A6 - Unsicker / Kalcheim

The classic perception of neural crest (NC) development suggests that neuroendocrine chromaffin cells and sympathetic neurons originate from a common sympathoadrenal (SA) progenitor cell, which, under the influence of glucocorticoids (GC), differentiates into a chromaffin cell. Our previous and current work in the SFB 488 indicates that both the “common progenitor” and the “glucocorticoid” hypotheses must be revised. Analyses of mice lacking glucocorticoid receptor (GR) signalling (GR-/-) or an adrenal cortex (SF1-/-) reveal largely normal chromaffin cells. Several lines of evidence suggest that the chromaffin cell fate is likely to be determined prior to the colonization of the adrenal anlagen. Thus, transcription factor and signalling networks generating sympathetic neurons are distinctly different from those in chromaffin cell development. Furthermore, we have found that some SA progenitors show characteristics of chromaffin cells prior to migration into the adrenal anlagen. So far, we have not been able to identify a signal in adrenal anlagen, which specifically induces a chromaffin phenotype. BMP4, because of its persistent expression in embryonic adrenal cortical cells and peri-adrenal mesenchyme, seemed to be a promising candidate for specifying chromaffin cell fate. However, grafting BMP4 overexpressing cells to sympathetic ganglia primordia did not shift neuronal to neuroendocrine cell fate. Since we found SA progenitors to be already phenotypically heterogeneous prior to invading the adrenal anlagen, we hypothesize that this diversity must be generated earlier in NC development. Using electroporation of NC cells with GFP-DNA we have begun to label single and small groups of NC cells at different time points of delamination from the neural tube (NT) and follow their migration to the sites of para- and prevertebral sympathetic ganglia and locations of intra- and extra-adrenal chromaffin cells. Preliminary data obtained in project A5 have suggested that progenitors for sympathetic ganglia, DRG, and melanocytes delaminate in successive waves from the NT. We have begun and will continue to investigate whether a dynamic spatiotemporal fate map of cells in the NT also exists for the progenitors of sympathetic ganglia and chromaffin cells. Following the strategy outlined in A5 and in close collaboration, we will aim at elucidating the putative molecular code underlying specification to the various sympathetic neuronal and chromaffin subtypes. We will explore the relationship between cell specification and cell migration and ask whether the respective homing sites, e.g. adrenal anlagen, when grafted homo- and heterochronously next to the site of delamination from the NT, may influence cell fate.

In a second part of the project we will continue to analyze the postnatal fate of extra-adrenal chromaffin cells in the organ of Zuckerkandl, investigate cellular and molecular details of their death, which reveals features of autophagy, and, in collaboration with the Schütz laboratory (D6), define the role of GR signalling in autophagic cell death pathways. In initial studies we have “re-discovered” the organ of Zuckerkandl and shown that conditional deletion of the GR (GR^DBHCre) dramatically accelerates chromaffin cell death in this largest accumulation of extra-adrenal chromaffin cells.

In previous studies we have established the protective roles of members of the neurotrophin (NT), FGF, TGFß, and CNTF families in the maintenance of target-deprived preganglionic sympathetic neurons in the intermediolateral column (IML) of the spinal cord that innervate adrenal chromaffin cells. We have shown that postnatal IML neurons, contrary to somatic motor neurons, die when deprived of their target cells. Furthermore, concerning the physiological rather than the pharmacological relevance of these factors we have used respective knockout mice and found that, amongst others, NT4 and cardiotrophin-1 play major roles in IML development and function. Thus, in a third part of this grant application we propose an in-depth investigation of the role of GDNF family members and their receptors for the generation and maintenance of IML neurons and the innervation of the adrenal gland as a major target organ. Particular emphasis will be put on a comparison with somatic motor neurons.

Unsere bisherigen Arbeiten im SFB 488 haben u.a. gezeigt, dass chromaffine Zellen bereits vor der Einwanderung in die Nebennierenanlage determiniert sind. Ob diese Festlegung schon auf Neuralleisten-Ebene geschieht werden wir in diesem Projekt untersuchen. Weiterhin werden wir die Bedeutung von Glucocorticoiden für die postnatale Entwicklung von extra-adrenalen chromaffinen Zellen sowie die Rolle der GDNF Familie und ihrer Rezeptoren für die Entwicklung und Erhaltung präganglionärer sympathischer Neurone klären.

