ATP signaling in the integrative neural center of Aplysia californica
ATP and its ionotropic P2X receptors are components of one of the most ancient signaling systems. However, little is known about the distribution and function of purinergic transmission in invertebrates. Here, we cloned, expressed, and pharmacologically characterized P2X receptors in the sea slug Aplysia californica - the prominent model in cellular and system neuroscience. These functional P2X receptors were successfully expressed in Xenopus oocytes and displayed activation by ATP (EC50=306 M) with two-phased kinetics as well as Na+-dependence. The ATP analog, Bz-ATP, was a less effective agonist (~20%) than ATP, and PPADS was a more potent inhibitor of the P2X receptors than the suramin. We showed that P2X receptors are uniquely expressed within Aplysias cerebral bioenergetic center (also known as F-cluster). Using RNA-seq, we found that the F-cluster contains more than a dozen unique secretory peptides, including three insulins, interleukins, and potential toxins, as well as ecdysone-type receptors and a district subset of ion channels. This structure is one of the most prominent integrative centers in the entire CNS and remarkably different from the morphologically similar neurosecretory center (bag cluster) involved in egg-laying behavior. Using RNA-seq, we also characterized the expression of P2X receptors across more than a dozen Aplysia peripheral tissues and developmental stages. We showed that P2X receptors are predominantly expressed in chemosensory structures and during early cleavage stages. The localization and pharmacology of P2X receptors in Aplysia highlight the evolutionary conservation of bioenergetic sensors and chemosensory purinergic transmission across animals. This study also provides a foundation to decipher homeostatic mechanisms in development and neuroendocrine systems. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=127 SRC="FIGDIR/small/392001v1_ufig1.gif" ALT="Figure 1"> View larger version (47K): org.highwire.dtl.DTLVardef@1ea3ab5org.highwire.dtl.DTLVardef@b45aadorg.highwire.dtl.DTLVardef@ad1851org.highwire.dtl.DTLVardef@17abbc_HPS_FORMAT_FIGEXP M_FIG We show that ATP and its ligand-gated P2X receptors are essential signaling components within both the chemosensory systems and the unique integrative neurosecretory center, present in the CNS of the sea slug Aplysia - a prominent model in neuroscience. Expression and pharmacology of P2X receptors in Aplysia confirms the preservation of evolutionary conserved bioenergetic sensors across animals and provide new tools to decipher homeostatic mechanisms in neuro-endocrine systems in general. C_FIG