Ir-rat-ability and Anxiety in the Amygdala Team Four: Megha - - PowerPoint PPT Presentation

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Ir-rat-ability and Anxiety in the Amygdala Team Four: Megha - - PowerPoint PPT Presentation

Ir-rat-ability and Anxiety in the Amygdala Team Four: Megha Andrews, Madlyn Kates, Christine Lu, Katherine Miao, Archana Raghunath, Angeli Sharma, Grace Shen, Michael Tai, Mary Tresvalles, Elena Wei, Destiny West, Audrey Zhou Anxiety/Anxiety


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Team Four: Megha Andrews, Madlyn Kates, Christine Lu, Katherine Miao, Archana Raghunath, Angeli Sharma, Grace Shen, Michael Tai, Mary Tresvalles, Elena Wei, Destiny West, Audrey Zhou

Ir-rat-ability and Anxiety in the Amygdala

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Anxiety/Anxiety Disorders

  • Anxiety-like behaviors vs. anxiety

disorders

  • One of the most prevalent classes of

mental illness

  • Fear, uneasiness, dread, irritability
  • GAD, Social Anxiety, Phobias, PTSD
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Purpose

INVESTIGATE THE ROLE OF THE AMYGDALA/AMYGDALOID COMPLEX IN ANXIETY LIKE BEHAVIORS Predator odor used as model for anxiety disorders

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Why are we interested in the amygdala?

  • Motivation, making

decisions, and processing emotions.

  • Area of interest:

corticomedial amygdala (CMA).

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Phasic VS Sustained Fear:

Phasic Fear Sustained Fear

Fades quickly Pervasive Visible Threat Potential Unseen Danger

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The amygdala receives input from multiple olfactory systems:

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Experiments

Open Maze Acoustic Startle Test Electrophysiology

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  • Analyze aversion
  • Predator pheromone odors, Preferential

Pheromone, & Control Odor

Experiment 1: Maze

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Experiment 1: Maze Procedure

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The rats surprisingly spent more time in the zones containing the predator pheromones.

Experiment 1: Maze Data

* * *

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Experiment 1: Maze Data

In the general the rats moved in greater speeds in the presence of the

  • dors.

* * * *

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On average the rats stayed closer to the points containing the predator odors. Overall it seems that the rats displayed the most aversion to the pheromones from the cat urine and fox urine.

Experiment 1: Maze Data

* * * *

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Experiment 1: Maze Results

  • Rats exhibited greater time in zone, faster speed, and

lesser distance from point for cat urine and fox urine

  • No significant difference in zones with propyl butyrate
  • Does not show aversion to rat and cat urine but rather

suggest that rats investigated the odorants

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Experiment 2: Startle Test

  • Monitor the rats’ acoustic

startle responses

  • Compare the standard,

“baseline” response to when the pheromones/ odorants are released (presence of anxiety).

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Experiment 2: Startle Procedure

3 Handling Sessions Baseline Test 1 Baseline Test 2 Baseline Test 3 Olfactory Startle Test 1 Olfactory Startle Test 2 Baseline Test 4 Olfactory Startle Test 3 Olfactory Startle Test 4

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Experiment 2: Baseline Startle Test Procedures

▪ Purpose) To test equipment as well as record initial startle responses of each subject without odor present ▪ Acclimation period of 2 minutes ▪ 40 sound pulses of varying intensities played in a pseudo-random order ▫ Played in 15 second intervals ▫ Order kept constant for all subjects

3 Handling Sessions Baseline Test 1 Baseline Test 2 Baseline Test 3 Olfactory Startle Test 1 Olfactory Startle Test 2 Baseline Test 4 Olfactory Startle Test 3 Olfactory Startle Test 4

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Experiment 2: Baseline Startle Data

shows that startle amplitude is influenced by pulse intensity. Habituation is evident by the decreasing amplitude.

* *

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The decrease in response across sessions shows evidence of habituation

Experiment 2: Baseline Startle Results

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Experiment 2: Olfactory Startle Test Procedures

▪ Odorants) fox urine, cat urine, female rat urine, propyl butyrate ▪ Acclimation period of 5 minutes ▪ 70 sound pulses of 95 dBA played in 15 second intervals ▫ On 31st sound pulse, odorant is first introduced ▪ Every 4th sound pulse after 31, odorant is reintroduced into chamber (35th pulse, 39th pulse, etc.) ▪ Order of odorants counterbalanced to avoid repeated testing 3 Handling Sessions Baseline Test 1 Baseline Test 2 Baseline Test 3 Olfactory Startle Test 1 Olfactory Startle Test 2 Baseline Test 4 Olfactory Startle Test 3 Olfactory Startle Test 4

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Trial Structure

5 minute acclimation 30 baseline trials 40 odorant trials

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Olfactory sessions represented as a percentage of the baseline.

Cat and female rat urine showed similar results as the control.

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Fox urine was the only odorant that produced a noticeable effect.

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Experiment 2: Olfactory Startle Conclusion

  • Cat urine and female rat urine

cause no significant anxiety induced response.

