Targeting of the enhanced green fluorescent protein reporter to adrenergic cells in mice.

Adrenaline and noradrenaline are important neurotransmitter hormones that mediate physiological stress responses in adult mammals, and are essential for cardiovascular function during a critical period of embryonic/fetal development. In this study, we describe a novel mouse model system for identifying and characterizing adrenergic cells. Specifically, we generated a reporter mouse strain in which a nuclear-localized enhanced green fluorescent protein gene (nEGFP) was inserted into exon 1 of the gene encoding Phenylethanolamine n-methyltransferase (Pnmt), the enzyme responsible for production of adrenaline from noradrenaline. Our analysis demonstrates that this knock-in mutation effectively marks adrenergic cells in embryonic and adult mice. We see expression of nEGFP in Pnmt-expressing cells of the adrenal medulla in adult animals. We also note that nEGFP expression recapitulates the restricted expression of Pnmt in the embryonic heart. Finally, we show that nEGFP and Pnmt expressions are each induced in parallel during the in vitro differentiation of pluripotent mouse embryonic stem cells into beating cardiomyocytes. Thus, this new mouse genetic model should be useful for the identification and functional characterization of adrenergic cells in vitro and in vivo.

Physiological and genomic consequences of adrenergic deficiency during embryonic/fetal development in mice: impact on retinoic acid metabolism.

Adrenergic hormones are essential for early heart development. To gain insight into understanding how these hormones influence heart development, we evaluated genomic expression changes in embryonic hearts from adrenergic-deficient and wild-type control mice. To perform this study, we used a mouse model with targeted disruption of the Dopamine ß-hydroxylase (Dbh) gene, whose product is responsible for enzymatic conversion of dopamine into norepinephrine. Embryos homozygous for the null allele (Dbh(-/-)) die from heart failure beginning as early as embryonic day 10.5 (E10.5). To assess underlying causes of heart failure, we isolated hearts from Dbh(-/-) and Dbh(+/+) embryos prior to manifestation of the phenotype and examined gene expression changes using genomic Affymetrix 430A 2.0 arrays, which enabled simultaneous evaluation of >22,000 genes. We found that only 22 expressed genes showed a significant twofold or greater change, representing ~0.1% of the total genes analyzed. More than half of these genes are associated with either metabolism (31%) or signal transduction (22%). Remarkably, several of the altered genes encode for proteins that are directly involved in retinoic acid (RA) biosynthesis and transport. Subsequent evaluation showed that RA concentrations were significantly elevated by an average of ~3-fold in adrenergic-deficient (Dbh(-/-)) embryos compared with controls, thereby suggesting that RA may be an important downstream mediator of adrenergic action during embryonic heart development.