February 20th lecture Embryology   Biology 441   Spring 2008   Albert Harris   Amphibian Axis Creation and Induction Lecture: Until fertilized, frog and salamander eggs have radial symmetry. Where the sperm enters causes that side to become anterior, by causing the opposite side (180 degrees around the other side) to invaginate and form the blastopore, which will later become the site of the anus of the embryo & animal. The causal mechanism is a mass rotational shift of cortical cytoplasm toward where the sperm entered. This creates a "grey crescent" of cytoplasm (not usually visible) where the blastopore will invaginate. The grey crescent was long assumed to be a special kind of cytoplasm, but its special property turned out to be that certain cortical layers of cytoplasm come into contact here, that don't touch elsewhere. If you hold a newly-fertilized egg upside down for many minutes, then gravity will shift heavier vegetal cytoplasm downward, and this cytoplasmic flow can bring those cortical layers into contact at some second location. That will cause a second invagination, and a second body axis. This method of producing conjoined twins was discovered in The late 1800s (& called Shultzian twinning) And many people published research papers about it; But then the phenomenon was forgotten (except by scientists who happened to own old textbooks, such as the book by Thomas Hunt Morgan) Éuntil rediscovered in the late 1900s, & its mechanism discovered. Embryonic Induction (discovery of which earned the Nobel Prize for Hans Spemann) Another way to cause formation of a second blastopore, And a second body axis, Is to wait until early or mid gastrula stages (when gastrulation, the inward migration of future mesoderm & endoderm is going on, and then surgically graft tissue from the dorsal lip of the blastopore into the blastocoel of a second embryo (even of another species, which helps trace cell fates!) The grafted tissue differentiates into notochord (& somites) and 'recruit' surrounding cells of the "host embryo" to change their differentiation & cell fates, and form an entire extra body from what would Have been belly skin (-> a second neural tube) And lateral plate mesoderm (-> more somites & kidneys). Any time a signal from some cells causes nearby cells to change their differentiated cell fate, and become different cell types than they otherwise would, Biologists called this "Induction" Notice that the word induction has very different meanings In physics and in philosophy, & other subjects, too. Ms. Hildegard Proesholdt discovered embryonic induction Of amphibian body axes by grafting dorsal lips of blastopores Of newt (salamander) gastrula-stage embryos. (she was a young graduate student, & this was her PhD research) She soon married Otto Mangold, and the discovery is credited to Spemann & Mangold. Spemann put his name first on the paper they published. Two other graduate students working for PhDs In Spemann's lab at that time (just after World War I) in Freiburg im Breisgau, Germany, between the Black Forest and the southern Rheinland, also made great discoveries, & became famous. Johannes Holtfreter & Victor Hamburger. =========================================== Many other examples of induction were then discovered For example, an American had already discovered that the lens of the eye is induced to form from what would otherwise become skin (somatic ectoderm). In this case, the inducing signal comes (mostly!) From the optic cup (which is the out-folding from the brain, that becomes retina.) And the lens induces overlying somatic ectoderm to differentiate into cornea. And the differentiation of the neural retina is induced by contact with somatic ectoderm. Without that contact, all the retina differentiates as pigmented retina. The two layers of teeth each induce differentiation of the other. Liver, pancreas, lungs and salivary glands are each induced by inductive signals from adjacent mesoderm. Endoderm that would have differentiated into one of these can be induced to differentiate into any of the others, by grafting of mesoderm. Induction of a neural tube by signals from notochord was called "Primary Induction" and for 70 years the conceptual assumption was like rows of dominos being knocked over. Thousands (I would estimate) of research biologists spent major parts of their careers trying to use bioassay methods to discover "the inducing substance" on the assumption that it was one simple chemical, analogous to acrasin or auxin. One reason they failed is that lots of abnormal chemicals can also induce neural tubes if grafted into the blastocoel of amphibian embryos. For example, Holtfreter induced an extra neural tube using a piece of salami from the edge of a sandwich he had eaten for lunch that day. When I was a graduate student, some other students and I found that salami is a more effective inducer that bologna, But that considerable induction can be induced by either. ------------------------------------------------------------------------ In the 1930s, the Englishman Conrad Waddington achieved primary induction in chicken embryos. He got this result by grafting the primitive node (Hensen's node) [which is the functional equivalent of the dorsal lip of the blastopore, because its cells are future notochord] Waddington made this discovery in the same room at Cambridge where others later discovered such Phenomena as that proteolytic enzymes can dissociate Cells, and that elastic substrata can map cell traction. ------------------------------------------------------------------------- Waddington then was the major designer of allied anti-submarine tactics in World War II, And is credited with inventing operations research. His papers on warfare were classified top secret for many years, but can now be bought in book form. He smoked a pipe, and at seminars, repeatedly blew smoke rings at whoever was speaking. When I was your age, I found this disconcerting, and dodged them by stepping sideways to me a moving target. It distracted me to wonder if maybe he was testing new anti-sub tactics on me. I just thought you might be interested. &&&&&&&&&&&&&&&&&&&&&&&&&&&&& Much more has since been learned about the proteins That cause induction, by the combinations of genetic methods Which our textbook calls... FIND IT... LOSE IT... MOVE IT...

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