INTRODUCTION The neural crest and its derivatives have long occupied and continue to play a paradigmatic role in studies designed to unravel molecular mechanisms that underlie the specification of neural cell fates. The sympathoadrenal (SA) cell lineage is a major derivative of the neural crest, which gives rise to sympathetic neurones, adrenal and extra-adrenal chromaffin cells, and small intensely fluorescent cells of sympathetic and paraganglia (Landis and Patterson, 1981; Anderson, 1993; Unsicker, 1993). An essential role in triggering chromaffin as opposed to sympathetic neuronal fate has been attributed to glucocorticoid hormones (Unsicker et al., 1978; Doupe et al., 1985; Anderson and Axel, 1986; Anderson and Michelsohn, 1989; Michelsohn and Anderson, 1992). Evidence based on in vitro studies with mammalian SA progenitors has suggested that glucocorticoids are necessary for two important sequential steps in chromaffin cell development: first, to suppress neuronal markers in SA progenitors channelling them towards a chromaffin cell phenotype and, second, to induce the adrenaline-synthesizing enzyme, PNMT, in a subpopulation of chromaffin cells (Bohn et al., 1981; Anderson and Axel, 1986;. Anderson and Michelsohn, 1989; Michelsohn and Anderson, 1992). Moreover, the ability of nerve growth factor (NGF) and other growth factors to induce a neuronal phenotype in young chromaffin cells, and the capacity of glucocorticoids to prevent 2935 Development 126, 2935-2944 (1999) Printed in Great Britain © The Company of Biologists Limited 1999 DEV1324 Molecular mechanisms underlying the generation of distinct cell phenotypes is a key issue in developmental biology. A major paradigm of determination of neural cell fate concerns the development of sympathetic neurones and neuroendocrine chromaffin cells from a common sympathoadrenal (SA) progenitor cell. Two decades of in vitro experiments have suggested an essential role of glucocorticoid receptor (GR)-mediated signalling in generating chromaffin cells. Targeted mutation of the GR should consequently abolish chromaffin cells. The present analysis of mice lacking GR gene product demonstrates that animals have normal numbers of adrenal chromaffin cells. Moreover, there are no differences in terms of apoptosis and proliferation or in expression of several markers (e.g. GAP43, acetylcholinesterase, adhesion molecule L1) of chromaffin cells in GR-deficient and wild- type mice. However, GR mutant mice lack the adrenaline- synthesizing enzyme PNMT and secretogranin II. Chromaffin cells of GR-deficient mice exhibit the typical ultrastructural features of this cell phenotype, including the large chromaffin granules that distinguish them from sympathetic neurones. Peripherin, an intermediate filament of sympathetic neurones, is undetectable in chromaffin cells of GR mutants. Finally, when stimulated with nerve growth factor in vitro, identical proportions of chromaffin cells from GR-deficient and wild-type mice extend neuritic processes. We conclude that important phenotypic features of chromaffin cells that distinguish them from sympathetic neurones develop normally in the absence of GR-mediated signalling. Most importantly, chromaffin cells in GR-deficient mice do not convert to a neuronal phenotype. These data strongly suggest that the dogma of an essential role of glucocorticoid signalling for the development of chromaffin cells must be abandoned. Key words: Sympathoadrenal cell lineage, Glucocorticoid signalling, Chromaffin phenotype, Mouse SUMMARY Analysis of mice carrying targeted mutations of the glucocorticoid receptor gene argues against an essential role of glucocorticoid signalling for generating adrenal chromaffin cells Susetta Finotto 1, *, Kerstin Krieglstein 1, *, Andreas Schober 1 , Frauke Deimling 1 , Karin Lindner 1 , Barbara Brühl 1 , Konstantin Beier 1 , Jürgen Metz 1 , José E. Garcia-Arraras 3 , José L. Roig-Lopez 3 , Paula Monaghan 2 , Wolfgang Schmid 2 , Timothy J. Cole 2 , Christoph Kellendonk 2 , Francois Tronche 2 , Günther Schütz 2 and Klaus Unsicker 1,‡ 1 Neuroanatomy, University of Heidelberg, D-69120 Heidelberg, Germany 2 Division of Molecular Biology of the Cell I, German Cancer Research Center, D-69120 Heidelberg, Germany 3 Department of Biology, Box 23360, UPR Station, University of Puerto Rico, 00931-3360 Rio Piedras, Puerto Rico *These authors have contributed equally ‡ Author for correspondence (e-mail: klaus.unsicker@urz.uni-heidelberg.de) Accepted 14 April; published on WWW 7 June 1999