Midterm 2 Discussion Papers Flashcards
What methods are being used here?
Describe the key results of the graph.
qPCR
Looking for expression of Fprs in different tissues. Fprs are localized only in the VNO, which indicates they most likely have a role in olfaction. Had these receptors been strongly expressed elsewhere, it would lend less support to the hypothesis that Fprs are olfactory receptors
What methods are being used here?
Describe the key results of the graph.
Why was V2R not examined?
Using co-in situ hybridization
Testing for the expression of different olfactory receptor families in the same neurons in the VNO. Since these neurons expressed only one family of receptors (V1r or Fpr) this lends support to Fprs being olfactory receptors
Did not examine V2rs because they are expressed in a functionally distinct, different layer of the VNO
What methods are being used here?
Describe the key results of the graph.
Co-in situ hybridization
Looking for coexpression of different Fpr members (same family) in VNO neurons. Since they are not coexpressed, they are likely olfactory receptors
What methods are being used here?
What kind of cells are being tested on here (cell line)?
Describe the key results of the graph.
What is the control?
Fura-2 calcium imaging
Human embryonic kidney (HEK) cells
When Fpr-rs3-expressing cells are exposed to ligand CRAMP, Ca2+ enters the cell. CRAMP is an anti-microbial-associated ligand. CRAMP is the only ligand of Fpr1 and Fpr2 cells which causes a response from Fpr-rs3 receptors.
ATP and bath solutions are controls
What methods are being used here?
Describe the key results of the graph.
Calcium response, fura-2 of HEK cells expressing Fpr receptors (dissociated VNO cells)
All Fpr-rs receptors respond to at least one ligand of Fpr1 and Fpr2. There is a wide tuning of responses, some receptors are responsive to multiple ligands
What methods are being used here?
Describe the key results of the graph.
En-face confocal imaging of VNO: intact VNE signals - system is intact to create more “natural” responses of different VNO neurons
Confirm results from previous dissociated neuron experiments. Fpr receptors respond to immune function-related ligands, cells respond to a variety of ligands
What methods are being used here?
Describe the key results of the graph.
Antibody staining/immunohistochemistry
Found that Fpr receptors were localized in dendritic tips of VNO neurons, which is evidence that these proteins are in fact receptors (had they been localized in the soma, would not be evidence of sensory receptors)
What methods are being used here?
Describe the key results of the graph.
In situ hybridization of Fpr receptors
Fpr receptors are expressed by VNO neurons: seem to be localized in dendritic tips
What are the key takeaways of this figure?
See RFP in the dentate gyrus, don’t see RFP in the olfactory bulb yet because this image was taken early on, right after the birth of 4D+ neurons. This is a good thing because it proves the validity of the system
What are the key takeaways of this figure? What methods were used to obtain this image?
In situ hybridization for RFP mRNA
No more RNA when doxycycline is taken away: this is good because it proves that the 4D+ construct was inducible, and goes away when administration of the substance ends
What are the key takeaways of this figure?
4D induction increased the presence of interneurons in all progenitor cell types
What are the key takeaways of this figure?
The spine density of 4D+ was not significantly different than the spine density of 4D- cells - nothing is anatomically different about the more proliferative cells
What are the key takeaways of this figure?
The electrophysiology activity of 4D+ neurons was normal, and the same as 4D- neuron electrophys
What are the key takeaways of this figure?
When the olfactory discrimination tests were easy enough, 4D+ and 4D- mice were equally capable of discriminating between odours
What are the key takeaways of this figure?
When olfactory stimuli were signficantly diluted, 4D+ mice were better able to discriminate between which odor they had received
4D+ mice were better at discriminating and more confident in their licks for water
What is the key takeaway from this figure?
No information about coexpression of bitter taste receptors can be determined from this image
Seeing presence of receptors in human airway epithelium, not even just in the ciliated cells, just in all the cells of the airway
What is the key takeaway from this figure?
What are the controls?
Confirmation via pcr that T2rs are expressed in airway epithelium (bc microarray [where they first got info from] are not that reliable)
b-actin and water are the controls (+ve and -ve, respectively)
What is the key takeaway from this figure? (2)
What methods are being used here?
Immunostaining
Based on the figure can tell that T2rs being expressed on the motile cilia of the cells: evidence for functional receptor role
a-tubulin and gustducin are present: evidence for further functionality
What is the key takeaway from this figure?
What methods are being used here?
What is the control?
Fura-2 imaging
Washing bitter ligands over the cells in varying concentrations
Control = PBS, phosphate buffer saline
Bitter ligands can induce Ca++ entry into the cells
What is the key takeaway from this figure?
Ciliated cells respond to ligands faster (smaller latency): measuring the time of cells to react
Other cells still react, likely through gap junction interactions
If the ligand binds to the cilia first then this makes sense: confirms this hypothesis
What is the key takeaway from this figure?
Cilia beat faster in response to bitter ligands
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