Effect of neonatal treatment with mifepristone or tamoxifen on the binding capacity of the thymic glucocorticoid or uterine estrogen receptor of adult rats: data on the mechanism of hormonal imprinting

For studying the mechanism of perinatal hormonal imprinting newborn rats were treated with a single injection of the antihormones, mifepristone (RU486) or tamoxifen (100 microg each). Glucocorticoid receptors of thymi of 6 weeks old male and female, and uterine estrogen receptors of 2 months old female rats were studied for dexamethasone or estradiol binding, respectively. Tamoxifen caused faulty imprinting both in the thymic and uterine receptors, increasing affinity and density of males, and decreasing females' glucocorticoid receptors as well, as decreasing the density of uterine estradiol receptors. Neonatal mifepristone treatment was indifferent to the thymus, and decreasing to density of uterine estrogen receptors. Males' body weight significantly decreased 6 weeks after tamoxifen treatment. The results suggest that imprinting can not be provoked by a molecule (hormone antagonist) which can bind to the receptor without any postreceptorial events (mifepristone/glucocorticoid receptor), in the presence of some postreceptorial effects the reaction takes place, however the strongest reaction can be observed by the hormone analogue (tamoxifen) with postreceptorial (agonist) effect, not considering that the receptor is the direct target of the molecule or a cross-reaction is present.     

Effect of single neonatal treatment with the soy bean phytosteroid,genistein on the sexual behavior of adult rats

Hormonal imprinting develops during the perinatal critical period, when the target hormone meets the yet unmatured receptor. As a consequence of imprinting the receptor accomplishes its maturation reaching the binding capacity characteristic to adults. In this period in the presence of foreign molecules similar to the target hormone faulty imprinting may occur with life-long consequences. Soy bean contains phytosteroids which can mimic estrogen effects. In the present experiments single genistein (20 microg) or combined genistein + benzpyrene (20 microg) treatments were done neonatally and the sexual behavior of male and female adult animals was studied. Genistein significantly increased the lordosis quotient of females, which was compensated by neonatal benzpyrene treatment. Genistein also enhanced the sexual activity of males, and this was significantly not reduced by parallel benzpyrene treatment. The results show that neonatal genistein exposure can imprint sexual activity for life and the presence of a second imprinter can modify genistein's behavioral effect.     

The environmental pollutant aromatic hydrocarbon, benzpyrene has deleterious effect on hormone receptor development

In this review, works to clear the effect of perinatal and pubertal benzpyrene imprinting to the steroid hormone receptors of adult animals are summarized. There is a comparison between the perinatal and adolescent effects and between the effects on the receptors of males and females. On the basis of the experiments benzpyrene is a general and dangerous imprinter which can influence durably the development of different steroid receptors given directly to the animals studied (perinatally) of given to the nursing mothers or influencing the offspring generations of the perinatally treated parents or grandparents. The results call attention to the outstanding deleterious effects of benzpyrene pollution.     

Prolonged impact of pubertal serotonin treatment (hormonal imprinting) on the later serotonin content of white blood cells

The first encounter between the developing receptor and its target hormone establishes the hormonal imprinting which is needed for the normal function of the cell. In the presence of foreign-however able to bind-molecules, faulty imprinting develops with lifelong consequences. Hormonal imprinting influences not only the receptors, but also the later hormone production of cells. The critical time of hormonal imprinting is the perinatal period, however it can be executed sometimes (in continuously differentiating cells) also at puberty. As in earlier experiments single neonatal serotonin treatment caused a life-long alteration of white blood serotonin content in female rats, the early (10-19 day) and late (8 weeks) effect of single pubertal serotonin treatment was studied presently, by using flow cytometry. In contrast to the earlier (neonatal) results, pubertal treatment caused a radical reduction of serotonin content in male's lymphocytes, monocytes, granulocytes and mast cells, independent on the time of study. The effect in females was rather increasing, however uncertain. The experiments call attention to the possible different effects of neonatal and pubertal hormonal imprinting and to the imprintability of blood cells in adolescence.     

Endorphin excess at weaning durably influences sexual activity, uterine estrogen receptor's binding capacity and brain serotonin level of female rats.

