The murine Hoxc cluster contains five neighboring AbdB-related Hox genes that show unique spatially coordinated expression in posterior embryonic subregions.

A common feature of the murine Abdominal B (AbdB) -related Hox genes, located in the 5' regions of the four Hox clusters, appears to be a function in patterning the developing limb. As a prerequisite for studying the role of the AbdB-related Hoxc genes during limb development, we have isolated and mapped the three predicted AbdB-related Hoxc-11, -12, and -13 loci, thus defining the 5' end of the Hoxc cluster. Sequence comparisons based on the homeobox sequences of presumably all murine AbdB-related Hox genes strongly support the concept of a two step process in their evolution. As expected, Hoxc-11, -12 and -13 exhibit nested and extremely posteriorly restricted expression domains, whose anterior boundaries reflect their map positions, in accordance with the colinearity rule. A limited comparison of the primary expression domains of all five AbdB-related Hoxc genes in the developing hindlimb revealed nested and increasingly restricted domains of expression in the mesenchyme for only Hoxc-9, -10 and -11. However, separate localized expression was detected for Hoxc-9, -10, -11, -12 and possibly -13 in distal epidermal regions of the developing hind- and forelimb, whereas no expression of any of the five genes was observed in mesenchymal tissues of the developing forelimb. These data suggest a specific role for the AbdB-related Hoxc genes in patterning the hindlimb and pelvic girdle, which is separate from a second role relevant for both hind- and forelimb development.

High affinity binding and regulatory actions of dexamethasone-type I receptor complexes in mouse brain.

It is often implied that the various molecular, physiological, and behavioral responses to the glucocorticoid dexamethasone (DEX) are mediated in brain exclusively via the interactions of this synthetic steroid with the classical glucocorticoid (type II) receptor. The results reported in this study, however, suggest this generalization may, at least for the female mouse, be too restrictive. In the first experiment we compared the efficacy of the mineralocorticoid aldosterone (ALDO) with that of DEX to measure the classical mineralocorticoid (type I) receptor in brain cytosol. Since both of these steroids also bind to type II receptors, our assays included the type II receptor-selective ligand, RU26988. Whereas the specific binding of ALDO to type I receptors was largely unaffected by a 10-fold increase in the concentration of RU26988 (50- vs. 500-fold excess), there was a dramatic reduction in the specific binding of DEX. In a follow-up experiment, Scatchard analyses were used to confirm the differential affinity of RU26988 for DEX- vs. ALDO-type I receptor-binding sites and to reveal that the affinity of type I receptors for DEX (Kd approximately 0.83 nM) was nearly as high as it was for ALDO (Kd approximately 0.46 nM). A series of competition studies indicated that the competitive affinity (Kdc) of DEX for the ALDO-binding site was equivalent to the Kd computed in the saturation analyses, thus suggesting that the high affinity binding sites for DEX and ALDO on type I receptors may be equivalent or at least overlapping. The binding of DEX to these high affinity sites may prove to be important, since the systemic administration of this steroid was found to down-regulate both type I and type II receptors in a number of brain regions. Because coadministration of the type I receptor antagonist RU26752 was shown to block these actions on type I, but not type II receptors, the formation of the DEX-type I receptor complex appears to be required for DEX-induced type I receptor down-regulation. An analysis of the in vitro efficacy of ALDO- vs. DEX-type I receptor transformation suggests that whereas there is a significant increase in the binding of both complexes to DNA-cellulose after treatment with thiocyanate, there is also a dramatic decrease in the stability of DEX- but not ALDO-type I receptor binding. We contend that it is this decrease in binding stability that mediates the DEX-induced down-regulation of type I receptors.(ABSTRACT TRUNCATED AT 400 WORDS)

Aldosterone-stimulated down-regulation of both type I and type II adrenocorticosteroid receptors in mouse brain is mediated via type I receptors.

The concentrations of type I and type II adrenocorticosteroid receptors in brain cytosol obtained from adrenalectomized-ovariectomized female mice were measured with five different assay conditions. Among the five brain regions studied, hippocampus had the highest concentration of type I receptors, whereas cerebral cortex had the highest concentration of type II receptors. The value of properly correcting for dexamethasone cross-binding to type I receptors when type II receptors are being assayed was demonstrated using the type II receptor-selective ligand RU28362. A time-course study revealed a transient up-regulation of both receptor classes in most brain regions after adrenalectomy-ovariectomy, with maximal values achieved 3-5 days postsurgery and a reduction to near-intact levels by 16 days postsurgery. A single sc injection of aldosterone given to adrenalectomized-ovariectomized mice produced a profound down-regulation of type I receptors in hippocampal, cerebral cortex, hypothalamic, brain stem, and cerebellar samples, whereas it down-regulated type II receptors only in hippocampal and cerebral cortical samples. A similar injection of RU28362 failed to down-regulate type I receptors in any brain region, but it did reduce the concentration of type II receptors in all brain regions except cerebellum. The actions of aldosterone appear to be mediated solely through type I receptors, since injections of the type I receptor antagonist RU26752 prevented aldosterone-induced down-regulation of both type I and type II receptors, whereas RU26752 had no effect on the down-regulatory actions of RU28362. The ability of aldosterone to down-regulate type I, but not type II, receptors in hypothalamic, brain stem, and cerebellar samples suggests that type I and type II receptors are concentrated in separate populations of cells in these brain regions, whereas in hippocampus and cerebral cortex there is a sufficient degree of colocalization to permit type II receptor down-regulation via the action of aldosterone-type I receptor complexes. We speculate that this action is mediated at least in part at the genomic level by the suppression of type I and type II receptor mRNA synthesis brought about by the interactions of transformed aldosterone-type I receptor complexes with the DNA regulatory elements upstream from the genes for these receptors.