Own project-related publications since the last application

Gut, P.#, Huber, K.#, Lohr, J., Brühl, B., Oberle, S., Treier, M., Ernsberger, U., Kalcheim, C., and Unsicker, K. (2005). Lack of an adrenal cortex in sf1 mutant mice is compatible with generation and differentiation of chromaffin cells. Development 132: 4611-4619 #equal first authors

Lohr, J., Gut, P., Karch, N. Unsicker, K., and Huber, K. (2006). Development of adrenal chromaffin cells in sf1 heterozygous mice. Cell Tiss. Res. 325: 437-444

Ernsberger, U., Esposito, L., Partimo, S., Huber, K., Franke, A., Bixby, J.L., Kalcheim, C., and Unsicker, K. (2005). Expression of neuronal markers suggests heterogeneity of chick sympathoadrenal cells prior to invasion of the adrenal anlagen. Cell Tiss. Res. 319: 1-13

Huber, K., Karch, N., Ernsberger, U., Goridis, C., and Unsicker, K. (2005). The role of Phox2B in chromaffin cell development. Dev. Biol. 279: 501-508

Oberle, A., Schober, A., Meyer, V., Holtmann, B., Henderson, C., Sendtner, M., and Unsicker, K. (2006). Loss of leukaemia inhibitory factor receptor ß or cardiotrophin-1 causes similar deficits in pre-ganglionic sympathetic neurons and adrenal medulla. J. Neuroscience 26: 1823-1832

Huber, K.#, Franke, A.#, Brühl, B., Krispin, S., Ernsberger, U., Schober, A., von Bohlen und Halbach, O., Rohrer, H., Kalcheim, C., and Unsicker, K. (2008). Persistent expression of BMP-4 in embryonic chick adrenal cortical cells and its role in chromaffin cell development. Neural Development (3) 28, doi:10.1186/1749-8104-3-28, #shared first authorship.

Parlato, R., Otto, C., Stotz, S., Grone, H.J., Unsicker, K., and Schütz, G. (2009). Chromaffin cell-specific ablation of the glucocorticoid receptor affects their postnatal development. Endocrinology 150, 1775-1781, Epub 2008 Nov 26

Own project-related publications prior to 2005

Finotto, S.#, Krieglstein, K.#, Schober, A., Deimling, F., Lindner, K., Brühl, B., Beier, K., Metz, J., Garcia-Arraras, J.E., Roig-Lopez, J.L., Monaghan, P., Schmid, W., Cole, T.J., Kellendonk, C., Tronche, F., Schütz, G., and Unsicker, K. (1999). Analysis of mice carrying targeted mutations of the glucocorticoid receptor gene argues against an essential role of glucocorticoid signalling for generating chromaffin cells. Development 126, 2935-2944   #equal first authors

Huber, K., Brühl, B., Guillemot, F., Olson, E.N., Ernsberger, U., and Unsicker, K. (2002). Development of chromaffin cells depends on MASH1 function. Development 129: 4729-4738

Roosen, A.#, Schober, A.#, Strelau, J., Böttner, M., Faulhaber, J., Bendner, G., McIllwrath, S.L., Seller, H., Ehmke, H., Lewin, G.R., and Unsicker, K. (2001). Lack of neurotrophin-4 causes selective structural and chemical deficits in sympathetic ganglia and their preganglionic innervation. J. Neuroscience 21: 3073-3084 #equal first authors

Schober, A., Minichiello, L., Keller, M., Huber, K., Layer, P.G., Roig-Lopez, J.L., Garcia-Arraras, J.E., Klein, R., and Unsicker, K. (1997). Reduced acetylcholinesterase (AChE) activity in adrenal medulla and loss of sympathetic preganglionic neurons in TrkA-deficient, but not TrkB-deficient mice. J. Neuroscience 17: 891-903

Schober, A., Wolf, N., Huber, K., Hertel, R., Krieglstein, K., Minichiello, L., Kahane, N., Widenfalk, J., Kalcheim, C., Olson, L., Klein, R., Lewin, G.R., and Unsicker, K. (1998). TrkB and neurotrophin-4 are important for development and maintenance of sympathetic preganlgionic neurons innervating the adrenal medulla. J. Neuroscience 18: 7272-7284

Schober, A., Hertel, R., Arumae, U., Farkas, L., Jaszai, J., Krieglstein, K., Saarma, M., and  Unsicker, K. (1999). Glial cell line-derived neurotrophic factor rescues target-deprived sympathetic spinal cord neurons but requires transforming growth factor-ß as cofactor in vivo. J. Neuroscience 19: 2008-2015

