% Group meeting paper template - (c) Dan O'Shea 2021 @djoshea
% This is a template for presenting figures from a paper enabling zooming
% into indivdual figure panels and quoting the caption below.
%
% Instructions:
% - insert the full size figure first, and commit to its size. Note that we refer to the figure file in 3 locations in order to get dimmed versions of it with spy.
% - uncomment the grid lines to see where the subpanels end up
% - the tikz spy on nodes are used to zoom into individual panels. The size is the magnified size (times the magnification)
% and the coordinates for on (x, y) are for the center of the rectangle. generally it's easiest to get the center location first and then tweak the size
% - you can tweak the frametitle and caption nodes to appear on specific slides to present one or more panels simultaneously.
% - After getting the spy on nodes working, it's basically copy and paste from the figure captions.
% - You can also use beamerbutton links to jump to supplemental figures as I do here.
%
% Hope you find this helpful. Enjoy!
% Twitter/Github: @djoshea
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\title[]{A midbrain - thalamus - cortex circuit reorganizes cortical dynamics to initiate planned movement}
\author[]{\texorpdfstring{%
Hidehiko K. Inagaki, Susu Chen, Margreet C. Ridder, Pankaj Sah, \\ Nuo Li, Zidan Yang, Hana Hasanbegovic, Zhenyu Gao, \\ Charles R. Gerfen, Karel Svoboda \\[5mm] \href{https://www.biorxiv.org/content/10.1101/2020.12.16.423127v1}{bioR$\chi$iv doi:10.1101/2020.12.16.423127}%
}{}}
\institute[]{\bf{NPSL / CTD Group Meeting}}
\date[]{\today}
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%%%%%%%%%% DOCUMENT BEGINS HERE %%%%%%%%%%
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\section{Figure 1}
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\centering
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\node<1>[figtitle] at (current page.north west) {Activity modes for motor planning and movement initiation in anterior lateral motor cortex};
\node<2>[caption] at (current page.south) {A. Tactile delayed-response task. An object (pole, red / blue circle) is presented within reach of the whiskers during the sample epoch to instruct lick direction.};
\node<3>[caption] at (current page.south) {B. Side view of a behaving mouse recorded with high-speed videography (400 Hz). Left, trajectories of nose (blue), tongue (green) and jaw (red) movement overlaid on an image at time 0 ms (onset of the Go cue). Middle, the first frame at which the tongue appears.};
\node<4>[caption] at (current page.south) {C. Example neurons in ALM. Top, spike raster. Bottom, mean spike rate. Blue, correct lick right trials; red, correct lick left trials. Time is aligned to the timing of the Go cue. Dashed lines separate behavioral epochs. S, sample epoch; D, delay epoch; R, response epoch.};
\node<5>[caption] at (current page.south) {D. Pearson’s Correlation of the population activity vector in ALM (bin: 10 ms; n = 5226 neurons). Dashed lines separate behavioral epochs. White dotted square, correlation between time points in the delay epoch; magenta dotted square, correlation between time points in the delay and the response epoch.};
\node<6>[caption] at (current page.south) {E. Projections of activity along CDdelay, CDgo, and GD. Line, median. Shade, S.E.M.};
\node<7>[caption] at (current page.south) {F. Selectivity explained by each direction in activity space. Black, square sum of selectivity of all recorded ALM neurons; Green, magenta, and cyan, the square sum of selectivity along CDdelay, CDgo, and their sum, respectively.};
\end{tikzpicture}
\end{frame}
\section{Figure 2}
\begin{frame}[label=fig2]
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\centering
\vspace{-5mm}
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\node<1>[figtitle] at (current page.north west) {ALM motor commands are necessary for movement initiation};
\node<2>[caption] at (current page.south) {A. Silencing ALM neurons that project to orofacial motor centers in the medulla (PTlower). Inset, coronal section showing PTlower neurons in ALM expressing GtACR1 fused with FusionRed; Blue, DAPI.};
\node<3>[caption] at (current page.south) {B. Raster plot of lick timing in an example animal (75 trials shown per trial type). Cyan box, laser on.};
