The research areas of Zoology group are Developmental Biology with special reference to cellular interactions during vertebrate developement, and Genetic Toxicology.
Developement Biology
Development of a three-dimensional organism from a unicellular zygote remains one of the most challenging problems in contemporary biology. Beginning of pattern formation in the developing embryo is driven not only by cytoplasmic determinants but also, to a great extent, by cellular interactions. Cells interact with one another through direct physical contacts as well as through diffusible signaling molecules. We are interested in both these aspects of cell-cell interactions. In addition to looking at the role of the cell surface components in such interactions, we have also been analyzing the possible role of cytoskeletal elements in early morphogenesis. Over the past few years, the focus of work has shifted to identifying the signaling molecules that initiate early embryonic inductive interactions. We use a combination of gross morphological, ultrastructural and molecular techniques for this purpose
The work is presently focused on the possible role of different growth factors in early embryonic induction and pattern formation. The candidate molecules being analyzed are insulin, activin, fibroblast growth factor and human seminal plasma inhibin-related molecules. These have been short-listed based on the work already carried out by us. For example, we have demonstrated, for the first time in an amphibian embryo, that insulin plays an essential role in prepancreatic development. Insulin seems to play a similar role in the chick embryo. In both these systems, the action of insulin is associated with modulation of expression of developmentally crucial genes, such as, Brachyury, goosecoid and noggin. In addition, activin, FGF and inhibin seem to play important roles in different stages of mesoderm and neural induction and development. The current work is directed towards designing experiments to dissect out the precise roles of these molecules in early morphogenesis. The classical but powerful embryological techniques, such as, embryo culture, micromanipulation and grafting of discrete embryonic tissues are being used in combination with tools in molecular biology. In addition to the work outlined above, we have begun to use our experience and expertise to address diverse questions, such as, evolution of molecular mechanisms of development using the diploblastic hydra, possible effect of tidal inundation on developmental gene expression in frog embryos and the early development of chick heart.
Over the next few years, we hope to unequivocally establish the precise roles of some of these molecules in early stages of pattern formation in the vertebrate embryo. Simultaneously, it will allow us to elucidate interactions, if any, between these signaling molecules.
Morphogenesis of the visual system in vertebrates is the other area of focus under which the role of cellular determinants of organogenesis like proliferation, migration, apoptosis, determination and differentiation is studied. The inquiry is based on photoperiodic behaviour of the select animal models coupled with preponderance of photoreceptor cell types and consequent complexity of neuronal circuitry in their retina. In these models, the connections to and from the brain will be explored to identify the pathway involved in colour vision. We study developmental mechanisms in detail that operate in formation of various ocular tissues, temporal synchronization of tissue assembly, interdependent inductive interactions and morphological sequence of their pattern formation. The progress in cell differentiation is monitored using whole-mount tracer techniques, histology and biochemical detection of tissue specific markers. The quantitative analysis of lipid subclasses is being made using suitable models and TLC-HPLC techniques from early embryonic stages till landmark stages in development of eye and brain. Molecular techniques will be employed in studying expression of genes like Pax (particularly 6), rx (1 & 2), BMP4, sonic-hedgehog, connexins and cine oculis which are important in morphogenesis of eye and brain. The biophysical simulation studies continue, using glass-gelatin diffraction gratings to understand colour vision fundamentals and using spectrophotometry to interpret the role of lenticular and macular yellow in wavelength modulation of optical input to the retina.
Genetic Toxicology
To cope up with the task of identifying ‘genetic risk’ from ever increasing number of chemicals, several genetic toxicology detection tests have been introduced during last two decades. Several schemes of the test system batteries have been recommended by regulatory authorities. Debate has been already initiated on predictive potential of these test systems in extrapolation to human risk. Serious attempts are also being made to improve predictive potential of more popular short term tests by using computerized classification methods and by estimating unbiased comparative parameters like sensitivity, specificity and accuracy. Furthermore, the uncontrolled presence of genotoxins in any compartment of the natural environment is an unwanted situation, in particular, also from human point of view. The earlier studies have shown that there is a high correlation (90%) between mutagens and carcinogens. There is enough evidence to support a belief that the initiation of cancer involves mutation in somatic cells. Many in vivo and in vitro assays have been developed for detection of genotoxic chemicals. In our earlier studies, we have compared popular in vitro Ames Salmonella test system with promising in vivo Drosophila wing spot test system. For first time, we have established a database of chemicals assessed in both the test systems. Furthermore, we have analyzed predictive potential of these systems. Both these systems ultimately provided an account of change in frequency of induced point mutations with respect to specific marker mutant genes. To assign the mutagenic status to any chemical, apart from the other requirements, it is essential to test the suspected mutagen in a mammalian test system. Therefore, we planned to use mammalian in vivo or human in vitro test system by applying a novel technical approach which allows detection of induced chromosome aberrations. Keeping this in mind, we are using the mouse bone marrow micronucleus assay for the detection of potential genotoxic agents in combination with the fluorescent in situ hybridization technique. Use of in vivo mammalian assay allows the extrapolation of the results to human situation while use of somatic cells will help in correlating the data with the carcinogecity studies. Application of FISH technique gives an advantage of differentiating a chromosome breaking agent (clastogen) from a spindle poison (aneugen) in the same preparation. Further, these techniques are applied for identification of the potential clastogenic agents of plant origin in pure chemical form. Plants produce toxins to protect themselves against fungi , insect, pest attack and animal predators. When plants are stressed or damaged they may greatly increase their natural pesticide levels. The human intake of these toxins varies markedly with the diet and would be higher in vegetarians. Various reports are available wherein about 72 natural pesticides are assessed for their clstogenic potential out of which 48% were found to be positive. With a view to detect potential genotoxic agents of plant origin, pure forms of both chlorogenic acid and canavanine are being screened presently. With the expertise gained over past several years and with excellent facilities at hand, we are also in position to provide genotoxicty testing service on commercial basis which includes i) Ames Salmonella assay, ii) Drosophila wing spot test, iii) In vivo rodent micronucleus assay, iv) Chromosome aberration assay and Dominant lethal assay.