THU121 Dopamine Regulates Neuroendocrine Cell Expansion In The Developing Lung

Disclosure: T. Watanabe: None. M. Stupnikov: None. Y. Zhou: None. X. Ke: None. J. Qien: None. A. Ozawa: None. M. Horigome: None. M. Yamada: None. M. Mori: None. W.V. Cardoso: None. Pulmonary neuroendocrine cells (PNECs) are rare airway epithelial cells in the lung that have function as neurosensory and endocrine cells. They arise from naive epithelial progenitor cells in mice on embryonic day (E) 12.5, migrate to branch point of proximal airway and rare found as solitary or in clusters of 5 to 20 cells termed neuroepithelial bodies (NEBs). PNECs can be aberrantly increased in a variety of human diseases, including asthma, neuroendocrine hyperplasia of infants (NEHI) and sudden infant death syndrome (SIDS). Ascl1, an Achaete-Scute family bHLH transcriptional factor is the earliest marker and master regulator of PNEC fate specification. No PNECs form in Ascl1 null mice. In an effort to identify Ascl1 downstream targets involved in PNEC specification and expansion, we performed transcriptional analysis of E14.5 lungs from Ascl1 null mouse. Gene Ontology analysis showed a number of genes associated with the catecholamine pathway downregulated in mutants, compared to WT. These included tyrosine hydroxylase (Th) dopa decarboxylase (Ddc), dopamine beta-hydroxylase (Dβh). To gain initial insights into how catecholamine signaling influence PNEC development, we cultured E12.5 lung explants with various inhibitors of the catecholamine biosynthesis pathway and examined the effects on cell differentiation and proliferation. Treatment with Metyrosine (Met) and Benserazide (Benz) which inhibits dopamine production, resulted in a striking increase in PNEC number in NEBs as seen by expansion in Ascl1+ cells, without affecting cell proliferation. By contrast, no effect was observed with Zamicastat (Zami) which inhibits norepinephrine production. To examine whether dopamine regulated PNEC differentiation, we used the D1-like receptor antagonist, SCH-23390 and the D2-like receptor antagonist, Haloperidol in lung explant cultures, as before. Only Haloperidol increased PNECs in cultured lungs. This effect was similarly observed by maternal administration of Haloperidol intraperitoneally from E9.5 to 13.5 and analysis of harvested embryos at E14.5 and 18.5. Current studies mapping expression of other components of the dopamine pathway in the developing airway epithelium suggests a complex interaction of PNECs with neighbor cells in the regulation of the proper number of NEBs and