\node<4>[caption] at (current page.south) {C. Proportion of licks within 600 ms after the Go cue. Blue, lick right trials; red, lick left trials. Circle, mean; lines, each animal, n = 4 mice. P < 0.001 in both lick right and left trials (hierarchical bootstrap with a null hypothesis that proportion of trials with licks in silencing trials are the same or higher than that in control).};
\node<5>[caption] at (current page.south) {D. Schema showing PTlower cells and other cell types (e.g. ‘PTupper’, brainstem-projecting neurons that do no innervate the medulla; intratelencephalic cells) (Li et al., 2015; Economo et al., 2018)analyzed in Figure 2 and S2. PTlower cells (magenta) in the deep layers of ALM project to medulla. They indirectly inhibit PTlower-inhibited cells (green).};
\node<6>[caption] at (current page.south) {E. Spike rate of individual neurons with or without PTlower silencing. Circle, individual neuron; magenta; significantly decreased neurons (putative PTlower cells); green, significantly increased neurons (putative PTlower inhibited cells).
};
\node<7>[caption] at (current page.south) {F. Example putative PTlower cell and putative PTlower-inhibited cell. Top, spike raster. Middle, PSTH in control trials. Bottom, PSTH in trials with PTlower silencing. Blue, all lick right trials; red, all lick left trials: cyan bar, laser on.};
\node<8>[caption] at (current page.south) {G. Projection of activity along CDdelay, CDgo, and GD with and without PTlower silencing. Line, grand median across sessions (n = 24 sessions; 4 mice); shading, S.E.M. (hierarchical bootstrap); cyan bar, laser on.};
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\section{Figure 3}
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\node<1>[figtitle] at (current page.north west) {Short latency Go cue signals in ALM-projecting thalamus};
\node<2>[caption] at (current page.south) {A. Short-latency Go cue responses of ALM (top) and thalALM neurons (bottom) sorted by their latency. Neurons with increases (left, go-up cells) or decreases (right, go-down cells) in spike rate are shown separately. The fastest 20\% and 3\% cells are shown for go-up and go-down cells, respectively. Spike rates are normalized by the baseline (spike rate before the Go cue, 100 ms window) for each neuron.};
\node<3>[caption] at (current page.south) {B. Cumulative distribution function (c.d.f.) of latency to the Go cue across neurons in ALM and thalALM. };
\node<4>[caption] at (current page.south) {C. Recordings in thalamus. Top, recording sites in Allen Common Coordinate Framework (CCF). Each colored region corresponds to a different thalamic nucleus. White contour, thalALM. Black dots, location of individual recorded neurons. Green, neurons with < 20 ms latency. Middle, the density of neurons with latencies < 20 ms. Bottom, the density of neurons with delay selectivity. AP, posterior to Bregma.};
\node<5>[caption] at (current page.south) {D. Recording in thalALM during ALM silencing. Left, schema. Right, mean activity of thalALM with or without ALM silencing. Cyan bar, photoinhibition of ALM.};
\node<6>[caption] at (current page.south) {E. Spike rate during the delay epoch in thalALM with or without ALM silencing. Circle, individual neuron in thalALM (n = 58 cells). Filled circle, significantly modulated cells (p < 0.01, ranksum test). Cross, median activity. P-value, signrank test comparing spike rate across neurons with or without silencing. \\ F. The amplitude of Go cue activity (spikes per s; change in spike rate after the go cue; 100 ms window) in thalALM with or without ALM photoinhibition. The same format as in E.};
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\section{Figure 4}
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\node[figtitle] at (-3.5cm, 9cm) {PPN/MRN projects to ALM-projecting thalamus};
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\begin{frame}{fig. 4 --- PPN/MRN projects to ALM-projecting thalamus}
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A. Projections from ipsilateral PPN/MRN to thalALM (coronal view). Top, Allen Reference Atlas. Each colored region corresponds to a different thalamic nucleus. White contour, thalALM. Bottom, the intensity of projection from PPN/MRN, registered to the Allen CCF (mean of 4 mice). AP, posterior to Bregma.
B. Quantification of anterograde labeling in thalALM from subcortical structures (Methods).
C. Top, projection-type non-specific labeling of PPN/MRN neurons. Bottom, signal at the injection site, in the thalamus, and in the medulla. The image gains and contrasts are identical between the images of the thalamus and the medulla. Green, YFP; Blue, Nissl staining.
D. Top, labeling thalamus-projecting PPN/MRN neurons. Bottom, same as in C for thalamus-projecting PPN/MRN neurons. Red, retrograde tracing from ALM.
E. Enlarged image of the thalamus in D.
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\section{Figure 5}
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\node<1>[figtitle] at (current page.north west) {Short latency Go cue signals in PPN/MRN};
\node<2>[caption] at (current page.south) {A. Recordings in the midbrain. Top, recording sites in Allen CCF. Each colored region corresponds to a different midbrain nucleus. Black dots, location of individual recorded neurons. Green, location of neuron with <15 ms latency. Bottom, the density of neurons with latency to the Go cue shorter than 15 ms. White contour indicates MRN and PPN. CUN, cuneiform nucleus. PRN, pontine reticular nucleus.};
\node<3>[caption] at (current page.south) {B. Four example PPN/MRN neurons. Top, location of recorded neuron (green circle) in the Allen CCF. Middle, spike raster. Bottom, mean spike rate.};
\node<4>[caption] at (current page.south) {C. Go cue response of PPN/MRN sorted by their latency to the Go cue. Neurons with an increase or decrease in spike rate after the Go cue are shown separately. Spike rates are normalized by the baseline (spike rate before the Go cue, 100 ms window) for each neuron.};
\node<5>[caption] at (current page.south) {D. Cumulative distribution (c.d.f.) of latency to the Go cue across neurons in each brain area (see Methods for number of neurons analyzed). Each color indicates a different brain area (box next to F).
E. Top, projection of activity along GD in each brain area. Bottom, the latency of post-Go cue increases in activity along GD (Methods). Central line in the box plot is the median. Top and bottom edges are the 75\% and 25\% points, respectively. The whiskers show the lowest datum within 1.5 interquartile range (IQR) of the lower quartile, and the highest datum within 1.5 IQR of the upper quartile.
F. Same as E for activity along CDgo.};
\node<6>[caption] at (current page.south) {G. Increase in spike rate of PPN/MRN neurons in response to the Go cue and different tone (top) or no sound at the expected timing of the Go cue (bottom). Circles, neurons (n = 178).};
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\section{Figure 6}
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\frametitle{fig. 6\only<2>{a,b,c}\only<3>{d}\only<4>{e}}
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\centering
\vspace{-5mm}
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\node<1>[figtitle] at (current page.north west) {Stimulation of thalamus-projecting PPN/MRN neurons triggers planned movement};
\node<2>[caption] at (current page.south) {A. Schema of PPN/MRNth stimulation experiment.
B. Proportion of trials with lick after PPN/MRNth stimulation. Circle, mouse (n = 20 mice). Filled circle, mice with unilateral virus injection (n = 4 mice). P-value, hierarchical bootstrap with a null hypothesis that the proportion of trials with licks in stimulation trials are the same or lower than that in control.
