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##Fluorescence Microscopy
## Brain lesion patients
* Lesions in brains or degenerative diseases help us understand brain function
* Phineas Gage– Railroad spike through frontal lobes changed his personality
<div><img src="figs/image7_0e1af20.png" height="200px"><figcaption></figcaption></div>
<div><img src="figs/image8_c3232ea.png" height="200px"><figcaption></figcaption></div>
Note:
Furthermore, studies of patients with brain lesions has historically been key to localizing parts of the brain that affect emotional states and learning and memory.
e.g. Phineas Gage in 1848 his whole personality changed after the spike went through his brain.
Harlow wrote: "the equilibrium... between his intellectual faculties and his animal propensities seems to have been destroyed"
---
## Model organisms— C. elegans
* The nematode worm *C. elegans* is great for genetic engineering and has a tiny nervous system (just 302 neurons)
<div><img src="figs/Adult_Caenorhabditis_elegans_d76c553.jpg" height="150px" title="CC from wikipedia https://commons.wikimedia.org/w/index.php?curid=2680458"><figcaption>C. elegans– commons.wikimedia.org/w/index.php?curid=2680458</figcaption></div>
<div><img src="figs/c-elegans-connectome_2_9548c95.jpg" height="150px"><figcaption>C. elegans wiring diagram– [openworm.org](http://www.openworm.org), neuroconstruct.org</figcaption></div>
Note:
It is difficult to visualize and record neurons and manipulate genes in humans so neuroscientists study a number of different model organisms.
Now to do neuroscience research we have to use model organisms of course. Small number of neurons, can be labeled using green fluorescent protein or other means.
C. elegans is a nematode or roundworm. It is non-infectious and non-parasitic organism just 1 mm long and it can be easily genetically engineered. That means you can introduce mutations to genes or express fancy inert proteins that allow you to track the function of genes and cells in living animals making it a great model organism.
For neuroscientists it has only 302 total neurons making it a great way to dissect neural circuits underlying simple behaviors. Many mutant worms have been isolated that affect nervous system function allowing us to learn about the function of those genes. And you can engineer the worms to express fluorescent proteins so that the animal's neurons glow under a microscope. How many of you have heard of green fluorescent protein?
Having just 302 neurons is great for for some types of studies, however we have more than a million neurons in each of our eyes just alone
More than 1 million neurons that just form the optic nerve from each of our eyes!
---
## Model organisms— squid
Squids have unusually large axons (1 mm diameter)
<div style="width:250px; float:left;"><img src="figs/20000_squid_holding_sailor_f98a242.jpg" height="300px"><figcaption>20000 Lieues Sous les Mers, J. Verne</figcaption></div>
<div style="width:500px; float:left;"><img src="figs/Squid_Loligo_pealei_cbafe46.jpg" height="300px"><figcaption>Atlantic squid, *Loligo pealei*</figcaption></div>
<!-- <div><img src="figs/axon_large_9a8a930.jpg" height="300px"><figcaption>Squid giant axon, R. Hanlon MBL Woods Hole</figcaption></div> -->
Note:
Jules Verne provided inspiration for the space age but also neuroscientists in the 1940s.
Squids are arguably the most important model organism in the history of neuroscience. They are rarely studied anymore but their large axons which are 1mm in diameter-- 1000x bigger than our axons-- made their axons amenable to sticking electrodes inside them in the 1930s-50s and allowed neuroscientist to discover the biophysical and mathematical basis of neuronal signaling. We will discuss squid giant axons in much more detail soon.
Other important invertebrate organisms in neuroscience research include sea slugs and fruit flies and zebrafish. Some of these are very amenable to genetic engineering like C. elegans and have nervous systems more similar to our own.
Phylum: Mollusca
Class: Cephalopoda
Order: Teuthida
Family: Loliginidae
Genus: Loligo
Atlantic squid (Loligo pealei)
Phylum: Mollusca
Class: Cephalopoda
Order: Sepiida
Family: Sepiidae
Genus: Sepia
---
## Model organisms— *Mus. musculus*
The mouse is a common model in neuroscience research.