Selected publications from other projects outside the SFB since 2005

Subramaniam, S., Shahani, N., Strelau, J., Laliberté, C., Brandt, R., Kaplan, D., and Unsicker, K. (2005). IGF-1 inhibits ERK to promote neuronal survival via the Phosphatidylinositol-3 kinase/Protein kinase 1/c-Raf pathway. J. Neuroscience 25: 2838-2852

von Bohlen und Halbach, O., Minichiello, L., and Unsicker, K. (2005). Haploinsufficiency for trkB and trkC receptors induces cell loss and accumulation of a-synuclein in the substantia nigra. FASEB Journal 19: 1740-7042. For full text see fasebj.org/cgi/doi/ 10.1096/fj.05-3845fje; doi: 10.1096/fj.05-3845fje IF 6.820

von Bohlen und Halbach, O., Krause, S., Medina, D., Sciaretta, C., Minichiello, L., and Unsicker, K. (2005). Regional- and age-dependent reduction in trkB receptor expression in the hippocampus in associated with altered spine morphology. Biol. Psychiatry 59: 793-800

von Bohlen und Halbach, O., Zacher, C., Gass, P., and Unsicker, K. (2005). Age-related alterations in hippocampal spines and deficiencies in spatial memory in mice. J. Neuroscience Res. 83: 525-531

Zechel, S., Jarosik, J., Kiprianova, I., Schober, A., Unsicker, K., and von Bohlen und Halbach, O. (2005). FGF-2 deficiency does not alter vulnerability of the dopaminergic nigrostriatal system towards MPTP intoxication. Eur. J. Neuroscience 23: 1671-1675

Egorov, A. V:, Unsicker, K., and von Bohlen und Halbach, O. (2006). Muscarinic control of graded persistent activity in lateral amygdala neurons. Eur. J. Neuroscience 24: 3183-3194 IF 3.820

Schober, A., Peterziel, H., von Bartheld, S., Simon, H., Krieglstein, K., and Unsicker, K. (2007). GDNF applied to the MPTP-lesioned nigrostriatal system requires TGF-ß for its neuroprotective action. Neurobiology of Disease 25: 378-391

Jarosik, J., Legutko, B., Unsicker, K., and von Bohlen und Halbach, O. (2007). Antidepressant-mediated reversal of abnormal behavior and neurodegeneration in mice following olfactory bulbectomy. Exp. Neurol. 204: 20-28

Peterziel H., Paech T., Strelau J., Unsicker K. and Krieglstein K. (2007). TGFß/GDNF-family signalling crosstalk in the regulation of ciliary ganglionic neuron survival depends on the nature of GFR alpha. J. Neurochem. 103: 2491-2504

Bachmann, I., Jakubick, V.C., Shaked, M., Unsicker, K., Tucker, K. L. (2007). A simple slice culture system for the imaging of nerve development in embryonic mouse. Dev. Dyn. 236: 3514-3523

Zechel, S., Schober, A., Meinhardt, A., Unsicker, K., von Bohlen und Halbach, O. (2008). Distribution of leucine-rich-repeat-kinase-2 (LRRK2), a novel member of the Ras/GTPase superfamily, in the developing and adult murine brain. Exp. Neurol.., in press

Reviews

Unsicker K., Reuss B. and Bohlen und Halbach, O.v. (2005) Fibroblast growth factors in brain functions. In: Handbook of Neurochemistry and Molecular Neurobiology. Volume “Neuroactive Proteins and Peptides, 2^nd Edition (R. Lim, ed). Oxford University Press

Subramaniam S., and Unsicker K. (2006). Extracellular signal-regulated kinase as an inducer of non-apoptotic neuronal death. Neuroscience 138: 1055-1065 (invited review)

Unsicker K., and Krieglstein K. (Editors) (2006). Cell Signaling and Growth Factors in Development, Two Volumes, 978 pages. Wiley-VCH Weinheim

Kuhn, G., Brüstle, O., Martens, U., Wobus, A., and Unsicker, K. (2008). Stem cells: established facts, open issues, and future directions. Cell Tiss. Res. 331:1-3.

von Bohlen und Halbach, O., and Unsicker, K. (2008). Neurotrophic factors for dopaminergic neurons. In: Development and Engineering of Dopamine Neurons (R.J. Pasterkamp, M.P. Smidt, and J.P.H. Burbach, eds) Landes Bioscience and Springer Science+Business Media, in press

Klaus Unsicker

Institut für Zoologie

Universität Heidelberg

Im Neuenheimer Feld 230

69120 Heidelberg, Germany

Phone:  06221 - 54 8228
Fax:      06221 - 54 5604
email:   Klaus.Unsicker@urz.uni-heidelberg.de

Chaya Kalcheim

Dept. of Anatomy and Cell Biology

Hebrew University of Jerusalem- Hadassah Medical School

Jerusalem 91120, Israel

Phone:  00972-2-6758438
Fax:      00972-2-6758438
email:    kalcheim(at)nn-shum.cc.huji.ac.il