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Published in final edited form as: Curr Biol. 2020 May 21;30(13):2602–2607.e2. doi: 10.1016/j.cub.2020.04.056

Population Density Modulates the Duration of Reproduction of C. elegans

Spencer S Wong 1,2, Jingfang Yu 3, Frank C Schroeder 3, Dennis H Kim 1,4,*
PMCID: PMC7343598  NIHMSID: NIHMS1589972  PMID: 32442457

Summary

Population density can modulate the developmental trajectory of Caenorhabditis elegans larvae by promoting entry into dauer diapause, which is characterized by metabolic and anatomical remodeling and stress resistance [1,2]. Genetic analysis of dauer formation has identified the involvement of evolutionarily conserved endocrine signaling pathways, including the DAF-2/insulin-like receptor signaling pathway [3–7]. Chemical and metabolomic analysis of dauer-inducing pheromone has identified a family of small molecules, ascarosides, which act potently to communicate increased population density and promote dauer formation [1,8–10]. Here, we show that adult animals respond to ascarosides produced under conditions of increased population density by increasing the duration of reproduction. We observe that the ascarosides that promote dauer entry of larvae also act on adult animals to attenuate expression of the insulin peptide INS-6 from the ASI chemosensory neurons, resulting in diminished neuroendocrine insulin signaling that extends the duration of reproduction. Genetic analysis of ins-6 and corresponding insulin-signaling pathway mutants showed that the effect of increased population density on reproductive span was mimicked by ins-6 loss-of-function that exerted effects on duration of reproduction through the canonical DAF-2-DAF-16 pathway. We further observed that the effect of population density on reproductive span acted through DAF-16-dependent and DAF-16-independent pathways upstream of DAF-12, paralleling in adults what has been observed for the dauer developmental decision of larvae. Our data suggest that under conditions of increased population density, C. elegans animals prolong the duration of reproductive egg laying, which may enable the subsequent development of progeny under more favorable conditions.

Keywords: Population density, Reproductive span, Pheromone, Insulin, Neuroendocrine signaling

eTOC:

Wong et al. show that population density, signaled through ascaroside pheromones, modulates the expression of an insulin peptide in a pair of sensory neurons of adult C. elegans, leading to an extension in reproductive egg laying. An extended period of egg laying may enable the development of progeny in more favorable conditions.

Results and Discussion

The sensory nervous system of C. elegans orchestrates the developmental plasticity and changes in organismal physiology that take place in response to environmental and endogenous cues [6,11]. The dynamic activities of neuropeptide ligands that can be expressed by, or act on, sensory neurons have been shown to have a major influence on the regulation of organismal physiology [12]. In particular, the dynamic expression of insulin genes, which encode approximately 40 insulin-like peptides in C. elegans, influences a multitude of phenotypes including dauer entry and exit, olfactory learning, lifespan, stress resistance, and reproductive span through the DAF-2/insulin-like receptor [7,13–16]. Members of the insulin gene family exhibit redundancy and crosstalk among peptides [13,17]. The insulin-like peptide INS-6 has been previously shown to regulate exit from dauer diapause and olfactory learning [15,16]. The role of ins-6 in the regulation of dauer diapause is associated with dynamic expression—ins-6 is expressed throughout larval development in the two ASI chemosensory neurons. Upon exposure to dauer-inducing conditions, ins-6 expression in ASI is attenuated during larval development and expression in the ASJ neuron pair is observed during dauer diapause, when ins-6 has a role in promoting dauer exit [15].