Perinatally, the first encounter between the maturing receptor and its target hormone results in hormonal imprinting, which adjusts the binding capacity of the receptor for life. In the presence of an excess of the target hormone or foreign molecules than can be bound by the receptor, faulty imprinting carries life-long consequences. In cytogenic organs, imprinting could also be provoked in other periods of life (late imprinting). Imprinting also durably influences the production of the imprinter and related hormones. In the present study, single beta-endorphin doses was given to three-week old female rats at 3 microg/animal, and the serotonin in five brain regions (frontal cortex, striatum, hippocampus, hypothalamus and brain stem) and uterine estrogen receptor content were determined, thymic glucocorticoid receptor binding capacity was measured, and sexual behavior was tested at five months of age. Brain serotonin levels highly significantly decreased, while sexual activity (Meyerson index and lordosis quotient) increased. At the same time, uterine estrogen receptor affinity decreased. There was no change in receptor binding capacity in the thymus. We will go on to discuss interrelations between the results. The experiments demonstrate that a non-perinatal treatment with a molecule acting at receptor level (late imprinting) can also lastingly influence various indexes in non-cytogenic organs. The results call attention to the possible long-lasting influence of an endorphin surge (caused, for example, by pain) on brain serotonin content and sexual behavior.     

Effect of single neonatal vitamin K1 treatment (imprinting) on the binding capacity of thymic glucocorticoid and uterine estrogen receptors of adolescent and adult rats.

Neonatal single treatment with vitamin K1 (50 microg/animal) significantly increased the density (Bmax) of thymic glucocorticoid receptors of the adolescent (6 weeks old) and uterine estrogen receptors of adult (10 weeks old) females. The same tendency was observed in the thymus of males and adult females, however without significance. Receptor affinity was (not significantly) influenced in the same direction. Considering that the steroid receptor imprinting effect of vitamins A and D as well as the imprinting-like effect of vitamin E was demonstrated earlier, the ability for neonatal steroid receptor imprinting of the whole lipid-soluble vitamin group is now justified.     

Effect of tamoxifen treatment at adolescent age on the sexual behaviour and steroid hormone receptor binding of adult female rats

Hormonal imprinting takes place perinatally, at the first encounter between the target hormone and its developing receptor. However, there is a secondary critical period of imprinting at puberty. In these periods molecules similar to the hormones (members of the same hormone family, antagonists, certain environmental pollutants, etc.) can cause faulty imprinting with lifelong consequences. In the present experiments 5+2 days of tamoxifen treatment (120 microg/day) at adolescent age dramatically (from approx. 40% to 10%) reduced the sexual activity (Meyerson index and lordosis quotient) of female rats, soon after the finishment of the treatment and between four to six weeks after treatment. Similar results were observed in animals neonatally treated with allylestrenol and tamoxifen treated at puberty. Thymic glucocorticoid receptor and uterine estrogen receptor binding capacity were not influenced.     

Molecules acting on receptor level at weaning, durably influence liver glucocorticoid receptors

In the experiments the effect of late hormonal imprinting to the liver glucocorticoid receptors were studied. Three-week-old (weanling) female rats were treated with five molecules acting at receptor level and four weeks later receptor kinetic analysis was done on liver glucocorticoid receptors. The tricyclic antidepressant, histamine and serotonin receptor blocker mianserin positively influenced receptor density and negatively receptor affinity. Vitamin D3 and the environmental pollutant benzpyrene elevated receptor density. Mifepristone (RU 486) which is bound by progesterone- and glucorticoid-receptor without postreceptorial effects was ineffective as well, as the H1 receptor blocker chlorpheniramine. The results demonstrate that receptor-level-acting foreign molecules can durably influence the binding capacity of glucocorticoid receptors, however, this is not a general phenomenon and it is not dependent on the type of receptors (membrane or cytosol). Those molecules were effective which 1. have receptor in the same receptor family (vitamin D3) and have postreceptorial effect, or 2. have a structure similar to steroids (benzpyrene) or 3. deeply influenced steroid receptors in earlier experiments (mianserin). This effect should be considered before administering such type of medicaments.     

