Ch 5 Flashcards
1
Q
Why might the post-mortem assessment affect the analysis of GENUS?
A
As we only have a single read-out post-mortem. As plaque pathology is different across mice, we would need to see a pre- and post-measure to illustrate an actual change in pathology.
2
Q
Why would the singular approach be different?
A
The original papers that complete the multisensory stimulation show that this modulated neuronal activity across wider brain regions with pronounced effects. Therefore, it may be more effective at reducing plaque pathology.
3
Q
What is the hypothesis and aims?
A
Hypothesis: novel approach allows monitoring of plaques caused by GENUS
Aim: to expose AD models to GENUS and monitor changes in AD pathology
4
Q
Why were the same animals used as before?
A
for 3Rs. They were used adter PK to ensure they are being used maximally.
5
Q
Why were the electrophysiological recordings not completed on the FP mice?
A
There has been papers, including my group, that have done dual photometry and ephys experiments by mounting an electrode along the fibre shank.
However, currently, TFs are just too fragile for this approach for this to be feasible. However, there are now being TFs made with electrode channels along this.
Another consideration is the 40-Hz noise that would occur due to the LEDs, which required an extra internal faraday cage which would not be feasible for long-term recordings as it is not freely behaving.
6
Q
Why were walls lined with black tape?
A
So that it limits reflection off the walls and ensures the light being delivered to the mouse is the 40hz light.
This is the same as the landmark study.
7
Q
Why white LED light?
A
Broad Spectrum: White LEDs emit light across a broad spectrum, covering a wide range of wavelengths. This broad spectrum ensures that the flicker stimulation affects a wide range of photoreceptors in the eye, including both rods and cones. This comprehensive stimulation can be beneficial for certain research or applications where a broad response is desired.
8
Q
Why did you choose 20-cm and 70-dB for the speaker?
A
20-cm was the same as the original study
70-dB is high enough for the c57 background to hear according to shuzo previous study that monitors the ABR in c57 mice.
9
Q
Why did you use a 50% duty cycle?
A
So, during a 40 Hz stimulation with a 50% duty cycle, the LED is on for 0.0125 seconds (or 12.5 milliseconds) during each cycle.
This is like previous papers.
10
Q
What does 100kHz mean for stimulation?
A
Full range of mouse hearing so we are activating the whole tonotopic auditory pathway
11
Q
Why can the NIDAQ not provide the sufficient current?
A
Because the NIDAQ has current safety limits which are below what is required for dlivery of the 5V LED with ~3A current flow.
12
Q
What is a
- MOSFET
- gate resister
- pull down resister?
A
Mosfet is an electrical component that is used as a switch to control the current flow through a circuit. This was used to control the current flow to the LEDS. It has a gate, source and drain. The date is the gate resistor, the source is the DAQ connection and the drain is the pull down resistor.
Gate resistor is used to control the overcurrent of the system. So the impedence is chosen so that it limits the current flow to protect tge system from damage.
Pull down resistor works as the drain of the mosfet as its connected to the pin it will have some voltage so this works to a low voltage value.
13
Q
What is the function of the gate resister? Why 1kOhm?
A
prevents overcurrent from teh NIDAW as this can damage the components
This minimises the current range to only a few milliAmps.
14
Q
What is the function of the pull-down resistor? Why 10kOm?
A
switch to the MOSFET, where if the DAQ is on, it will allow the current ro flow
as its higher than the gate resistor, allowing a good amount of voltage at the gate - it needs to be high enough to allow sufficient voltage flow.
15
Q
Can you explain how this electrical breadboard works?
A
Each pin on the board allows a connection to be formed.
The two outer channels represent power rails which are positive and negative to allow power flow and this power will flow horizontly.
Then to transfer the power to the terminals, a connection must be made and this allows power accross horizontal channels.
Then, the DAQ connection will contact the pulldown resister. THE DAQ ground on MOSFET and the LED on the MOSFET.
This voltage flow will then traveel along the row to turn on LED drip.
16
Q
What are broadband noise and puretones?
A
Broadband noise contains all frequecies of sound.
Pure tones is a sound that has just one frequency.
