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Neuroscience Cyber Technology Laboratory Manual

Terence Bazzett

SUNY Geneseo

2013

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© Terence Bazzett Page 1


Table of Contents


Chapter 1: Organelle Staining

Introduction: A wide range of cell staining techniques are used in Neuroscience. Simple staining techniques can be used to visualize neuronal cell bodies, neuronal projections, or glial cells. More sophisticated techniques can also be used to visualize functional elements of cells. In particular, most organelles within the cell can be individually stained allowing researchers to visualize, and in some cases quantify, the integrity of intracellular structures. Characterizing changes within neurons during the progression of neurodegenerative diseases can be invaluable for verification of a diagnosis. Furthermore, if those changes are found to be contributing factors to the degenerative process, they may become a starting point for testing treatments to slow or halt progression of an illness. As such, research designed to elucidate organelle changes in degenerative disorders is a potentially valuable form of applied research in Neuroscience. Read the following article, paying particular attention to the Introduction and Discussion sections, which provide the rationale for the methods used. Within the Experimental Procedures section focus on the Immunocytochemistry section which gives a brief summary of the methodology required for staining of the Golgi apparatus and the mitochondria within the neuron.

Background reading: Nakagomi, Barsoum, Bossy-Wetzel, Sütterlin, Vivek Malhotra, and Liptona (2008). A Golgi fragmentation pathway in neurodegeneration. Neurobiology of Disease, 29, 221-231.


ERIN resource:Authors: InvitrogenTopic / Subtopic: B. Neural Excitability, Synapses, and Glia: Cellular Mechanisms; I. Neuroanatomy / I.05. Anatomical Techniques NeuroanatomyResource Type: Image - Photograph, SimulationEducational Level: 1 Introductory undergraduate, 2 Intermediate undergraduate, 3 Advanced undergraduateListing Date: October 23, 2012http://www.invitrogen.com/site/us/en/home/support/Research-Tools/Cell-Staining-Tool.html?CID=fl-cellstaintool

Although the images are not from a neuron, this clever and realistic simulation allows students to stain specific cellular structures using fluorescent antibodies. Up to four structures can be stained simultaneously. Stains can be deleted so that other structures can be stained instead, a flexibility that encourages exploration. Cellular structures include the membrane, cytoskeletal components, mitochondria, the nucleus, Golgi, ER, and others. Using real images rather than diagrams makes the simulation especially persuasive.

Lab exercise: Consider the immunocytochemistry methods described by Nakagomi et al. Use the resource link above to select appropriate stains for this laboratory preparation. Note: you do not need to use the same stains cited in the Nakogomi et al. article, but you must select stains that would be appropriate for your sample.

Student quiz questions:1. When is Golgi fragmentation anticipated and possibly even functional within the cell?

Mitosis

2. Nakagomi and colleagues cite Gonatas et al., 2006 when noting that Golgi fragmentation is believed to be associated with neuronal degeneration in what two disorders?a. Alzheimerb. ALS

3. Using the immunocytochemistry methods of Nakagomi et al., approximately how much time would you anticipate needing to complete the staining procedure used in their paper?a. 5 minutesb. 15 minutesc. 1 hourd. 14-16 hours e. 16-19 hours


http://www.invitrogen.com/site/us/en/home/support/Research-Tools/Cell-Staining-Tool.html?CID=fl-cellstaintool

http://www.invitrogen.com/site/us/en/home/support/Research-Tools/Cell-Staining-Tool.html?CID=fl-cellstaintool

4. In Nakagomi et al. fragmentation was quantified after Golgi staining. What was the effect of NMDA on the fragmentation process?a. Increased fragmentationb. Decreased fragmentationc. Had no effect on fragmentation

5. In your own words, describe how the image of the Golgi apparatus from the control group differs from that of the image of the Golgi apparatus from the NMDA group.

The staining is more diffuse (fragmented) in the NMDA image.

6. What is general effect of NMDA on cell activity?a. Increases activityb. Decreases activityc. Has no effect on activity

7. In the experiment by Nakagomi et al., NMDA exposure was used to mimic which of the following? (circle all that apply)a. Effects of a neurodegenerative disorderb. Excitotoxicityc. Neuroprotectiond. Neuronal sproutinge. Mitosis

8. Which green Golgi stain(s) available from Invitrogen would be appropriate to use for this experiment?

Golgin-97 is the only stain for fixed tissue as describe in the article.

9. Which orange-red mitochondrial stain(s) available from Invitrogen would be appropriate to use for this experiment?

The two Oxphos stains are the only stains for fixed tissue as described in the article.

10. What doe Nakagomi and colleagues suggest can be accomplished by inhibiting Golgi fragmentation such as that seen in response to NMDA stimulation?

They believe it can dlay apoptiotic-like cell death

11. Generate a useful, concise question from the background reading for this Chapter.

12. Generate a useful, concise question from the cyber laboratory exercise for this Chapter.


Chapter 2: Epigenetics

Introduction: There has been long standing debate about the relative importance of genetics versus the importance of environment on phenotype expression. Since the origins of this “nature vs. nurture” debate, many researchers have argued vehemently that the one or the other of these factors reigns as most influential. In particular, the famous behavioral psychologist John B. Watson is oft cited for his 1930 statement promoting the importance of environmental influences:

"Give me a dozen healthy infants, well-formed, and my own specified world to bring them up in and I'll guarantee to take any one at random and train him to become any type of specialist I might select–doctor, lawyer, artist–regardless of his talents, penchants, tendencies, abilities, vocations and race of his ancestors"

More recently, there seems to be a growing consensus among researchers that, for most behaviors, it may be difficult to determine a comparable difference in contributions of genetics and environmental influences. Perhaps more importantly, researchers are now finding that an equally influential factor might, in some cases, be the effect of a an interaction between these two elements. Epigentics is the study of modifications to the genome that do not involve a change in the nucleotide sequence. Such modifications are brought about by variables in the environment in which the genome resides. Such variables may, in turn, be directly influenced by the environment in which an organism resides, behaviors of the organism, experiences of the organism, and so on.

