Connectomic analysis of the Drosophila lateral neuron clock cells reveals the synaptic basis of functional pacemaker classes
- Advanced Science Research Center, The City University of New York, United States
- Center for Theoretical Neuroscience, Zuckerman Institute, Columbia University, United States
- Department of Neuroscience and Behavior, Barnard College, United States
- Department of Electrical Engineering, Columbia University, United States
Figures
Figure 1
The circadian clock neuron network and identified clock neurons in the hemibrain.
(A) The circadian clock network is shown in both hemispheres. Currently identified cells are shown in color (s-LNvs red, l-LNvs light red, LNds in orange, DN1a in purple, a subset of DN1p in blue) and known clock cells not yet identified within the hemibrain connectome data are shown in gray (DN2, DN3, and a subset of DN1p). The pars intercerebralis (PI) and optic lobe (OL) are indicated in gray. (B) Identified clock neurons in the hemibrain. Neuropils are shown in gray. Color codes as indicated in (A). (C) Heatmap indicates synaptic connections strength among all identified circadian clock neurons, including weak (weight <3) connections. (D) The timekeeping lateral neurons that compose the morning (M) and evening (E) Oscillators: s-LNvs (M cells), and 5th LNv and LNds (E cells).
Figure 2 with 1 supplement
Connectivity patterns of the s-LNvs.
(A–C). Synaptic connections of the four pdf + s LNvs, s-LNv_R_1 through s-LNv_R_4. Neuronal morphology is shown in gray. In A and B, inputs to the s-LNvs are shown in blue, outputs are shown in magenta. (A) All connections, including non-clock cells, (B) Connections to/from clock cells only, excluding connections to s-LNvs, (C) Connections among the four pdf + s LNvs. Input and output sides coincide and are indicated in orange. (D–E) Sankey diagram indicating the strong synaptic partners of all s-LNvs, including the 5th LNv. The total weight of synapses formed by each cell with its inputs (D) or outputs (E) is shown. (D) Presynaptic partners (inputs) of s-LNvs. No shared connections were found between the pdf + s LNvs and the 5th LNv. (E) Post synaptic partners (outputs) of s-LNvs. Only one cell (Body ID 571372889) receives synaptic input from the 5th LNv plus a pdf + s LNv. (F–G) Synaptic connections of the 5th LNv. (F) All connections, including non-clock cells, (G) connections to clock cells only. (H) Connectivity map of the four pdf + s LNvs indicating their peptide and receptor expression.
Figure 2—figure supplement 1
Connections of individual s-LNvs.
(A–D). Synaptic connections of each of the four pdf + s LNvs, s-LNv1 (A), s-LNv2 (B), s-LNv3 (C), and s-LNv4 (D), either to all neurons (left columns), to other clock cells except s-LNvs (middle columns), or to other s-LNvs (right columns). Neuronal morphology is shown in gray. Inputs to the s-LNvs are shown in blue, outputs are shown in magenta. Body IDs of each s-LNv are indicated on the right.
Figure 3 with 2 supplements
Connectivity patterns of the LNds.
(A–C). Synaptic connections of the six LNds. Neuronal morphology is shown in gray. In A and B, inputs to the s-LNvs are shown in blue, outputs are shown in magenta. (A) All connections, including non-clock cells, (B) Connections to clock cells only, excluding connections to LNds. Input and output sides coincide and are indicated in orange. (C) Connections within LNds. (D) Connectivity map of the six LNds indicating their peptide and receptor expression. (E–F) Sankey diagram indicating the strong synaptic partners of all LNds. The total weight of synapses formed by each cell with its inputs (E) or outputs (F) is shown. (E) Presynaptic partners of LNds. No shared connections were found between LNd1-3 and LNd4-6. Most of the shared connections are between LNd4 and LNd5 (X cells provide strong inputs to both). (F) Postsynaptic partners of the LNds.
Figure 3—figure supplement 1
Connections of individual LNds.
(A–F). Synaptic connections of each of the six LNds, LNd1 (A), LNd2 (B), LNd3 (C), LNd4 (D), LNd5 (E), and LNd6 (F) either to all neurons (left columns), to other clock cells except LNds (middle columns), or to other LNds (right columns). Neuronal morphology is shown in gray. Inputs to the LNds are shown in blue, outputs of the LNds are shown in magenta. Body IDs of each LNd are indicated on the right.
Figure 3—figure supplement 2
Connections of E cell classes.
