Lateral Line Sensitivity: An Experimental Evolution Approach

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Lateral Line Sensitivity An Experimental
Evolution Approach
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The Lateral Line System (LLS)

• The Sixth Sense of aquatic organisms
• Series of mechanosensory hairs
• Detect displacement
• Two varieties of sensing units
• Superficial Neuromasts
• Canal Neuromasts
• Purpose
• Serve as either velocity or acceleration sensors.
• Predator avoidance, prey detection, near field
  flow regimes etc.

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Research Questions

• What genetic factors influence lateral line
  sensitivity?
• Is the lateral line in fish tuned to certain
  frequencies such that they elicit differing
  responses?

www.termpro.com/ articles/hearing.html
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Model System The Zebrafish

• Known genome
• Commercially available pre-printed mirco-arrays
  for multiple stains.
• Vertebrate model of intense research
• Hardy animal
• Breeds rapidly in captivity

www.nbb.cornell.edu/ neurobio/Fetcho/ dev.biologis
ts.org/.../ vol132/issue2/cover.shtmlwww.hci.utah
.edu/.../ trede/zebrafishFacility.jsp
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Previous Studies

• Nicolson T, Rusch A, Friedrich RW, Granato M,
  Ruppersberg JP, Nusslein-Volhard C (1998) Genetic
  analysis of vertebrate sensory hair cell
  mechanosensation the zebrafish circler mutants.
  Neuron Feb 20(2)271-83.
• Driever W, Solnica-Krezel L, Schier AF, Neuhauss
  SC, Malicki J, Stemple DL, Stainier DY,
  Zwartkruis F, Abdelilah S, Rangini Z, Belak J,
  Boggs C. (1996) A genetic screen for mutations
  affecting embryogenesis in zebrafish.
  Development. Dec 12337-46.
• Gompel N, Dambly-Chaudiere C, Ghysen A. (2001)
  Neuronal differences prefigure somatotopy in the
  zebrafish lateral line. Development.
  Feb128(3)387-93.

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Mutant Lines

• Morpholinos are now available for highly
  selective gene knockdown in zebrafish.
• Mutant lines which affect the LLS include
• Sputnik, Mariner involve mechanosensory hair
  bundle integrity
• Orbiter, Gemini, and Mercury involve receptor
  potentials
• Astronaut, Cosmonaut Everything seems fine, but
  still signals are not being correctly processed

www.eb.tuebingen.mpg.de/.../ nicolson/home.html
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Problems with the knockdown method

• LLS or inner ear?
• Binary, no gauge on sensitivity
• Ignores emergent properties
• Blind to evolutionary framework
• Of lesser ecological importance

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Experimental Evolution another possibility

• Would correct for many of the missing pieces from
  knockdown experiments.
• Has proved successful in organisms of similar
  reproductive lifespan
• Garlands mice 2 months to maturaty 2 week
  gestation
• Zebrafish 3 months to maturity after
  fertilization
• Currently in practice with zebrafish regarding
  oxygen stress
• Moore, FBG, Bagatto, B (2003) Evolutionary
  response to exygen stress in zebrafish (Danio
  rerio) An experimental evolution approach. Int.
  Comp. Bio., 43 (6) 1071-1071

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Selection Criteria

• Artificial Truncation Selection for LLS
  sensitivity
• Startle Response
• Fast Start
• Stimuli
• Frequency specific to LLS
• Variable amplitude

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Experimental Setup
Larvae Fish

• Selection arena
• Selection of Larvae
• Two Lines
• No fast start response at high amplitude
• Response at lowest amplitude
• As response sensitivities change signal intensity
  will be adjusted
• Continue Selection for 20 generations

Vibrating Probe
Barrier
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Confounding Factors

• Even if a change in response (via fast start) is
  found we cannot, a priori, know with any
  certainty that this has anything to do with the
  LLS
• How can we tell if our fish just got paranoid or
  dull?

http//www.disney.fr/FilmsDisney/nemo/
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Behavioral Tests

• Test startle response in non-LLS involved ways
• Visual startle
• Test other LLS behavioral functions
• Schooling efficiency
• Detection of Flow change

Larvae Fish
Cylinder With Black Stripe
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Morphological Tests

• Are there any differences in the morphology of
  the lateral line?
• Differences in size, number, spacing, location
  etc.
• Check for similar responses to those seen in the
  mutant lines.

www.eb.tuebingen.mpg.de/.../ nicolson/home.html
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Physiological Tests

• Test the relative neural responses to stimulus in
  the two lines of evolved zebrafish.

Coombs, S., Hastings, M., Finneran J. (1995). J.
Comp. Physiol. 178, 359-371.
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Microarray Analysis

• Extract RNA from LLS cells and other neural
  centers.
• Competitively hybridize two evolved lines onto a
  commercially available printed microarray for
  original zebrafish line.
• Locate genes of up and down regulation
• Identify corollaries to mutated lines

www.research.vt.edu/.../s_maroof_lab.html
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Selected References

• Driever W, et. al. (1996) A genetic screen for
  mutations affecting embryogenesis in zebrafish.
  Development. Dec 12337-46.
• Coombs, S., Görner, P., Münz, H. editors (1989).
  The Mechanosensory Lateral Line. Springer-Verlag.
  
• Coombs, S., Hastings, M., Finneran J. (1995). J.
  Comp. Physiol. 178, 359-371.
• Gompel N, et. al. (2001) Neuronal differences
  prefigure somatotopy in the zebrafish lateral
  line. Development. Feb128(3)387-93.
• Moore, FBG, Bagatto, B (2003) Evolutionary
  response to exygen stress in zebrafish (Danio
  rerio) An experimental evolution approach. Int.
  Comp. Bio., 43 (6) 1071-1071
• Nicolson T, et. al. (1998) Genetic analysis of
  vertebrate sensory hair cell mechanosensation
  the zebrafish circler mutants. Neuron Feb
  20(2)271-83.
• Nüsslein-Volhard, C., and Dahm, R. editors
  (2002). Zebrafish A Practical Approach. Oxford
  Univeristy Press.

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Questions?