17
Q
Why does the sound need to be amplified before reaching the speaker?
A
It is required to amplify the analog signals into sound.
18
Q
Why does the microphone require an amplifier?
A
The microphone picks up small vibrations that are converted to a voltage signal that is amplified and recorded.
19
Q
Why do you need to use the root mean squared voltage to play a broadband noise?
A
When playing a broadband noise from a speaker, you need to control the voltage that drives the speaker to ensure that the sound produced has the desired characteristics, such as amplitude and intensity.
Audio signals are dynamic and can vary in amplitude over time. The RMS voltage takes into account these variations by averaging the square of the instantaneous voltages over time and then taking the square root. This provides a measure of the effective voltage level that drives the speaker, which is more representative of the overall power delivered than simply using the peak voltage. Therefore, it represents the average voltage signal.
20
Q
What is a bipolar electrode and why did you use it?
A
This is two conductive electrodes that are mounted together. They have a small offset and allow recording of electrical signals at the same time.
We did this as it provides a hippocamal measurement at two regions, allowing improvment of targeting success.
21
Q
Why did you do EEG over the PFC?
A
PFC provides a hub of brain functionality and activity.
Also, previous papers discuss the spread of gamma oscillations to the PFC.
22
Q
Why did you have EMG?
A
These mice were used for other recordings for a seperate project.
This was a sleep project where they required the measurement of muscle tone.
23
Q
Why did you have two ground connections?
A
Incase one is faulty, the other one can be used. This prevents the loss of the animal without recording.
24
Q
What is the difference in EEG/ground and EMG wires?
A
EMG are coated in silver to allow better preservation as they are implanted into the muscle.
25
Why do you use stainless steel wire instead of the copper wire for electrodes?
Copper will oxidise within the brain and can be toxic. Whereas stainless steel is conductive, non corrosive and non toxic.
26
Why was an offset of 0.1 cm used?
The target was the CA1 and so to ensure the targeting was appropriate.
27
Why was the electrode wire attached to the pin using epoxy instead of soldering?
Because the stainless steel doesnt conduct with solder, instead the epoxy allows good connection to the pin.
28
Why do you cover the connections with dental cement?
This works as extra insulation of the pins to reduce electrical noise.
29
Why were the electrodes superglued together?
To maintain the shape of the electrodes and to make insertion into the brain easier with the electrodes being sturdier.
30
Why do the electrodes need to be within a certain impedance?
The resistence controls the current floe through the electrode.
If the resistence is too high this will prevent the current flow and reduce signal strength.
If it is too low, it may be noisy.
31
Why were ephys animals housed alone?
For the seperate project, they were recorded from their home cages.
31
Why were the ground screws above the cerebellum?
Because this has the least neuronal activity so will not influence the recorded signals.
32
Why was that age range used for ephys recordings?
Because these mice on the c57 background begin to become deaf from around 5-months.
33
Why do you use a preamp and amp for plexon?
Because the signals that we are recording are on a miniscule range and are therefore greatly susceptible to noise.
Therefore, they need amplified as qucikly as possible. Thus, they are amplitified on the skull by preamp.
Then, they are amp 100 times by amp.
34
Why were the signals amplified 1000 times?
As signals are so low so they need maximised so they are on an identifiable range.
35
Why was ephys recorded at 1-kHz?
Gamma oscillations are fast rhythmic fluctuations in neural activity, with cycles occurring within milliseconds. A higher sampling frequency provides better temporal resolution, allowing you to capture the rapid dynamics of gamma oscillations with greater precision.
And to preserve the signal by oversampling
36
Why was the internal faraday cage required?
Because the treatment chamber has the LEDs which will provide 40-Hz noise. So the mice are within the internal chamber so ground from them.
37
How can you ensure the signals were not noise from 40-Hz?
First we tested with the connections in salineinside the internal faraday cage and did not find this.
Then, we found that the signal intesntiy varied acrtoss the duration of treatment on which suggests this is not due to noise as we would expect that to be more stable.
38
Why did you perform the treatment for ephys in a randomised order and how was it randomised?
To be sure that the effect being seen was not due to the same order being presented.
Using a built in labview generator.
