The history of embryology

History Of Embryology

The field of heredity includes three disciplines: embryology, regeneration, and genetics. Discussions of genetics necessarily entail a theory of development, and any theory of development must show how the eggs of different species develop in different ways. The hereditary theories of William Keith Brooks or August Weismann did not distinguish separate genetic and embryological domains (Pereda and Motta). The developmental mechanics of His, Roux and Driesch likewise contained explicit genetic components whereby the hereditary determinants thought to reside either within the cytoplasm or inside the nucleus were seen to direct the processes of organ formation and cell differentiation.

The split between genetics and embryology emerged gradually, largely through the investigations of Thomas Hunt Morgan and his laboratory (Murillo-Gonzalez). Whereas most American and German experimental embryologists followed Boveri in thinking that the nucleus was the site of the hereditary determinants, Morgan was convinced that these determinants lay in the cytoplasm. Morgan had collaborated with Driesch on a project that involved the removal of cytoplasm from the uncleaved ctenophore egg. The result of such operations was defective embryos. Morgan declared that there was "no escape from the conclusion that in the protoplasm and not in the nucleus lies the differentiating power of the early stages of development." However, in 1905, E.B. Wilson and Nettie Stevens both provided evidence that the nucleus did indeed contain the determinants of genetics and development (Leperchey and Barbet). They both correlated the XX chromosome composition with female animals and the XO or XY chromosome complement with male animals. If this were true, then the nucleus determined the sex of the individual.

Morgan responded by investigating a parthenogenetic species of aphids, eventually correlating chromosome number and sex. However, he interpreted his results as still being consistent with the cytoplasm having the controlling role in development. However, by 1910, Morgan had found mutations in Drosophila that could be best interpreted as segregating with the X chromosome. Although he initially resisted this interpretation, he eventually came to see the genes as physically linked on the chromosomes. What had begun as an investigation as to whether the nucleus or the cytoplasm controlled development ended in the founding of the gene theory.

In 1911, genetics arose as a discipline within experimental embryology (Kuratani, Kuraku and Murakami), but it soon evolved its own techniques, favored organisms, rules of evidence, and specialized vocabulary, which separated it from the rest of embryology (Churchill). In his 1926 book, The Theory of the Gene, Morgan formalized the split by declaring that genetics dealt exclusively with the transmission of hereditary traits, while embryology concerned the expression of those traits. He claimed that the sorting out of characters in successive generations can be explained without reference to the way in which the gene affects the developmental process, and that confusion had arisen from confusing the problems of genetics with those of development. Genetics and embryology began to go their separate ways.

Morgan publishing Experimental Embryology the year after The Theory of the Gene (Weaver and Hogan). When he left Columbia University to head the Biology division at the California Institute of Technology, he returned to study the problems of ascidian development. Thus, when Morgan published Embryology and Genetics in 1934, many biologists hoped that it would reunite these disciplines. However, this was not to be the case. In 1939, Richard B. Goldschmidt and Ernest E. Just published their respective attempts to unify the fields. Goldschmidt would have had embryology subsumed under genetics, while Just saw genetics as a rather minor subset of embryology. At the same time, at least three other researchers, Salome Gluecksohn-Schoenheimer, Conrad Hal Waddington, and Boris Ephrussi were attempting more balanced syntheses of the two disciplines.

The path from experimental embryology to developmental genetics from the Freiburg laboratory of Hans Spemann to the Columbia University of Leslie C. Dunn, was first traveled by Salome Gluecksohn-Schoenheimer (Van Speybroeck, De Waele and Van de Vijver).

Spemann, like many other embryologists of his day, had no interest in the new science of genetics. However, although he did not believe that genes played any major role in embryonic development, two members of his laboratory did perceive that genetics had some critical things to say concerning how organisms developed. One of these was Spemann's assistant, Viktor Hamburger. Hamburger supervised Gluecksohn-Schoenheimer's thesis and was the only one who provided students with some introduction to the principles of genetics. The second person was Conrad Hal Waddington, a Cambridge graduate student who came to Germany in 1932 to study Organizer phenomena and to learn the techniques of tissue grafting. He became one of Gluecksohn-Schoenheimer's closest friends, and the two of them had several discussions concerning the possible roles of genes in development. Gluecksohn-Schoenheimer decided that when she completed her dissertation, she would attempt to uncover the roles that genes played in the development of the embryo.