<div style="width:225px; float:left;"><img src="figs/adult_mouse_jax_ec76ad4.jpg" height="200px"><figcaption>Common house mouse *Mus. musculus*, jax.org</figcaption></div>
<div style="width:300px; float:left;"><img src="figs/abi_adult_mouse_brain_e79e400.jpg" height="200px"><figcaption>Mouse brain 3D rendering, [Brain Explorer 2](http://mouse.brain-map.org/static/brainexplorer)</figcaption></div>
<div style="width:430px; float:left;"><iframe src="https://www.youtube.com/embed/stPThgZ2Y5o" width="420" height="315"></iframe><figcaption>Green fluorescent protein (GFP) labeled neurons inside a mouse brain</figcaption></div>
Note:
But mammals are the only animals that have evolved a convoluted superficial part of the brain called the neocortex. And it is the cerebral neocortex is crucial for our highest cognitive functions, even if it sometimes seems that in election years that humans have lost their cerebral function.
Thus for research pertaining to the structure and function of the mammalian brain and human disease we turn to rodents like the common house mouse. Mice are small with a brain 2 cm in length, develop fairly quickly, and their genome has long been one of the most amenable to genetic engineering though this is quickly changing newer molecular biology techniques (like the CRISPR/Cas9 system).
* Mouse brain is about 2 cm in length
* genetically tractable
* [https://www.youtube.com/watch?v=stPThgZ2Y5o](https://www.youtube.com/watch?v=stPThgZ2Y5o)
---
## Model organisms– other mammals
Higher mammals are used to study more complex brain functions.
<div><img src="figs/1f412_3fc8278.svg" height="200px"><figcaption>Cats– visual system function, locomotion</figcaption></div>
<div><img src="figs/1f412_f738dec.svg" height="200px">
<figcaption>
Non-human primates– attention, decision
making, vision, brain machine interfaces
</figcaption></div>
<div><iframe src="https://www.youtube.com/embed/UJ_rFMYDbAE" width="420" height="315"></iframe><figcaption>Rhesus monkey mind controlled wheelchair</figcaption></div>
<!--
<div style="float:left;"><figure style="display:inline-table;"><img src="figs/1f412_3fc8278.svg" height="200px"><figcaption style="display:table-caption; caption-side:bottom;">Cats– visual system function, locomotion</figcaption></figure></div>
<div style="float:left;"><figure style="display:inline-table;"><img src="figs/1f412_f738dec.svg" height="200px"><figcaption style="display:table-caption; caption-side:bottom;">Non-human primates– attention, decision making, vision, brain machine interfaces</figcaption></figure></div>
<div style="width:430px; float:left;"><iframe src="https://www.youtube.com/embed/L2O58QfObus" width="420" height="315"></iframe><figcaption>Rhesus monkey mind controlled wheelchair</figcaption></div>
-->
Note:
put new 2018 duke study in here. [interbrain cortical synchronization and mirror neurons between monkeys](http://dx.doi.org/10.1038/s41598-018-22679-x)
Research with cats was critical for work from the 1950s to 1980s that allowed neuroscientist to learn how visual signals are processed in the highest circuits of the mammalian brain.
And research with rhesus monkeys has been essential for learning about perceptual, attentional, and decision making in the mammalian brain together with research into brain-machine interfaces that have direct clinical applications for human patients.
3rs: Replacement, Reduction, and Refinement
---
## Fluorescence Microscopy
* Fluorescent molecules absorb light at one wavelength and emit it at another-longer wavelength.
* Uses optical filters to allow only light of a given wavelength in and out.
@@ -76,7 +221,7 @@ Here expressed in fly peripheral neurons
Note:
Remember that GFP is a gene that encodes a protein. You can put it behind the promoter to detect which cells express a given gene.
Remember that GFP is a gene that encodes a protein. You can put it behind the promoter to detect which cells express a given gene.
---
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## Tumor detection
MRI
CT-SCAN
MRI
CT-SCAN
<div><img src="figs/image29_a6736bb.png" height="100px"><figcaption></figcaption></div>
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## Magnetic resonance imaging (MRI)
<div><video height=400px controls src="figs/Animation01-01MagneticResonanceImaging.mp4"></video><figcaption>Neuroscience 5e Animation 1.1</figcaption></div>
<div><video height=400px controls src="figs/Animation01-01MagneticResonanceImaging.mp4"></video><figcaption>Neuroscience 5e Animation 1.1</figcaption></div>
Note: