4/16/13 9:50 PM 4/3/12 Lecture 1 Issues at hand: 1. Fertilized eggà a region becomes NS, what’s the mechanism and mapping of where NS will be. Specifying the region of the embryo that gives rise to the NS. Region called neuro-ectoderm. Three germ layers: • Inner layer: endoderm: gut/intestinal tubes • Middle layer: mesoderm: muscle, fat, blood • Outer layer: ectoderm: two components o Epidermis: skin o Neuroectoderm: NS - The mechanism by which you establish the dorsal-ventral axis for the embryo determines the tissue partitioning. -anterior-posterior axis gets separated into various segments. -Maternal vs. zygotic gene functions. • Maternal: completely from maternal genes. Ex dorsal region of embryo is from the egg(mother). Mother’s dorsal ventral axis can be through transferred to the egg to have dorsal-ventral axis. • Zygotic: genes provided by both parents. Activated at a certain point and the zygotic genes then take over development 2. Pathfinding, how cells wire together. • Axon’s follow a few critical choice points at which there are several independent cues that are informative. 3. Learning: • experience based rewiring • overlap between developmental and learning pathways and associative mechanism by which coincidences are registered (brain is designed to remember coincidences as the essence of how you learn) Model Systems: - Humans: o Advantages: many diseases are self reported so information comes from there.§ Genome sequence is complete and can identify many defects in the genome § Genetics comparison in animal studies vs. human models is different b/c in order to detect subtle mutations in animals is harder than subtle relations in humans because the subtle abnormalities in humans (such as the leg tripping ex) are self-reported. o Can record behavior and can ask subjects about state of minds and record self-reported mutants. 7X10^9 humans in the world. Thought that around 5000 genes that can be mutants. • Disadvantages: o No experimental access and fetal material hard to get - Non-Human Primates: • Advantages: o Most like human in terms of physiological and some behavioral aspects o Can show theory of mind: can estimate what other animals are thinking and what they want them to do or not to do. • Disadvantages: o Fetal experiments difficult and expensive o No genetics: there’s no systematic genetic analysis available - Carnivores: dogs, cats, ferrets • Advantages: o Humanlike brains, visual systems are born immature so NS develops much more after birth. o Advanced mammals o Fetal work is easier o Dog genome (including for diseases such as epilepsy and behaviors such as sheep eye) and well established breeds o Some behaviors are determined purely by single genes • Disadvantages: o No genome sequenced for cats and ferrets 4/5/12 - Rats and Mice: Advantages: • Mice:o still somewhat similar to humanlike brains o Reverse genetics: many mutants exists and possible to do targeted gene-knockouts: introduce into embryonic stem cell of mouse and knockout gene of interest upto a single AA change and use the mouse may carry a single or even two copies of the gene. Can even introduce a human gene mutation in the mouse to study effects. o Genetic mosaics: trimeric mice. Ex mouse that trembles may be a defect in the nerve or a defect in the muscles. Take a wild type and mix with cells of a mutant (can tell them apart by using gfp or some other bacteria to detect gene). If knockout in NS and still trembles then it’s probably a muscle defect. • Rats: o Advantages: Rats are smarter than mice so better for maze experiments o Disadvantages: difficult to do embryonic studies because if a mutant is severely deformed, then mother will resorb it. Harder to do knockout and development is relatively slow. Chicks/Quails: Advantages: • The genome for chickens is known. • Genes can be crossed between chicks and quails to see which regions control which behavior (ex quail can be transgenic to make chicken sounds) • Chicken embryos are accessible outside of mother in eggs; cut a hole in the egg and can cut out regions of the embryo and place that region in an egg of a quail. Grafting experiment. Disadvantages: 300 million years of separation difference between birds and mammals. There’s little systematic genetics available. Amphibia: Frogs and Newts Advantages: • The best for grafting experiments • Still vertebrate• Accessibility of embryo; tadpoles develop outside of mother and there’s not shell. • Can do gene knock-down by suppressing it’s function but not completely knocking it out. Disadvantages: • Not so good for genetic studies because harder to knock genes out and make transgenic frogs. Frogs are tetraploid so it’s harder to control genetics. Zebrafish: Advantages: • excellent genetics, many mutants exist because they are transparent and can find mutants much more easily. Embryos are easy to work with to make knockout fish. • -gene-knock down from RNAi and morpholinos. • Easier to do grafting. Disadvantages: • Gene knockouts are hard. Invertebrates: dorsal-ventral axis is also inverted in embryo Drosophila: -Advantages: • Genetics are very well known • Can actually knockout/knockdown/mis-regulate (take a gene and make it active only in certain areas). Transgenesis is very easy • Can add a peptide tag to a fruit fly gene so that they glow. • Gene manipulation is very flexible. Fast generation time ~2 weeks. -Disadvantages: • neurons are small so hard to do physiological experiments (i.e sticking electrodes in) • not a vertebrate. 550 million years of separation. C. Elegans: Advantages: - excellent genetic known -RNAi knockdown is possible for every gene- 959 cells/302 neurons/56 glial. Less neurons so can see experimental effects of individual neurons. - laser ablations of specific cells that have defined lineage. Cell-cell communication known. -entire wiring diagram of neural circuits is known. -faster life cycle Disadvantages: - can’t do physiological experiments. - even more separation from humans.-! - Gurdon Exp: frog embryo removed of nucleus and inserted another frog nucleus in its cytoplasm. Could do this with any organ system such as a nucleus from a skin cell. the skin cell nucleus had all the genes necessary to make the fully developed frog in the irradiated egg.-