Recent studies have revealed that ascarosides can also have diverse effects on adults animals as well, including roles in mediating mate attraction and regulating lifespan [18–23]. Considering the potent effects of ascarosides in signaling increased population density to induce entry into dauer diapause during early larval development, we hypothesized that increased population density could also influence animals at a later stage of life, after the point when the dauer decision can be made. We observed that increased population density resulted in the marked downregulation of ins-6 expression from the ASI neuron pair in adult hermaphrodites (Figure 1A–E). Diminished fluorescence intensity was observed from the ASI neurons of transgenic animals carrying an ins-6p::GFP transcriptional reporter under increased population density (Figure 1E). These data suggest that the sensory nervous system of adult animals detects changes in population density. Neuronal expression of daf-28, another insulin peptide that acts to increase insulin receptor signaling, was unaffected by population density (Figure 1F). Another secreted neuropeptide, daf-7, is also expressed from the ASI sensory neurons and regulated by dauer pheromone during pre-dauer development [2,24], but expression of daf-7 from the ASI neurons in adult animals was unaffected by population density (Figure 1G).

Figure 1. Neuronal insulin expression is regulated by a population density-dependent ascaroside signal in reproductive adult C. elegans.

Figure 1.

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(A–D) Representative images showing animal density and ins-6 expression pattern. Brightfield microscopy for 1mL lawn (A) and 20 μL lawn (C), where n = 100 animals for each lawn condition. Representative DIC images at 100X magnification with ins-6p::gfp and DiI lipophilic chemosensory neuron stain for the 1 mL lawn (B) and 20 μL lawn (D). Arrowhead denotes the ASI neuron. Scale bar for (A,C) 1 mm. Scale bar for (B,D) 5 μm. (E) Quantification of fluorescence of ins-6p::gfp in the ASI neurons of age-matched Day 1 adult hermaphrodites. A.B.U. = arbitrary brightness units. **** p<0.0001 as determined by unpaired Welch’s t-test. Error bars represent SD. Mean fluorescence by column (left to right): 932, 3822. (F) Quantification of daf-28p::gfp fluorescence in both ASI and ASJ neurons of Day 1 adults. High density: 2.5 μL OP50, low density: 250 μL OP50. Results are not statistically significant as determined by ordinary one-way ANOVA followed by Sidak’s multiple comparison test. Error bars represent SD. Mean fluorescence by column (left to right): 12715, 14177, 4086, 5250. (G) Quantification of daf-7p::gfp fluorescence in ASI neuron in Day 1 adults. High density: 2 μL OP50, low density: 200 μL OP50. Results are not statistically significant as determined by ordinary unpaired t-test with Welch’s correction. Error bars represent SD. Mean fluorescence by column (left to right): 1887, 1738.

Because of the known activities of ascarosides in signaling population density during larval development [1,10,15,25], we sought to confirm that the observed density-dependent effects on expression of ins-6 in adult hermaphrodite animals were mediated by ascarosides. Ascarosides are synthesized through a pathway terminating with the C. elegans homolog of the mammalian peroxisomal 3-ketoacyl-CoA thiolase SCPx, DAF-22. We observed that daf-22(ok693) mutant animals, which are devoid of short-chain ascarosides, exhibited increased expression of ins-6, which was unaffected by the density of daf-22 mutant animals (Figure 2A). To show that the observed effect of the daf-22 mutation was due to the abrogation of a density-dependent signal produced and secreted by animals in a DAF-22-dependent, density-dependent manner, and not because of the effects of the daf-22 mutation in a mutant animal altering expression of ins-6 independent of population density, we incubated 10 wild-type or 10 daf-22 mutant animals on a lawn (radius of 5 mm) of E. coli for 6 h to “condition” the plates by enabling any secreted molecules to accumulate on the plates, followed by removal of the animals. Then, one wild-type “tester” animal carrying the ins-6p::gfp transgene was placed onto each conditioned lawn (Figure 2B). We observed that conditioning plates with wild-type animals, but not daf-22 mutant animals, caused diminished expression of ins-6 in the ASI neurons (Figure 2C). These data suggest that the decrease in ins-6 results from the presence of daf-22-dependent secreted molecules with increased population density.

Figure 2: Small molecule ascaroside pheromone compounds inhibit ins-6 expression.