39
Why did you only supply the treatment fo 60s? As this is different to the treatment recordings will this not be different?
Because the change in brain state can occur rapidly.
We do not expec it to be
40
What is int16?
Its a data type for storing infomration.
This approach allows lowest memory to be used and as these files are large we use this.
41
Why was the sync pulse 0.5 v used?
Becuase the speaker was driven at 0.77V and the led at 5V which meant that both sync pulses will show a voltage greater than 0.5v in their on periods.
Allows detection of the stimulation in the ephys channels.
42
Why do you have three different frequency ranges for ephys analysis?
The first was to show if we see an increase in 40-Hz over the duration of the treatment and therefore focussed surrounding the 40-Hz band which is why we choose 35-45.
Second was to see changes in the frequyency bands across treatments which is why we choose a wider band - 10-60.
Lastly, we wanted to quantify a specific target of 40-Hz to see the exact change in 40-Hz power which is why we choose 39.9-40.1
43
Was the mean not calculated for the heatmap analysis for ephys?
Yes it was, it shows the mean PSD analysis across mice.
44
Why did you use shapiro-wilk tests and what does this do?
We wanted to check the normality ofg the data in order to present and analyse it properly. It tests to see the goodness of fit of the data to a normal distribution
We choose this normality test because this is more suitable for smaller groups of data.
45
Why did you do a one-way anova on ephys data?
Because I wanted to compare the 40-Hz power across each treatment, which is only one indepdentn vairable.
As the data was normally distributed this was the best choice.
46
Why did you do bonferroni corrections?
If the one-way anova showed that the test was significant, we wanted to determine what treatment groups showed a statistically significant difference to one another.
bonferroni is a good way to do so as it is conservative and will adjust the p-values appropriately to avoid false positives.
47
How was the treatment order randomised if the same two treatments had to be completed as the last recordings?
A recording schedule was planned before the recordings began, always placing the AC or NS as the last rreatment. We then used a random gonline generator.
48
Do you worry that your older mice cannot hear the sound?
We did consider this when designing the experiment as our group has previously assessed the hearing abilities of c57 mice.
They found that around 6-months their hearing threshold should be around 35dB so should hear it well. Also, the orifinal paper has mice this age and also see an effect.
49
Why do you only end on AV or NS?
Because as our approach has not been fully confirmed for real-time assessment using a treatment, we wanted to confirm the effects seen with histology.
For this, due to our low sample size, we choose to have a non treatment control group and a treatemnt group.
As we hypothesise that the multisensory approach should be most effective, we choose this.
50
Will the fact the mice have underwent all treatments not effect the histology analysis?
The original paper that show the effects on soluble AB levels with ELISA show that the effects recover betweeen 12-24 hours. As all treatments were seperated by at least 48-hours, this should not effect this?
51
This will require alot of redosing of M04... what will this affect?
The clearance of m04 is unclear.
However, we maintained fluorescence and so it is liekly it will continue to bind to new oodiogmers in the brian.
52
Why did you use different powers?
At first, we had a spell where we were concerned that the higher powers were causing bleaching of m04 and therefore we reduced the powers.
However, this minimised our signal intesntiy and we increased this again.
53
Why are the number of reps different?
Because we have issues with laser stability, this means our signals can contain noise and therefore, variability across our recordings.
To increase the iterations to allow a better median signal and to try reduce this noise, we increased the number of reps.
54
Why did you not continue the day 0 recording for all?
At first, I thought having the baseline day 0 signal with no m04 would be benefiical as a control. As for RD exps we wused to see an increase from this signal and did this like so.
However, here we are monitoring the changes throughout the recording so our goal was slightly different. Therefore, the bleching of M04 role throughout the recording was likely to be important and so we would normalise against day 1.
Therefore, day 0 was not a necessity.
55
Why was the post-baseline changed?
After a discussion ont the conflicting findings suggested that the differences the brain was taken post-treatment may have an effect on the differing results, we wanted to make this as consistent with the previous acute study as posisble.
56
Were the mice scared, what was their behaviour like throughout the treatment?
They were startled when the treatment first started. After they would be around the chamber and may be in active rest.
hey tended to be pretty immobile, but this was not uncommon for these recordings.