The effect of perinatal hormonal imprinting with 13-cis-retinoic acid (isotretinoin) on the thymic glucocorticoid receptors of female and testosterone level of male adult rats.

In earlier experiments, the long-term effect of perinatal treatment (hormonal imprinting) with all-trans-retinol and all-trans-retinoic acid on the thymic glucocorticoid and uterine estrogen receptors was studied and was found effective. In the present experiments, the imprinting effect of four retinoids (13-cis-retinaldehyde, 13-cis-retinoic acid, 9-cis-retinaldehyde and 9-cis-retinoic acid) was investigated, using receptor kinetic analysis and sexual hormone (testosterone and progesterone) level determinations. Exclusively 13-cis-retinoic acid (isotretinoin) had an effect, significantly decreasing glucocorticoid receptor affinity and increasing serum testosterone level. Relationships with RAR-RXR receptor binding and teratogenicity is discussed.     

Impact of single neonatal serotonin treatment (hormonal imprinting) on the brain serotonin content and sexual behavior of adult rats

Hormonal imprinting takes place perinatally at the first encounter between the developing receptor and its target hormone. As a consequence of imprinting the receptor accomplishes its maturation and reaches the binding capacity characteristic to the adult age. In the excess of target hormone or presence of molecules similar to the target hormone, which are able to bind to the unmatured receptors, faulty imprinting develops with life-long consequences. At present, serotonin was given to neonatal rats and their sexual activity, brain serotonin level and steroid receptor's binding capacity was measured in adult age. Brain serotonin level was significantly reduced in male's striatum and parallel with this, male's sexual activity significantly increased. In other regions of the male brain (prefrontal cortex, hypothalamus, hippocampus) there was a statistically non-significant tendency for a decrease in serotonin level. No significant differences were detected in female brain values, and there was only slight change in female's sexual activity. There was also no change in the binding capacity of thymic glucocorticoid and uterine estrogen receptors. The experiments call attention to the possibility of perinatal imprinting by a neurotransmitter causing changes in brain neurotransmitter level for life, which is manifested in altered sexual activity.     

Binding capacity of rat liver glucocorticoid receptor in different periods after single neonatal benzpyrene treatment (imprinting)

Newborn rats of both sexes were treated (imprinted) with 20 microg of benzpyrene. Two hours, 2 days, 1, 2, 3 weeks, 1 month and 2 months after imprinting the liver glucocorticoid receptors were studied for binding of dexamethasone. Two-hour and 2-day values were not appreciable. One week after treatment the receptor's affinity was extremely low both in control and treated treated animals. Two weeks after imprinting a significant difference in density (lower) and affinity (higher) was observed between the male treated and control animals. At 3 weeks and one month the binding capacity of treated and control animals was equal however, at 2 months Bmax of males increased and that of females decreased significantly in the neonatally benzpyrene treated animals. This means that for the development of perinatal imprinting effect a long time is needed, and the effect is manifested after a period of lability.     

Effect of treatment at weaning with the serotonin antagonist mianserin on the brain serotonin and cerebrospinal fluid nocistatin level of adult female rats: a case of late imprinting

Four weeks old (weanling) female rats were treated with the tricyclic antidepressant and histamine/serotonin receptor blocker mianserin for studying its faulty hormonal imprinting effect. Measurements were done four months later. Brain serotonin levels significantly decreased in four regions (hippocampus, hypothalamus, striatum and brainstem), without any change in the cortex. Sexual activity of the treated and control rats was similar. Cerebrospinal fluid nocistatin level was one magnitude higher in the treated rats, than in the controls. The density of uterine estrogen receptors was significantly reduced, while binding capacity of glucocorticoid receptors of liver and thymus remained at control level. The results call attention to the possibility of 1. a broad spectrum imprinting at the time of weaning by a receptor level acting non-hormone molecule 2. imprinting of the brain in a non-neonatal period of life and 3. a very durable (lifelong?) effect of the late imprinting with an antidepressant.     