Figure 2:

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(A) Expression of ins-6 in wild type animals and daf-22 (ok693) mutants defective in ascaroside biosynthesis. Five animals per plate. High density: 2.5 μL OP50, low density: 250 μL OP50. **** p<0.0001, *** p<0.001 as determined by ordinary one-way ANOVA followed by Sidak’s multiple comparison test. Error bars represent SD. Mean fluorescence by column (left to right): 2649, 4935, 8200, 8256. (B) Schematic showing the preconditioning protocol for wild-type and daf-22(ok693) mutants. (C) The effect of secreted daf-22-dependent molecules on attenuation of ins-6 expression. **** p<0.0001 as determined by ordinary one-way ANOVA followed by Dunnett’s multiple comparison test. Error bars represent SD. n.s., not significant. Mean fluorescence by column (left to right): 3196, 1315, 3078. (D–E) Representative image of animals treated with control solvent ethanol (D) or 1.5 μM ascaroside blend (E). Scale bar indicates 50 μm. Arrowhead denotes the ASI neuron. (F) Quantification of ins-6p::gfp fluorescence values in ASI with the addition of synthetic C. elegans pheromone. A.B.U. = arbitrary brightness units. **** p<0.0001 as determined by Welch’s unpaired t-test. Error bars represent SD. Mean fluorescence by column (left to right): 3649, 378. See also Figure S1.

Next, we directly examined the ability of ascr#2, ascr#3, ascr#5, and ascr#8, to affect ins-6 expression, and we observed that the addition of an equimolar mixture of these ascarosides abrogated ins-6 expression from the ASI neurons of adult hermaphrodite animals, mimicking the effect of increased population density (Figure 2D–F). The ascaroside compounds may act individually, additively, or synergistically. Therefore, the individual components were tested for their ability to suppress ins-6 expression (Figure S1A). Interestingly, ascr#2, ascr#3, and ascr#8 repressed ins-6 strongly, whereas ascr#5 exhibited less potent effects on ins-6 expression. The ascarosides ascr#2, ascr#3, and ascr#8 each produced a strong dose-response curve for ins-6 expression, with strong effects observed on ins-6 expression when the tested ascarosides were in the 20 nM –400 nM range (Figure S1B–D), within the physiological range in which the density-dependent secretion of ascarosides have been shown to promote dauer entry in larvae [26]. Previous reports have shown that daf-37, daf-38, srbc-64, srbc-66, srg-36, and srg-37 mutants have decreased formation of dauers in the presence of high concentrations of pheromone [9,27]. Because of this, we tested if these function in the pathway leading to ins-6 expression in ASI. These canonical ascaroside receptors do not seem to diminish the responsiveness of ins-6 to ascr#2 and ascr#3 at the highest concentrations tested and also the minimum inhibitory concentration for the ins-6 expression assay (Figure S1E–F). Redundancy in pheromone receptors may exist for the effect on ins-6 expression, or additional uncharacterized receptors may have a role. However, we did observe that the repressive effect of ascr#5 is mitigated in daf-38(tm4150), srg-36(tm6454), and srg-37(tm6502) mutants, suggestive of a functional contribution from each of these receptors in the relatively weaker effects on ins-6 expression observed in the presence of ascr#5 (Figure S1E).

We next evaluated adult animals for physiological changes that might be affected by dynamic ins-6 levels during conditions of increased population density. In particular, we observed an increase in the number of progeny produced at late timepoints of adulthood when animals were incubated in high population density (Figure 3A), an effect that was also observed in the presence of a mixture of ascr#2, ascr#3, and ascr#5 (Figure S2A).

Figure 3: High population density extends reproductive span of C. elegans, which is mimicked by effects of diminished insulin signaling at low population density.

