Developmental change in the steroid hormone signal for cell-autonomous, segment-specific programmed cell death of a motoneuron.

During metamorphosis of the hawkmoth, Manduca sexta, accessory planta retractor (APR) motoneurons undergo a segment-specific pattern of programmed cell death (PCD): e.g., APRs from abdominal segment six [APR(6)s] die at pupation in direct response to the prepupal rise in 20-hydroxyecdysone (20E), whereas APR(4)s survive through the pupal stage and die at eclosion (adult emergence). The hypothesis that the death of APR(4)s is triggered by the decline in 20E at eclosion was supported by findings that injection of 20E into developing pupae to delay the fall in 20E delayed APR(4) death. Furthermore, abdomen isolation to advance the fall in 20E caused precocious APR(4) death. In other experiments, APR(4)s were placed in primary cell culture 4 days before eclosion in medium containing 1 microg/ml 20E. A switch to hormone-free medium induced PCD in a significant proportion of APR(4)s, compared to APR(4)s that remained in 20E. Process fragmentation was a reliable early indicator of PCD. These results show that a decline in 20E triggers cell-autonomous PCD of APR(4)s, in contrast to the rise in 20E that triggers cell-autonomous PCD of APR(6)s. Thus, the PCD of homologous motoneurons in different body segments at different developmental times is triggered by different steroid hormone signals.

Profound differences in the transport of steroids by two mouse P-glycoproteins.

There are two mouse P-glycoproteins that convey multidrug resistance, mdr1 (mdr1b) and mdr3 (mdr1a), by serving as drug efflux transporters. These proteins each exhibit tissue-specific expression. There is relatively high expression of the mdr1 gene in the adrenals, the site of glucocorticoid and mineralocorticoid hormone synthesis. We previously demonstrated that mdr1 gene expression in murine thymoma cells correlated well with a decrease in their ability to accumulate the glucocorticoid dexamethasone and their increased resistance to glucocorticoid-induced apoptosis. Additional evidence is presented that supports the proposition that the mdr1 P-glycoprotein can transport glucocorticoids. Specifically, introduction and expression of the mouse mdr1 gene in the human HEK 293T cell line conveys a multidrug resistance phenotype that includes a reduced capacity to accumulate dexamethasone. Moreover, isolation of additional mdr1-expressing mouse lymphoid cells, without using steroids in the selection, confirms the linkage between multidrug resistance conveyed by the mdr1 P-glycoprotein and resistance to dexamethasone. In contrast, two newly isolated lymphoid lines, selectively expressing the mdr3 gene, were not found to have increased dexamethasone resistance or the capacity to accumulate significantly lower levels of hormone. The results support the concept that the mdr1 and mdr3 P-glycoproteins may serve alternative roles in the transport of endogenous substances such as steroids.

Estrogen receptor-dependent sexual differentiation of dopaminergic neurons in the preoptic region of the mouse.

Although it has been known for some time that estrogen exerts a profound influence on brain development a definitive demonstration of the role of the classical estrogen receptor (ERalpha) in sexual differentiation has remained elusive. In the present study we used a sexually dimorphic population of dopaminergic neurons in the anteroventral periventricular nucleus of the hypothalamus (AVPV) to test the dependence of sexual differentiation on a functional ERalpha by comparing the number of tyrosine hydroxylase (TH)-immunoreactive neurons in the AVPV of wild-type (WT) mice with that of mice in which the ERalpha had been disrupted by homologous recombination (ERKOalpha). Only a few ERalpha-immunoreactive neurons were detected in the AVPV of ERKOalpha mice, and the number of TH-immunoreactive neurons was three times that of WT mice, suggesting that disruption of the ERalpha gene feminized the number of TH-immunoreactive neurons. In contrast, the AVPV contains the same number of TH-immunoreactive neurons in testicular feminized male mice as in WT males, indicating that sexual differentiation of this population of neurons is not dependent on an intact androgen receptor. The number of TH-immunoreactive neurons in the AVPV of female ERKOalpha mice remained higher than that of WT males, but TH staining appeared to be lower than that of WT females. Thus, the sexual differentiation of dopamine neurons in the AVPV appears to be receptor specific and dependent on the perinatal steroid environment.

Hormonal regulation of CREB phosphorylation in the anteroventral periventricular nucleus.