Background reading:


Roth and Sweatt (2011). Annual Research Review: Epigenetic mechanisms and environmental shaping of the brain during sensitive periods of development. J Child Psychology and Psychiatry 52, 398–408.

ERIN resource:Authors: Genetic Science Learning Center, University of UtahTopic / Subtopic: A. Development / A.02. Neurogenesis and Gliogenesis, A.03. Postnatal Neurogenesis, A.06. Synaptogenesis and Activity-Dependent Development; C.Disorders of the Nervous System / C.16. Cognitive, Emotional, and Behavioral State Disorders; E. Homeostatic and Neuroendocrine Systems / E.01. Neuroendocrine Processes, E.03. Behavioral Neuroendocrinology, E.04. Autonomic Regulation, E.05. Stress and the Brain; F. Cognition and Behavior / F.02. Language; G. Novel Methods and Technology Development / G.07. Computation, Modeling, and SimulationResource Type: Animation, SimulationEducational Level: 1 Introductory undergraduate, 2 Intermediate undergraduateListing Date: March 22, 2012http://learn.genetics.utah.edu/content/epigenetics/

A series of web links to readings and interactive videos describing epigenetics. Links provide information directly related to the assigned reading. Emphasis is on developmental periods in animals, but information about lifespan changes and human research is also included.

Lab exercise: After completing the background reading, begin by watching the video “The Epigenome at a Glance” followed by the interactive lab “Gene Control”. These two exercises will give you some basic information on epigenomics. Next, view the video “Insights from Identical Twins” and finish the lab by completing the interactive lab entitled “Lick Your Rats”. In this exercise, take the time to lick several rat pups with a range of vigor. Begin with what you feel would be a “normal” level of licking. Then try another pup with very vigorous licking. Finally, try a pup with only a few licks. See what differences occur in the epigenome, and listen to the explanation given for your licking activity in each case.

Student quiz questions:1. From the Roth and Sweatt (2011) article, a rat model of an “abusive” caretaker, is described.

In this model, what behaviors exhibited by mother rats toward their pups that are considered, and quantified as, abusive.

Frequent stepping on, dropping, dragging, actively rejecting, rough handling.


http://learn.genetics.utah.edu/content/epigenetics/

2. From the Roth and Sweatt (2011) article, rats raised by mothers who displayed high levels of pup licking/grooming had significantly less methylation of glucocorticoid receptor DNA in their ____ than did rats raised by mothers who displayed low levels of pup licking/grooming.a. Amygdalab. Hypothalamusc. Hippocampusd. Thyroid glande. Entire brain

3. have been found to show decreases in which of the following?a. Glucocorticoid receptor mRNAb. Dopamine receptor mRNAc. Cell numberd. Tissue volumee. All of the above

4. From the movie, The Epigenome at a Glance, you learned that epigenome does which of the following? a. Changes the DNA codeb. Alters the sequence of genes within the DNAc. Stimulates DNA self-replicationd. Shapes the physical structure of the DNAe. All of the above

5. While the epigenome in identical twins may initially be nearly identical, factors throughout life can create significant differences between the epigenome of twins as they age. As a result, while identical twins may continue to share physical features established during early development, behavioral characteristics tend to be less similar over time. From the laboratory video, list the four categories of environmental factors that were cited as potentially contributing to such epigenomic changes.

1. Diet2. Physical Activity3. Toxins4. Stress

6. From the interactive lab Gene Control, you learned that growth of cancerous tumors is mediated in part by out of control cell growth that has two primary effects on gene coding. Describe those two effects in very basic terms.


Out of control growth leads to cancer by both turning off genes coding for proteins that slow cell growth, and turning on genes coding for proteins that speed up cell growth.

7. From the interactive lab Gene Control, list the effects on the following factors when you increase and decrease the external signal using the control knob.

Stimulation Increase Decrease

GFP Gene Structure coils/tightens uncoils/loosens

GFP mRNA Increases Decreases

GFP Increases Decreases

Cell image Brightens Dims

8. List from highest percentage (1) to lowest percentage (5), the traits listed below that may be shared by identical twins. 2 Autism 4 Bipolar disorder 1 Height 3 Hypertension5 Stroke

9. In adulthood, rats that were vigorously licked as pups tend to have more GR protein in the hippocampus. This is particularly important because GR protein binds circulating _______________ released during stressful periods, and helps to reduce the brain’s stress response resulting in a calmer, generally more adaptive, animal.a. Dopamineb. Glutamatec. Cortisold. Adrenalinee. Oxytocin

10. After completing the Lick Your Rat interactive lab, briefly describe the difference in appearance of the epigenome in a pup that was licked vigorously versus a put that received very little licking. Also describe the primary behavioral difference that will be seen between these two rats as a result of this licking, when they become adults.


