4809 Introduction The floor plate of the vertebrate embryo develops at the ventral midline of the neural tube. It acts as an embryonic organiser and plays an essential role both in the generation of specific neuronal subtypes along the dorsoventral axis of the brain and spinal cord and in the guidance of axons (Giger and Kolodkin, 2001; Jessell, 2000). Although the role of the floor plate in patterning the neural tube is well accepted, its ontogeny has been a controversial subject (Le Douarin and Halpern, 2000; Placzek et al., 2000). The prevailing view, for which there is extensive evidence in the chick, is that axial mesodermal notochord cells are the source of an instructive inducing signal that mediates floor plate differentiation in medial cells of the overlying neural plate (Jessell, 2000; Placzek et al., 2000). A large body of evidence, moreover, suggests that the secreted signalling molecule Sonic hedgehog (Shh) mediates the ability of notochord to induce floor plate differentiation. Shh is expressed in the node and the notochord prior to floor-plate differentiation. Gain-of-function experiments show that Shh can induce the ectopic differentiation of floor-plate cells in the neural plate in vitro (Marti et al., 1995; Roelink et al., 1994), while blockade of Shh in the notochord eliminates its ability to induce floor plate cells (Ericson et al., 1996). In support of studies in the chick, mutations in the Shh gene, and in components of the Shh signalling pathway, in mouse, block ventral midline differentiation (Chiang et al., 1996; Ding et al., 1998; Matise et al., 1998; Wijgerde et al., 2002). Several recent studies, however, have questioned this model of floor plate induction. In particular, analysis of chick-quail chimaeras in which quail cells from the chordoneural hinge (CNH), a derivative of the Node, are grafted into chick embryos, have led to the proposal that medial floor plate cells are derived from a population of precursors that are initially situated in the Node, can segregate into either notochord or floor plate, and are already pre-specified within this region (Le Douarin and Halpern, 2000; Teillet et al., 1998). In this model, floor-plate cells thus derive from pre-specified cells that intercalate from the node into the neural midline. A further challenge to the paradigm of notochord/Shh- mediated floor plate induction arises through observations of zebrafish embryos. Floor-plate cells persist in embryos in which notochord precursors are surgically ablated, demonstrating that a normally developed notochord is not a pre-requisite for floor plate differentiation in this species (Shih and Fraser, 1995). Analyses of zebrafish mutant embryos further supports this contention. Mutations in both no tail (ntl) and floating head (flh) affect notochord formation (Amacher and Kimmel, 1998; Halpern et al., 1993; Schulte Merker et al., 1994; Talbot et al., 1995). Despite this, in ntl mutant embryos the most medial set of floor plate cells are present, To begin to reconcile models of floor plate formation in the vertebrate neural tube, we have performed experiments aimed at understanding the development of the early floor plate in the chick embryo. Using real-time analyses of cell behaviour, we provide evidence that the principal contributor to the early neural midline, the future anterior floor plate, exists as a separate population of floor plate precursor cells in the epiblast of the gastrula stage embryo, and does not share a lineage with axial mesoderm. Analysis of the tissue interactions associated with differentiation of these cells to a floor plate fate reveals a role for the nascent prechordal mesoderm, indicating that more than one inductive event is associated with floor plate formation along the length of the neuraxis. We show that Nr1, a chick nodal homologue, is expressed in the nascent prechordal mesoderm and we provide evidence that Nodal signalling can cooperate with Shh to induce the epiblast precursors to a floor-plate fate. These results indicate that a shared lineage with axial mesoderm cells is not a pre-requisite for floor plate differentiation and suggest parallels between the development of the floor plate in amniote and anamniote embryos. Movies available online Key words: Floor plate, Prechordal mesoderm, Chick, Nodal, Sonic hedgehog Summary Distinct modes of floor plate induction in the chick embryo Iain Patten 1, *, Paul Kulesa 2,† , Michael M. Shen 3 , Scott Fraser 2 and Marysia Placzek 1,‡ 1 Centre for Developmental Genetics, Department of Biomedical Science, University of Sheffield, Sheffield S10 2TN, UK 2 Beckman Institute 139-74, California Institute of Technology, Pasadena, CA 91125, USA 3 Center for Advanced Biotechnology and Medicine, UMDNJ-Robert Wood Johnson Medical School, 679 Hoes Lane, Piscataway, NJ 08854, USA *Present address: Departament de Genetica, Facultat de Biologia, Universitat de Barcelona, Avenida Diagonal 645, Barcelona 08028, Spain † Present address: Stowers Institute for Medical Research, 1000 East 50th Street, Kansas City, MO 64110, USA ‡ Author for correspondence (e-mail: m.placzek@sheffield.ac.uk) Accepted 19 June 2003 Development 130, 4809-4821 © 2003 The Company of Biologists Ltd doi:10.1242/dev.00694 Research article