neuroscience - physiology assignment

Student Name_____________________________9

MODULE 2 EXAMINATION.

May 12, 2017

you should be able to complete the examination in ~3-4 hours (although you can take as much time as you need until the deadline 17TH May). 

Instructions

Deadline is 17h May

If you want to use hand drawn pictures that is fine, but then please scan them into the computer text and images must be your own work, i.e. no cutting and pasting of text or images is allowed.

You MUST work independently, so while you can consult your lecture notes, books, and the research literature, you cannot consult with other students, post-docs, faculty, spouses, or anyone.
It is also very important that you turn your exam in on time 17TH May.

1. Lecture S1, Dr. Lechletier, 20 points total

After a stroke, potassium ion homeostasis is disrupted due to energy depletion.

A. (10 points) Describe how this disruption leads to glutamate excitotoxicity and cytotoxic edema?

B. (10 points) Describe two mechanisms by which astrocytes regulate potassium homeostasis after a stroke.

2. Lecture S2,    Dr. Clarke, 30 points total

A. (10 points) G protein coupled receptors (GPCR), and probably all receptors, can adopt at least one active conformation capable of regulating signaling pathway activity in the absence of a ligand to activate the receptor (an agonist). What experimental approaches might you use to demonstrate that a GPCR (any receptor) is constitutively active? You can use any system (in vivo, in vitro, etc.) or combination of systems. Discuss how the outcome of the experiment(s) you design will allow you to conclude that the receptor is constitutively active.

B. (5 points) What cellular factors dictate the magnitude of constitutive activity of a GPCR in a particular cell?

C. (5 points) Evidence has been accumulating to suggest that 7 transmembrane-spanning receptors can form homodimers and heterodimers. What aspects of receptor function would be different when comparing heteromeric receptors (two different 7-TMS receptors bound together) versus monomeric receptors (a single 7-TMS receptor)?

D. (10 points) In class, we discussed a paper that contained evidence that the alpha and beta/gamma subunits of a Gi protein do not dissociate from one another as previously thought, but instead undergo a receptor-activation mediated conformational rearrangement of the subunits.

1) In reviewing the data, what is your opinion about this conclusion?

2) Are you convinced that the authors are correct?

3) If so, what is the most compelling evidence that influenced your opinion? If not, what additional evidence would be required to convince you?

4) If you were one of the investigators on this project, what would you do next?

3. Lecture S3,    Dr. Kim, 20 points total

A. (10 points) In the following AP train (50 Hz train of APs) at the presynaptic terminal, (1) describe the change in AP wave-forms between 1st AP (red) and the last AP (blue), (2) explain the underlying mechanisms of the change in AP wave-forms between 1st AP and the last AP. (10pt)

B. (10 points) Describe the relationship between presynaptic AP shape and Na+ channel location (or Na+ current density) at either unmyelianted presynaptic boutons or myelinated axon terminals (5 pt), and how does the amplitude of presynaptic AP affect Ca2+ influx to presynaptic terminals (5 pt)?

* No more than one page for two answers above (1 page limit).

4. Lecture S4, Dr. Pugh, 20 points total

While studying mouse behavior you find that prolonged exposure to images of Martha Stewart induces increased grooming behavior. Intrigued, you decided to investigate the mechanism of this effect. You begin by recording excitatory synaptic currents from acute brain slices of the striatum, a brain region involved in grooming behavior.

In control mice you observe very little short-term plasticity during a pair of synaptic stimuli. However, in the Martha-exposed mice you observe significant paired-pulse depression.

Control Martha

A. (6 points) Describe one presynaptic and one postsynaptic mechanism that could produce the paired-pulse depression observed in the Martha mice.

B. (3 points) You then apply cyclothiazide (which prevents AMPA receptor desensitization) to the brain slices and observe the following synaptic responses in the Martha mice.

What does this tell you about the mechanism of depression in (A)?

C. (5 points) A lab mate has been working on a novel drug which specifically blocks synaptotagmin7, so you decided to put some on your slice. How would you expect this drug to change the synaptic responses in (B)? Why? (Feel free to draw the responses if you prefer)

D. (6 points) Finally, if you wash a high calcium solution (4 mM) onto the brain slices, how would this change the release probability of the synapses in (B)? And what change would you expect in the paired-pulse ratio?

5. Lecture S5,    Dr. Beckstead, 20 points

A. (10 points) Explain what is meant by the term “silent synapse,” and mechanistically how silent synapses can be functionally altered by pairing pre- and postsynaptic activity.

B. (10 points) In the Valtcheva & Venance manuscript the authors described a bidirectional spike timing-dependent plasticity (STDP) with discrete timing windows. They go on to describe how the timing and mechanisms are disrupted by either acutely inhibiting or chronically overexpressing EAAT2. Please explain in detail what these results tell us about the role of EAAT2 on STDP under basal conditions.