C. Same as B for the proportion of trials with correct lick direction (correct rate) after the Go cue (control) or stimulation.};
\node<3>[caption] at (current page.south) {D. Top, cumulative distribution of the first tongue detection (with high-speed videography) after time 0. Dotted line, data in Go cue omitted condition for comparison. Bottom, jaw movement (black), and nose movement (green). Trials are classified as follows: Go cue, trials with the Go cue; Go cue omitted, trials without the Go cue or stim; Stim w.o. lick, trials with stimulation but without lick; Stim followed by lick, trials with stimulation followed by lick. Tongue detection onset: (Go cue) 64.3 (56.0-75.0) ms; mean (2.5-97.5\% confidence interval); (stim followed by lick) 75.5 (50.0-118.0) ms; p = 0.194 (hierarchical bootstrap). Jaw movement onset: (Go cue) 33.2 (20.0-42.5) ms; (stim followed by lick.) 69.7 (8.8-102.5) ms; p = 0.114 (hierarchical bootstrap). Nose movement onset: (Go cue) 43.0 (32.5-50.0) ms; (stim followed by lick) 61.9 (7.5-117.1) ms; p = 0.139 (hierarchical bootstrap). The null hypothesis for p-value is that the onset of Go cue trials is shorter than that in stim trials.};
\node<4>[caption] at (current page.south) {E. Projection of activity along CDdelay (left), CDgo (middle), and GD (right) across trial types. Cyan dashed line, photo-stimulation. Line, grand median of sessions (n = 21 sessions; 12 mice); shading, S.E.M. (hierarchical bootstrap).};
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\section{Figure 7}
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\centering
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\node<1>[figtitle] at (current page.north west) {Activity in thalamus-projecting PPN/MRN neurons is required for cue-triggered movement initiation};
\node<2>[caption] at (current page.south) {A. Perturbation of PPN/MRNth before and during the Go cue.
B. Behavioral effects of perturbing Th-projecting hsyn+ neurons (left; n = 4 mice), Th-projecting CamKII+ neurons (middle; n = 4 mice), and Chat+ neurons (right; n = 2 mice; note Chat+ cells are not necessarily projecting to thalamus) in PPN/MRN. P-value, hierarchical bootstrap with a null hypothesis that the proportion of trials with licks in perturbation trials are the same or higher than that in control.};
\node<3>[caption] at (current page.south) {C. Go cue response sorted by their latency to the Go cue. Neurons with increase in spike rate before movement (within 50 ms after the Go cue) are shown (45/292 cells in PPN/MRN and 44/635 cells in ALM). Activities of the same neurons in control (left) and perturbation (right) trials. Spike rates are normalized by baseline (spike rate before the Go cue in control trials, 100 ms window). In C-F, data of perturbing Th-projecting CamKII+ neurons is shown.};
\node<4>[caption] at (current page.south) {D. Projection of activity along CDdelay, CDgo, and GD across trial types. Cyan, laser on. Line, grand median of sessions (n = 17 sessions; 4 mice); shading, S.E.M. (hierarchical bootstrap).};
\node<5>[caption] at (current page.south) {E. Trial-by-trial relationship between proportion of trials with lick, activity along GD at the Go cue (go cue response) and at the stim onset (onset response). Left, schema of activity analyzed in the regression analysis. Mean activity within the green dotted lines were analyzed (window size, 200 ms). Right, estimated coefficients of logit regression. P-value, hierarchical bootstrap (n = 17 sessions; 4 mice).
F. Correlation between activity along GD at the Go cue and at the stim onset. P-value, hierarchical bootstrap with a null-hypothesis that coefficient is lower than 0 (n = 17 sessions; 4 mice).};
\node<6>[caption] at (current page.south) {G. Multi-regional flow of information underlying the cue-triggered movement initiation. Left, preparatory activity (CDdelay) is maintained in a cortico-thalamocortical loop. Middle, the Go cue (speaker) activates PPN/MRN, which then activates neurons in thalALM, which are different from neurons that maintain preparatory activity (green circles). This induces activity along GD in ALM. Right, the GD activity then causes a collapse of activity along CDdelay and an emergence of motor command (CDgo), which engages medulla (Med.) circuits to initiate planned movements.};
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\section{Supplemental Figures}
\subsection{Supp. Figure 1}
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