Vigorous Licking Little Licking

Epigenome appearance Loosely coiled Tightly coiled

Adult Behavior Easily relaxes Does not relax easily




Chapter 3: Pedigree Mapping

Introduction: Understanding inheritance patterns of disorders is one of the first, and most basic, steps in establishing genetic influences that may cause, or contribute to a disorder. Generating a family pedigree chart allows researchers to summarize information about inheritance patterns over several generations. The pedigree chart is also an easily interpreted visual aid that offers a concise summary of basic and essential information. In simple terms, a pedigree chart is a great example of the old adage “a picture is worth a thousand words”. In reviewing a simple pedigree chart, researchers can identify, at a glance, family members as living or deceased, male or female, affected or unaffected, and so on. Pedigree charts are generally considered a staple for studying qualitative traits, such as single gene disorders. However, they also provide useful information for studying quantitative traits, including diseases that are influence by multiple genes. With the advent of the Human Genome Project, and the subsequent increase in the prevalence and utility of pedigree charts, the National Council of Genetic Counselors assembled a Pedigree Standardization Task Force (PSTF) in the early 1990s and charged its members with creating a universal coding system of symbols to be used in pedigree charts. The standardization of pedigree symbol usage helped legitimize the use of these charts for medical and research purposes. Today, regardless of the area of neuroscience, the pedigree chart is a useful if not essential tool for understanding inheritance patterns, co-morbidity, genetic contributions and more.

Background readings: Ahearn, Steffens, Cassidy, Van Meter, Provenzale, Seldin, Weisler, & Krishnan (1998). Familial leukoencephalopathy in bipolar disorder. American Journal of Psychiatry, 155, 1605-7.


Bennett, Steinhaus, Uhrich, O'Sullivan, Resta, Lochner-Doyle, Markel, Vincent, & Hamanish (1995). Recommendations for standardized human pedigree nomenclature. American journal of human genetics, 56, 745-52.

ERIN resource:Authors: Genetic Science Learning Center, University of UtahTopic / Subtopic: B. Neural Excitability, Synapses, and Glia: Cellular Mechanisms / B.01. Neurotransmitters and Signaling Molecules, B.02. Ligand Gated Ion Channels, B.03. G-Protein Linked Receptors, B.06. Neurotransmitter Release, B.07. Synaptic Transmission; C.Disorders of the Nervous System / C.17. Drugs of Abuse and Addiction, C.18. Behavioral Pharmacology; H. Teaching, History and Societal Impacts of Neuroscience / H.03. Public Awareness of NeuroscienceResource Type: AnimationEducational Level: 1 Introductory undergraduate, 2 Intermediate undergraduate, 3 Advanced undergraduateListing Date: March 22, 2012http://learn.genetics.utah.edu/content/addiction/genetics/

This web site is expansive, with many related web links. You are encouraged to browse about some of the other links when your assignment is complete. There are audio links, some video links of related lectures, several animated exercises (including the Pedigree Investigator lab assigned for this chapter), and links to other related sites.

Lab exercise: Read the Ahearn et al., article as background information and to help you understand how familial studies are conducted and how pedigree charts can be used to represent results. You do not need to read the Bennett et al. article, but instead you should use it to help develop a greater understanding of the depth and utility of pedigree charts. Understanding the possible complexity of such charts is important for this lab assignment, since Ahearn et al., and the cyber lab both use highly simplistic representations that may give an overly simplistic impression of this area of research. After reading the Adhearn et al. article and reviewing Bennett et al., visit the web link listed above and from there the Pedigree Investigator link. Read all of the information associated with the Pedigree Investigator lab exercise and view all interview videos. After completing the Pedigree Investigator lab exercise, return to the original web link page, and listen to both audio recordings by Dr. Glen Hanson.

Student quiz questions:


http://learn.genetics.utah.edu/content/addiction/genetics/

1. Adhearn et al. (1998) note the use of SCID in their research. What does SCID stand for?

Structured Clinical Interview for DSM III-R

2. From the Ahearn et al. (1998) article, it was noted that Dupont, Jernigan and Butters were the first to report white matter hyperintensities in a large number of young patients diagnosed with bipolar disorder. In what year was that early report published?a. 1980b. 1985c. 1990d. 1995e. The year of that publication is not known

3. In reviewing the pedigree chart from Adhearn et al. (1998), indicate the following:Number of males showing Unipolar Disorder symptoms __3___Number of deceased females showing Bipolar symptoms __1__Number of living females showing Unipolar Disorder symptoms __4__Number of sets of twins evaluated__0__

4. What was the earliest age of Bipolar Disorder symptom onset for a participant in this study? At what age was that participant interviewed for this pedigree. Is that participant male or female?

Age of onset __8 years old__Age at time of interview __20 years old__Sex of participant __Female__

5. When gathering information from a client about severe combined immune deficiency syndrome (SCIDS) you are told that she had elected to have an earlier pregnancy terminated when it was discovered through a genome analysis that the fetus she was carrying tested positive for SCIDS. With consideration to Bennett et al. (1995), what would be the appropriate symbol to use in your pedigree chart to note this pregnancy termination if the fetus was female? What would be the symbol used if the fetus was male? (note, symbols can be created using the Insert Shapes function of your Word program).

Female Male

6. In some cases, dizygotic twins may closely resemble each other in physical appearance, making it difficult to determine if they are dizygotic or monozygotic. Actresses Mary Kate


?

and Ashley Olsen, for example, are often believed to be monozygotic twins, but are reportedly dizygotic. Without genetic testing to positively establish twin status, pedigree charts may necessarily include some level of uncertainty. With consideration to Bennett et al. (1995) what would be the appropriate way to represent twin sisters if zygote status is unknown? Also indicated that one of the sisters is affected and the other unaffected for the condition of interest.