Tge activity of thr mouse throughout the treatment andn baseline recordings could be monitored to see if there is a change in stressful behaviour.
57
Why were FAD- mice undergo only AV treatment?
This was a control group as they do not have plaques so we should not see any singal.
Therefore, we choose the multisensory to see if this caused an other changes in autofluroescnece.
58
Why did you record at the photodetectors at 5000 hz?
Because we stimulate on a ms range and by applying 5000 Hz this allows us to sample ~ every 0.2 ms which allows us to have a good sampling around our laser ON periods.
59
Why was data at 120 and 440 used?
440 nm provided us our m04 signals. as we seen good signals here, we did not use the 550 nm channel.
120 was the hiest power across all recordings and allows a fair comparison across recordings.
60
What did the 550 channel show across these recordings?
It was dairly similar as in FF experiments where no change was seen upon addition of m04.
61
Why were galvo measures below 0V removed?
Because there was an initialisation lag so we recorded from -1G to make this more accurate and therfore we did not require this.
Also, we did not align below 0V so this would not be characterised.
Also, this will just span back up the fibre which will have already been measured.
62
Why was there an initialisation lag with the galvo?
We were modulating the galvo and laser together at very rapid speeds which is feasible with this equipment. But commonly when modulating with DAQ it may take several ms before the appropriate power is reached, which we moniored with the sync channels and modified appropritately.
63
Why did you only show in vitro and Norm data for GENUS?
Because in chapter 4 this combinatiom provided the strongest signals in the depth-resolved manner and so this would be the optimal way to monitor a change in fluroescence over times.
64
Why were only 3 mice used for histological analysis?
For FAD+ mice we only had a total of 6 mice foe GENUS experiments. As we spolit this into two groups for histological assessment, this resulted in only two groups of 3.
65
Why did you not do ELISA and do you think this would have changed the results you would have seen?
On these mice it was not feasible for us to do ELISA as for this we would need to take at least one hemisphere for processing.
However, our main goal would be also to confirm that the phtootometry signals are correlated with histology signals as investigated in the previous chapter.
Therefore, we requried both hemispheres for histological analysis.
I think it may have provided a change in plaque pathology as ELISA is more sneisive at detecting changes in olgiomer fragments, while histology better represents the clusters of AB pathology.
66
Why can you not use amasine to quantify plaque pathology?
Because we only have a set amount of sections which will vary across mice. In this, we will not always have the full area of the specific brain region we want to monitor.
Therefore, we cannot calculate the volume of the brain region that we have processed.
67
Why was photometry data tested using the kruskal-wallis test?
Because the data was not normally distributed and we wanted to complete a reapeat measures test comparing two variables such as time and depth with the dependent variable being fluorescence.
We would use this over friedman as it allows for groups to be unequal in size as we gve some time points with more measurements due to differing lengths in recordings.
68
Why was histology quantified using wilcoxon signed rank test?
Because the data was not normally distributed and this allowed testing of the two independent treatment groups.
69
Why is there some with more recrodings?
In some mice, if we had time or we thought the signals were average, we would repeat the recording in order to ensure we captured somethign good before perfusion. Therefore, some have 2 recordings on the one mouse.
70
What do the ages for histology quantification represent?
This shows the age at which they were perfused.
71
Why do you mention a treatment-specific increase if there was no statistical power between groups?
Upon quantification of the change in gamma power is evident at a stronger value at AV compared to the others. Also, previous studies suggest this combined approach allows a greater spread across the brain.
I believe the lack of significance is most likely due to the small sample size.
72
Why do you think the increase in gamma power is not as intense in the hippocampus?
Because this is a deeper brain structure and as this stimulation should directly effect the sensory cortices,
73
Do you think there is some sex-specific differences in the way they respond to GENUS?
Across this data, in some recordigns it did appear that the FAD- may have a slightly increased power compared to the FAD+, suggesting that there is a less response. But this was not consistent across recordings and we have a low sample size.
74
Do you think the slighlt lower increase in 40-Hz in A is due tot he hearing abiltiies of mice?
Potentially as in some recordings the response at A stimulation is low. As all mice were same age we tried to limit this effect but of course some may present with worse hearing than the others.