Figure 3:

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(A) Assessment of persistent egg-laying phenotypes beyond 72 h post-L4 under highly dense conditions (100 animals grown on 20 μL lawn) or low density (100 animals grown on 1 mL lawn) for wild-type animals. **** p<0.0001 as determined by ordinary one-way ANOVA followed by Sidak’s multiple comparison test. Error bars represent SD. n.s. not significant. Mean progeny by column (left to right): 32.6, 6.4. (B) Amount of progeny animals continue to produce after 72 h post-L4 under low population density. tm2416 is a 316 bp deletion in the first exon of ins-6. * p<0.05 as determined by ordinary one-way ANOVA followed by Sidak’s multiple comparison test. Error bars represent SD. Mean progeny by column (left to right): 2.6, 8.2, 2.8. (C) Under low population density, double mutant singled adults of ins-6 and insulin-like signaling pathway components do not further extend reproductive span. Progeny production measured 72 h post-L4. *p<0.05, n.s. not significant as determined by ordinary unpaired t-test with Welch’s correction. Error bars represent SD. Mean progeny by column (left to right): 0.8, 5.5, 49, 4.2, 60, 0.7. (D) High population density is transduced through both daf-16-dependent and daf-16-independent pathways to extend progeny production. Measurement of progeny produced under conditions of high population density in various dauer-deficient mutant backgrounds after 72 h post-L4. WT, first column, is under low population density (1 mL lawn); all others 20 μL lawn. **** p<0.0001, n.s. not significant as determined by ordinary one-way ANOVA followed by Dunnett’s multiple comparison test. Mean progeny by column (left to right): 7.8, 48.5, 50.4, 33.5, 18. See also Figures S2, S3.

Considering the marked effect of population density on both decreasing ins-6 expression and extending the reproductive period, we sought to determine whether a loss-of-function mutation in ins-6 could mimic the effects of increased population density on the duration of reproduction of animals under conditions of low population density. We observed that Day 3 ins-6(tm2416) adults exhibited extended progeny production compared with wild-type animals, phenocopying the effects of high population density (Figures 3B, 3C). The extended duration of progeny production observed in ins-6(tm2416) mutants at low population density was rescued by the introduction of a genomic ins-6p::ins-6 transgene (Figure 3B, S2B).

We confirmed that neither ins-6 mutants nor animals subjected to increased population density during larval development exhibited delays in reaching the last larval stage or the beginning of egg laying (Figure S2C–F). This suggests that the effects on the duration of reproduction are not due to delayed development or onset of reproduction. In addition, we observed that under conditions of increased population density, the duration of reproduction of ins-6 mutants was comparable to that observed for wild-type animals, suggesting the involvement of other insulin-like peptides (Figure S2G). We observed that daf-28 mutants indeed continued to lay eggs beyond the time when egg-laying was completed by wild type animals, underscoring the role of insulin-signaling in modulating duration of reproduction (Figure S3A). As insulins have been previously shown to function in an interrelated regulatory network, double mutants of ins-6(tm2416);daf-28(tm2308) were tested for further elevated ability to produce late stage progeny. Indeed, the ins-6(tm2416);daf-28(tm2308) double mutants produced more progeny beyond Day 3 adults than daf-28 or ins-6 single mutants (Figure S3B). In contrast to the dynamic expression that we observed for ins-6 in response to changes in population density, we observed that the expression of daf-28 in the ASI or ASJ neurons of wild-type animals was not affected by population density, suggesting a specificity in the role for ins-6 in regulating the density-dependent effects on reproductive span (Figure 1F).

This phenotype of ins-6 mutants is reminiscent of, albeit weaker than, the previously reported extended progeny production of daf-2 mutants, which we also confirmed (Figures 3C, S3C). Long-lived daf-2 mutants exhibit delayed reproductive aging, with concomitant increased reproductive span and perdurance of robust germ cells in older adults [5,14,28–33], suggestive of a mechanism by which decreased ins-6 activity may influence the duration of progeny production. The ins-6 mutation did not further extend the reproductive span of daf-16;ins-6 or ins-6;daf-2 double mutants, consistent with the interpretation that the effect of the ins-6 mutation on the duration of reproduction is mediated by the canonical DAF-2 insulin-like pathway (Figure 3C). The lack of extension in the daf-16 mutant background is notable, even as the interpretation of the data on the ins-6;daf-2 mutant is limited by the non-null nature of the daf-2(e1370) allele.