7. From the cyber technology laboratory entitled Pedigree Investigator, which of the following most closely represents the explanation for why researchers typically examine multiple affected families when studying genetic influences on a complex trait such as addiction?a. It is possible that multiple genes contribute and these genes may vary between familiesb. Families may withhold information, particularly about sensitive traits such as addiction. c. When multiple families are used, they may benefit from forming social support groups.d. Multiple families increase the number of participants and therefore the statistical powere. All of the above are noted in the lab exercise

8. From the cyber technology laboratory entitled Pedigree Investigator, are Todd and Alex monozygotic or dizygotic twins? What was their age at the time of the interview? Which of the twins shows signs of the addiction trait? Do either of the twins report treatment for ADHD?

Monozygotic or DizygoticAge __31__Addiction trait ___Both show this trait__ADHD treatment __Both were treated for ADHD__

9. From the cyber technology laboratory entitled Pedigree Investigator, what is the approximate number of cigarettes Tess reports smoking in a day? What is her age at the time of the interview? Which of her parent’s reported being a tobacco user?

Number of cigarettes ___A pack a day__Age __16__Parent who uses tobacco __Father__

10. In the audio clips associated with this laboratory exercise, Dr. Glen Hanson notes that genetics may contribute to the addiction process. He then continues that thought, suggesting


a particular brain region that might be associated with the decision making process that leads to drug usage, and continuation of drug use. What is the brain regions cited by Dr. Hanson as possibly contributing to drug use and addiction in this way?

Frontal Cortex




Chapter 4: Basic Neurophysiology

Introduction: A clear understanding of basic neurophysiological mechanisms is essential when trying to learn, or teach, elements of more complex neural functioning. Included in this basic information is a simplistic description of neural membrane structure, an explanation of how voltage and ligand sensitive protein channels contribute to neural signaling, and a description of agonist and antagonist effects. Beyond these elements, a well prepared student should also be familiar with the concept of functional plasticity in the nervous system. This concept was eloquently represented in the seminal work of Eric Kandel who demonstrated the physiological basis of rudimentary learning (i.e. habituation, sensitization) in the aplysia.

One feature common to all areas of study in neurophysiology is the dynamic nature of the phenomenon that are described. Written descriptions may clearly present the elements of the activity, but it is in fact the activity itself that is focus of physiological function. As an analogy, imagine writing a paper, or giving a lecture, trying to describe to students the elements of classical ballroom dancing, traditional tap dancing, or modern jazz dancing. Such descriptions, whether written or presented orally, would be profoundly improved if the students were actively engaged in the dance forms, or at the very least could view examples of the each form of dance.

The present exercise has no require background reading, but you are strongly encouraged to use one of your own textbooks that describes basic neurophysiological functions as a guide. Read a basic description of the activities presented in the animations for this lab. Imagine the activities that are described in the written work. Then view the video animations. Evaluate that animations to see how they compare to your imagined explanation. Try to identify strengths and weaknesses in the animation series. Think actively about how you could integrate these animations into a lecture designed to teach basic neurophysiological functioning to your peers.


Background reading: For background or supplemental reading, students should use a basic Neuroscience, Biopsychology, or Neurophysiology textbook.

ERIN resource:Authors: Sumanas, Inc.Topic / Subtopic: Neural Excitability, Synapses, and Glia: Cellular Mechanisms / B.00. Action Potentials, B.01. Neurotransmitters and Signaling Molecules, B.02. Ligand Gated Ion Channels, B.04. Ion Channels, B.07. Synaptic Transmission, B.08. Synaptic Plasticity, B.10. Intrinsic Membrane PropertiesResource Type: AnimationEducational Level: 1 Introductory undergraduate, 2 Intermediate undergraduate, 3 Advanced undergraduate, 4 GraduateListing Date: March 20, 2012http://www.sumanasinc.com/webcontent/animations/neurobiology.html

A set of animations that accompany Purves et al. (2008) "Neuroscience" (4th edition). This site provides a very nice series of animations, several of which describe basic elements of neurophysiology. You are encouraged to view the other animations to determine if they have any value as supplemental materials for other courses you may be taking.

Lab exercise: For the present exercise, view the following video animations. It may be helpful, though not essential, to view the videos in the following order: 1) The Resting Membrane Potential 2) The Action Potential 3) Synaptic Transmission 4) AMPA and NMDA Receptors 5) Sensitization in the Aplysia. Ideally, you should view each animation in full, and then answer the quiz questions associated with that video. After completing the quiz questions, review the video to check your answers. In doing this, you will be able to identify areas where you may require additional information for a complete understanding. For any questions that remain unclear after reviewing the video animations, refer to your background textbook for a written explanation that may be more complete than the information provided in the animation.

Student quiz questions:1. By definition, when is equilibrium achieved? Are ions in neuronal intracellular fluid at

equilibrium with ions in the extracellular fluid? If not, describe the imbalance.

Equilibrium is reached when ion concentration is equally distributed. The intracellular and extracellular fluids are not at equilibrium for ions. The intracellular fluid has a net negative charge compared to the extracellular fluid


http://www.sumanasinc.com/webcontent/animations/neurobiology.html

2. Neuronal membrane protein channels designated as “potassium” channels also allow which of the following ions to pass through the membrane?a. Sodiumb. Chloridec. Calciumd. All of the above since these channels are relatively nonselectivee. None of the above since these channels are highly selective

3. In a resting neuron, potassium channels may be in an open state. Considering that potassium is in much higher concentration inside of the cell relative to outside of the cell, why does diffusion fail to bring intracellular and extracellular potassium to equilibrium?

Intracellular electrical gradient keeps intracellular potassium in higher concentration. Sodium potassium pump may also contribute.

