Functional integration of a serotonergic neuron in the Drosophila antennal lobe
- University of Maryland, United States
Figures
Figure 1
Olfactory stimulation hyperpolarizes serotonergic neurons innervating the AL.
(A) Schematic representation of the AL circuitry showing excitatory connections from ORNs to PNs, and lateral inhibition from both GABAergic and glutamatergic interneurons. Serotonergic input onto LNs and PNs is inferred from previous studies across other model systems (see text). (B) An anterior to posterior Z-projection of a Drosophila brain expressing GFP in the R60F02-Gal4 (CSD -Gal4) promoter line to illustrate the innervation of the CSDn (green) in the antennal lobe (white-dashed circles). Serotonergic neurons are labeled with a 5-HT antibody and co-localize with the soma of the CSDn (white arrows). Neuropil (magenta) is labeled with the nc82 antibody. (C) Whole-cell recordings from a CSDn showing excitatory and inhibitory responses to odors. Horizontal black line denotes period of odor presentation (500 ms). (D) A raster plot from one experiment showing that most odors inhibit the CSDn. Each tick represents one action potential from a CSDn. Odors are grouped and colored according to chemical class. Ammonia, CO2, and cVA, which activate very few ORNs types are grouped together. All odors are diluted 100-fold in paraffin oil except cVA and methyl laurate, which are undiluted. (E) CSDn responses are sorted by increasing strength of hyperpolarization. Each open circle represents one preparation. Horizontal black bar is the mean of 10 preparations. (F) Schematic representation of EAG recording paradigm. (G). Regression analysis shows correlation between EAG responses and hyperpolarization of the CSDn. Insert shows sample EAG responses to ammonia and pentyl acetate. R2 = 0.69, p=0.00007.
Figure 2 with 1 supplement
Inhibitory responses in the CSDn arise at the level of the AL.
(A) Odor responses in the CSDn in normal saline and in the presence of the GABAA and GABAB receptor antagonists, picrotoxin (5 μM) and CGP54626 (50 μM) respectively. (B) GABA antagonists blocked the inhibition of the CSDn. n = 5, paired t-test, p=5.59 × 10−5. (C) The same protocol as in A except in the presence of 100 μM picrotoxin, which blocks inhibitory glutamatergic transmission in the fly. (D) Blocking inhibitory glutamate receptors also blocks the inhibtion of the CSDn. n = 5, paired t-test, p=5.12 × 10−4. (E) dSerT-eGFP is expressed selectively in the CSDn using the GAL4/UAS system. Presynaptic 5-HT release sites are seen as GFP signal in the AL. Neuropil is labeled as in Figure 1A. Scale bar = 20 µm in C and D. (F) The postsynaptic dendrite marker, DenMark, is expressed in the CSDn and visualized in green. (G) A cartoon representation of our protocol to reveal functional CSDn synapses within the AL. A sole CSDn neuron is drawn in green onto an schematic of the Drosophila brain. Cells are targeted using GFP and their neurites are stimulated with neurotransmitters delivered via pressure injection into the contralateral antennal lobe. To block polysynaptic and network contributions, TTX (1 μM) is added to the recording saline. (H) Top, sample hyperpolarizations of the CSDn in response to pressure injection of GABA in saline (black trace) and saline containing GABA antagonists (red trace). Antagonists as in A. The horizontal bar above the trace denotes the duration of pressure injection, and the coincident brief upward transient is an artifact from the opening and closing of the pressure injector’s valve. (H) bottom. A summary of recordings with GABA injection. Each gray circle represents one preparation. Black horizontal line is the mean across preparations and black vertical line shows the SEM. n = 4, paired t-test, p=0.002. (I) Same as in H, but for the application of glutamate and 100 μM picrotoxin. n = 4, paired t-test, p=0.002. (J) Same as in H, but for the application of acetylcholine and mecamylamine (100 μM). n = 4, paired t-test, p=0.008. *p<0.05, **p<0.01, ***p<0.001, N.S. = not significant. Same symbols used in all figures.