Hormonal imprinting: phylogeny, ontogeny, diseases and possible role in present-day human evolution

Hormonal (chemical) imprinting which was first observed (and named) by us in the seventies of the last century, is a general biological phenomenon which takes place when the developing receptor meets its target hormone for the first time. Under the effect of imprinting, receptors mature and reach their maximal binding capacity. It also influences the cells' hormone production and different functions depending on receptors and hormones. Hormonal imprinting is present already at the unicellular level causing the development of specific receptors and helping the easier recognition of useful or harmful surrounding molecules. The phenomenon is an important factor in the survival of the species, as the effect of imprinting is transmitted to the progeny cell generations. At the same time it possibly helps the selection of molecules which are suitable for acting as hormones in higher ranked animals. In mammals, hormonal imprinting takes place perinatally and determines the function of receptor-signal-transduction systems as well as hormone production for life. However, there are other critical imprinting periods for continuously developing cells. Excess of the target hormones or presence of foreign molecules which are able to bind to the receptors, provoke faulty imprinting in the critical periods with life-long morphological, biochemical, functional or behavioural consequences. As many receptor-bound foreign molecules are used as medical treatments and many such molecules are present around us and inside us as environmental pollutants, they--causing faulty imprinting--are able to predispose the (human) organism to cardiovascular, endocrine, metabolic and cancerous diseases. It seems likely that this effect is connected with disturbance of DNA methylation process in the critical periods of life. There are some signs of the transgenerational effect of faulty imprinting and this could be manifested in the evolution of humans by an epigenetic route.     

Interaction of acetochlor with estrogen receptor in the rat uterus. Acetochlor--possible endocrine modulator?

The aim of the present study was to investigate the ability of the chloroacetanilide herbicide acetochlor to interact with the endocrine system. The modulation of the binding of [3H]estradiol-17beta to protamine sulphate-precipitated uterine nuclear and cytoplasmic estrogen receptors was analysed for this purpose. Two different stages of reproductive life cycle of female rats, virgin and uniparous, were used. Our results demonstrate that acetochlor interacts in a specific manner with high-affinity limited-capacity uterine estrogen receptors. A significant difference (p < 0.001) in estrogen receptor density was observed between two control groups: uniparous rats (Bmax = 43.634 +/- 9.516) and virgin rats (Bmax = 154.375 +/- 21.462), suggesting an intrinsic difference in the nuclear estrogen receptor levels between female rats in different reproductive life cycle stages. Consequently, a different response to acetochlor treatment was noted. Exposure to acetochlor significantly (p < 0.001) increased estrogen receptor binding activity in the nuclear fraction of uniparous female rats (Bmax = 123.324 +/- 5.666) in comparison to control (Bmax = 43.634 +/- 9.516). In exposed virgin female rats, no significant difference was detected when compared to the corresponding control group. These results should prompt us to more thoroughly evaluate potential hazards of acetochlor to human and wildlife endocrine systems.     

Effect of typical and atypical antipsychotic drugs on 5-HT2 receptor density in rat cerebral cortex

The effect of acute treatment with seven atypical antipsychotic drugs and four typical antipsychotic drugs on serotonin2 (5-HT2) receptor binding sites in rat cerebral cortex was studied. Among the atypical antipsychotic drugs examined, clozapine, fluperlapine, RMI-81582 and setoperone decreased the density of 5-HT2 receptors, but ticspirone, amperozide and melperone did not. None of the drugs affected the Kd value. Among the typical antipsychotic drugs, loxapine decreased Bmax and increased the Kd of 5-HT2 receptor binding sites, whereas chlorpromazine and cis-flupenthixol had no effect. Clothiapine, a typical antipsychotic drug of the same chemical class as clozapine, decreased Bmax without increasing Kd. The downregulation of 5-HT2 receptor binding sites following a single injection of clozapine, 20 mg/kg, remained almost unchanged during the first 72 hrs and was still significantly decreased for up to 120 hrs. There was no relationship between the affinity for the downregulation of rat cortical 5-HT2 receptor binding site and 5-HT2 receptor density. Coadministration of the D1 dopamine agonist, SKF-38393, did not affect the clozapine-induced downregulation. It is suggested that rapid and prolonged downregulation of 5-HT2 receptor sites is characteristic of some but not all atypical antipsychotic drugs and is not specific to atypical antipsychotic drugs. Dibenzo-epines (clozapine, loxapine, amoxapine, chlothiapine) consistently downregulate 5-HT2 receptors in frontal cortex after acute treatment.     