But neither Hamburger, who was studying the innervation of embryonic limbs, nor Waddington, who was trying to isolate the molecules responsible for Organizer function, were truly geneticists. To study animal genetics in Germany meant studying in Richard Goldschmidt's laboratory at the Kaiser Wilhelm Institute in Berlin-Dahlem. So in 1932, as she finished her dissertation, Gluecksohn-Schoenheimer went to see the "Lieber Gott von Dahlem."

In 1933, Gluecksohn-Schoenheimer and her husband, the noted physiologist Rudolph Schoenheimer, fled to America. Hamburger and Stern left the same year, eventually followed by Goldschmidt in 1936. While her husband had an appointment in Columbia University's College of Physicians and Surgeons, Salome Gluecksohn-Schoenheimer worked as a technician in the laboratory of Samuel Detwiler. This was an obvious place to work since Detwiler was interested in those problems of limb innervation, which the Freiburg laboratory had helped identify. This employment did not last long. Work by Dunn and his graduate student Paul Chesney had shown that the heterozygous (T/+) condition resulted in a shortened tail due to a constriction in the neural tube. Homozygous embryos died at 11 days in utero with the posterior half of their body missing. Chesney's work pointed to an earlier defect in the notochord as being responsible for the lack of a posterior neural tube. It appeared that the T. mutation was involved in axial determination. It was also possible that the wild-type T gene controlled the posterior inducer substance of the notochord, itself.

The first papers on the T-locus mutants make it clear that Gluecksohn-Schoenheimer interpreted these phenotypes as being caused by a genetic defect in the induction of the posterior neural tube by the notochord. In 1938, she concluded that our data do not give conclusive evidence for conceiving the malformations of the neural tube as secondary to the disorders of the notochord, but they point in this direction. By 1940, Gluecksohn-Schoenheimer was able to state more assuredly that in the heterozygous Brachy (T+) mouse the notochord in the posterior region of the embryo is defective and as a result the Brachy phenotype develops.

In doing these studies on the T-locus, Gluecksohn- Schoenheimer made a virtue out of necessity and founded the first version of developmental genetics (Schoenwolf). Unable to manipulate the mammalian embryo inside the uterus and placenta, she had looked to nature's own experiments. She gave a rationale for the emergence of developmental genetics out of experimental embryology. First, one could not study mammalian development as one had studied amphibian embryogenesis. It was not possible yet to use transplantation, isolation, or vital staining methods on mammalian embryos as they have been used on amphibian embryos. Second, instead of manipulating the embryo and seeing its affect on the phenotype, Gluecksohn-Schoenheimer proposed to look at the phenotypes produced by mutant genes and relate them back to their embryologic causes. A mutation that causes a certain malformation as the result of a developmental disturbance carries out an experiment in the embryo by interfering with the normal development at a certain point. By studying the details of the disturbed development it may be possible to learn something about the results of the 'experiment' carried out by the gene. Moreover, this program bridged the gap between genetics and embryology (Van Speybroeck, De Waele and Van de Vijver). Most American embryologists in the late 1930s did not think that genes acted during the early stages of development.

Third, Gluecksohn-Schoenheimer declared that this type of research was to be done by a new type of scientist, the developmental geneticist (Horder). While the experimental embryologist carries out a certain experiment and then studies its results, the developmental geneticist first has to study the course of the development and can then sometimes draw conclusions on the nature of the 'experiment' carried out by the gene.

Between 1938 and 1949, Gluecksohn-Schoenheimer pursued a research program explicitly linking embryonic organizers and specific genes in the mouse. The first series of these investigations looked at the interactions between dominant and recessive alleles at the T. locus. The dominant T. allele was interpreted as affecting the notochord's ability to induce the neural tube. The recessive t-alleles, moreover, were interpreted as being involved in more general mesoderm-forming processes. In to/to embryos, the mesoderm failed to form at all, while in T/to embryos, the mesoderm and notochord of the posterior regions were seen to be defective. These mutant genes, however, were considered to be alleles at the same locus. The answer to this genetic quandary was to be found in the embryology of the mouse (Burian et al.).

