Role of the calcium modulated cyclases in the development of the retinal projections Xavier Nicol, 1,2, * Mohammed Bennis, 3, * Yoshihiro Ishikawa, 4 Guy C.-K. Chan, 5 Jacques Repe ´rant, 6 Daniel R. Storm 5 and Patricia Gaspar 1,2 1 INSERM, U616, IFR Neurosciences, Ho ˆ pital Salpe ˆ trie ` re, Paris, France 2 Universite ´ Pierre et Marie Curie-Paris-06, Paris, France 3 Equipe Neurobiologie, University Cadi Ayyad, Marrakech, Morocco 4 Department of Cell Biology and Molecular, Medicine and Medicine (Cardiology), Newark, NJ, USA 5 Department of Pharmacology, University of Washington, Seattle, USA 6 UMR-5166, USM 501, MNHN, Paris, France Keywords: adenylyl cyclase, cAMP, dLGN, RGC, visual system Abstract Transmembrane isoforms of adenylate cyclases (AC) integrate a wide variety of extracellular signals from neurotransmitters to morphogens and can also regulate cAMP production in response to calcium entry. Based on observations in the barrelless mouse strain, the Adcy1 gene (AC1) was involved in the segregation of binocular retinal inputs. To determine the potential role of other AC isoforms we localized the Adcy genes in the visual centres during development, using in situ hybridization. Six different AC subtypes were found in the developing retinal ganglion cell layer (RGC; AC1, AC2, AC3, AC5, AC8, and AC9), and three AC subtypes were expressed in the central brain targets, the dorsal lateral geniculate nucleus (AC1 and AC8), the ventral lateral geniculate nucleus (AC2 and AC8) and the superior colliculus (AC1, AC2, AC8). Using a genetic approach we tested the role of the calcium modulated cyclases AC1, AC5 and AC8 for the segregation retinal fibres. Ipsilateral retinal axons remained exuberant in the AC1 – ⁄ – mice, with overlapping retinal projections from both eyes in the superior colliculus and the visual thalamus. These abnormalities were similar to those of barrelless mouse mutants. No abnormalities were detectable in the AC5 – ⁄ – or the AC8 – ⁄ – mice. Similar abnormalities were noted in the single AC1 – ⁄ – and the AC1 ⁄ AC8 double-knockout mice (DKO). Thus, only AC1 is required for the maturation of the retinal axon terminals whereas AC5 and AC8 are not needed. The specificity of AC1¢s action is linked to its cellular localization in the RGCs and to its distinctive functional profile, compared with the other cyclases expressed in the same cells. Introduction In mammals, retinal inputs from both eyes terminate into distinct cell layers in the primary visual relays, the dorsal lateral geniculate nucleus (dLGN) and the superior colliculus (SC), forming the anatomical basis of binocular vision. The development of this system has been widely used as a model to unravel mechanisms involved in developmental plasticity. Axons arising from each retina are initially intermixed and gradually separate from one another into complementary nonoverlap- ping layers. This occurs during early postnatal life in rodents (Land & Lund 1979; Godement et al. 1984). The eye-specific segregation in central targets appears to be driven by spontaneous neural activity, which exists in the retinal ganglion cells prior to the opening of the eyes and before the maturation of the photoreceptors (Maffei & Galli- Resta, 1990, Wong, 1993). Within the terminal targets, segregation involves competitive interactions among retinal afferents from both eyes (reviewed in Torborg & Feller, 2005) and it is influenced by neurotransmitters such as serotonin (Mooney et al., 1998; Upton et al., 1999), glutamate (Simon et al., 1992) and by neurotrophic factors (Menna et al., 2003). cAMP signalling is important for the segregation of eye-specific inputs (Stellwagen et al., 1999; Pham et al., 2001; Ravary et al., 2003), and could be a common downstream signal for the action of several neurotransmitters or trophic molecules. The adenylate cyclases (AC), the enzymes that produce cAMP from ATP, exist in at least ten different isoforms (nine transmembrane AC and one cytoplasmic AC), which are all expressed in the brain. Each isoform has unique regulatory properties, being modulated by G-proteins (Gas, Gai, Gbc or Gq), by Ca ++ , or by a combination of both signals (Hanoune et al., 2001). In a mouse strain that carries a spontaneous invalidating mutation of the AC1 gene, the barrelless mutation (Welker et al., 1996; Abdel-Majid et al., 1998), we previously established the role of AC1 for the normal separation of the ipsi- and contralateral retinal axons and for the refinement of the retinotopic map (Ravary et al., 2003, Nicol et al., 2006). Likewise, excessive stimulation of the 5-HT1B receptor, which is negatively coupled to adenylate cyclase via G-proteins of the Gai subtype (Lin et al., 2002), prevents the segregation of binocular inputs, possibly by modulating cAMP signals. cAMP-mediated signalling can in turn control numerous developmental processes such as gene transcrip- tion, via CREB (Pham et al., 2001), spontaneous neural activity (Stellwagen et al., 1999), synaptic plasticity (Villacres et al., 1998; Wong et al., 1999), or responses of retinal axons to guidance molecules (Song et al., 1998; Nicol et al., 2006). All these cellular Correspondence: Dr P. Gaspar, 1 INSERM, U616, as above. E-mail: gaspar@chups.jussieu.fr * X.N. and M.B. contributed equally to this work. Received 4 February 2006, revised 5 October 2006, accepted 9 October 2006 European Journal of Neuroscience, Vol. 24, pp. 3401–3414, 2006 doi:10.1111/j.1460-9568.2006.05227.x ª The Authors (2006). Journal Compilation ª Federation of European Neuroscience Societies and Blackwell Publishing Ltd