Prolonged impact of five imprinters on the serotonin content of white blood cells and mast cells of weanling rats: outstanding effect of benzpyrene and chlorpheniramine

In previous experiments, treatment at weaning or adult age with endorphin, serotonin or an antihistamine (late hormonal imprinting) durably influenced the serotonin content of white blood cells and mast cells of rat. In the present experiments, five molecule (approved imprinters in other indexes) were studied for imprinting effect of immune cells, 3 weeks after a single treatment at weaning. Three steroid hormone-like molecule (vitamin D3, mifepristone and dexamethasone) were ineffective (except dexamethasone in 1/4 indexes), while benzpyrene (aromatic hydrocarbon) and chlorpheniramine (H1-receptor blocker antihistamine) were highly effective (5/6 and 4/4 respectively). The results indicate: (1) a prolonged (late imprinting) effect of a single treatment with certain molecules acting at receptor level; (2) non-generality of late imprinting, and (3) the very extensive effects of benzpyrene, which in earlier experiments was one of the strongest imprinter at receptorial and behavioral level at any periods of life studied.     

Effect of endorphin exposure at weaning on the endorphin and serotonin content of white blood cells and mast cells in adult rat

Hormonal imprinting takes place perinatally at the first encounter between the developing receptor and its target hormone, resulting in the accomplishment of normal receptor development. In the presence of an excess of target hormone or the absence of it, or an excess of related molecules which can be bound by the receptor, faulty imprinting develops with life-long consequences. In previous experiments neonatal endorphin exposure caused a decrease in endorphin and serotonin content of peritoneal mast cells of adult animals. In the present experiment 25-day-old (weaned) female rats received 2 microg endorphin, and the endorphin as well as serotonin content of adult mast cells and white blood cells was studied by flow cytometry and confocal microscopy. Peritoneal lymphocytes and blood monocytes contained significantly (p<0.01) less endorphin and peritoneal mast cells less serotonin (p<0.07, i.e. of questionable significance) than the untreated control. The results bring attention to the possibility of durable imprinting of differentiating cells later in life and to the durable (possibly life-long) effect of an endorphin excess (perhaps caused by injury) manifested in the change of endorphin and serotonin content of immune cells.     

Characterization of the human T lymphocyte adenosine receptor: comparison of normal and systemic lupus erythematosus cells

To determine whether a defect in the T cell response to adenosine exists at the level of the adenosine receptor in systemic lupus erythematosus (SLE) we measured the binding affinity and maximum binding of T cell membranes from both normal and SLE T cells by utilizing radiolabeled adenosine ligands. Normal T lymphocyte membranes possess a single class of [3H]5-N-ethylcarboxamide adenosine binding sites with a Kd of 0.61 microM, a Bmax of 23.5 pmol/mg protein, and a Hill coefficient of 0.98, which indicates the presence of noncooperative sites. In contrast, T cell membranes do not bind significant amounts of either [3H]cyclohexyladenosine or [3H]phenylisopropyladenosine. These data indicate that T lymphocyte membranes have only A2, and not A1, adenosine receptors. Similarly, T cells from both active and inactive SLE subjects also express only A2 receptors with a Kd of 0.93 microM, a Bmax of 20.4 pmol/mg protein, and a Hill coefficient of 0.85, which is consistent with the presence of noncooperative sites. There is no difference in the on-rate, affinity, or density of T cell A2 receptors from active SLE patients, inactive SLE patients, or healthy controls. We conclude that T lymphocytes from both healthy and SLE subjects express A2, but not A1, receptors. Thus, the inability of SLE T cells to respond to adenosine does not reflect a decreased density of A2 (stimulatory) receptors, diminished A2 receptor binding, or an increased affinity or number of A1 (inhibitory) adenosine receptors. These observations support the conclusion that the defect in the T cell cAMP-dependent pathway may occur at a point distal to the adenosine receptor.