(A–C). Synaptic connections of the three E cell classes, E3a (A), E1 (B), and E2 (C) either to all neurons (left columns), to other clock cells except LNds (middle columns), or to other LNds (right columns). LNd1 alone (E3b) is shown in Figure 2—figure supplement 1. Neuronal morphology is shown in gray. Inputs to the E classes are shown in blue, outputs are shown in magenta (left and middle columns). Within each group (right columns), input and output sides coincide and are indicated in green.
Figure 4 with 2 supplements
M cells are homogeneous while E cells can be clustered in three distinct groups.
(A–B) Jaccard indices for overlap in synaptic partners of M and E cells. Only includes synaptic partners that make medium or strong connections. Higher index values indicate more similarity in either inputs (A) or outputs (B). (C) Total input and output Synapse counts for M and E cells. (D–E) Strong shared connections of the four pdf +s LNvs. Only cells that share one connection with at least two M cells are shown. The strength (weight) of the connection is indicated. Only medium and strong connections are included. (D) The two cells that send strong connections to at least two M cells send strong connections to all four. (E) The six cells that receive strong connections from at least two M cells receive strong connections from all four. (F–G) Strong shared connections of the six LNds plus the 5th LNv (collectively referred to as E cells). Only cells that share a strong connection with at least two E cells are shown. The strength (weight) of the connection is indicated. Only medium and strong connections are included. (F) Cells that send strong connections to at least two E cells are included in the heatmap. (G) Cells that receive strong connections from at least two E cells are included in the heatmap.
Figure 4—figure supplement 1
Comparison of medium and strong inputs of 5th s-LNv relative to the pdf +s LNvs and the LNds.
(A–B) Supervenn diagrams indicate the number of shared inputs between all combinations of neurons, either among the 5th LNv and the LNds (A) or among the 5th LNv and the pdf +s LNvs (B). Each neuron is represented in a row, boxes in color indicate inputs to that neuron. The numbers on the right y-axis indicate the total number of neurons that provide input to that cell. For example, 74 neurons provide medium and strong inputs to LNd3. The numbers on the x-axis indicate the number of shared inputs between different combinations of cells.
Figure 4—figure supplement 2
Comparison of medium and strong outputs of 5th s-LNv relative to the pdf +s LNvs and the LNds.
(A–B) Supervenn diagrams indicate the number of shared outputs between all combinations of neurons, either among the 5th s-LNv and the LNds (A) or among the 5th LNv and the pdf + s LNvs (B). Each neuron is represented in a row, boxes in color indicate inputs to that neuron. The numbers on the right y-axis indicate the total number of neurons that are being contacted by a specific cell. For example, 172 neurons receive medium or strong contacts from LNd3. The numbers on the x-axis indicate the number of shared outputs between different combinations of cells.
Figure 5
Neuropils innervated by M and E cells.
(A–B). Percentage of connections located in each of the indicated neuropils. Medium and strong connections are included. (A) Neuropils in which the outputs of each of the four pdf + s LNvs are located. (B) Neuropils in which the outputs of each of the LNds and the 5th s-LNv are located.
Figure 6 with 2 supplements
Strong shared outputs of M cells.
(A–C) The six cells that are strong shared outputs of the four M cells involve three different neuronal types (SPL316-R, SLP403-R, and SMP232-R). The M cells outputs onto each representative neuron of each type are shown in magenta. Representative target neurons are shown in green. (A) M cells contact three SPL316 neurons. Left, all four M cells are shown in gray and their contacts to 355453590 (neuronal morphology shown in green) are shown in magenta. Right: 355453590 is shown in gray and its outputs to clock cells are shown in magenta. (B) M cells contact two SPL403-R neurons. Left, all four M cells are shown in gray and their contacts to 325455002 (neuronal morphology shown in green) are shown in magenta. Right: 325455002 is shown in gray and its outputs to clock cells are shown in magenta. (C) M cells contact one SMP232-R neuron. Left: all four M cells are shown in gray and their contacts to 325455002 (neuronal morphology shown in green) are shown in magenta. Right: 325455002 is shown in gray and its outputs to clock cells are shown in magenta. (D) Jaccard index of the six M cell shared output cells. The index is based on the similarity of their outputs, the more similar their outputs are the higher the index value. Y and x-axis indicate the cell body ID of each of the six cells. Their neuronal type is indicated to the left of the body ID on the y-axis. Only indices ≥ 0.01 are shown. (E) All strong shared outputs of M cells in turn contact clock neurons. On the x-axis, the clock neurons that receive contacts from each cell are indicated. The values on the cells represent the weight of each connection. Medium and strong connections are included. (F) Percent of strong shared outputs of each neuronal class that in turn sends contacts to clock cells. Medium and strong connections are included. E1=LNd4 and LNd5, E2=LNd6 and the 5th s-LNv, E3=LNd1, LNd2, and LNd3.