6. Lecture S6    Dr. Nicholson, 20 points

A. (10 points) It has become increasingly recognized that gap junctions do not only serve as electrical synapses to rapidly conduct impulses in circuits that mediate escape responses, but that they also can modulate the behavior of neural networks and circuits. Choose TWO examples of such modulatory actions of gap junctions and briefly explain how they affect the circuit/network and the underlying mechanism, if known.

B. (10 points) The variable C-terminal domain of connexins has been shown to play many roles in the function and modulation of their functions. In which of the following functions is the C-terminus directly implicated (choose all that are correct)?

(a) Regulation of hemichannel opening by Ca++

(b) Gating of gap junctions in response to mitogenic signals (e.g. EGF/MAPK, src, etc.)

(c) Regulation of connexin oligomerization into heteromeric or homomeric assemblies.

(d) Regulation of neuronal or astrocyte motility

(e) Modulation of electrical synapse strength in response to repetitive firing of neurons (e.g. LTP).

For ONE of the above functions, explain how the C-terminal domain is involved in the mechanism

7. Lecture S7/S8/S9     Dr. Eaton, 60 points

A. (20 points) Dr. Eaton has obtained adult cerebellar cortex from an interesting mutant mouse harboring a targeted gene mutation in a gene of unknown function that has been implicated in cerebellar function in humans. Analysis of the gene sequence predicts that the gene encodes a trans-membrane protein, and sequencing of the mutant mouse line confirms that the mutation is a null allele. He prepares the tissue for standard immunocytochemistry (ICC) and stains wild type and mutant cerebellar cortices with an antibody against CaMKIV. Microscopic inspection of the stained tissue reveals that there are significantly more CaMKIV positive cells in the external granule layer (EGL) of the mutant compared to the wild type cerebellums. Wild type cerebellums have a higher proportion of CaMKIV granules cells in the internal granule layer (IGL) compared the mutant which have few granule cells in the IGL. Provide a model for the function of this trans-membrane protein during brain development. (400 word or less)

B. (20 points) Retinal starburst amacrine cells (SACs) form large dendritic fields that suggest self-avoidance. SACs also make inhibitory synaptic contacts with their neighbors, which improve the function of the retinae in determining directional movement. The inhibitory synaptic connections between two neighboring SACs can be directly measured using dual patch-clamp electrophysiology. Under normal conditions, the inhibitory contacts between neighboring SACs are present and can be measured. Autaptic (self) synapses are rarely observed. Recently, Kostadinov and Sanes manipulated the protocadherin gene cluster (Pcdhg) in the retinae using a SAC specific CRE mouse line. They found that SACs lacking any Pcdhg gene products (Pcdgh0) formed increased connections with neighbors and had many autaptic synapses. The authors present evidence that Pcdhg function is required for synaptic elimination during development explaining the increase in connectivity between neighbors. This group then attempted to rescue these defects by putting a singular Pcdgh isoform in all SACs. What would you predict for the connectivity between neighboring SACs and the number of autaptic synapses under these conditions? What do you base your prediction on? (400 words or less)

C. (20 points) Retinal ganglion cells (RGCs) send long axonal projections from the retinae of the eye to the visual cortex within the brain. It has been observed that there is a small group of cells that express netrin-1 near where the optic nerve exits the eye.

i. What role do you think netrin-1 could be playing there? (200 words or less)

ii. RGCs from the same eye project both to the ipsi-lateral LGN and the contra-lateral LGN. All retinal axons project to the optic chiasm where some cross the midline and project contra-laterally and others do not cross the midline but rather project ipsi-laterally. It has been observed that there exists high expression of Slit1/2 at the chiasm and mutations in Slit1/2 result in a severe reduction in ipsi-lateral projections to the LGN. Provide a molecular mechanism for the control of RGC axon crossing at the chiasm by Slit. (200 words or less)

TEXTBOOKS 

Recommended: The major textbook for the course is Squire et al. 4th edition (i). Readings in Byrne & Roberts 2nd edition (ii) and Byrne et al. 3rd edition (iii) are required for selected lectures. We suggest that you purchase Squire (available to buy or rent from amazon), which will also be used in Fundamentals of Neuroscience II, although copies are on reserve in Briscoe library.

1. Fundamental Neuroscience; 4th edition, Eds. Squire et al.; Academic Press, 2012, http://www.amazon.com/Fundamental-Neuroscience-Fourth-Edition-Squire/dp/0123858704 

1. From Molecules to Networks: An Introduction to Cellular and Molecular Neuroscience; 2nd edition, Eds. Byrne J & Roberts J; Academic Press, 2009. http://www.amazon.com/Molecules-Networks-Second-Introduction-Neuroscience/dp/0123741327/ref=sr_1_10?s=books&ie=UTF8&qid=1359428204&sr=1-10&keywords=roberts+neuroscience

1. From Molecules to Networks: An Introduction to Cellular and Molecular Neuroscience; 3rd edition, Eds. Byrne et al.; Academic Press, 2014. http://www.amazon.com/From-Molecules-Networks-Third-Edition/dp/0123971799/ref=pd_cp_b_0