Read through the Engisch paper on homeostatic synaptic plasticity at the NMJ.

1. Describe the 3 aspects of activity sensed to trigger 3 distinct mechanisms that contribute to homeostatic plasticity.

2. Look at the article on homeostatic synaptic plasticity at the NMJ. Were any of your
questions answered by experiments/studies described in this paper? If so, what did
those studies find? If not, why do you think that it hasn’t been done yet?

Read the article Activity–Dependent Presynaptic Regulation of Quantal Size at the
Mammalian Neuromuscular Junction in Vivo. And answer the following questions.

1. Look through the figures and the main findings, create a list of bulleted points of your overall interpretations and conclusions for the experiments. These should be points that you think would be worth including in a Discussion section of the article if you were the author.

2. Read the Discussion section of the article. After reading it, make a list of the author’s interpretations of their findings and their conclusions. Compare the list that you created with the list that you created based on the author’s interpretations. How are they similar? Where do they differ? [Hint: You should have a minimum of 5–6 points for each list.]

3. Describe the evidence that blocking presynaptic action potentials can trigger a change in quantal size.

4. Are there any questions that still remain unanswered? Identify at least TWO question that the authors still would like to answer. Then identify at least FOUR questions that YOU still would like to have answered that weren’t answered in this article. Why do you think that these questions are important to answer?

5. In the article, the authors try to separate the contributions of blocking the action
potentials in the presynaptic cell while leaving activity in the muscle intact. How did they accomplish this experimentally? What tool did they use? How did this allow them to block presynaptic action potentials without blocking postsynaptic activity? What did they find when presynaptic activity was blocked but muscle activity was not? Why was this finding important?

6. Define homeostatic synaptic plasticity. What happens when a network of neurons has been treated with TTX for several days then the TTX is removed? What do the
electrophysiological (current clamp) recordings from those cells look like? How do they compare to current clamp recordings made prior to the activity block?

7. What homeostatic mechanisms likely occur to help a neuron regain target signaling
ranges after activity is blocked for a few days?

8. What homeostatic mechanisms likely occur to help a neuron regain target signaling
ranges after activity is increased for a few days?
9. If a person takes an SSRI (a drug that blocks the reuptake of 5–HT) for a prolonged
period of time, homeostatic mechanisms may occur to try to return the synaptic
transmission at that site to the neuron’s target zone. Sometimes these changes cause
the SSRI to become less effective and dosages must be adjusted. What types of
homeostatic changes do you hypothesize may occur at the 5–HT synapse to
compensate for the chronic blockade of SERT?

Read textbook chapter 14 – 17
molecular and cellular synaptic plasticity
1. Experiments using Actinomycin D (DNA transcription/mRNA synthesis inhibitor) which
showed that activity–dependent, de novo RNA transcription is involved in LTP. The data
shown below is related to these experiments.
Note: The Y–axis measure (fEPSP slope as a % of baseline) was not discussed in detail.
For our purposes, the greater the fEPSP slope relative to baseline, the stronger the
synapse/LTP. (100% means strength is equal to what it was at the start; 200% means
strength is twice as strong as it was at the start).
a. Figure A shows the neuronal response to a
single tetanic stimulation (black arrow). This
was done with no other manipulation (black trace),
with Actinomycin D (blue trace; the DNA
transcription inhibitor [preventing mRNA
synthesis]), or with Anisomycin (red trace; an
mRNA translation inhibitor [preventing protein
synthesis]). The black bar at the bottom shows
when the inhibitors were present (before, during,
and shortly after stimulation). The initial peak is a
direct response to the depolarizing stimulus and should not be interpreted as a
change in synaptic strength.
i. Following the stimulation, is there an enhancement in synaptic strength?
1. If so, what group(s) display this enhancement?
2. If so, for how long does it persist?
ii. Based on this data, is transcription required for induction of LTP?
1. Why or why not?
iii. Based on this data, is translation required for induction of LTP?
1. Why or why not?
iv. Based on your knowledge of the mechanisms underlying early LTP, what
is the mechanism of a drug/toxin that you could add to block it? (do not
need a specific drug, just how it would act to block early LTP)

