FEB 25

Question 1

paper: “stress disrupts _______ ________ in ________ ______ to generalize threat memory in mice”

Answer 1

engram ensembles

lateral amygdala

Question 2

how do neurons become allocated to engrams?

Answer 2

neurons that are MORE ACTIVE at the TIME OF AN EVENT are more likely to be allocated to the engram representing that event

Question 3

what type of coding do engrams use?

Answer 3

sparse coding

^ this is critical for the SPECIFICITY of the memory (and efficiency - allows for more experiences to be stored in the brain because there are MORE UNIQUE COMBOS)

Question 4

competitive engram allocation

Answer 4

maintains sparse coding

Question 5

during fear engram encoding, what increases?

Answer 5

1. increase in activity in ENGRAM NEURONS
2. overall increase in INHIBITION

^ increased inhibition actively prevents less excitable cells from becoming part of the engram

Question 6

what happens to parvalbumin labeling around pyramidal neurons during fear conditioning?

Answer 6

there’s a TRANSIENT INCREASE

Question 7

parvalbumin interneurons

Answer 7

GABAergic interneurons (inhibitory)

powerfully control the activity of pyramidal neurons

they’re the specific cell type that’s important in MAINTAINING SPARSE CODING

Question 8

stress leads to overgeneralization of what?

Answer 8

of AVERSIVE MEMORIES

this is a hallmark of many psychiatric disorders

Question 9

paper hypothesis

Answer 9

stress mediates threat memory generalization by INCREASING THE SIZE OF AN ENGRAM ensemble

via an unknown mechanism

Question 10

mice experiment - threat discrimination task

Answer 10

exposed all mice to a tone paired with a shock (CS+) and a tone paired with nothing (CS-)

conditioned these on separate days

then one group of mice experience restraint stress IMMEDIATELY BEFORE CS+

Question 11

what does acute restraint stress increase?

Answer 11

the stress hormone corticosterone (CORT)

CORt increases during restraint stress, and foot shock etc

Question 12

what does blocking CORT do?

Answer 12

it prevents stress-induced increases in threat memory generalization

Question 13

CORT has been shown to increase what?

Answer 13

threat memory generalization

(administering CORT results in similar increases in fear generalization to those seen in restraint stress manipulations)

Question 14

systemic CORT injection increases the overall size of what?

Answer 14

the LA engram ensemble

this supports a threat memory

Question 15

tags: TRAP2-TdT mouse injected with AAV-RAM-GFP virus in the LA

Answer 15

they label Fos expressing neurons during THREE DIFF EVENTS

TAG 1: TdT - labels neurons expressing Fos during training

TAG 2: GFP - labels cells active during CS+ recall test

TAG 3: endogenous Fos - labels cells during the CS- recall test

Question 16

TdT

Answer 16

the first tag

labels cells expressing Fos during fear/reinforcement training

Question 17

GFP

Answer 17

the second tag

labels cells active during CS+ recall test

Question 18

endogenous Fos

Answer 18

labels cells active during CS- recall test

Question 19

in both CORT and vehicle mice, what was found pertaining to reactivated cells during CS+ recall?

Answer 19

similar numbers of cells were reactivated

so CORT isn’t necessarily changing how much the threat memory is reactivated

what it’s changing is how much the threat memory is reactivated when the stimuli that’s being tested ISN’T THE OG THREAT (but is instead the CS-)

Question 20

are more cells reactived in the CORT mice during CS+ recall?

Answer 20

no

similar numbers are reactivated

because CORT isn’t changing how much the threat memory is reactivated in response to the original stimulus

Question 21

In CORT mice, more cells are active compared to vehicle mice when?

Answer 21

during the CS- test

more of the cells that were active during the CS+ TRAINING are REACTIVATED during the CS- test

more of the cells that were active during the CS- TEST

Question 22

in CORT mice, more of which cells are active during the CS- test?