4. Depolarizing a cell membrane to its threshold level triggers events that evoke an action potential. What is the response of the cell when the membrane is depolarized beyond threshold (suprathreshold) level?a. An action potential of greater amplitude is seenb. An action potential of longer duration is seenc. An action potential comparable to that produced by a threshold depolarization is seend. The membrane enters a tonic state, and no action potential is seene. The membrane enters a state of reverse polarization and hyperpolarizes

5. List the six stages of the action potential.1. Resting phase 2. Rising phase 3. Overshoot 4. Falling 5. Undershoot 6. Recovery

6. Not all vesicles in the synapse are free floating and ready for immediate use. Some vesicles are actually held in reserve. Where are these reserve vesicles docked, or tethered?a. On the myelinb. Within the somac. On the cytoskeletond. On membrane transporter proteinse. On the mitochondria


7. Synaptic vesicles are drawn together at the terminal membrane by protein complexes known as “SNAREs”. SNARE is an acronym for what?

SNARE is an acronym derived from "SNAP (Soluble NSF Attachment Protein) REceptor"

8. CAM kinase has two primary effects on AMPA receptors. One of these effects is to phosphorylate the receptors which ultimately has what effect on ion conductance?a. Increase conductance to sodium ionsb. Decrease conductance to sodium ionsc. Increase conductance to potassium ionsd. Decrease conductance to potassium ionse. Increase conductance to all ionsf. Decrease conductance to all ions

9. The sensory neuron that transmits neural signals from the aplysia siphon utilizes which of the following neurotransmitters?a. Dopamineb. Serotoninc. Acetylcholined. Glycine e. None of the above

10. Interneurons that modulate the sensitization response in the aplysia utilize which of the following neurotransmitters?a. Dopamineb. Serotoninc. Acetylcholined. Glycine e. None of the above


12. Generate a second, concise question from the cyber laboratory exercise for this Chapter.


Chapter 5: Rat Brain Anatomy – Modeling Parkinson Disease

Introduction: Animal models have long been a research staple for increasing understanding of brain damage and neurological dysfunction. Among the most prevalent and enduring models are rodents with induced damage to the nigrostriatal pathway, designed to mimic Parkinson disease (PD). Because PD has a relatively well defined region of degeneration (the substantia nigra), early researchers found that nonselective cell damage to this structure, typically produced by electrolytic lesions, resulted in motor deficits that could be roughly equated to PD. More recently, neurotoxic lesion, using intracranially injected 6-hydroxy dopamine (6-OHDA) or systemically injected 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), have been used to produce more selective damage, destroying dopamine producing cells in the substantia nigra while leaving other cells and fibers of passage relatively intact. For several decades now, these neurotoxins have been used for modeling PD and testing potentially therapeutic drugs. In the case of intracranial injections, researchers have also developed a method by which behavioral effects of neurotoxic damage can be readily quantified. To do this, animals are subjected to unilateral lesions, which produce asymmetrical posturing and turning behavior when the animal walks. Turning behavior is easily quantified as the number of rotations an animal exhibits in a given time period. This paradigm can then be expanded upon by challenging the animal with drugs or environmental influences (e.g. amphetamine or stress) that exacerbate rotating behavior. Likewise, therapeutic compounds can be tested for effectiveness in reducing rotations. The simplicity of this behavioral model, combined with its intuitively applied results, explain why unilateral nigrostriatal lesions have endured for so long in neuroscience research.


Background readings: Creese, Burt, & Snyder (1997). Dopamine receptor binding enhancement accompanies lesion-induced behavioral supersensitivity. Science, 197, 596-8.

Ungerstedt, & Arbuthnott (1970). Quantitative recording of rotational behavior in rats after 6-hydroxy-dopamine lesions of the nigrostriatal dopamine system. Brain Res, 24, 485-493.

ERIN resource:Authors: Rose Feor, Davidson College; Nicole Mah , Davidson College; Donna Molinek, Davidson College; Mur Muchane, Davidson College; Todor Penev, Davidson College; Julio J. Ramirez, Davidson CollegeTopic / Subtopic: C.Disorders of the Nervous System / C.03. Parkinson's DiseaseResource Type: Animation, Laboratory ExerciseEducational Level: 2 Intermediate undergraduate, 3 Advanced undergraduateListing Date: March 20, 2012

http://www.davidson.edu/neuroscience/neuronconnection/default.aspx

Lab exercise: You are going to perform a virtual experiment using the unilateral 6-OHDA model. Your rats will be given neurotoxic 6-OHDA injections to the substantia nigra of the right hemisphere. The toxin will selectively destroy dopaminergic neurons beginning within 24 hours, leading to Parkinson-like symptoms. You will select the amount of denervation you would like to induce, choose a drug with which to challenge your animals, and view graphs of the rats' rotational behavior both pre- and post- lesion.

Student quiz questions:1. Creese and colleagues (1977) pointed out that like rats with 6-OHDA lesions, rats subjected

to long-term treatment with neuroleptics seemed to develop apomorphine supersenstivitity. They noted that this rat model may be useful for studying which of the following?a. Hallucinations reported by patients receiving chronic treatment for Parkinsonismb. Hallucinations reported by patients receiving chronic treatment for schizophreniac. Tardive dyskinesia reported by patients receiving chronic treatment for Parkinsonismd. Tardive dyskinesia reported by patients receiving chronic treatment for schizophreniae. Tremor reported by patients receiving chronic treatment for Parkinsonismf. Tremor reported by patients receiving chronic treatment for schizophrenia

2. From the Cresse et al. (1977) reading, what did the authors identify as the major finding of their study?

“Haloperidol binding sites increase in rats with lesions of the nigrostriatal dopamine pathway”


http://www.davidson.edu/neuroscience/neuronconnection/default.aspx

3. What was the rate of delivery and concentration of 6-OHDA used by Ungerstedt and Arbuthnott (1970)?a. 4 microliters per minute @ 8 micrograms per microliter b. 16 microliters per minute @ 16 micrograms per microliterc. 8 microliters per minute @ 8 micrograms per microliterd. 4 microliters per minute @ 4 micrograms per microlitere. 1 microliter per minute @ 2 micrograms per microliter

4. It seems reasonable to assume that even without disruption of the nigrostriatal dopamine system, an animal stimulated by amphetamine might be inclined to exhibit rotational behavior if it is put into a spherical chamber. Was this concerned addressed by Ungerstedt and Arbuthnott (1970)?

Yes. They noted that though animals were highly active, there were very few rotation counts recorded in sham lesioned animals.

5. From the information provided in the cyber technology laboratory exercise, approximately what percentage of striatal dopamine depletion is required before behavioral symptoms become apparent?a. 20%b. 40%c. 60%d. 80%e. 100%

6. When viewing coronal sections of the rat brain, what is anterior/posterior reading that is given at the point where you are able to visualize the most anterior portion of the superior colliculus? At this same point, what structure can be visualized in an adjacent ventral/medial location relative to the superior colliculus?

-5.6 mm, PAG

7. From a horizontal perspective, what structure appears in a lateral location relative to you lesion at all levels from bregma?

Hippocampal region

8. In a group of 9 animals with 60% damage, what is the prominent direction of the rotation and the mean number of post surgery rotations per minute in that direction, after treatment with amphetamine?

Ipsilateral (mean number varies)


9. What is the percentage of damage that must be apparent in the substantia nigra for the simulation to show mean apomorphine induced rotations in excess of 70 per trial session?

90%

10. The authors of this laboratory exercise suggest that when testing rats for apomorphine and amphetamine induced rotational behavior, it may be desirable to randomly administer these drugs, so that rats do not rotate in the same direction on several consecutive trials. What is the reason for avoiding successive trials using the same drug to induce the same rotational direction?

It reduces the likelihood of “learned rotation” in rats being tested




Chapter 6: Cognitive Neuroscience

Introduction: Neuroscientists have long been interested in the processes of mental imagery and in particular how the brain functions when interpreting complex images. Research in this realm is filled with challenges. First, most questions on this topic are specific to humans, limiting the use of animal models. Related to the lack of animal models, is a significant restriction in the use of experiments that allow direct observation of neural functioning, such as intracranial recording. These limitations in research methods have required researchers to use techniques that rely largely on observation of human responding to perception of external stimuli. For example, if asked “How many windows are on your family home?”, most people report creating a mental image of their home, from which they then count windows using a systematic movement around that mental image. Related to this, if asked to compare two similar objects presented from different visual orientations, observers are believed to create mental images of the objects and to then systematically move, or rotate, those images into equivalent orientations as a strategy for making accurate comparison. The original theories suggesting humans use such manipulation of visual imagery have been researched extensively. Among the most convincing evidence supporting the “mental rotation” hypothesis has been the finding that the amount of time required to compare objects is highly correlated to the degree of difference in orientation between the images of objects being compared. For example, an object needing to be rotated 20o to be brought into congruence with a second object can be evaluated more quickly than an object needing to be rotated 40 o which can be evaluated more quickly than an object needing to be rotated 80 o and so on. Classic experiments used to evaluate mental rotation demonstrate the utility of basic procedures being used to provide evidence for highly complex questions.


Background readings: Shepard & Metzler (1971). Mental rotation of three-dimensional objects. Science, 171, 701-3.

ERIN resource:Authors: John H. Krantz, Hanover CollegeTopic / Subtopic: D. Sensory and Motor Systems / D.02. AuditoryResource Type: AnimationEducational Level: 1 Introductory undergraduateListing Date: March 8, 2012

http://psych.hanover.edu/Krantz/sen_tut.html

Lab exercise: From the Krantz home page, that is loaded with interesting exercises and labs, click on the “Cognition Laboratory Experiments” link. From the new page, select the “mental rotation” link listed under the “mental imagery” subtitle. Carefully review the instructions page for the mental rotation experiment. Once you are comfortable with your understanding of these instructions, click on the link to start the experiment (a little more than halfway down the page). Activate the Java plug in to begin the experiment. The Java page will show you the parameters for the study. You need change only the first setting (to original 3D) for the stimulus type, the rest of the default settings work well for this exercise. Your parameters page should be set up as follows:

Stimulus type: change to “original 3D”All other settings can be left at the following default settings:

Number of Rotations = 7Stimulus should “Rotate”

Stimulus size = 100Number of trials per level = 10

Show “No Lines”Distance from fixation = 0.150

Delay before stimulus = 1000 msecStimulus on Till Response = yes

Click on the “done” button at the bottom of the screen to begin the experiment. The first screen of the experiment requires a prompt (space bar) to initiate the sequence of stimuli used for this study.

Complete the experiment and record your own personal results. Then, if you are male, recruit a female to complete this experiment. If you are female, recruit a male to complete this experiment. Record their results. Student quiz questions:


http://psych.hanover.edu/Krantz/sen_tut.html

1. Metzler and Shepard (1971) concluded that when a mental image is rotated within the plane of a picture, _________ time is needed compared to when a mental image is rotated in depth.a. significantly moreb. significantly lessc. about the same amount of

2. What was the explanation given by Metzler and Shepard (1971) for using mirror images of objects for the “different” pair category, rather than just using objects that had random differences in shape?

This prevented participants from identifying distinctive features of to establish noncongruence without requiring them to carry out the process of mental rotation.

3. The results presented by Metzler and Shepard (1917) indicate that there was ___________ variability in reaction time scores for objects that required a greater degree of mental rotation when compared to those that required a lesser degree of mental rotation. a. lessb. greaterc. about the same amount of

4. How many scientific articles are referenced in the Metzler and Shepard (1971) paper?

None

5. According to the Metzler and Shepard (1971) article, approximately how long, on average, would it take a participant to mentally rotate an object 150o?a. 1.0 secondb. 1.5 secondsc. 2.0 secondsd. 2.5 secondse. 3.0 seconds

6. According to the Metzler and Shepard (1971) article, approximately how long does it take a participant, in their paradigm, to identify two objects as the same when no mental rotation is required?

1 second

7. From the mental rotation exercise, when viewing the results, what number would represent a “perfect score” for accuracy?a. 1.0


b. 10c. 100d. 1000e. A perfect score cannot be achieved on this test

8. From the mental rotation exercise, which of the following is not an option in the experimental set-up for a “stimulus type”?a. Original 2Db. Original 3Dc. Random 2Dd. Number e. Letter

9. From the mental rotation exercise, Did you find a sex related difference in either speed or accuracy of completing this task?

Yes No (circle one)

Which subject did you recruit Male Female (circle one)

When your score was compared to the participant you recruited, it was Faster Slower (circle one)

When your score was compared to the participant you recruited, it wasMore accurate Less accurate (circle one)

10. List an article reference (in APA format) that might be useful for evaluating the results you circled for responses to the previous inquiries (responses to #9).



Chapter 7: Transduction of Sensory Input - Auditory


Introduction: From the neuroscience perspective transduction is, in general, the conversion of environmental energy into neural energy. The processes of auditory transduction are particularly interesting for a number of reasons. Among these is the role of hair cells that act as mechanoreceptors within the inner ear. Mechanoreceptors are rapidly responding cells that require no chemical processes to initiate depolarization or hyperpolarization. In addition, basic hair cell functioning in the auditory system is nearly identical to that of the vestibular system. Thus, to understand transduction processes of the auditory system, is to likewise understand the basic processes used in transducing energy used for balance and kinesthetic function. Beyond basic transduction, the auditory system is designed with an intricate systematic arrangement of receptor cells within the inner ear. The tonotopic representation of cells within the cochlea is a unique example of how a basic process, such as transduction, can be integrated into a highly complex process, such as pitch perception. In this case, the complex process of pitch perception utilizes relative location and distribution of receptors, physical properties of the inner ear, and timing of neural signals. In general to understand the working of the auditory system is to understand a relatively simple yet elegant neural process that has evolved so that organisms may interact more readily with the environment around them.

Background reading: Hudspeth A.J (2005). How the ear's works work: mechanoelectrical transduction and amplification by hair cells. C R Biologies, 328,155-62.Background video viewing:http://www.youtube.com/watch?v=PeTriGTENoc


http://www.youtube.com/watch?v=PeTriGTENoc

ERIN resource: Authors: John H. Krantz, Hanover CollegeTopic / Subtopic: D. Sensory and Motor Systems / D.02. AuditoryResource Type: AnimationEducational Level: 1 Introductory undergraduatePublication Date: 0001Listing Date: March 8, 2012http://psych.hanover.edu/javaTest/NeuroAnim/audition.physiology.html

This website gives an overview of behavioral, anatomical, and computational aspects of working memory. A major feature is an interactive computer model. The example here is tuned persistent delay activity in the prefrontal cortex of non-human primates observed during oculomotor delayed-response tasks. Some parameters in the simulation of the network model can be changed interactively to study how the strength of inhibition and excitation in the microcircuit affect the capacity of the model to store information in working memory.

A series of Java applets that animate aspects of audition. Components include: 1) Illustration of the sound stimulus including the effects of intensity and frequency on perception of sound and various features of a sound wave, e.g, compression, rarefaction and phase. 2) Reception and transduction concepts illustrated for middle ear including the working of the ossicles and how their motions depend upon the frequency of sound. Cochlea illustrations show functions of the inner including how sound stimuli affect hair cells to cause transduction.

Lab exercise:From the Hanover College audition animation home page, visit each of the web links. The sound basics link allows you to visualize compressions and rarefactions of air molecules produced by sounds of different frequencies. Frequency of sound can be adjusted using the slide bar on the left of the screen. Sound corresponding to the different frequencies is also produced by this link. There are four links listed under reception and transduction, but if the link for “the ear” is opened, the other three links can be accessed from this one web page and movement from the entire ear to the individual components allows better visualization of the workings of the entire ear. From “the ear” page, sound frequency and amplitude can be adjusted to see how these changes affect reception and transduction processes within the ear. The youtube video gives a nice continuous animation of the processes shown on the Hanover College web page, as well as a nice descriptive explanation of these processes.


http://psych.hanover.edu/javaTest/NeuroAnim/audition.physiology.html

Student quiz questions:1. Hudspeth (2005) notes that the auditory system utilizes a general principle whereby objects

may be “tuned” to certain frequencies, causing them to vibrate when that frequency is emitted. What is the analogy he uses to describe this example?a. A person playing a pianob. A plane causing a window pane to rattlec. A dog whistle that is not perceived by humansd. A person playing a pipe organe. None of the above, Hudspeth describes this process as unique to the auditory system.