The anteroventral periventricular nucleus (AVPV) is a nodal point in neural circuits regulating secretion of gonadotropin and contains sexually dimorphic populations of hormonally regulated dopamine-, dynorphin-, and enkephalin-containing neurons. Because the tyrosine hydroxylase (TH), prodynorphin (PDYN), and proenkephalin (PENK) genes contain cAMP response elements that control their expression in their promoters, we used histochemical methods to determine whether ovarian steroids alter expression of the cAMP response element-binding protein (CREB) in the AVPV. Because the ability of CREB to activate transcription depends on phosphorylation at Ser133, we also evaluated the effects of acute steroid treatment on levels of phosphorylated CREB (pCREB) in AVPV neurons by using an antibody that differentiates between CREB and pCREB. Treatment of ovariectomized rats with estradiol treatments caused a significant induction in the number of pCREB-immunoreactive nuclei within 30 min that was maintained for at least 4 hr, but did not alter CREB immunostaining in the AVPV. Pretreatment with the estrogen antagonist Nafoxidine blocked this induction. In contrast, acute administration of progesterone to estrogen-primed animals suppressed and then increased pCREB staining in the ASVPV at 30 and 60 min, respectively; no significant differences between experimental and control animals were apparent by 2 hr after progesterone treatment. Double-labeling experiments showed that pCREB was colocalized with PDYN, PENK, or TH mRNA in the AVPV, suggesting that pCREB may mediate the effect of steroid hormones on gene expression in these neurons.

Ovarian steroid regulation of estrogen and progesterone receptor messenger ribonucleic acid in the anteroventral periventricular nucleus of the rat.

The anteroventral periventricular nucleus of the preoptic region (AVPV) represents a key site for hormonal feedback on gonadotropin secretion. It plays a critical role in the neural control of luteinizing hormone secretion and contains high densities of neurons that express receptors for estrogen and progesterone. In this study in situ hybridization was used to examine the expression of mRNAs encoding the estrogen (ER) and progesterone (PR) receptors in the AVPV during the estrous cycle. ER gene expression fluctuated during the cycle with the lowest levels of ER mRNA observed in animals killed on the afternoon of proestrus, and the highest levels present in animals killed during metestrus. This apparent inverse relationship between circulating levels of estradiol (E2) and ER mRNA levels in AVPV neurons was supported by the observation that treatment of ovariectomized rats with E2 suppressed expression of ER mRNA in the AVPV. The influence of progesterone (P4) on ER expression was less pronounced, but a significant increase in ER mRNA in the AVPV was detected 3 h after treatment with P4. In contrast, PR mRNA levels were highest in the AVPV during diestrus and lowest on the morning of proestrus suggesting that PR expression in the AVPV is regulated in a complex manner that may reflect the combined regulatory effects of E2 and P4. E2 treatment caused a dramatic induction of PR mRNA in the AVPV, but P4 did not affect PR mRNA expression acutely, although PR mRNA appears to be attenuated in the AVPV 27 h after P4 treatment. These findings suggest that ovarian steroid hormones regulate ER and PR gene expression in the AVPV during the estrous cycle, which may represent molecular events that contribute to cyclic changes in the responsiveness of AVPV neurons to steroid hormones.

Reversal of multidrug resistance by RU 486.

P-Glycoproteins represent a family of drug efflux proteins that convey multidrug resistance to cells in which they are expressed. This phenomenon can lower the efficacy of drugs used in chemotherapy. The steroid progesterone has been shown to bind P-glycoproteins and inhibit their drug efflux. We report that the antiprogestin RU 486 can reverse multidrug resistance in cells overexpressing the mouse mdr1 gene. Using flow cytometry to measure inhibition of P-glycoprotein-dependent efflux of rhodamine 123, RU 486 was found to be considerably more effective than progesterone and one-half as effective as verapamil. The results suggest a valuable new use for RU 486.

[A patient with hypersensitive carotid sinus syndrome]. / Een patiënt met het hypersensitieve-sinus caroticus-syndroom.

We describe a patient with carotid sinus hypersensitivity that is cardiodepressive as well as vasodepressive. He was treated with AV-sequential pacing after which his complaints were reduced considerably. The various forms of carotid sinus hypersensitivity and their therapeutic consequences are discussed.