Ligand-dependent stimulation of introduced mammalian brain receptors alters spicule symmetry and other morphogenetic events in sea urchin embryos

Abstract

Zygotes of S. purpuratus were injected with synthetic mRNAs encoding rodent brain neurotransmitter receptors, and specific developmental phenotypes were produced on addition to the sea water of the respective ligands. Most of these experiments were carried out with a mouse serotonin receptor (5HT-R) mRNA, though exactly comparable results were obtained with a rat muscarinic acetylcholine receptor (MAChR) mRNA; these receptors are expected to couple to the same endogenous signal transduction system. We show by whole mount in situ hybridization that the injected mRNAs diffuse to all of the early blastomeres, and that they are translated in vivo. Three specific phenotypes were reproducibly observed. The most severe, occurring at highest levels of injected mRNA, was a cleavage arrest phenotype in which no overtly differentiated cells ever appear, though the embryos remain alive for at least 72 h. A gastrular arrest (GA) phenotype is generated in appreciable fractions of embryos developing from eggs injected with lower levels of mRNA. In GA embryos the blastocoel is filled with disorganized mesenchyme cells, including pigment cells and skeletogenic cells; there is no archenteron; and the entire ectoderm expresses an oral ectoderm cell surface marker. The least severe phenotype that we recognized displays an altered arrangement of spiculogenic foci (RSE phenotype), generating a ring of extra spicules that are properly positioned with respect to the animal/vegetal axis, but that lack any reference to the oral/aboral axis. However, use of cytological and molecular markers demonstrates that RSE embryos retain normal spatial patterns of aboral and oral ectoderm. They develop a fully formed archenteron, but fail to form either a stomodaeum or a ciliated band. RSE embryos can be produced in embryos expressing the 5HT-R by exposure to serotonin, beginning as late as 12 h postfertilization (pf). All of the morphogenetic processes affected in RSE embryos depend in normal embryos on intercellular interactions occurring at the blastula-gastrula stages of development.

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