Neuropeptide feedback modifies odor-evoked dynamics in Caenorhabditis elegans olfactory neurons

Journal name:
Nature Neuroscience
Volume:
13,
Pages:
615–621
Year published:
DOI:
doi:10.1038/nn.2526
Received
Accepted
Published online

Abstract

Many neurons release classical transmitters together with neuropeptide co-transmitters whose functions are incompletely understood. Here we define the relationship between two transmitters in the olfactory system of C. elegans, showing that a neuropeptide-to-neuropeptide feedback loop alters sensory dynamics in primary olfactory neurons. The AWC olfactory neuron is glutamatergic and also expresses the peptide NLP-1. Worms with nlp-1 mutations show increased AWC-dependent behaviors, suggesting that NLP-1 limits the normal response. The receptor for NLP-1 is the G protein-coupled receptor NPR-11, which acts in postsynaptic AIA interneurons. Feedback from AIA interneurons modulates odor-evoked calcium dynamics in AWC olfactory neurons and requires INS-1, a neuropeptide released from AIA. The neuropeptide feedback loop dampens behavioral responses to odors on short and long timescales. Our results point to neuronal dynamics as a site of behavioral regulation and reveal the ability of neuropeptide feedback to remodel sensory networks on multiple timescales.

At a glance

Figures

  1. AWC releases NLP-1, which acts on NPR-11 in AIA.
    Figure 1: AWC releases NLP-1, which acts on NPR-11 in AIA.

    (a) AWC sensory neurons, downstream interneurons, and relevant glutamate receptors (from this work (AIA) and ref. 9). (b,c) Local search behavior 7–12 min after removal from food. RevOmega, coupled reversal-omega behaviors characteristic of local search. Analysis of nlp-1 mutants (b) and npr-11 mutants (c). In all figures, WT indicates control N2 strain, AWCdouble colonnlp-1 indicates nlp-1 cDNA under AWC-selective odr-3 promoter, AWCdouble colonnlp-1(OE) indicates the same plasmid injected at high concentrations, AIAdouble colonnpr-11 indicates npr-11 cDNA under AIA-selective gcy-28.d promoter. Error bars, s.e.m.; *P < 0.05 by t-test or t-test with Bonferroni correction, as appropriate; NS, not significant. Complete behavioral data with all genotypes and time points are in Supplementary Table 1. (d,e) Response of npr-11- and Gα16Z-, npr-11 or Gα16Z- transfected HEK 293 cells to an NLP-1 peptide and a scrambled NLP-1 peptide (sNLP-1). (d) Pseudocolor images of fura2-labeled cells indicating fluorescent ratio intensities. Scale bar, 100 μm. (e) Average calcium response of all cells in the window (n = 10 fields for npr-11 and Gα16Z, n = 8 for npr-11 and n = 7 for Gα16Z). Means and s.e.m. are shown.

  2. Calcium responses in AIA interneurons require AWC glutamate and NLP-1.
    Figure 2: Calcium responses in AIA interneurons require AWC glutamate and NLP-1.

    (a,b) Heat maps showing the ratio of change in fluorescence to total fluorescence in AIA neurons expressing GCaMP2.2b15; addition (a) and removal (b) of odor stimulus at t = 10 s in each recording (n = 18). (c,d) Average G-CaMP fluorescence change in AIA neurons in wild-type (WT; n = 18) and wild-type AWC-ablated worms (n = 12) on addition (c) and removal (d) of odor. (eg) Mutant AIA responses. (e) eat-4 (n = 18, WT n = 18). (f) glc-3 (n = 16, WT n = 16). (g) nlp-1 (n = 18, WT n = 18) and AWCdouble colonnlp-1 cell-selective rescue (n = 18). In all imaging figures, odor is a 10−4 dilution of isoamyl alcohol. Light gray shading indicates s.e.m. *Significantly different from wild type; **significantly different from nlp-1 mutant (P < 0.05, t-test with Bonferroni correction).

  3. Altered AWC calcium responses in nlp-1 and npr-11 mutants.
    Figure 3: Altered AWC calcium responses in nlp-1 and npr-11 mutants.

    (a,b,e,f) Heat maps showing ratio change in fluorescence to total fluorescence in AWC neurons expressing G-CaMP1.0. Odor was removed at 10 s in each recording. (a) Wild type (n = 32); (b) nlp-1 (n = 32); (e) wild type (n = 18); (f) npr-11 (n = 18). (c,g) Representative AWC calcium responses from individual wild-type worms, nlp-1 (c) and npr-11 mutants (g), and rescued strains. (d,h) Fourier power analysis of AWC calcium responses in nlp-1 (d) and npr-11 mutants (h). Left, normalized energy density spectrum averaged across all calcium traces of each genotype; arrows indicate range of the middle frequency band (color code on right). Right, the average power ratio of the middle frequency band (0.033–1 Hz) across all calcium traces of each genotype; error bars, s.e.m. *P < 0.05 (t-test with Bonferroni correction).