75
What does group comparison mean in this stats table?
This row shows the p-value of the group comparison using the one-way anova test.
76
Why do you think you would see a stronger effect with the multisensory stimulation?
Because its activating two sensory cortical pathways which will enhance the spread of gamma oscillations.
77
It looks liek some mice have no response to the treatment, what do you think is happening in these cases?
In some it may represent the fact that they were sleeping throughout the recording which limits the effect of the stimulation.
78
The FAD- AV has a strongest power, why would this not be significnat?
It appears that we have an outlier which has little response to the treatment which may effect the overall contribution to the effect seen.
79
Why did you not show histology to confirm the location of ephys implant?
Histology on these mice were still being completed by my colleague.
80
Why do you expect a bigger change in the VC?
Because this treatment is a sensory treatment and therefore, its stronger effects should be in the sensory cortex.
81
Why would you see these signal variations from the previous day?
These signal variations can be due to small differences in the system state.
For example, the laser power could be slightly higher. The autofluorescence may be slightly higher than before. The brain fluorescence and blood flow may be different.
These are relatively small differences and are also seen in RD exps.
82
Why do you think you see this increase in fluorescence at these higher galvo voltages for A and V treatment?
For V, the majority of recordings remain flat and there is not a consistent trend of increased fluorescence. I believe this increase is due to the differing post-tme points which were at the lower fluorescence range which scews the last data points.
For A, at 4V galvo the trend of increase fluorescence. This may be due to AF or an increase over time of laser power.
83
What does H and (*) mean?
In summary, the H statistic in the Kruskal-Wallis test quantifies the degree of difference among the medians of the groups being compared, helping to determine whether these differences are statistically significant.
* represents the number of groups.
84
Why do you see the decline in fluorescence for control treatments?
Combination of a lack of full recording times which may be skewing the data at end time points.
Also, some may be vleaching of surrounding AF tissue.
85
Why are all the colour ranges different?
Because the variation from day to day can be different across each recording.
86
Why is there an increase in V heatmap?
The increase in fluorescence at higher galvos was always most seen at higher galvos, so this could represent a recovery of AF which was not fully accounted for with this analytical approach.
87
Why do you see stronger signals at specific depths in the A and AV examples?
These small differences show a increase in fluorescence at these depths from the previous day. This could be due to a change in AF of the system and the brain tissue.
88
Why are there such big outliars?
Some appear to have a substantial difference from the previous day.
89
What so the specs on the box plot represent?
Middle bar is the median.
Upper and lower bars are the IQR - intraquartile range. The IQR is the difference between the upper and lower medians.
The top and bottom lines show the minimum and maximum value.
The outliers are values that are more than 1.5 IQR away from the box.
90
Why is there such a big variation in fluorescence values across recordings?
Differences in system state across days
91
Why is there such big differences from day 1 to 2? Does this not go against the redose finsings where you claim its consistent across days?
No, we do see some small variations in the RD experiment which is due to the reasons as explained. However, it is not a substantial value that showed not be significant.
92
Did you try normalise against the first 30-mins of treatment?
No, but this was something that I have considered. But as we were trying to show the analytical approaches we stuck to this method for simplicity.
93
Why are there some missing timepoints for NS?
There are some with a shorter post-baseline measurement.
Also, there is one recording that was stopped prematurely due to environmental conditions.
94
In AV- the decline is steep in some, why?
Because this is just brain tissue so it is likely the high power is bleaching surrounding tissue.
95
Why would uou see small rises and falls trhoughout these recordings, what would this represent?
The range of change is quite low so might represent such low noisy fluctuations.
96
How did you choose the example data?
A best example that had the same parameters for all recordings.
97
What do previous histological quantification show?
After chronic, they seen reduction in plaque number and area.
More recently they show this after acute stimulation.
98
Why do you get a p value that is =1?
If there is no stastical power across groups. This may be due to low sample size, no difference in distribution of mean between groups or also due to the conservative nature of the post-hoc tests which can increase the p-value.
99
Why do you see a decrease in plaque pathology in this histology example?
It could be due to the treatment.