These data suggest that the decrease in ins-6 expression from the ASI neurons under conditions of high population density can extend the duration of reproduction through modulation of the insulin-like signaling pathway and increased activation of DAF-16. We next examined whether signaling through DAF-16 acts solely or redundantly with other pathways to modulate the duration of reproduction. We observed that under conditions of high population density, animals carrying daf-16 loss-of-function mutation continued to exhibit an extended duration of reproduction (Figure 3D), indicating the role of DAF-16-independent pathways likely acting in parallel to DAF-16-dependent mechanisms to mediate the extended duration of reproduction. Interestingly, however, we observe that whereas a mutation in daf-3(mgDf90) also did not suppress the effects of high population density on duration of reproduction, a mutation in daf-12(m20) suppressed the extended duration of reproduction observed under conditions of high population density (Figure 3D), paralleling what is observed for daf-12 suppression of the density-dependent induction of dauer diapause.

Our data suggest that population density can modulate neuroendocrine insulin signaling, influencing the duration of reproduction of individuals within a population. Prior studies have shown that diminished insulin signaling through DAF-2 can modulate reproductive aging, with improved oocyte quality and maintenance [28,29,33], providing a plausible mechanism for the increased duration of reproduction that results from the neuroendocrine modulation of insulin receptor signaling in response to population density. In an adaptation of parental investment theory, the species as a whole attempts to maximize the chances of success of their progeny by holding out for, or roaming to better conditions in the future, rather than modifying progeny size [34]. An extended window of reproduction may enable gravid hermaphrodites to lay eggs over a wider range, providing a selective advantage in environments where progeny may otherwise face increased competition for resources. A prior study showed that pheromone activates the GnRH reproductive neurons in mice [35], and our data underscore how individual reproduction, in a simple but social animal, may be influenced by population-dependent environmental cues.

STAR Methods and Materials

RESOURCE AVAILABILITY

Lead Contact

Further information and requests for resources and reagents should be directed to and will be fulfilled by the Lead Contact, Dennis H. Kim (dennis.kim@childrens.harvard.edu).

Materials Availability

There are no restrictions to the availability of reagents.

Data and Code Availability

The published article includes all datasets generated or analyzed during this study.

EXPERIMENTAL MODEL AND SUBJECT DETAILS

Nematode strains

Nematode strains were grown on nematode growth medium (NGM) plates seeded with live E. coli OP50 [36]. Alleles and transgenes used were as follows: CF1588 daf-16(mu86), daf-2(e1370), muIs84[sod-3p::gfp + rol-6(su1006)]; DR20 daf-12(m20); FK181 ksIs2[daf-7p::gfp + rol-6(su1006)]; GR1311 daf-3(mgDf90) 3x BC; GR1455 mgIs40[daf-28p::gfp]; HT1701 unc-119(ed3), wwEx65[ins-6p::gfp; unc-119(+)]; N2; RB859 daf-22(ok693); QZ81 ins-6(tm2416); ZC239 ins-6(tm2416), yxEx175[ins-6p::ins-6::ins-6 3’UTR; ofm-1p::gfp]; ZD699 daf-28(tm2308); ZD1022 daf-16(mu86) 6x backcrossed; ZD1153 daf-2(e1368) 6x backcrossed; ZD1155 daf-2(e1370) 6x backcrossed; ZD2539 daf-22(ok693), wwEx65[ins-6p::gfp; unc-119(+)]; ZD2583 daf-37(tm11887), wwEx65; ZD2584 daf-38(tm4150), wwEx65; ZD2585 srbc-64(tm1946), wwEx65; ZD2586 srbc-66(tm2943), wwEx65; ZD2587 srg-36(tm6454), wwEx65; ZD2588 srg-37(tm6502), wwEx65; ZD2589 daf-2(e1370), ins-6(tm2416); ZD2590 daf-16(mu86), ins-6(tm2416); ZD2602 ins-6(tm2416), daf-28(tm2308). Nematodes were provided by the Caenorhabditis Genetics Center, except for QZ81 and ZC239, which were kindly provided by Dr. Yun Zhang and described in [16]. Strains were maintained at 20°C.