Figure 2—figure supplement 1
The CSDn expresses cellular markers of pre- and postsynaptic release sites throughout its neurites.
(A) A z-projection of the whole brain showing expression of the postsynaptic marker, DenMark, in the AL and the lateral horn. Scale bar for A and D = 50 µm. (B) A higher magnification view showing DenMark expression in the AL and throughout most olfactory glomeruli. Scale bar for B,C,E and F = 20 µm. (C) A higher magnifcation view of the lateral horn showing DenMark expression. (D) A whole brain image showing expression of dSerT through the arbors of the CSDn. (E) A high magnification view of dSerT expression within the AL. (F) A high magnification view of dSerT within the lateral horn.
Figure 3 with 4 supplements
CSDn stimulation monosynaptically inhibits LNs and polysynaptically excites them.
(A) Schematic representation showing optogenetic stimulation of the CSDn and whole-cell recording of GABAergic LNs. bottom, stimulation of CSDn results in an action potential in an LNs. (B) LNs were held at −60 mV and CSDn stimulation resulted in a fast depolarization followed by a delayed hyperpolarization. Methysergide (50 μM, red) bocked the delayed hyperpolarization but has no effect on the depolarization. Mecamylamine (100 μM, blue) blocked the depolarization. (C,D). Summary statistics for B. Methysergide has no effect on the peak depolarizing response, n = 11, ANOVA, F = 58.93, p=4.13 × 10−9, saline versus methysergide p=0.42. The addition of mecamylamine eliminated the depolarization from CSDn stimulation, methysergide versus methysergide plus mecamylamine p=9.4 × 10−8. Methysergide did block the delayed hyperpolarization, ANOVA, F = 11.01, p=0.0006. Saline vs methysergide p=5.02 × 10−4. Mecamylamine had no effect the hyperpolarization p=0.3403. (E) An LN was depolarized to −30 mV to magnify the CSDn evoked inhibition. This inhibition is blocked by methysergide. (F) A raster plot showing the inhibition of LN spikes. Black horizontal line above raster denotes period of depolarization to −30 mV. CSDn stimulation occurred during the 40 ms red bar. (G) Summary of such experiments at −30 mV. n = 10, paired t-test p=8.47 × 10−4. (H) Schematic representation of NaChBac experiments. NaChBac and Chrimson are co-expressed in the CSDn. TTX is used to block all action potentials in the brain except in the CSDn. (I) NaChBac potentials in the CSDn cause a fast depolarization and delayed hyperpolarization in the LNs. The hyperpolarization is blocked by methysergide (50 μM, red). Mecamylamine (200 μM, blue) did not block the fast depolarization. (J) Summary statistics for experiments in I. ANOVA, n = 6, F = 44.25, p=1.08 × 10−5, saline vs methysergide p=2.70 × 10−5, methysergide vs methysergide plus mecamylamine p=0.999.
Figure 3—figure supplement 1
Demonstration and calibration of Chrimson activation of the CSDn.
(A) top. A 2 ms flash of red light elicits approximately 2 action potentials from the CSDn, while a 50 ms stimulus a barrage of spikes (bottom). (B) Quantification of the relationship between light input and firing of the CSDn with Chrimson expression.
Figure 3—figure supplement 2
Stimulation of the CSDn depolarizes LNs via acetylcholinergic transmission and subsequently inhibits LNs via serotonin.
(A) Schematic representation showing optogenetic stimulation of the CSDn and whole-cell voltage clamp recording of GABAergic LNs. (B) CSDn stimulation (red arrow) evokes dynamic responses in LNs. Red dashed line shows mean holding current prior to stimulation. Responses were divided into early, mid, and late responses. Data is the mean current from 19 LNs. (C) Time series of the peak amplitude of the early inward current in saline (black) and mecamylamine (100 μM, red). Data shown is mean of one experiment. (D) Mean current traces from LNs in response to CSDn stimulation across six flies. Subtraction of currents in mecamylamine (Meca, middle) from saline-recorded currents (top) reveals the total current that is sensitive to mecamylamine (bottom). (E) Summary plot for the early, mid, and late phase mecamylamine sensitive currents. Early = p=0.004, Mid = p=0.507, Late = p=0.034, n = 6, one-sample t-test. (F) Mean current traces from seven flies in saline (top) and methysergide (methy, middle). The methysergide-sensitive current is shown on bottom. (G) Summary plot for the early, mid, and late phase methysergide-sensitive currents. Early = p=0.032, Mid = p=0.0007, Late = p=0.128, n = 7, one-sample t-test.
Figure 3—figure supplement 3
The CSDn may release acetylcholine as a co-transmitter.
(A) Membrane-bound GFP is expressed via the promoter, ChAT, to reveal cholinergic neurons and its co-localization with the 5-HT antibody. The CSDn is identified as the sole 5-HT positive neuron adjacent to the antennal lobe. nc82 = magenta, mcd8-GFP = green, and 5-HT = yellow. Scale bar = 20 μm for all panels. (B–D) A single slice of the brain is shown at the level of the CSDn with GFP and 5-HT labeling. (E) The CSDn is labeled with mcd8-GFP and the brain is stained with a ChAT antibody. (F–H) A single slice through the brain at the level of the CSDn reveals that it is labeled with the ChAT antibody. (I) The CSDn is labeled with mcd8-GFP and the brain is stained with the VAchT antibody.(J–L) A single slice through the brain at the level of the CSDn reveals that it is labeled with the VAchT antibody.
Figure 3—figure supplement 4
Co-expression of the TTX-insensitive sodium channel, NaChBac, and Chrimson can be used to effectively test mono-synaptic versus poly-synaptic connections.
(A) A recording from a CSDn that co-expresses NaChBac and Chrimson. The CSDn displays normal action potentials and is inbibited by the presentation of odors (black trace). The addition of TTX (1 μM) to the saline bath blocks all typical firing in the CSDn and blocks the odor-evoked inhibtion (red trace). This is presumably due to the prevention of action potentials in ORNs and LNs. (B) Red light drives subthreshold activity in the CSDn via Chrimson expression even in TTX. Higher levels of Chrimson activation result in a broad NaChBac-mediated plateau potential. (C) Design of a proof-of-principle experiment to demonstrate that NaChBac plateau potentials can mediate synaptic transmission. NaChBac and Chrimson are co-expressed in ORNs using the Orco-Gal4 promoter and an LN in the AL is patched. TTX is used to block all activity in the brain, thus blocking all poly-synaptic contributions. The ORNS are stimulated with Chrimson and ORN APs are mediated by NaChBac. (D) Chrimson stimulation of NaChBac-expressing ORNs results in a large post-synaptic depolarization in an LN even in the presence of TTX. TTX blocked all action potentials in the LN both at rest and during ORN stimulation. This connection is blocked by the acetylcholine antagonist mecamylamine (200 μM).
Figure 4 with 1 supplement
CSDn makes similar connections onto PNs as LNs.
(A) PNs were held at −60 mV. Stimulation of the CSDn depolarizes PNs briefly and results in a delayed hyperpolarization (gray trace, saline). (B) The early depolarization could not be blocked by methysergide (50 μM) but was blocked by mecamylamine (100 μM), ANOVA, n = 11, F = 27.6, p=1.77 × 10−6, saline vs methysergide p=0.99, methysergide vs methysergide plus mecamylamine p=8.67 × 10−6. (C) The delayed hyperpolarization was fully blocked by methysergide, while mecamylamine had no further effect on the delayed part of the response, ANOVA, n = 11, F = 13.32, p=0.0002, saline vs methysergide p=2.70 × 10−4, methysergide vs methysergide plus mecamylamine p=0.63. (D) PNs were depolarized to −30 mV to induce spiking and to amplify the effects of the hyperpolarization. At −30 mV CSDn stimulation significantly reduced PN firing, n = 11, p=0.031. (E) PNs were patched in saline containing TTX to block all activity in the brain. NaChBac and Chrimson were co-expressed in the CSDn to selectivity restore activity only in this neuron to probe monosynaptic connections with randomly selected PNs. (F) CSDn stimulation rapidly depolarized the PNs and then hyperpolarized them in TTX. (G) The hyperpolarization was blocked by methysergide, ANOVA, n = 14, F = 5.58, p=0.0096, saline (TTX) vs methysergide p=0.0072, methysergide vs methysergide plus mecamylamine p=0.35.
Figure 4—figure supplement 1
Stimulation of the CSDn results in a fast acetylcholine-dependent inward current and a delayed serotonin-mediated outward current.
(A) The mean macroscopic current recorded from 14 PNs in response to CSDn stimulation. The red arrow indicates CSDn stimulation (50 ms). Currents were measured in normal saline and in the presence of mecamylamine. Mecamylamine-sensitive current is the difference of the currents measured in mecamylamine and saline. (B) Summary of mecamylamine-sensitive currents. Ealy, mid, and late components as defined in Figure 3. n = 14, Early p=0.0002, Mid p=0.416, Late p=0.0008, one-sample t-test. (C) The mean macroscopic current recorded in saline and methysergide. (D) Summary plot of each component of the methysergide-sensitive current. n = 14, Early p=0.019, Mid p=0.004, Late p=0.314, one-sample t-test.
Figure 5 with 1 supplement
Increasing serotonergic transmission decreases PN responses in vivo.
(A) A schematic of a serotonergic synapse showing vesicles and postsynaptic receptors. The receptors are blocked by the antagonist methysergide. (B) A mean PSTH of the DA1 PN responses to a 500 ms pulse of cVA in saline and methysergide. The shaded regions show the standard error of the mean. (C) A schematic representation of a serotonergic synapse showing serotonin reuptake transporters blocked by fluoxetine (10 μm). Blockade of reuptake transporters concentrates 5-HT in the synaptic cleft. (D) DA1 PN responses to a 500 ms pulse of cVA in saline and fluoxetine. (E) Quantification of DA1 responses. Data are normalized to the mean of the responses in saline. Serotonergic transmission increases from left to right (methysergide, saline, fluoxetine). n = 6 for saline vs. methy and n = 8 for saline vs fluox. ANOVA, p=8.7 × 10− 4, F = 9.46. Tukey-Kramer post-hoc test was used for panels E,F, and G. Methy vs saline p=0.0055, saline vs fluox p=0.38, methy vs fluox p=0.0008. (F) DM6 PN responses to valeric acid (10−6) under the same protocol. n = 7 for each condition, repeated measures ANOVA, p=3.5 × 10−5, F = 27.10. Methy vs saline p=0.0021, saline vs fluox p=0.036, methy vs fluox p=3.0 × 10−5. (G) DL5 PN responses to trans-2-hexenal (10−7) under the same protocol. n = 7 for each condition, repeated measures ANOVA, p=0.0025, F = 10.32. Methy vs saline p=0.049, saline vs fluox p=0.20, methy vs fluox p=0.0019. (H,I,J) PN responses from the same three glomeruli are compared in saline vs exogenous serotonin application (104 M), n = 5 for each glomeruli. DA1 p=0.005, DM6 p=0.019, DL5 p=0.008.
Figure 5—figure supplement 1
Exogenous application of 5-HT (10−4) boosts PN responses to odors.
(A) The mean odor responses of 5 DA1 PNs in saline (black) and in serotonin. (B), top. A sample LN recording and its response to exogenous 5-HT (10−4) application. The first break in the recording represents a gap of 5 min. The second break in the recording during washout represents a 10 min gap in the recording. Bottom. A similar recording from a DA1 PN in exogenous 5-HT. Breaks in the recording are as described for B, top.