Or it is more likely that this is due to natural variation in plaque pathology across mice and as we do not have a before image to go off this is misleading.
100
Why did you choose these brain regions?
This is where the fibre is recording from.
Also, the VC is the sensory visual cortex which should be most activated by ther V and AV treatment and should have some effect there.
The subiculum is in hippocampal areas which have been tested before but have not seen much effect.
101
What does each dot on the histology histogram represent?
A density measure. For each mouse, two sections that had the fibre track on it were quantified.
102
Why are you showing sex differences?
Becuase it is known that females have a more aggressive pathology than males so it is important to show incase this may cause some trend.
Also, while not largely studied, this treatment may have sex-specific differences.
103
What is the scale in the zoomed images?
the same
104
For Post and PreS there is a lower median for AV, what do you think this means?
This is most likely due to the low sample size.
105
How well was image J at labelling hthe plaques?
The overlap was near perfect and there was little false positives.
106
How were the eilcoxin tests corrected for mlticomparisons?
using bonferroni correction where we multiplied the p value by the number of iterations.
107
Why do you not show the multicomparison results for TF or histology data?
If I do not show the multicomparisons, it is because the multicomparison test was not completed because the initial test was not significant or because no group significance was found between variables.
108
Why is there 6 measurements from 3 mice for histology quantification?
Each mouse had 2 sections quantified.
109
How do you choose the images for histological assessment?
I took the images that had the TF track present.
110
Why do you think Soula has conflicting findings?
It is unclear if their elisa was measuring soluble or insoluble AB wich may be a reason.
Also, they are monitoring plaque area and number which will be insoluble fibrils which may not be as dramatic change after acute stimulation.
111
What do you think of Soula approach for real-time monritroing using M04?
I think its a good way to monitor plaques as it allows a pre and post measurement which overcomes limitations with histology.
112
Out of the papers you state are consistent with your findings, what ones show an increase in the PFC?
The multisensory paper shows that with AV treatment but not A alone, they have a better increase in LFP power was very small in mPFC, the median distribution of mean firing rate differences during A+V stimulation, compared to no stimulation, differed significantly from zero
113
What brain regions do they see an increase in c-fos in?
40-Hz visual stimulation increased the number of c-Fos+ cells in V1, somatosensory cortex (SS1), hippocampal area CA1, and dentate gyrus (DG), and cingulate cortex (CC) of the prefrontal cortex (PFC)
114
Why do you think you seen an increased 40-Hz band in the PFC with V stimulation when previous papers did not?
Potentially because the effect is stronger because of the way the LEDs are arranged along the floor of the chamber which means the animals cannot avoid the stimulation.
115
How does your treatment chamber vary from previous studies and why?
We have the LED strip on the bottom of the chamber, instead of on a wall 60-cm away from the chamber. We done this to try equalise the light power along the whole chamber so there would not be areas which have less activation.
116
What differences in anumal model, age, set-up are there for ephys which may result in these differences?
age: hearing abilities
animal model: differences in PV vell viability
set-up: the light power and sound of treatment
117
Why would this comparison be misleading?
Because we are monitoring fibrils which are insoluble, while this acute measurmeent is soluble ab.
118
Has there been any recent advances in the field which may effect this finding?
Yes, recent paper showing acute GENUS and quantify plaque pathology on confocal z-stacks. They show reduction of plaque pathology.
119
Do you think having M04 bound to the plaques would effect the clearance mechanisms?
I am not completely sure as the clearance of M04 from the brain by iteself remains unknown.
120
Why would you not see the large decline in plaque pathology prevoously seen in acute conditions?
Previously in acute experiments, they monitor the change in AB levels by using elisa to test the change in soluble ab and did not see an effect with insoluble AB. However, soluble AB should not contain beta-sheets. Which is consistent with our findings.
However, more recently a paper has shown that with acute stimulation they saw a decreased signal intensity of plaques. However, their methods are lacking and the approach can introduce some bias if they have not blinded and randomised appropriately.
121
Is the way you monitor plaques in histological images consistent with the other papers?
In some.
Others do confocal imaging - recent paper - where they create z-stacks and create a dense core and halo which the quantify.