The effect of the two alleles T. And to on notochord and mesoderm might suggest that the two alleles act on two different structures. However, if considered from the embryological point-of-view, the notochord and mesoderm of the mouse have the same origin, namely in the tissue of the wall of the primitive gut. This brought the problem back to what had been thought of as the mammalian equivalent of the dorsal blastopore lip. Gluecksohn-Schoenheimer was trying to do with mutants what Spemann and the Mangolds had done by transplantations. As she would summarize in 1949, "The study of this material makes it very likely that in mammals the notochord plays a role in processes of early organization similar to that of the notochord in amphibians as analyzed with the techniques of experimental embryology." More than that, Gluecksohn-Schoenheimer thought that she could do with the T-locus what Spemann's group and the Cambridge laboratory of Needham and Waddington could not do: Find the inducer molecule, itself. Gluecksohn-Waelsch would later write, "It was therefore hoped that the identification of the mode of action of T-locus genes -- and the nature of their gene products-might provide leads towards the molecular analysis of normal inductive mechanisms."

The T-locus alleles weren't the only mutations that appeared to control induction. The phenotypes caused by another, closely linked, mutation, Kinked were interpreted in terms derived directly from Spemann's work on amphibian embryonic regulation. Homozygous mutants of Kinked were found to have duplications of their dorsal axis, sometimes forming twin embryos.

Their striking resemblance to the double-monsters obtained by constriction experiments of amphibian embryos at the two-cell stage led to the suggestion that an "organizer" region analogous to that identified experimentally in amphibians existed in mammalian embryos and that its normal functioning was severely affected in FuKi / FuKi embryos. There is no doubt that all these interpretations of mutational effect on the developmental mechanism was strongly influenced by the orientation of the particular investigators and their view of development as depending on a series of inductive interactions.

Gluecksohn-Schoenheimer interpreted all three genes (T, to, and FuKi) as disturbing "specific organizer relationships." She interpreted the action of the Kinked gene as causing constrictions analogous to those done experimentally by Spemann and Holtfreter on salamander eggs (Van Speybroeck, De Waele and Van de Vijver). These famous studies had shown that the constriction of the egg down the medial plane caused the formation of two organizers, each of which formed embryonic axes, thereby creating twin larvae. Constriction in the frontal plane, however, caused the formation of one normal larva and one BauchstYch, an amorphous tissue mass consisting chiefly of endoderm and blood cells. Partial constrictions, moreover, caused conjoined larvae, an observation that Spemann had related to mammalian teratology.

According to Gluecksohn-Schoenheimer, the Kinked mutants had an inducing mesoderm that was divided in two, just like Spemann's and Holtfreter's constricted embryos. The duplicated axes formed when this constriction was in the medial plane, and the BauchstYch-like mass seen in several of the Kinked embryos also "might well be the result of a frontal constriction."

Gluecksohn-Schoenheimer was aware of her integrating embryology and genetics. She announced that her research on the Kinked gene "was undertaken both from the point-of-view of the embryologist interested in the causal analysis of development and that of the geneticist interested in the analysis of gene effects." Linking organizers to genes meant linking embryology to genetics.

During this investigation of axial development, other tailless or short-tailed mutant mice were found. One of these tailless mutants was due to the Sd/Sd genotype that also caused the lack of kidneys. Gluecksohn-Schoenheimer wrote that when confronted with such cases, the developmental geneticist must reverse the order of the experimental embryologist and work backward from effect to cause. She demonstrated that the ureteric bud normally grew into the area of the metanephrogenic mesenchyme. When that occurred, the ureter continued to grow and branch, and the mesenchyme condensed into tubules. In the Sd/Sd mutant, however, the ureteric bud failed to reach the mesenchyme and no kidney was formed. In Sd/+ heterozygotes, some tips of the short ureteric bud did find there way into the metanephrogenic mesenchyme, and a small kidney resulted. These findings indicate strongly the existence of an inductive relationship between the ureter and kidney -- such as has been shown experimentally to exist in other vertebrates.

In Gluecksohn-Schoenheimer's work during this period, there is a reciprocity between genetics and embryology. Genetics could be used to analyze development in areas where experimental techniques had not yet been perfected. Embryology could identify the effects of these genes whose functions were necessary for the construction of the embryo. These early embryonic abnormalities "represent the end result of a chain of events at the beginning of which stands the gene. The analysis of the action of the gene is our ultimate goal."