Answer 22

more cells generally are more active during the CS- test

specifically:

1. cells that were ACTIVE DURING CS+ TRAINING
2. cells that were ACTIVE DURING CS+ TEST

(points to generalization of the fear memory)

Question 23

how can we force cells into an engram?

Answer 23

by using HSV-NpACY (a viral tool allowing for BIDIRECTIONAL CONTROL of neural activity)

it allows us to activate subsets of cells and force them into engrams

Question 24

HSV-NpACY

Answer 24

viral tool that allows for BIDIRECTIONAL CONTROL of neural activity

it’s a virus that’s injected and sparsely infects neurons - and allows for subsequent CONTROL of a SUBSET of neurons

(activating these cells will FORCE THEM INTO ENGRAMS)

Question 25

they find that RED LIGHT does what?

Answer 25

it suppresses freezing

Question 26

the effect of the red light is stronger in which mice?

Answer 26

the vehicle mice (weaker effect of red light in the CORT mice) (and the red light suppresses freezing)

Question 27

why is the effect of the red light weaker in CORT mice?

Answer 27

in the CORT mice, additional neurons that were active were RECRUITED TO THE ENGRAM (beyond the NpACY-expressing neurons) so more fear cells are being reactivated and therefore freezing is being suppressed less

Question 28

does CORT modulate the activity of PV neurons?

Answer 28

recall: PV neurons inhibit activity and therefore maintain sparse coding they test this with FIBRE PHOTOMETRY in PV-Cre mice use GCaMP

Question 29

GCaMP

Answer 29

a LIGHT-SENSITIVE calcium indicator can be TARGETED to PV neurons to get a SPECIFIC READOUT OF ACTIVITY in these cells

Question 30

what does fibre photometry do?

Answer 30

it's an IMAGING TECHNIQUE - it's just observational provides a readout of neural activity (in this study, use GCaMP to get a specific readout of activity in PV neurons)

Question 31

GCaMP and fibre photometry

Answer 31

allow us to get a readout of neural activity in the PV interneurons to see how CORT affects them

Question 32

what does CORT do to activity of LA PV+ inhibitory neurons during threat conditioning?

Answer 32

CORT DECREASES the activity of these cells

Question 33

footshock does what to PV neurons?

Answer 33

increases their activity but this effect is BLUNTED in CORT treated mice

Question 34

separate experiments show that footshock increase activity in pyramidal neurons and that CORT...

Answer 34

CORT further increases this activity

Question 35

CORT didn't change what things?

Answer 35

1. baseline activity without shock 2. activity in other interneurons (so CORT is specifically blunting the PV increase after the footshock - specific neurons, specific timing)

Question 36

CORT did change what things?

Answer 36

1. CORT specifically decreases activity of LA PV neurons 2. and then increases LA pyramidal activity to produce a less sparse engram

Question 36

how does CORT produce a less sparse engram?

Answer 36

1. decreasing inhibitory LA PV+ neuron activity 2. increasing excitatory pyramidal neuron activity

Question 36

if CORT decreases PV neuron activity, does manipulating PV neuron activity interact with the effects of CORT?

Answer 36

use CHEMOGENETIC TOOLS to increase or decrease activity of PV interneurons 1. increasing PV neuron activity DECREASES GENERALIZATION in CORT mice 2. decreasing PV neuron activity INCREASES GENERALIZATION in vehicle mice (mimics CORT's activity)

Question 37

what does increasing PV neuron activity do in CORT mice?

Answer 37

it decreases generalization

Question 38

what does decreasing PV neuron activity do in vehicle mice?

Answer 38

it increases generalization

Question 39

how does stress suppress PV activity?

Answer 39

we know that CORT suppresses PV neuron activity - but CORT is just a proxy for stress that was directly manipulated in the experiment how does this work in real life? ENDOCANNABINOIDS

Question 40

endocannabinoids (eCBs) are implicated in...