Figure 6—figure supplement 1
Connections of individual strong shared outputs of s-LNvs.
(A–F). Synaptic connections of each of the six cells that receive strong shared inputs from all four pdf +s LNvs. Each cell is indicated in gray. Inputs from clock neurons to each cell are indicated in blue, and outputs from each cell to any clock neurons are indicated in magenta. (A–C) Three SLP316 receive the strongest contacts from all pdf +s LNvs and are indicated in gray. (D–E) Two SLP403 are indicated in gray. (F) SMP223 is indicated in gray.
Figure 6—figure supplement 2
Neuroanatomy of the three SLP316 cells and identified DN1s.
(A) Five DN1ps (green) that exhibit contralateral as well as dorsal projections have been identified, and are referred to as DN1pA. (B) Two DN1ps (magenta) that lack a contralateral or dorsal projection but extend ventrally and medially have been identified and are referred to as DN1pB. (C) The three SPL316 neurons (gray) have both dorsal and ventral projections and a short medial projection. (D) Overlap of DN1pA, DN1pB, and SLP316.
Figure 7
Strong shared outputs of E1.
(A–C) The cells that are strong shared outputs of E1 involve multiple neuronal types. E1 outputs onto three representative neurons are shown. E1 neurons (LNd4 and LNd5) are shown in gray in the left panels, where representative output neurons are shown in green. On the right panels, each representative cell is shown in gray and its contacts to clock cells are shown in magenta (A) E1 cells contact two SMP368-R neurons. Left, E1 neurons (shown in gray) contacts to SMP368-R neuron 390331583 (shown in green) are shown in magenta. (B) E1 cells contact two AVLP075-R neurons. Left, E1 neurons (shown in gray) contacts to AVLP075-R neuron 702152113 (shown in green) are shown in magenta. (C) E1 cells contact two SMP315-R neurons. Left, E1 neurons (shown in gray) contacts to SMP315-R neuron 5813040712 (shown in green) are shown in magenta. (D) Jaccard indices indicating overlap among the output synaptic partners of the top 10 E1 strong shared output cells. The index is based on the similarity of their outputs, the more similar their outputs are the higher the index value. Y and x-axis indicate the cell body ID of each of the six cells. Only indices ≥ 0.01 are shown. (E) Most shared outputs of E1 cells that contact clock cells send strong contacts to both E1 neurons. On the x-axis, the clock neurons that receive contacts from each cell are indicated. The values on the cells represent the weight of each connection. Medium and strong connections are included.
Figure 8
Strong shared outputs of E2.
(A–C) The cells that are strong shared outputs of E2 involve multiple neuronal types. E2 outputs onto three representative neurons are shown. E2 neurons (LNd6 and the 5th s-LNv) are shown in gray in the left panels, where representative output neurons are shown in green. On the right panels, each representative cell is shown in gray and its contacts to clock cells, if present, are shown in magenta (A) Left, E2 neurons (shown in gray) contacts to SMP368 neuron 329732855 (shown in green) are shown in magenta. (B) Left, E2 neurons (shown in gray) contacts to SLP249 neuron 356140100 (shown in green) are shown in magenta. (C) Left, E2 neurons (shown in gray) contacts to aMe22 neuron 5813021192 (shown in green) are shown in magenta. (D) Jaccard indices for the outputs of the top 10 E2 strong shared output cells. The index is based on the similarity of their outputs, the more similar their outputs are the higher the index value. Y and x-axis indicate the cell body ID of each of the ten cells. One of the strong shared outputs of E2 is LNd4. Only indices ≥ 0.01 are shown. (E) Shared outputs of E2 that contact clock cells contact different clock subclasses. On the x-axis, the clock neurons that receive contacts from each cell are indicated. The values on the cells represent the weight of each connection. Medium and strong connections are included.
Figure 9
Strong shared outputs of E3.
