used in simulation of the dorsal striatum.

Regional difference in uptake greatly impact DA dynamics.

(a) Representative snapshots of steady state dynamics at 4 Hz tonic firing with parameters mirroring the dorsal (left) and ventral striatum (right). (b) Cross-section of temporal dynamics for data shown in a. Bottom row show concentrations of the dashed lines in the top panels. (c) Normalized density of DA concentration of simulations in (a). Thick lines are for the entire space, thin lines are across time for five randomly sampled locations. Dashed red line is for simulation of the ventral striatum with lowest reported innervation density in the literature. (d) Same data as in (c), but for concentration percentiles. Note that even the lowest percentiles of VS were above 10 nM in [DA]. (e) Convoluted model response (Figure S1c) to mimic FSCV measurements mirroring the experimentally tested stimulation paradigm in (May & Wightman, 1989) for the dorsal (left) and ventral striatum (right) (f) DA release during three burst activity scenarios for all release sites in a 10×10×10 µm cube (black boxes) and spill-over into the surrounding space. Burst simulated as increase in firing rate on top of continued tonic firing of the surrounding space. Traces on top are average DA concentrations for the marked cubes, with bursts schematized by coloured lines below. First image row is at the end of the burst, second row another 100 ms after. Scale bars for traces are 200 ms and 500 nM. Scale bar for the images is 20 µm. (g) Top: representative [DA] trace 1 µm away from a release site during pacemaker and burst activity. Bottom: Occupancy of D1Rs and D2Rs for same site. Occupancy data from corresponding DS simulation on Figure 1k shown in as dotted line. (h) Peak occupancy at different distances from the area bursting normalized to maximal and minimum occupancy.

Sensitivity of the model to parameter changes.

(a) Schematic of the fraction of active release sites. Black dots are inactive sites and green dots indicate actively releasing sites. (b) Effect of changing fraction of active release sites on DA concentrations. Blue line, DS peak DA concentration (99.5th percentile); Red line, VS peak DA concentration (99.5th percentile); Dotted blue line, DS tonic DA concentration (50th percentile); Dotted red line, VS tonic DA concentration (50th percentile). (c) Ratio between peak (99.5th percentile) and tonic (50th percentile) concentrations across fractions of active release sites in the DS (blue line) and VS (red line) as a measure of DA signal focality. (d) Schematic of changing quantal size (Q). (e) Effect of changing quantal size on tonic and peak DA concentrations in DS (blue lines) and VS (red lines). (f) Ratio between peak and tonic concentrations across various quantal sizes in in DS (blue line) and VS (red line). (g) Relative difference between the DS and VS for peak (black line) and tonic DA (dotted line) at different quantal sizes. (h) Schematic of changing DAT Km. (i) Effect of changing DAT Km on DA concentrations in DS (blue lines) and VS (red lines). j, Schematic of changing DAT Vmax. (k) Effect of changing DAT Vmax on DA concentrations. Shaded areas are median Vmax of the two regions (DS and VS) as found in the literature shown in Table S2 ± 50%. (l) Effect of changing DAT Vmax, with tonic (50th percentile) and peak (99.5th percentile) DA concentrations normalized to their value at 2 µm s-1 (median value for VS). Shaded area indicates median Vmax for VS found in the literature shown in Table S2 ± 50%.

DAT nanoclustering reduces uptake and shows regional variation.

(a) Schematic of dense DA cluster. White dots represent individual DAT molecules, and colour gradient the surrounding DA concentration. (b) Effective transport rate dependent on local concentration. (c) Top view of unfolded DA varicosity. Black shapes denote clusters of DAT. Dashed white line indicates placement of cross-section shown in (d). (d) Cross section showing DA concentration in space surrounding varicosity unfolded in c. Grey line at the bottom is surface of the varicosity. Colour-coded for DA concentration. (e) Top view from (c), but colour coded for DA concentration immediately above membrane surface at different DAT cluster sizes. (f) Mean [DA] over time for different DAT cluster sizes and unclustered (Un.). (g) Time to reach a mean DA concentration of 10 nM for different DAT cluster sizes and unclustered (Un.). (h) Difference between DA concentration at the centre of clusters and mean concentration throughout the simulation for different cluster sizes and unclustered (Un.). (i) Concentrations across a cross section of a surface with 80 nm diameter clusters. Shaded areas highlight cluster locations. (j) Location of images of the dorsal (DS) and ventral striatum (VS) in striatal slices from mice as imaged in (Sorensen et al., 2021) with direct stochastic optical reconstruction microscopy (dSTORM). (k) Two representative DA varicosities from DS and VS with VMAT2 in white and DAT in magenta. Images are 1.5×2 µm. (l) Individual DAT localizations from images in (k) coloured by clustering. Black indicates localization identified as clustered based on DBSCAN with parameters 80 nm diameter and 40 localizations. Grey indicates unclustered localizations. (m) Quantification of clustering across all images in (j) with parameters in (l). Welch’s two-sample t-test, P = 0.012(*), n = 12 (DS) and 13 (VS). (n) Absolute difference in percentage of clustering as assessed with DBSCAN across a range of parameters. VS has a higher propensity to cluster across cluster sizes typically reported for DAT clusters.