Answer 40

eCBs are implicated in effects of STRESS on BEHAVIOUR

Question 41

CB1 receptors

Answer 41

located on inhibitory neurons eCB signalling disrupts GABA (inhibitory) release from PV neurons in other brain regions

Question 42

what effect does eCB signalling have on PV neurons in other brain areas?

Answer 42

eCB signalling DISRUPTS GABA RELEASE from PV neurons in other brain regions so it disrupts inhibitory signalling (just like CORT does)

Question 43

what ligands bind CB1 receptors?

Answer 43

2 endogenous ligands 1. ANANDAMIDE (AEA) 2. 2AG retrograde signalling to pre-synaptic CB1 receptors mediate changes in NT release CB1 receptors = located on pre-synaptic neurons so, 2AG and AEA bind to CB1 and affect NT release

Question 44

what happens to 2AG and AEA after fear conditioning under CORT?

Answer 44

they're both INCREASED by CORT

Question 45

what does blocking 2AG synthesis do?

Answer 45

it DISRUPTS FREEZING to both CS+ and CS- in vehicle and CORT groups (so 2AG does something, but it's not specific to threat memory generalization)

Question 46

does 2AG do something specific to threat memory generalization?

Answer 46

no, because blocking 2AG disrupts freezing TO BOTH CS+ and CS- in BOTH VEHICLE and CORT groups

Question 47

what does inhibiting AEA in CORT treated mice do?

Answer 47

it REDUCES GENERALIZATION inhibiting AEA restores memory specificity!

Question 48

inhibiting AEA restores what?

Answer 48

memory specificity

Question 49

AEA inhibition doesn't alter anxiety-like behaviour in other tests - what does this suggest?

Answer 49

suggests that AEA is SPECIFIC to THREAT MEMORY GENERALIZATION

Question 50

does increasing AEA reproduce the effect of CORT on threat memory generalization?

Answer 50

yes simply increasing AEA mimics the effects of CORT (increases fear generalization)

Question 51

CORT-induced memory generalization is mediated by...

Answer 51

increased AEA levels in the LA (can mimic effect of CORT by increasing AEA, and can block effect of CORT by reducing AEA)

Question 52

t/f: effects of stress on threat memory generalization are mediated by eCBs in the LA

Answer 52

true! simply increasing AEA (an endogenous CB1 receptor ligand) mimics the effects of CORT (leads to threat generalization)

Question 53

use optogenetic stimulation to elicit responses from PV neurons and record the response in excitatory neurons - they do this to do what? and what do they find?

Answer 53

to test the role of CB1 receptors in controlling GABA release from PV neurons find that activating CB1 receptors disrupts GABA release

Question 54

CORT's effect on pyramidal activity and eCB signalling

Answer 54

CORT increases pyramidal activity CORT increases eCB signalling

Question 55

which occurs first: CORT's effect on eCB signaling or CORT's effect on pyramidal signalling?

Answer 55

change in eCB signalling occurs first suggests that the eCB signalling is CRITICAL to the effect of CORT ^ change in endogenous signalling precedes changes inn pyramidal activity - suggests that endo changes in CORT lead to changes in neural activity

Question 56

what does knocking down CB1 in LA PV+ neurons do?

Answer 56

1. sufficient to RESTORE MEMORY SPECIFICITY 1 a) knocking down CB1 receptors in PV prevents CORT-induced generalization in freezing 1 b) also restores the sparsity of the threat engram 2 c) also prevents stress induced generalization in freezing

Question 57

find that knocking down CB1 in LA restores memory specificity - could this be clinically relevant?

Answer 57

(we don't do targeted manipulations in humans) 1. a GLUCOCORTICOID RECEPTOR ANTAGONIST administered systemically prevents stress-induced generalization 2. an AEA synthesis inhibitor administered systemically also prevents stress induced generalization so yes, it can be clinically relevant

Question 58

summary of the effects

Answer 58

1. stress > 2. increased LA AEA signalling > 3. decreased GABA release (decreased PV neuron activity) > 4. increased excitatory activity > 5. increased engram size > 6. threat generalization