Research Summary
Developmental studies of the gut nervous system, implicating GI tract disorders

The hypothesis that bone morphogenetic proteins (BMPs), which act early in gut morphogenesis, also regulate specification and differentiation in the developing enteric nervous system (ENS) was tested. Expression of BMP-2 and -4, BMPRIA, -IB, and -II, and the BMP antagonists, noggin, gremlin, chordin, and follistatin, was found when neurons first appear in the primordial bowel at E12. When applied to immunopurified E12 ENS precursors, BMP-2 and –4 induced the nuclear translocation of phosphorylated Smad-1. The number of neurons developing from these cells was increased by low concentrations and decreased by high concentrations of BMP-2 or BMP-4. BMPs also induced the precocious appearance of TrkC-expressing neurons, and induced dependence of the TrkC neurons on NT-3 for survival. BMP-4 interacted additively with GDNF to enhance neuronal development and to limit GDNF-driven expansion of the precursor pool. BMP-2 and –4 also promoted smooth muscle development from mesenchymal cells. To determine the physiological significance in vivo of these observations, the BMP antagonist noggin was overexpressed in the developing ENS of transgenic mice under the control of the neuron specific enolase promoter. Neuronal numbers were increased by more than 50% in the myenteric plexus in all regions of the bowel and by 30% -50% in the submucosal plexus. In contrast, TrkC-expressing neurons decreased in both the myenteric and submucosal plexuses of the noggin-overexpressing animals. BMPs-2 and/or –4 thus limit the size of the ENS but promote the development of specific subsets of enteric neurons, including those that express TrkC.

Service Activities
Editorial and Consultative
1980-present: referee for articles submitted to: J. of Neuroscience; Developmental Biology; J. of Neuroscience Research; Molecular Brain Research; J. of Neurobiology; Experimental Neurology; Neurobiology of Aging; Cell& Tissue Research; J. of Cell Science.

1980-present: reviewer for grants submitted to NSF, SBIR tissue culture/NIH; NIMH; Hunter College internal grants CUNY; Research Grants Council Hong Kong University, China; The Eli and Edythe L. Broad Foundation Los Angeles, CA.

Selected Publications:

1. Chalazonitis, A. Neurotrophin-3 In the Development of the Enteric Nervous System In “NGF and Related Molecules in Health & Disease”, eds Aloe/Calza Elsevier, Amsterdam, The Netherlands Progress in Brain Research 146:243-263 (2003).

2. Chalazonitis, A., Pham,T.D., Rothman T.P., DiStefano P.S., Bothwell M., Blair-Flynn J., Tessarollo, L. and Gershon M.D. Neurotrophin-3 is Required for the Survival-Differentiation of Subsets of Developing Enteric Neurons. The J. Neurosci. 21:5620-5636, 2001.

3. Chalazonitis, A., Rothman, T.P. Chen, J., and Gershon, M.D. Age-dependent Differences in the Effects of GDNF and NT-3 on the Development of Neurons and Glia from Neural-Crest-Derived Precursors Immunoselected from the Fetal Rat Gut: Expression of GFRalpha1 in vitro and in vivo. Devel. Biol. 204:385-406,1998.

4. Chalazonitis, A., Rothman, T.P., Chen, J., Vinson, E.N., Maclennan, A.J. and Gershon, M.D. Promotion of Enteric neurons and Glia by Neuropoietic Cytokines: Interactions with Neurotrophin-3. Devel. Biol., 198:343-365 (1998)

5. Chalazonitis, A., Tennyson, V.M., Kibbey, M.C., Rothman, T.P., and Gershon, M.D. The alpha1 Subunit of Laminin-1 Promotes the Development of Neurons by Interacting with LBP110 Expressed by Neural Crest-Derived Cells Immunoselected from the Fetal Mouse Gut. J. Neurobiol. 33:118-138, 1997.

6. Chalazonitis, A., Rothman, T.P., Chen, J., Lamballe, F., Barbacid, M. and Gershon, M.D. Neurotrophin-3 induces neural crest-derived cells from fetal rat gut to develop in vitro as neurons or glia. The J. Neurosci. 14:6571-6584, 1994.

Current Projects

1. Regulatory Molecules in Enteric Nervous System
Specific aims are: (i) to identify the cells in the developing gut that express BMP2/4 signaling molecules (ligands, BMPRs, transductional substrates (Smads 1, 5, 8), antagonists) and analyze their developmental expression; (ii) to test the first hypothesis, that BMPs cause uncommitted crest-derived precursors (isolated from E12 fetal gut by immunoselection) to commit to neuronal or glial lineages, by means of experiments with mass clonal organotypic cultures; and (iii) to test, by in vitro studies, the second hypothesis, that BMP2/4 continue to act at E14 on some of the now-committed progenitors to enable them to become responsive to other factors, such as glial growth factor 2 (the neuregulin we have found to be expressed in the fetal gut), NT-3, and neuropoietic cytokines and, in so doing, to terminally differentiate and survive.
National Institute of Diabetes and Digestive and Kidney Diseases
4/1/2002-3/31/2007

Keywords

neural crest, neurotrophin receptors, GDNF, NT-3, Smad proteins, BMPs, enteric nervous system, gut, autonomic nervous system, tissue culture, transgenic mice

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