METHOD DETAILS

Measurement of gene expression in the ASI neurons

For the isolation of synchronous populations of nematodes for GFP quantification, adults were placed in hypochlorite solution to release eggs and the progeny allowed to hatch and arrest as L1 larvae in M9 media overnight. Animals were transferred to various lawn sizes (seeded the previous day and grown at room temperature) at the L4 larval stage and incubated overnight at 20°C for 16 h. Figure 1E has various amounts of animals, stated in the figure. All other experiments were performed with 5 L4 animals per plate. Animals were then mounted on agar pads on glass slides (Corning), anesthetized in sodium azide (50 mM), and imaged at 40X using a Zeiss Axioimager Z1 microscope. 15–20 animals were imaged for each condition or strain. For quantification, maximum intensity values of GFP inside the ASI neuron were calculated via FIJI [37,38]. Light microscopy images of the lawns (Figure 1) were taken on a Zeiss SteREO Discovery.V12 at 10X.

Lipophilic dye filling was achieved with 100 animals treated as above in 200 μL M9 with DiI stain (Life Technologies) (1 μg/mL) in dimethyl formamide for 3 hours. The nematodes were then allowed to destain for 1 h on a standard NGM plate seeded with OP50. Animals were then anesthetized as above and imaged at 100X on a Zeiss LSM 800 with the same settings for both images.

Preincubation experiments involved placing 10 L4 stage animals on a 20 μL lawn of E. coli for 6 h, removing them, and placing a single HT1701 L4 stage larva on each plate. The animals were then incubated for 16 h at 20°C and imaged as above.

Synthetic ascarosides ascr#2, ascr#3, ascr#5, and ascr#8 [39] were added to a final concentration of 1.5 μM of each ascaroside on 3.5 cm SKA plate. 250 μL E. coli OP50 was 20x concentrated and then seeded after the pheromone solution had dried off of the plate. Five L4 animals were then placed on for the GFP induction assays. Serial dilutions were performed in 10% ethanol prior to adding onto plates.

Egg-laying, brood size, and egg-lay extension assays

For the isolation of synchronous populations of nematodes for all egg lay assays, 10 adults were egg laid for 1 h at 20°C on lawns of E. coli OP50. For brood size over time assays, nematodes were singled onto their own plate every 24 h starting from the L4 stage. For the assay involving daf-2 and daf-16 alleles, all genotypes were grown at 16°C from egg for 72 h (L4 larval stage), then transferred onto their own plate and placed at 20°C, where daf-2 retains activity beyond temperature sensitive period [33]. For egg-lay extension assays, nematodes were grown at 20°C for 72 h after egg lay, then singled as day 1 adults for 48 h at 20°C. Finally, they were transferred to new plates at 20°C. Progeny were counted two days after active egg lay. For the differential density conditions, 20 μL or 1 mL of 20x concentrated OP50 were seeded on 10 cm NGM plates. Eggs were laid as above and then picked over to the lawns to guarantee 100 animals on each lawn. Animals were grown at 20°C for 120 hours. Finally, the adults were singled and used for the egg lay assay on standard NGM plates from beyond 120 h (72 h post-L4). These same density conditions were employed for assaying developmental rates.

QUANTIFICATION AND STATISTICAL ANALYSIS

For all experiments, the significance of differences between conditions was evaluated with t-test statistics or nonparametric tests as appropriate with Prism 7 (GraphPad). Statistical details can be found in the respective figure legend.

Welch’s t-test was applied when there are only two groups of data being compared or with wide variation in the experimental means of all conditions. With two or more groups, we applied ANOVAs followed by Sidak’s or Dunnett’s multiple comparison test. Dunnett’s is employed when all columns are compared to one control column (i.e. A-B, A-C, A-D, A-E, and so forth), while Sidak’s is employed when multiple specific columns are compared to each other (i.e. A-B, B-C, A-C, A-D, B-D, C-D, and so on). Sidak’s assumes that each comparison is independent of the others, while Dunnett’s does not.