2. What are the two properties of the basilar membrane that contribute to its ability to vibrate sympathetically in specific regions to specific frequencies?a. Length and girthb. Mass and tensionc. Fluid content and fiber densityd. Ionic concentration and protein contente. Ionic concentration and lipid content

3. Which of the following sound detecting organs/structures is not found in humans?a. Lateral line organb. Cochleac. Vestibular labyrinthd. All of the above are found in humanse. None of the above are found in humans

4. When the hair bundle is moved toward the short edge (the shorter hair cells), the result is a ________________ in _______________ causing a net _______________________.a. Decrease, cation influx, depolarizationb. Increase, cation influx, depolarizationc. Decrease, anion influx, depolarizationd. Increase, anion influx, depolarizatione. Decrease, cation influx, hyperpolarizationf. Increase, cation influx, hyperpolarizationg. Decrease, anion influx, hyperpolarizationh. Increase, anion influx, hyperpolarization


5. The auditory system is extremely responsive, in terms of speed, when interpreting high frequency sounds. In brief terms, explain why hair cells can emit signals more rapidly than many other neural systems when responding to stimuli (sound).

Their response uses rapid mechanical mechanisms rather than slower chemical release.

6. Hair bundles do not simply respond to sound waves, but also amplify the signal emitted by wave energy. This amplification process utilizes a unique “twitch” process in which a physical link between ____________ of one hair cell and an adjacent hair cell mechanically force movement that increases ion flow and thus amplifies the neural signal.a. The membraneb. Sodium/potassium pumpsc. Calcium channelsd. Dendritese. The nucleus

7. In the Youtube video, movement of the three bones of the middle ear is described as pivoting on an axis that is due to ___________________.a. A viscous fluid that bathes these bonesb. A series of ligaments that hold the bones in placec. A series of muscles that respond to and amplify movement of the bonesd. Four small bones that encase the three bones of the inner eare. A single axis bone that passes through the malleolus

8. From the Hanover College resource page, viewing the “Sound Basics” link, set the frequency (left side of screen) to 400 Hz and note how many compressions occur on the screen (time duration 625.00). Change the frequency to 800 Hz and note how many compressions now occur over the same duration. Click on the “play” button to listen to the differences in tone for the two frequencies.

Number of compressions: 400 Hz = ______________ 800 Hz = ____________________

9. From the Hanover College resource page, view the “the ear” link. Start the sound and slowly move the frequency from low to high. Describe what happens to the hair cell containing membrane within the cochlea as sound waves change from low frequency to high frequency.a. Membrane movement shifts from apex only to base onlyb. Membrane movement shifts from base only to apex onlyc. Membrane movement shifts from entire length of membrane to base onlyd. Membrane movement shifts from entire length of membrane to apex only


e. Membrane movement is consistent regardless of the frequency change

10. From the Hanover College resource page, view the “the ear” link. Start the sound and slowly move the amplitude from high to low. Describe what happens to the hair cell containing membrane within the cochlea as sound waves change from high amplitude to low amplitude.f. Membrane movement shifts from apex only to base onlyg. Membrane movement shifts from base only to apex onlyh. Membrane movement shifts from entire length of membrane to base onlyi. Membrane movement shifts from entire length of membrane to apex onlyj. Membrane movement does not shift, but does change in size




Chapter 8 : Working Memory

Introduction:

Background reading:

Background video viewing:

ERIN resource: Authors: Albert Compte, IDIBAPS, Barcelona, SpainTopic / Subtopic: F. Cognition and Behavior / F.01. Learning and MemoryResource Type: SimulationEducational Level: 2 Intermediate undergraduatePublication Date: 2012Listing Date: February 22, 2013http://wm.crm.cat/en/

This website gives an overview of behavioral, anatomical, and computational aspects of working memory. A major feature is an interactive computer model. The example here is tuned persistent delay activity in the prefrontal cortex of non-human primates observed during oculomotor delayed-response tasks. Some parameters in the simulation of the network model can


http://wm.crm.cat/en/

be changed interactively to study how the strength of inhibition and excitation in the microcircuit affect the capacity of the model to store information in working memory.

Student quiz questions:1. According to the cyber lab exercise, the first researcher to study working memory was

William James which he named primary memory.

2. According to the cyber lab exercise, which of the following brain structures would not show increased oxygen consumption during the delay period of the delayed matching task?a. Dorsolateral prefrontal regionsb. Superior frontal gyrusc. Superior frontal junctiond. Visual cortexe. Parietal anterior cortex

3. From the cyberlab exercise titled, “network memory during working memory trial” set the excitatory and inhibitory stimuli at equal levels of very low intensity (far left side of the bar for both). Count and report below the number of action potentials that occur in the response period of the third (bottom) neuron recording when these inputs are both set at the lowest level.

Number of action potentials during response period ___4-5____

Now increase the inhibitory input to the highest possible level (far right of the bar) while leaving the excitatory input at the lowest possible level (far left of the bar). Count and report below the number of action potentials that occur in the response period of the third (bottom) neuron recording when excitatory input is at this lowest level and inhibitory input is at this highest level.

Number of action potentials during response period _zero__


Chapter 9 : Sleep

Introduction:

Background reading:

Background video viewing:

ERIN resource: Authors: D.T. Max, National GeographicTopic / Subtopic: E. Homeostatic and Neuroendocrine Systems / E.08. Biological Rhythms and SleepResource Type: Article - News, Image - Diagram, Image - PhotographEducational Level: 1 Introductory undergraduate, 2 Intermediate undergraduatePublication Date: 2010Listing Date: March 8, 2012http://ngm.nationalgeographic.com/2010/05/sleep/max-text

The Secrets of Sleep From birth, we spend a third of our lives asleep. After decades of research, we’re still not sure why. Includes a feature article, photographs, and interactive diagrams.


http://ngm.nationalgeographic.com/2010/05/sleep/max-text

Chapter: Transduction of Sensory Input – Visual


commons.suny.educommons.suny.edu/.../uploads/iitg/Cyber_Technology_La… · Web viewNeuroscience . Cyber Technology . Laboratory Manual. Terence Bazzett. SUNY Geneseo. 2013

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