  • Propyl butyrate (the control
  • dorant) causes a significant

increased response which indicates that there was another factor involved.

  • Fox urine data is evident of a

phasic fear response.

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Experiment 3: Electrophysiology

▪ Objective: ▫ Record electrical activity of neurons in the corticomedial amygdala ▫ Determine its connection to pheromone processing

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Experiment 3: Electrophysiology Procedure

Olfactometer

  • Cat urine
  • Fox urine

Thermocouple Electrode E Oscilloscope Amplifier Computer

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Experiment 3: Electrophysiology Procedure

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Experiment 3:Electrophysiology Procedure

Olfactometer

  • Cat urine
  • Fox urine

Thermocouple Electrode E Oscilloscope Amplifier Computer Data Analysis

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Analysis of electrophysiology data yields two distinct waveforms.

Trough

  • f action

potential

Crest of action potential Unit A Unit B

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Results from a previous experiment that would corroborate our hypothesis that the detected cells process specific odors.

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Our results did not match. Neither control nor experimental odors elicited an obvious response from the detected cells.

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Experiment 3: Electrophysiology Conclusion

❖Sample Cell ➢showed both odor responsivity and specificity ❖Our 3 Tested Cells ➢Neither odor specificity or responsivity ❖Sample size did not permit an accurate estimation of the population of corticomedial amygdala cells.

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Conclusion

❖Most of the results did not align with our hypothesis, that predator pheromones would have an effect on the neurons of the corticomedial amygdala in processing

  • lfactory information.

❖Main Findings ➢Open Maze ➢Acoustic Startle ➢Electrophysiology ❖Possible Sources of Error ❖Future Implications

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References

1. Bind RH. The role of pheromonal responses in rodent behavior: future directions for the development of laboratory protocols. Journal

  • f the American Association for Laboratory Animal Science. 2013: 124–129. Available from:

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3624779/ 2. Curzon P, Zhang M, Radek RJ. The behavioral assessment of sensorimotor processes in the mouse: acoustic startle, sensory gating, locomotor activity, rotarod, and beam walking. Methods of Behavior Analysis in Neuroscience. 2009 [Internet]. [cited 2015 Jul 27] 2nd

  • edition. Boca Raton (FL): CRC Press; 2009. Chapter 8. Available from: http://www.ncbi.nlm.nih.gov/books/NBK5236/

3. Davis M, Walker D, Miles L, Grillon C. Phasic vs sustained fear in rats and humans: role of the extended amygdala in fear vs anxiety.

  • Neuropsychopharmacology. 2009: 105-135. Available from: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2795099/

4. Harris A, D'eath R, Healy S. Environmental enrichment enhances spatial cognition in rats by reducing thigmotaxis (wall hugging) during testing. Animal Behaviour: 1459-1464. 5. Koch M. The neurobiology of startle. Prog Neurobiol. 1999 Oct; 59(2): 107-28. Available from: http://www.sciencedirect.com/science/article/pii/S0003347209001183 6. Lowry R. One-way ANOVA. VassarStats. 2015; [Internet]. [cited 27 July 2015]. Available from: http://vassarstats.net/anova1u.html 7. Stoelting ANY-maze flexible video tracking for neuroscience experiments. [updated 2011]. [cited 2015 Jul 26]. Available from: http://www.anymaze.com/ 8. Takahashi L. Olfactory systems and neural circuits that modulate predator odor fear. Frontiers in Behavioral Neuroscience. 2014; [Internet]. [cited 2015 Jul 28]: 1-13. Available from: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3949219/ 9. Valsamis B, Schmid S. Habituation and prepulse inhibition of acoustic startle in rodents. Journal of Visualized Experiments. 2011; [Internet]. [cited 2015 Jul 27]: (55): 3446. Available from: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3217252/

  • 10. Xu, Wenjin, Donald A. Wilson. Odor-evoked activity in the mouse lateral entorhinal cortex. Neuroscience. 2012; [Internet]. [cited 2015

Jul 29]: 12–20 p. Available from: http://www.sciencedirect.com/science/article/pii/S0306452212008159

  • 11. [updated 2015]. Silver Spring (MD): Understanding the Facts | Anxiety and Depression Association of America, ADAA.: [cited 2015

Jul 29] Available from: http://www.adaa.org/understanding-anxiety

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Acknowledgements

  • Dr. Graham Cousens
  • Runi Patel
  • Dr. Cassano and Dr. Surace
  • Our Sprague-Dawley rats:

○ (MAZE) Elliot the Scrub Rat, Simon, Michael, George, Russell, Mercutio, Remy ○ (STARTLE) Ace, Felix, Tucker, Mrs. Frisby, Stitch, Jerry, Finnegan, Julius ○ (ELECTROPHYSIOLOGY) Ratsby, Sirius RIP

  • Funders & Supporter of NJGSS:

○ AT&T, Bayer Healthcare, Independent College Fund of New Jersey, Johnson & Johnson, The Overdeck Family Foundation, NJGSS Alumnae and Parents, Corporate Matching Funds, The State of New Jersey, Board of Overseers, New Jersey Governor’s School

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