Preliminary Observations on the Spawning Conditions of the European Amphioxus (Branchiostoma lanceolatum) in Captivity MICHAEL FUENTES 1 , MICHAEL SCHUBERT 4 , DIANA DALFO 2 , SIMONA CANDIANI 3 , ELIA BENITO 2 , JOSEP GARDENYES 2 , LAURA GODOY 2 , FREDERIC MORET 5 , MARGARITA ILLAS 2 , IAIN PATTEN 2 , JON PERMANYER 2 , DIANA OLIVERI 3 , GILLES BOEUF 1 , JACK FALCON 1 , MARIO PESTARINO 3 , JORDI GARCIA FERNANDEZ 2 , RICARD ALBALAT 2 , VINCENT LAUDET 4 , PHILIPPE VERNIER 5 , AND HECTOR ESCRIVA 4n 1 Laboratoire Arago, UMR 7628, CNRS and Universite´Pierre et Marie Curie, BP 44, F-66651 Banyuls sur Mer, France 2 Departament de Genetica, Facultat de Biologia, Universitat de Barcelona, 08028 Barcelona, Spain 3 Dipartimento di Biologia Sperimentale, Ambientale e Applicata, Universita di Genova, 16132, Genova, Italy 4 Laboratoire de Biologie Mole´culaire de la Cellule, UMR 5161 CNRS/ENS de Lyon, Ecole Normale Supe´rieure de Lyon, IFR128 BioSciences Lyon-Gerland, 69364 Lyon Cedex 07, France 5 Developpement, Evolution et Plasticite du Systeme Nerveux, UPR 2197, Institut de Neurobiologie A. Fessard, CNRS, 91118 Gif-sur-Yvette, France ABSTRACT Members of the subphylum Cephalochordata, which include the genus Bran- chiostoma (i.e. amphioxus), represent the closest living invertebrate relatives of the vertebrates. To date, developmental studies have been carried out on three amphioxus species (the European Branchiostoma lanceolatum, the East Asian B. belcheri, and Floridian-Caribbean B. floridae). In most instances, adult animals have been collected from the field during their ripe season and allowed (or stimulated) to spawn in the laboratory. In any given year, dates of laboratory pawning have been limited by two factors. First, natural populations of these three most studied species of amphioxus are ripe, at most, for only a couple of months each year and, second, even when apparently ripe, animals spawn only at unpredictable intervals of every several days. This limited supply of living material hinders the development of amphioxus as a model system because this limitation makes it more difficult to work out protocols for new laboratory techniques. Therefore we are developing laboratory methods for increasing the number of amphioxus spawning dates per year. The present study found that a Mediterranean population of B. lanceolatum living near the Franco-Spanish border spawned naturally at the end of May and again at the end of June in 2003. Re-feeding experiments in the laboratory demonstrated that the gonads emptied at the end of May refilled with gametes by the end of June. We also found that animals with large gonads (both, obtained from the field and kept and fed at the laboratory during several weeks) could be induced to spawn in the laboratory out of phase with the field population if they were temperature shocked (spawning occurred 36 hours after a sustained increase in water temperature from 191C to 251C). J. Exp. Zool. (Mol. Dev. Evol.) 302B:384–391, 2004. r 2004 Wiley-Liss, Inc. INTRODUCTION The invertebrate chordate amphioxus (Chorda- ta, Cephalochordata, Branchiostomidae) is of considerable importance for understanding the n Correspondence to: Hector Escriva, Laboratoire de Biologie Mole´culaire de la Cellule, UMR 5161 CNRS/ENS de Lyon, Ecole Normale Supe´rieure de Lyon, IFR128 BioSciences Lyon-Gerland, 46, Alle ´e d’Italie, 69364 Lyon Cedex 07, France. E-mail: hector.escriva. garcia@ens-lyon.fr Received 23 March 2004; Accepted 30 March 2004 Published online 28 June 2004 in Wiley Interscience (www. interscience.wiley.com). DOI: 10.1002/jez.b.20025 r 2004 WILEY-LISS, INC. JOURNAL OF EXPERIMENTAL ZOOLOGY (MOL DEV EVOL) 302B:384–391 (2004)