MCDB 423 Lecture 19: Lecture 19

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University of Michigan - Ann Arbor
Molecular, Cellular and Developmental Biology
MCDB 423
John Kuwada

Lecture 19 Slide 2 Most of the ganglion axons will cross the optical chiasm to go to the other side of the brain and will enter the tectum (in frogs) or the colliculus (in mamals) Within this tectum structure, there are layers of cells -These axons from synapses with neurons in specific layers So how do we get to the correct site of synatpic contact? Slide 3 The paper uses this technique Place a small piece of lipophlic dye I in one specific region of the tectum, it will fill the entire cell including all the axons and their projection points to the tectum -This is done in the mouse so it’s the colliculus -All these axons are going to the termination zone slide 4 We know already that there’s chemo specificity -There are molecules determining the location of the target -You’ll also see that there is a role for neural and synaptic activity in the final location of the synapses and the final selection of synaptic target slide 5 If we look early in development and look at those retinal ganglion cells that are forming specific synaptic targeting sites in one specific part of the colliculus, the axons are survialing a broad region of the colliculus -So the axon is touching a large portion of the colliculus and then they are reorganizing and retracting and ultimately settle down to make synapses in one area This is a major kind of remodeling that happens -This is an important part in selection of the target So what molecules determine the final location? Slide 6 So they wanted to identify molecules that guide axons to specific places in the midbrain structure by isolating biochemically from the membranes of different regions of the structure and testing the proteins in vitro -The retinal ganglion axons are making a choice between different regions (not chemoattractant to one place like the netrin) So the experiment is to make a strip assay where you’ll take a part of the retina where the retina ganglion cells are located at different parts of the retina and you’re going to in an explant culture present those retinal ganglion cells with either membranes from the anterior part or posterior part of the colliculus -In a strip assay, you’re setting it up so that the cells are close to both and they can choose which one to grow on or if they can make a choice (some are growing equally on both) They isolate the membrane proteins biochemically and constitute them into lysosomes Slide 8 This is the zoom up of what the axons look like growing on the strips A= anterior and alternate= posterior What you see: -If you look at the axons from the temporal part of the retina, you see them growing nicely on the anterior stripes and avoiding the posterior -But the nasal grow equally on all of them slide 9 Answer = either c or d (the experiment rules out a & b, but cannot distinguish c & d). This is the cartoon version of what they see We know that the temporal part of the retina are making some kind of choice; the nasal axons don’t discriminate -The temporal retinal ganglion cells could be recognizing an attractant in the anterior part of the tectum or they could be growing away that’s in the posterior part of the tectum To resolve between C and D -If we know what it is, we could have inhibit it -What they did is they probed to see which part (A or P) has functional molecules that helps make the choice slide 10 Answer = d. The posterior membrane has a component that is important for the choice, and this choice is avoidance! To
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