Figure 6
The CSDn does not modulate DA1 odor responses.
(A) DA1 odor responses to cVA with chronic optogenetic stimulation of the CSDn (CSDn-Gal4 > UAS-Chrimson) in saline (red circles) and methysergide (black circles). The CSDn was continuously activated with a 10 Hz sine wave at 660 nm. The sinewave was intrupted temporarily to test DA1 responses to cVA. (B) PSTH showing DA1 spiking response to cVA before (magenta) and during (green) CSDn stimulation in saline. (C) Summary statistics for data in A and B. DA1 responses to cVA were not statistically modulated by optogentically driven CSDn activity. Period 1 vs Period 2 in saline. n = 7, p=0.307. DA1 odor responses also did not change with CSDn stimulation in methysergide. n = 6, p=0.138. (D) DA1 odor responses were sampled at 80 s intervals and the CSDn was stimulated with pulses of ammonia every 10 s (red circles). Ammonia was not presented simultaneoulsy with cVA to avoid fast lateral inhibition and excitation. DA1 resonses to cVA were stable in the absense of intermittent ammonia stimulation (black circles). Period 1 represents timeframe before ammonia stimulation in experimental group and Period 2 represents time frame during ammonia stimulation. (E) PSTH showing DA1 spiking response to cVA before (magenta) and during (green) ammonia presentation. (F) Summary statistics for data in D and E. DA1 responses to cVA were not statistically modulated by ammonia driven CSDn activity. n = 7, Period 1 vs Period 2 with NH3 presentation. p=0.976, paired t-test. DA1 odor responses also did not change without ammonia stimulation. n = 6, p=0.105, paired t-test.
Figure 7
Serotonergic modulation of DA1 is governed by the network of serotonergic neurons and not the CSDn exclusively.
(A) Optogenetic stimulation of all 5-HT neurons suppresses DA1 odor responses in saline (red) but not methysergide (black). (B) PSTH showing DA1 spiking response to cVA before (magenta) and during (green) Trh stimulation in saline. Odor pulse is 500 ms for B,E, and H. (C) Summary statistics of data in A. Stimulation in saline conditions reduced DA1 odor responses, n = 7 p=0.014, paired t-test. Methysergide blocked the suppression seen in normal saline n = 8, p=0.413. (D) The CSDn is killed by the expression of a temperature-sensitive variant of diphthera toxin. The expected location of the CSDn is illustrated with yellow, dashed circles. The remaining 5-HT circuit remains intact. 5-HT positive soma are indicated with white arrows. Note the 5-HT fiber innervation of the subesophageal ganglion (SOG) and the ellipsoid body (EB). (E) PSTH's of DA1 responses to the odor cVA in flies without CSDns. Responses in normal saline are shown in black and in the presence of methysergide in red. (F) Summary of DA1 responses from E. p=0.037, n = 5, paired t-test. (G) As in D, but with expression mediated by the Trh promoter to target dipthera toxin in all 5-HT neurons. The imaging gain was elevated until a signal in the 5-HT channel was discerned giving rise to a visible background level. Note the lack of clear dense green labeled neurons. Staining in the AL, the SOG, and the EB is largely absent. (H) Odor responses of flies with killed 5-HT systems. Colors and scales bars are the same as those in E. (I) A summary of DA1 responses from brains with ablated serotonergic systems, n = 6, paired t-test. p=0.172.
Figure 8 with 1 supplement
Acute stimulation of the CSDn alters VA1d responses via cholinergic transmission.
