The effect of various ions on uptake2 of catecholamines

1. The effects of a decrease of the K gradient on the extraneuronal inward transport and outward movement of catecholamines were studied in rat heart, rabbit aortic rings and guinea-pig trachealis smooth muscle. Elevation of the extracellular K concentration caused a) inhibition of the corticosteroid-sensitive extraneuronal uptake (uptake) ofH-isoprenaline in rat heart and ofH-noradrenaline in rabbit aorta, and b) acceleration of efflux ofH-isoprenaline from rat heart,H-noradrenaline from rabbit aorta and adrenaline (measured by microphotometry) from guinea-pig trachealis muscle. 2. In rat heart and rabbit aorta, the acute omission of one or the other of the ions Na, Cl, K or Ca from the perfusion or incubation medium had no effect on initial rates of uptake of catecholamines, except that the absence of K had a small inhibitory effect in the rat heart. 3. The prolonged absence of Na, Ca or K from the perfusion or incubation medium caused a marked inhibition of uptake of catecholamines. These inhibitory effects developed more quickly in rat heart than in rabbit aorta. 4. These results are compatible with the possibility that either the K gradient across the cell membrane or the resting membrane potential is the force driving uptake

Enhancer trap integrations in mouse embryonic stem cells give rise to staining patterns in chimaeric embryos with a high frequency and detect endogenous genes.

We have generated mouse embryonic stem cell lines that carry lacZ enhancer trap constructs integrated in their genome. Fifty-nine cell lines were analysed for lacZ expression in undifferentiated stem cells and at day 7.5, 8.5 and 12.5 of development in chimaeric embryos obtained after blastocyst injection. In 13 cell lines the lacZ reporter gene was expressed in undifferentiated stem cells (\u27blue\u27, lines) as monitored by beta-galactosidase activity; 46 cell lines did not show detectable beta-galactosidase activity (\u27white\u27, lines). In chimaeric embryos one-third of the analysed 59 embryonic stem cell lines gave rise to a variety of patterns. Six out of the 13 \u27blue\u27 lines and 14 out of the 46 \u27white\u27 lines showed spatially and temporally regulated patterns of beta-galactosidase expression and were additionally analysed on day 9.5. The majority of patterns showed staining exclusively or predominantly in structures of the developing nervous system, three patterns were observed only or predominantly in non-neuronal structures and five patterns were found exclusively in extraembryonic tissues. The analysis of DNA from cell lines that gave rise to staining patterns in chimaeric embryos showed that in 11 out of 15 cases simple integrations had occurred at a single site while in the remaining four cell lines multiple copies had integrated either at a single or at multiple sites. Flanking sequences from five reporter gene integrations have been cloned. At present, three integration sites have been analysed further and in all three cases we have identified transcribed sequences in the flanking DNA and isolated corresponding cDNA clones. The expression patterns of two of these genes were analysed by RNA in situ hybridisation. In both cases, expression of the endogenous genes was more widespread than the corresponding beta-galactosidase staining, suggesting that the reporter gene responded to only a subset of the regulatory elements of the endogenous gene. Our results demonstrate that enhancer trap integrations in embryonic stem cells can be used to efficiently identify transcriptional activation patterns during mouse embryogenesis and to isolate endogenous genes expressed in spatially and temporally regulated patterns

The mouse Enhancer trap locus 1 (Etl-1): a novel mammalian gene related to Drosophila and yeast transcriptional regulator genes.

A novel mouse gene, Enhancer trap locus 1 (Etl-1), was identified in close proximity to a lacZ enhancer trap integration in the mouse genome showing a specific beta-galactosidase staining pattern during development. In situ analysis revealed a widespread but not ubiquitous expression of Etl-1 throughout development with particularly high levels in the central nervous system and epithelial cells. The amino acid sequence of the Etl-1 protein deduced from the cDNA shows strong similarity, over a stretch of 500 amino acids, to the Drosophila brahma protein involved in the regulation of homeotic genes and to the yeast transcriptional activator protein SNF2/SWI2 as well as to the RAD54 protein and the recently described helicase-related yeast proteins STH1 and MOT1. Etl-1 is the first mammalian member of this group of proteins that are implicated in gene regulation and/or influencing chromatin structure. The homology to the regulatory proteins SNF2/SWI2 and brahma and the expression pattern during embryogenesis suggest that Etl-1 protein might be involved in gene regulating pathways during mouse development