  4. Worms with mutations in nlp-1 and npr-11 are defective in olfactory adaptation.
    Figure 4: Worms with mutations in nlp-1 and npr-11 are defective in olfactory adaptation.

    (a) Schematic diagram of adaptation assay. (b) Adaptation in nlp-1 and npr-11 mutants, and cell-selective rescue. Error bars, s.e.m. *P < 0.05 (t-test with Bonferroni correction). (c,d) AWC calcium responses in wild-type, nlp-1 and AWCdouble colonnlp-1 transgenic rescued worms (c) and wild-type, npr-11 and AIAdouble colonnpr-11 transgenic rescued worms (d) adapted for 60 min (n = 12 each). Odor pulses are marked. Light gray shading indicates s.e.m. *Significantly different from wild type (P < 0.05, t-test with Bonferroni correction).

  5. ins-1 is a component of the nlp-1-npr-11 pathway.
    Figure 5: ins-1 is a component of the nlp-1-npr-11 pathway.

    (a) Local search behavior 7–12 min after removal from food. RevOmega, coupled reversal-omega behaviors characteristic of local search. AIAdouble colonins-1, ins-1 cDNA expressed under AIA-selective gcy-28.d promoter. Error bars, s.e.m. *P < 0.05, t-test with Bonferroni correction. (b) Fourier power analysis of AWC calcium responses in ins-1 mutants. Left, the normalized energy density spectrum averaged across all calcium traces of each genotype; arrows indicate range of the middle frequency band (color code on right). Right, the average power ratio of the middle frequency band (0.033–1 Hz) across all calcium traces of each genotype; error bars, s.e.m. *P < 0.05 (t-test with Bonferroni correction). (c) Adaptation in ins-1 mutants, and cell-selective rescue. *Different from unadapted control (P < 0.05, t-test). Error bars, s.e.m. (d) AWC calcium responses in wild-type, ins-1 and AIAdouble colonins-1 rescued transgenic worms adapted for 60 min (n = 12 each). Odor pulses are marked. Light gray shading indicates s.e.m. *Different from wild type at P < 0.05, t-test with Bonferroni correction.