122
How can you know it was not influenced by sex?
I do not see any trends of increased or decreased plaque density with sex, instead it is varied.
123
Why do you think the original papers have been able to see such differences with such a low sample size?
The elisa is much more sensitive way to test the amyloid-beta levels.
124
What is this photometry measure actually quantifying when it comes to plaques?
We are quantifying the level of beta sheets and therefore, amyloid fibrils in the brain. This could be fibrils, or plaques.
Therefore, this is insoluble AB.
125
What does neuner et al 2019 show?
we combined a well-established mouse model of AD with a genetically diverse reference panel to generate mice that harbor identical high-risk human mutations but differ across the remainder of their genome
that genetic variation modifies the impact of human AD mutations on both cognitive and pathological phenotypes in AD mouse models.
126
Why do you think the chronic treatment would be better?
The chronic treatment has shown to have a bigger effect that is not transient.
For example, in acute experiments they originally quantified the change in plaque pathology using ELISA to quantify the AB fragment levels but in chronic they use histological measure to quantify the plaque number and see an effect. They also needed the chronic V stimulation to have an effect on insoluble AB and acute only showed an effect on soluble AB. This suggests that to see an effect on plaque size, a chronic paradigm may be required.
127
How was these fibres implanted and did they correlate with histology?
These were targeted in the same region as previous chapter as these were completed on the same mice. Correlation data is included in the previous chapter where in these specific mice we again seen some inconsistensies with the profiles.
This is something that was out of the scope of this project but is being worked on with help from collaborators that have a better knowledge on the light profile within the brain from TF.
128
You mentioned varied parameters and age group may help, why?
A bigger age group may help with a modified implant site where the plaques surrounding the fibre in the sensory cortex should be higher to increase the probability of collecting a strong signal at these regions.
129
Do you think administeraion of another treatment may have allowed identification of a decrease in plaque pathology?
Yes, there are other pharmaceutical agents that have already been shown to have effects at reducing AB levels in AD mouse models, even 5xFAD.
This would have perhaps a more dramatic decrease than GENUS and it would be easier to confirm the success of the approach if we akready know this treatment should reduce plaque pathology.
130
What kind of therapeutic agents could have been applied?
The newly approved antibodies could be administered although I am unsure how easy they are to get or administer i.p.
Alternatively, there are other pharmaceuticals that are not approved
DAPT --> gamma secretase inhibitor which has shown effects on AB levels and memory on PDAPP mouse and Tg2576 mice after sc injection.
delta-secretase inhibtior 11 --> seen effect on plaque and memoryin 5xfad mouse
131
How would this approach show an increase in plaque pathology?
It cant. This approach is to monitor the decreases in plaque signals only as this would be the therapeutic goal.
If the plaque signals were increasing, we would exptect to see a maintenenace of signal.
132
Why are you confident that it is not 40-Hz noise?
- implant in saline and seen no 40Hz
- variation of intensity across the duration of the signal
- bipolar not subtracted so would be stronger
but yes,, need spiking activity to make solid conclusion
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How can you detect strong LFP increases in PFC and CA1 unlike previous studies? What do they show?
the previous papers show increases in firing rate and phase modulation and some increases in LFP but they note it is not always consistent across mice due to implants etc.
Our PFC electrode has big surface area so collecting alot of strong signals
HIP electrode in target
But these may account for inconsistenties in our signal s
Also becaue our set up may be stronger as they cannot avoid the treatment.
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Why do you see those variations on GENUS TF?
Because we did not have the day 0 without methoxy for normalisation meant we were unable to attain the depth resultion.
Therefore, these signals show variation from a noisy M04 channel which may have decayed at a different rate, resulting in these changes from D1.
This need an optimal approach and further investigation into how to reliably analyse this data.
135
Why no effect with GENUS?Why are there differing findings from GENUS?
M04 staining insoluble - original papers with acute show difference with only soluble, but most recent apper shows changes in plaque size - conflucts with previous study
potentially stress - tsai commented that the mice being avoidance could prevent the treatment working
sensitivity differences in the approach - if TF is in region with low plaque density in VC it may not have sufficient signal to monitor a change
Differences in approaches to monritor