(A–C) The cells that are strong shared outputs of E3 involve multiple neuronal types. E3 outputs onto three representative neurons are shown. E3 neurons (LNd1, LNd2, and LNd3) are shown in gray in the left panels, where representative output neurons are shown in green. On the right panels, each representative cell is shown in gray and its contacts to clock cells are shown in magenta. (A) Left, E3 neurons (shown in gray) contacts to SMP335 neuron 297243542 (shown in green) are shown in magenta. (B) Left, E3 neurons (shown in gray) contacts to SMP486 neuron 327933679 (shown in green) are shown in magenta. (C) Left, E3 neurons (shown in gray) contacts to SMP334 neuron 360254108 (shown in green) are shown in magenta. (D) Jaccard indices for the outputs of the top 11 E3 strong shared output cells. The index is based on the similarity of their outputs, the more similar their outputs are the higher the index value. Y and x-axis indicate the cell body ID of each of the cells. Only three cells are strong shared outputs of all E3 cells (E3a+E3 b). 8 cells are strong shared targets of E3a only. Only indices ≥ 0.01 are shown. (E) Shared outputs of any two E3 cells that contact clock cells. On the x-axis, the clock neurons that receive contacts from each cell are indicated. The values on the cells represent the weight of each connection. Medium and strong connections are included.
Figure 10
Connections within the clock neuron network.
(A) Combined weights of medium and strong connections among different classes of identified clock neurons. Arrows indicate the direction of the connection. Some classes, such as E2, are strongly interconnected, while other clusters are relatively isolated. (B) Representative strong shared output of E1 that in turn contacts clock neurons. E1 are indicated in gray, their strong shared target LHPV6m1 cell 388881226 is indicated in green, and its strong target neurons DNa1 and 2 are indicated in magenta (the weight of output contacts is 19 and 10, respectively). (C) Representative strong shared output of E2 that in turn contacts clock neurons. E2 are indicated in gray, their strong shared target SMP223 cell 417143726 is indicated in green, and its strong target neurons LPN are indicated in magenta (the weight of output contacts is 14 to LPN, 18 to LPN2, and 20 to LPN3). (D) Summary of ‘inter-clock’ neurons providing strong poly synaptic connections between clock neurons.
Tables
Table 1
Identification of Clock Neurons within the Hemibrain Volume.
| BodyId | Type | Sequential label | Subphase |
|---|---|---|---|
| 2068801704 | s-LNv | s-LNv1 | M |
| 1664980698 | s-LNv | s-LNv2 | M |
| 2007068523 | s-LNv | s-LNv3 | M |
| 1975347348 | s-LNv | s-LNv4 | M |
| 5813056917 | LNd | LNd4 | E1 |
| 5813021192 | LNd | LNd5 | E1 |
| 5813069648 | LNd | LNd6 | E2 |
| 511051477 | 5th LNv | 5th LNv | E2 |
| 296544364 | LNd | LNd1 | E3 |
| 448260940 | LNd | LNd2 | E3 |
| 5813064789 | LNd | LNd3 | E3 |
| 356818551 | LPN | LPN1 | |
| 480029788 | LPN | LPN2 | |
| 450034902 | LPN | LPN3 | |
| 546977514 | LPN | LPN4 | |
| 264083994 | DN1a | DN1a1 | |
| 5813022274 | DN1a | DN1a2 | |
| 5813010153 | DN1pA | DN1pA1 | |
| 324846570 | DN1pA | DN1pA2 | |
| 325529237 | DN1pA | DN1pA3 | |
| 387944118 | DN1pA | DN1pA4 | |
| 387166379 | DN1pA | DN1pA5 | |
| 386834269 | DN1pB | DN1pB1 | |
| 5813071319 | DN1pB | DN1pB2 | |
| 1884625521 | l-LNv | lLNv1 | |
| 2065745704 | l-LNv | lLNv2 | |
| 5813001741 | l-LNv | lLNv3 | |
| 5813026773 | l-LNv | lLNv4 |
Additional files
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Supplementary file 1
Inputs onto clock neurons.
List of Body IDs of all neurons that provide inputs onto currently identified circadian clock neurons and the weight of those pre-synaptic connections.
- https://cdn.elifesciences.org/articles/79139/elife-79139-supp1-v3.xls
-
Supplementary file 2
Outputs of clock neurons.
List of Body IDs of all neurons that receive inputs from currently identified circadian clock neurons and the weight of those post-synaptic connections.
- https://cdn.elifesciences.org/articles/79139/elife-79139-supp2-v3.xls
-
MDAR checklist
- https://cdn.elifesciences.org/articles/79139/elife-79139-mdarchecklist1-v3.pdf
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Connectomic analysis of the Drosophila lateral neuron clock cells reveals the synaptic basis of functional pacemaker classes
eLife 11:e79139.
https://doi.org/10.7554/eLife.79139