(A) VA1d odor responses to methyl laurate were sampled every 80 s before, during, and after stimulation of the CSDn with Chrimson. Stimulation consisted of a 10 Hz sine wave that was interrupted briefly to sample VA1d response. (B) Chronic stimulation of the CSDn did not significantly alter VA1d responses either in saline (n = 7, p=0.77) or methysergide (n = 6, p=0.78). (C) A mean PSTH of VA1d odor responses with (red trace) and without (black trace) simulatneous CSDn activation with Chrimson (40 ms). (D) A raster showing an increase in the odor respnse of VA1d PNs during brief CSDn stimulation. Methyl laurate was presented undiluted. (E) In saline, PN firing over the entire duration of the odor response was unchanged (left, n = 7, p=0.84) but during the 40 ms of CSDn stimulation, VA1d responses increased (right,, n = 7, p=0.046). (F) In methysergide, PN firing over the entire duration of the odor response was also unchanged (left, n = 6, p=0.16). During the 40 ms of CSDn stimulation, VA1d responses increased (right, n = 6, p=0.008).
Figure 8—figure supplement 1
Acute stimulation of the CSDn does not modulate DA1 odor respones.
(A) A PSTH of DA1 odor responses to cVA. Responses were sampled at 80 s intervals and the CSDn was stimulated via Chrimson with 40 ms pulses of red light. The timing of the light pulses was set to occur roughly during th peak of the odor responses. The delay seen between odor onset and odor response is due to the movement of odors through the olfactometer and to the preparation. (B) Left, DA1 odor respones were not significantly changed over the duration of the odor response, n = 6, p=0.280, paired t-test. This analysis window was 500 ms starting from the onset of the response seen in the PSTH. Right, DA1 responses were also not statistically different during the brief 40 ms of CSDn stimulation, n = 6, p=0.971. (C) PSTH of DA1 responses with co-activation of CSDn stimulation in the presence of methysergide. DA1 responses were not significantly altered during the entire stimulus window (left, n = 6, p=0.287) or the brief period of CSDn stimulation (right, n = 6, p=0.141).
Tables
Table 1
Odors used in the study.
| Odors | Supplier |
|---|---|
| 1-hexanol | Sigma-Aldrich CAS: 111-27-3 |
| 1-octanol | Sigma-Aldrich CAS: 111-87-5 |
| 1-pentanol | Sigma-Aldrich CAS: 71-41-0 |
| Acetic acid | Sigma-Aldrich CAS: 64-19-7 |
| Ammonium hydroxide | Sigma-Aldrich CAS: 1336-21-6 |
| Apple cider vinegar | Spectrum Naturals |
| Benzaldehyde | Sigma-Aldrich CAS: 100-52-7 |
| Beta-citronellol | Sigma-Aldrich CAS: 106-22-9 |
| Butyl acetate | Sigma-Aldrich CAS: 123-86-4 |
| Butyric acid | Sigma-Aldrich CAS: 107-92-6 |
| cVA | Pherobank, Wijk bij Duurstede, Netherlands |
| Ethyl acetate | Sigma-Aldrich CAS: 141-78-6 |
| Ethyl propionate | Sigma-Aldrich CAS: 105-37-3 |
| Geosmin | Sigma-Aldrich CAS: 16423-19-1 |
| Limonene | Sigma-Aldrich CAS: 5989-27-5 |
| Linalool | Sigma-Aldrich CAS: 78-70-6 |
| MCH | Sigma-Aldrich CAS: 589-91-3 |
| Methyl laurate | Sigma-Aldrich CAS: 111-82-0 |
| Methyl salicylate | Sigma-Aldrich CAS: 119-36-8 |
| Paraffin oil | J.T.Baker CAS: 8012-95-1 |
| Pentanoic acid | Sigma-Aldrich CAS: 109-52-4 |
| Pentyl acetate | Sigma-Aldrich CAS: 628-63-7 |
| Phenylacetaldehyde | Sigma-Aldrich CAS: 122-78-1 |
| Propyl acetate | Sigma-Aldrich CAS: 109-60-4 |
| Trans-2-hexen-1-al | Sigma-Aldrich CAS: 6728-26-3 |
Table 2
Drosophila genotypes used in the study.