References

  1. White, J.G., Southgate, E., Thomson, J.N. & Brenner, S. The structure of the nervous system of the nematode Caenorhabditis elegans . Phil. Transact. R. Soc. Lond. B 314, 1340 (1986).
  2. Chalfie, M. et al. The neural circuit for touch sensitivity in Caenorhabditis elegans . J. Neurosci. 5, 956964 (1985).
  3. Bargmann, C.I. Chemosensation in C. elegans. in WormBook (ed. The C. elegans Research Community) doi:10.1895/wormbook.1.123.1, http://www.wormbook.org (2006).
  4. Bargmann, C.I., Hartwieg, E. & Horvitz, H.R. Odorant-selective genes and neurons mediate olfaction in C. elegans . Cell 74, 515527 (1993).
  5. Wakabayashi, T., Kitagawa, I. & Shingai, R. Neurons regulating the duration of forward locomotion in Caenorhabditis elegans . Neurosci. Res. 50, 103111 (2004).
  6. Gray, J.M., Hill, J.J. & Bargmann, C.I. A circuit for navigation in Caenorhabditis elegans . Proc. Natl. Acad. Sci. USA 102, 31843191 (2005).
  7. Pierce-Shimomura, J.T., Morse, T.M. & Lockery, S.R. The fundamental role of pirouettes in Caenorhabditis elegans chemotaxis. J. Neurosci. 19, 95579569 (1999).
  8. Tsunozaki, M., Chalasani, S.H. & Bargmann, C.I. A behavioral switch: cGMP and PKC signaling in olfactory neurons reverses odor preference in C. elegans . Neuron 59, 959971 (2008).
  9. Chalasani, S.H. et al. Dissecting a circuit for olfactory behaviour in Caenorhabditis elegans . Nature 450, 6370 (2007).
  10. Nathoo, A.N., Moeller, R.A., Westlund, B.A. & Hart, A.C. Identification of neuropeptide-like protein gene families in Caenorhabditis elegans and other species. Proc. Natl. Acad. Sci. USA 98, 1400014005 (2001).
  11. Wenick, A.S. & Hobert, O. Genomic cis-regulatory architecture and trans-acting regulators of a single interneuron-specific gene battery in C. elegans . Dev. Cell 6, 757770 (2004).
  12. Etchberger, J.F. et al. The molecular signature and cis-regulatory architecture of a C. elegans gustatory neuron. Genes Dev. 21, 16531674 (2007).
  13. Mody, S.M., Ho, M.K., Joshi, S.A. & Wong, Y.H. Incorporation of Galpha(z)-specific sequence at the carboxyl terminus increases the promiscuity of galpha(16) toward G(i)-coupled receptors. Mol. Pharmacol. 57, 1323 (2000).
  14. Tallini, Y.N. et al. Imaging cellular signals in the heart in vivo: Cardiac expression of the high-signal Ca2+ indicator GCaMP2. Proc. Natl. Acad. Sci. USA 103, 47534758 (2006).
  15. Tian, L. et al. Imaging neural activity in worms, flies and mice with improved GCaMP calcium indicators. Nat. Methods 6, 875881 (2009).
  16. Lockery, S.R. & Goodman, M.B. The quest for action potentials in C. elegans neurons hits a plateau. Nat. Neurosci. 12, 377378 (2009).
  17. Suzuki, H. et al. In vivo imaging of C. elegans mechanosensory neurons demonstrates a specific role for the MEC-4 channel in the process of gentle touch sensation. Neuron 39, 10051017 (2003).
  18. O'Hagan, R., Chalfie, M. & Goodman, M.B. The MEC-4 DEG/ENaC channel of Caenorhabditis elegans touch receptor neurons transduces mechanical signals. Nat. Neurosci. 8, 4350 (2005).
  19. Clark, D.A., Biron, D., Sengupta, P. & Samuel, A.D. The AFD sensory neurons encode multiple functions underlying thermotactic behavior in Caenorhabditis elegans . J. Neurosci. 26, 74447451 (2006).
  20. Ramot, D., MacInnis, B.L. & Goodman, M.B. Bidirectional temperature-sensing by a single thermosensory neuron in C. elegans . Nat. Neurosci. 11, 908915 (2008).
  21. Mellem, J.E., Brockie, P.J., Zheng, Y., Madsen, D.M. & Maricq, A.V. Decoding of polymodal sensory stimuli by postsynaptic glutamate receptors in C. elegans . Neuron 36, 933944 (2002).
  22. Chronis, N., Zimmer, M. & Bargmann, C.I. Microfluidics for in vivo imaging of neuronal and behavioral activity in Caenorhabditis elegans . Nat. Methods 4, 727731 (2007).
  23. Horoszok, L., Raymond, V., Sattelle, D.B. & Wolstenholme, A.J. GLC-3: a novel fipronil and BIDN-sensitive, but picrotoxinin-insensitive, L-glutamate-gated chloride channel subunit from Caenorhabditis elegans . Br. J. Pharmacol. 132, 12471254 (2001).
  24. Lee, R.Y., Sawin, E.R., Chalfie, M., Horvitz, H.R. & Avery, L. EAT-4, a homolog of a mammalian sodium-dependent inorganic phosphate cotransporter, is necessary for glutamatergic neurotransmission in Caenorhabditis elegans . J. Neurosci. 19, 159167 (1999).
  25. Sieburth, D., Madison, J.M. & Kaplan, J.M. PKC-1 regulates secretion of neuropeptides. Nat. Neurosci. 10, 4957 (2007).
  26. Colbert, H.A. & Bargmann, C.I. Odorant-specific adaptation pathways generate olfactory plasticity in C. elegans . Neuron 14, 803812 (1995).