Supplementary Material

1

KEY RESOURCES TABLE.

REAGENT or RESOURCE SOURCE IDENTIFIER
Bacterial and Virus Strains
E. coli OP50 CGC RRID:WB-STRAIN:WBStrain00041971
Chemicals, Peptides, and Recombinant Proteins
ascr#2, ascr#3, ascr#5, ascr#8 F. Schroeder N/A
Sodium Azide Sigma Cat#S2002–5G
DiI Stain Life Technologies Cat#D282
Dimethyl Formamide Sigma Cat#319937–500ML
Ethanol Sigma Cat#E7023–500ML
Experimental Models: Organisms/Strains
C. elegans QZ81 ins-6(tm2416) Y. Zhang / J. Alcedo N/A
C. elegans ZC239 ins-6(tm2416);yxEx175 Y. Zhang / J. Alcedo N/A
C. elegans ZD699 daf-28(tm2308) D. Kim N/A
C. elegans ZD2602 daf-28(tm2308),ins-6(tm2416) D. Kim This Study
C. elegans N2 CGC RRID:WB-STRAIN:WBStrain00000003
C. elegans CF1588 daf-16(mu86), daf-2(e1370), muIs84 CGC N/A
C. elegans DR20 daf-12(m20) CGC N/A
C. elegans FK181 ksIs2[daf-7p::gfp + rol-6(su1006)] CGC N/A
C. elegans GR1455 mgIs40[daf-28::gfp] CGC N/A
C. elegans RB859 daf-22(ok693) CGC N/A
C. elegans HT1701 unc-119(ed3), wwEx65[ins-6p::gfp; unc-119(+)] Y. Zhang / J. Alcedo N/A
C. elegans ZD1153 daf-2(e1368) D. Kim N/A
C. elegans ZD1155 daf-2(e1370) D. Kim N/A
C. elegans ZD2539 daf-22(ok693); wwEx65 D. Kim This Study
C. elegans ZD2589 daf-2(e1370);ins-6(tm2416) D. Kim This Study
C. elegans ZD2583 daf-37(tm11887); wwEx65 D. Kim This Study
C. elegans ZD2584 daf-38(tm4150); wwEx65 D. Kim This Study
C. elegans ZD2585 srbc-64(tm1946); wwEx65) D. Kim This Study
C. elegans ZD2586 srbc-66(tm2943); wwEx65 D. Kim This Study
C. elegans ZD2587 srg-36(tm6454); wwEx65 D. Kim This Study
C. elegans GR1311 daf-3(mgDf90) 3x BC CGC N/A
C. elegans ZD1022 daf-16(mu86) 6x BC D. Kim N/A
C. elegans ZD2588 srg-37(tm6502); wwEx65 D. Kim This Study
C. elegans ZD2590 daf-16(mu86); ins-6(tm2416) D. Kim This Study
Software and Algorithms
Prism 7 GraphPad RRID:SCR_002798
Fiji Fiji RRID:SCR_002285
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Highlights:

  1. Neuronal insulin signaling is modulated by population density

  2. Ascaroside pheromones downregulate INS-6 expression in the ASI neuron pair

  3. High population density modulates insulin expression to extend reproductive span

Acknowledgments:

We thank members of the Kim laboratory for helpful suggestions. We thank the Caenorhabditis Genetics Center, H. R. Horvitz, J. Alcedo, and Y. Zhang for strains. This research was supported by NIH Grants R01GM084477 (to D.H.K.) and R01AT008764 (to D. H. K. and F. C. S.).

Footnotes

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Declaration of Interests: The authors declare no competing interests.

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Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

1
NIHMS1589972-supplement-1.pdf (323.4KB, pdf)

Data Availability Statement

The published article includes all datasets generated or analyzed during this study.

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