b. Figure B shows the neuronal response to a
series of tetanic stimulations (black arrows). As
with A, this was done alone (black trace), with
Actinomycin D (blue trace), or Anisomycin (red
trace) present before, during, and shortly after the
stimulation (black bar). The initial 4 peaks are
direct responses to the depolarizing stimulus and
should not be interpreted as a change in synaptic
strength.
i. Following the stimulation, is there an enhancement in synaptic strength?
1. If so, what group(s) display this enhancement? Does it last?
2. Why does the fEPSP slope of the control not decay as in Fig. A?
ii. Based on this data, is transcription required for maintenance of LTP?
Why or why not?
iii. Based on this data, is translation required for maintenance of LTP? Why
or why not?
iv. Why is the time course of decay different for Anisomycin versus
Actinomycin D?
Hint: We think of translation (mRNA → protein) as occurring after
transcription (DNA → mRNA), which is true, but spines have a unique
feature allowing more rapid changes in protein synthesis during LTP.
c. Figure C shows the neuronal response to BDNF
application (green bar). This was done alone
(black trace) or with Anisomycin (red trace)
present before, during, and shortly after the BDNF
application (black bar).
i. Following BDNF application, is there an
enhancement in synaptic strength? If so,
what group(s) display this enhancement?
ii. Based on this data, is translation required
for BDNF–induced LTP? Why or why not?
iii. Based on your knowledge of BDNF’s mechanism of action, would you
expect Actinomycin D (transcription inhibitor) to have similar or different
effects than Anisomycin? Why?
2. Compare and contrast ASSOCIATIVE LTP & LTD, including the concepts highlighted
below:
a. Describe models used to study associative LTP/LTD. Make sure you understand
the following terms: Unconditioned Stimulus (US), Unconditioned Response
(UR), Neutral Stimulus (NS), Conditioned Stimulus (CS), Conditioned Response
(CR) and their relationships to each other.
b. Strength of input signal from the US relative to NS

c. Timing of the US and NS/CS relative to each other during conditioning
d. Relative Ca2+ levels (high/low) in the postsynaptic neuron (neuron driving the
response) during conditioning
e. Activity of CaMKII and PP2B during conditioning
f. Magnitude of response to the CS after conditioning (relative to before)
g. AMPAR phosphorylation (higher/lower) after conditioning (relative to before)
h. AMPARs in membrane (higher/lower) after conditioning (relative to before)
i. Relative time scale of how long the changes in synaptic strength will persist
without additional training (if initial training occurred in a few sessions across a
single day): <20 minutes, <12 hours, ~1–7 days, >1 month
j. Changes in structural plasticity
3. We set up an experiment to examine short–term changes in synaptic plasticity at the
neuromuscular junction (NMJ), where we have the ability to stimulate the presynaptic
motor axon and record from the end plate.
a. We begin with low Ca2+ levels in the bath and deliver a
brief train of 4, equal strength, depolarizing stimuli to the
presynaptic terminal and record this response from the end
plate.
i. What is this phenomenon called?
ii. What molecular changes are occurring to cause this
(be specific)?
b. Next, we add curare and a large amount of Ca2+,
repeating the same stimulations and recordings as
described previously. We record the following.
i. What is this phenomenon called?
ii. What molecular changes are occurring to cause
this (be specific)?
c. In the experiments above, we isolated one process from
the other. Under conditions with normal Ca2+, we
record the tracing shown here.
i. Why is there an increase in response to the
second stimulus relative to the first?
ii. Why do the third and fourth stimuli produce
stronger responses than the first stimulus, but
not significantly different from each other?
iii. Why is there a decrease in response to the final
stimulus relative to the first?
4. Know the pathways by which LTP & LTD are induced and maintained, including the
concepts highlighted below:
a. Glutamate transmission: AMPAR & NMDAR opening; coincidence detection
b. Synaptic versus extrasynaptic NMDAR signaling pathway activation
c. Calmodulin, CaMKII (including discussed CaMKII targets), PP2B

d. Mechanisms of Ca2+ entry/release, (including localization and functional impact):
NMDARs, Ryanodine receptors, IP3 receptors, L–type Ca2+ channels
e. AMPAR insertion, AMPA:NMDA ratio and functional implications
f. Activation of silent synapses
g. Rapid, local protein translation
h. ERK/Jacob signaling, Activity–dependent transcription (CREB, immediate–early
genes and their general functions)
i. mRNA transport and late local protein translation
j. Maintenance of long–term changes (new transcriptional state/Neuroepigenetics
and the 3 mechanisms), transgenerational inheritance of neuroepigenetic traits
5. Synapses are constantly undergoing changes in synaptic strength, some last for very
short periods of time while others last for many years. The transition from short–term to
long–term plasticity requires molecular changes in proteins. List the three (3) processes
that underlie the transition to and maintenance of long–term plasticity in the order that
they occur. Be able to briefly describe a change in target/effector/outcome for each of
the three processes listed above.
6. In the illustration of the dendritic spine below, match each of the components (A–G) to
the descriptions listed in the table. Some letters may be used multiple times