  27. Colbert, H.A., Smith, T.L. & Bargmann, C.I. OSM-9, a novel protein with structural similarity to channels, is required for olfaction, mechanosensation, and olfactory adaptation in Caenorhabditis elegans . J. Neurosci. 17, 82598269 (1997).
  28. L'Etoile, N.D. et al. The cyclic GMP-dependent protein kinase EGL-4 regulates olfactory adaptation in C. elegans . Neuron 36, 10791089 (2002).
  29. Palmitessa, A. et al. Caenorhabditis elegans arrestin regulates neural G protein signaling and olfactory adaptation and recovery. J. Biol. Chem. 280, 2464924662 (2005).
  30. Matsuki, M., Kunitomo, H. & Iino, Y. Goalpha regulates olfactory adaptation by antagonizing Gqalpha-DAG signaling in Caenorhabditis elegans . Proc. Natl. Acad. Sci. USA 103, 11121117 (2006).
  31. Yamada, K., Hirotsu, T., Matsuki, M., Kunitomo, H. & Iino, Y. GPC-1, a G protein gamma-subunit, regulates olfactory adaptation in Caenorhabditis elegans . Genetics 181, 13471357 (2009).
  32. Kaye, J.A., Rose, N.C., Goldsworthy, B., Goga, A. & L'Etoile, N.D. A 3′UTR pumilio-binding element directs translational activation in olfactory sensory neurons. Neuron 61, 5770 (2009).
  33. Tomioka, M. et al. The insulin/PI 3-kinase pathway regulates salt chemotaxis learning in Caenorhabditis elegans . Neuron 51, 613625 (2006).
  34. Marder, E. & Bucher, D. Understanding circuit dynamics using the stomatogastric nervous system of lobsters and crabs. Annu. Rev. Physiol. 69, 291316 (2007).
  35. Nassel, D.R. & Homberg, U. Neuropeptides in interneurons of the insect brain. Cell Tissue Res. 326, 124 (2006).
  36. Burnstock, G. Cotransmission. Curr. Opin. Pharmacol. 4, 4752 (2004).
  37. Demb, J.B. Functional circuitry of visual adaptation in the retina. J. Physiol. (Lond.) 586, 43774384 (2008).
  38. Stein, W., DeLong, N.D., Wood, D.E. & Nusbaum, M.P. Divergent co-transmitter actions underlie motor pattern activation by a modulatory projection neuron. Eur. J. Neurosci. 26, 11481165 (2007).
  39. Pierce, S.B. et al. Regulation of DAF-2 receptor signaling by human insulin and ins-1, a member of the unusually large and diverse C. elegans insulin gene family. Genes Dev. 15, 672686 (2001).
  40. Kodama, E. et al. Insulin-like signaling and the neural circuit for integrative behavior in C. elegans . Genes Dev. 20, 29552960 (2006).
  41. Ivell, R. & Einspanier, A. Relaxin peptides are new global players. Trends Endocrinol. Metab. 13, 343348 (2002).
  42. Macosko, E.Z. et al. A hub-and-spoke circuit drives pheromone attraction and social behaviour in C. elegans . Nature 458, 11711175 (2009).
  43. Wachowiak, M., Wesson, D.W., Pirez, N., Verhagen, J.V. & Carey, R.M. Low-level mechanisms for processing odor information in the behaving animal. Ann. NY Acad. Sci. 1170, 286292 (2009).
  44. Gomez, C. et al. Heterogeneous targeting of centrifugal inputs to the glomerular layer of the main olfactory bulb. J. Chem. Neuroanat. 29, 238254 (2005).
  45. Ignell, R. et al. Presynaptic peptidergic modulation of olfactory receptor neurons in Drosophila . Proc. Natl. Acad. Sci. USA 106, 1307013075 (2009).
  46. Stein, C. et al. Peripheral mechanisms of pain and analgesia. Brain Res. Brain Res. Rev. 60, 90113 (2009).
  47. Li, C. & Kim, K. Neuropeptides. in WormBook (ed. The C. elegans Research Community) doi:10.1895/wormbook.1.142.1, http://www.wormbook.org (2008).
  48. Li, C., Nelson, L.S., Kim, K., Nathoo, A. & Hart, A.C. Neuropeptide gene families in the nematode Caenorhabditis elegans . Ann. NY Acad. Sci. 897, 239252 (1999).
  49. Kindt, K.S. et al. Dopamine mediates context-dependent modulation of sensory plasticity in C. elegans . Neuron 55, 662676 (2007).
  50. Brenner, S. The genetics of Caenorhabditis elegans . Genetics 77, 7194 (1974).

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Author information

Affiliations

  1. Howard Hughes Medical Institute, The Rockefeller University, New York, New York, USA.

    • Sreekanth H Chalasani,
    • Saul Kato,
    • Dirk R Albrecht,
    • Takao Nakagawa &
    • Cornelia I Bargmann
  2. Department of Neuroscience, Department of Physiology and Cellular Biophysics, Columbia University College of Physicians and Surgeons, New York, New York, USA.

    • Saul Kato &
    • L F Abbott

Contributions

S.H.C. conceived, conducted and interpreted experiments and co-wrote the paper; S.K., D.R.A. and L.F.A. performed and interpreted data analysis; T.N. performed HEK expression experiments; C.I.B. conceived and interpreted experiments and co-wrote the paper.

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The authors declare no competing financial interests.

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