| Genotypes (transgene with Bloomington number) | |
|---|---|
| Figure 1–2 | w-;; Gal4-R60F02 (48228), UAS-mCD8-GFP |
| Figure 2E | UAS-dSerT-GFP (24463); +; Gal4-R60F02 |
| Figure 2F | w-; UAS-DenMark (33062); Gal4-R60F02 |
| Supplementary for Figure 2A–C | w-; UAS-DenMark (33062); Gal4-R60F02 |
| Supplementary for Figure 2D–F | UAS-dSerT-GFP; +; Gal4-R60F02 |
| Figure 3A–G | CS; UAS-Chrimson/+; Gal4-R60F02, UAS-mCD8-GFP/+ |
| Figure 3H–J | w-; UAS-Chrimson/UAS-NaChBac (9466); Gal4-R60F02, UAS-mCD8-GFP/+ |
| Supplementary Figure 1– 2 and for Figure 3 | w-; UAS-Chrimson/+; Gal4-R60F02, UAS-mCD8-GFP/+ |
| Supplementary Figure 3A–D for Figure 3 | w-; ChAT-Gal4; UAS-GFP (6793) |
| Supplementary Figure 3E–L for Figure 3 | w-;; Gal4-R60F02 (48228), UAS-mCD8-GFP |
| Supplementary Figure 4A–B for Figure 3 | w-; UAS-Chrimson/UAS-NaChBac; Gal4-R60F02, UAS-mCD8-GFP/+ |
| Supplementary Figure 4D for Figure 3 | w-; UAS-Chrimson/Gal4-Orco (26818); UAS-NaChBac/+ |
| Figure 4A–D | CS; UAS-Chrimson/+; Gal4-R60F02, UAS-mCD8-GFP/+ |
| Figure 4E– G | w-; UAS-Chrimson/UAS-NaChBac; Gal4-R60F02, UAS-mCD8-GFP/+ |
| Supplementary for Figure 4 | w-/CS; UAS-Chrimson/+; Gal4-R60F02, UAS-mCD8-GFP/QF-GH146, QUAS-mCD8-GFP (30038) |
| Figure 5A– E,H | CS; QF-Mz19 (41573), QUAS-mCD8-GFP (30002)/Cyo |
| Figure 5I–J Figure 5F–G, | Gal4-NP3062, UAS-mCD8-GFP |
| Supplementary for Figure 5 | CS; QF-Mz19, QUAS-mCD8-GFP/Cyo |
| Figure 6A–C | w-/CS; QF-Mz19, QUAS-mCD8-GFP/UAS-Chrimson; Gal4-R60F02, UAS-mCD8-GFP/+ |
| Figure 6D–F | CS; QF-Mz19, QUAS-mCD8-GFP/Cyo |
| Figure 7A–C | w-/CS; QF-Mz19, QUAS-mCD8-GFP/UAS-Chrimson; Gal4-Trh (38389)/+ |
| Figure 7D–F | w-/CS; UAS-Diphthts (25039)/ QF-Mz19, QUAS-mCD8-GFP; Gal4-R60F02, UAS-mCD8-GFP/+ |
| Figure 7G–I | w-/CS; UAS-Diphthts/ QF-Mz19, QUAS-mCD8-GFP; Gal4-Trh/+ |
| Figure 8 | w-/CS; QF-Mz19, QUAS-mCD8-GFP/UAS-Chrimson; Gal4-R60F02, UAS-mCD8-GFP/+ |
| Supplementary for Figure 8 | w-/CS; QF-Mz19, QUAS-mCD8-GFP/UAS-Chrimson; Gal4-R60F02, UAS-mCD8-GFP/+ |
-
UAS-Chrimson was from Dr. Vivek Jayaraman, Janelia Farm, Ashburn, VA
-
Gal4-NP3062 was from Dr. Rachel Wilson, Harvard Medical School, Boston, MA
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Gal4-R60F02, Gal4-Trh, UAS-Chrimson, QF-Mz19 were backcrossed with Canton-S flies.
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Functional integration of a serotonergic neuron in the Drosophila antennal lobe
eLife 5:e16836.
https://doi.org/10.7554/eLife.16836
