Effects of an organophosphate pesticide, quinalphos, on the hypothalamo-pituitary-gonadal axis in adult male rats

The effects of chronic sub-lethal doses (7-14 mg kg-1 a day for 15 days) of quinalphos were evaluated in adult male rats for changes in testicular morphology, circulatory concentrations of hormones (LH, FSH, prolactin and testosterone), activities of acetylcholinesterase (AChE) and angiotensin converting enzyme (ACE) as well as metabolism of biogenic amines (dopamine, noradrenaline and 5-hydroxytryptamine (5-HT)) in the hypothalamus and pituitary. Hormones were assayed by radioimmunoassay or chemiluminescent immunoassay (testosterone). The enzymes were estimated after spectrophotometry and the biogenic amines by HPLC-electrochemistry. Sub-lethal chronic administration of quinalphos resulted in: decreased testicular mass and AChE activity in central as well as peripheral organs; increased serum LH, FSH, prolactin and testosterone concentrations; decreased pituitary or increased testicular ACE activity; severe disruption of spermatogenesis with increasing doses of pesticide; and no significant effects on dopamine, noradrenaline or 5-HT concentrations in the hypothalamus or pituitary. Administration of oestradiol (50 micrograms per rat a day) during pesticide treatment resulted in: a significant decrease in the mass of the testis and accessory sex organs; decreases in serum LH, FSH, testosterone concentrations; an increase in prolactin concentration; and a decrease in dopamine or an increase in noradrenaline and 5-HT in the hypothalamus or pituitary. Oestradiol had a marked effect: in pesticide-treated animals, the pesticide effects were significantly reversed. This indicates that in pesticide toxicity, the hypothalamo-pituitary-gonadal axis is operational. Since many of the observed pesticide effects could be inhibited by oestradiol, it is suggested that the pesticide acts directly on the gonadotrophins. In conclusion, quinalphos decreases fertility in adult male rats by affecting the pituitary gonadotrophins.     

Tyrosine content, influx and accumulation rate, and catecholamine biosynthesis measured in vivo, in the central nervous system and in peripheral organs of the young rat. Influence of neonatal hypo- and hyperthyroidism

The influence of neonatal hypo- and hyperthyroidism on different aspects of tyrosine metabolism in the hypothalamus, striatum, brainstem, adrenal glands, heart and brown adipose tissue (BAT) were studied in 14-day old rats. The synthesis rate of catecholamines (CA) was also determined in vivo after the injection of labelled tyrosine. Hypothyroidism increases tyrosinaemia and endogenous tyrosine concentration in the hypothalamus and BAT. Hyperthyroidism decreases tyrosinaemia and endogenous tyrosine levels in the striatum, adrenals and heart. The accumulation rate of tyrosine determined 30 min after an intravenous injection of the labelled amino acid has been determined in the organs, together with the influx of the amino acid, determined within 20s. Hypothyroidism increases tyrosine accumulation rate in all the organs studied, and tyrosine clearance is decreased in the striatum and brainstem; together with an increased tyrosinaemia, this leads to a normal influx. The influx of tyrosine is increased in the hypothalamus. Hyperthyroidism decreases tyrosine accumulation rate in all the organs except the adrenals. These results indicate that the thyroid status of the young rat can influence tyrosine uptake mechanisms, without modifying an organ's tyrosine content. The fact that hypothyroidism increases tyrosine influx in the hypothalamus without modifying it in the brainstem and striatum reflects an heterogeneous reactivity to the lack of thyroid hormones in different brain structures. Neonatal hypothyroidism decreases the CA synthesis rate in the striatum, the heart and the interscapular brown adipose tissue, while synthesis was enhanced in the brainstem and the adrenals. It is likely that these variations in CA synthesis are due to thyroid hormone modulation of tyrosine hydroxylase activity, the enzyme which catalyses the rate limiting step in CA biosynthesis.     

Postnatal ontogeny of uridine kinase in the cerebellum,hypothalamus, and cerebral cortex of the rat

Postnatal developmental patterns of uridine kinase were determined in crude subcellular fractions of the rat cerebellum, hypothalamus and cerebral cortex at ages 3 through 60 days. The highest specific activity and predominant distribution of enzyme was in the 105,000g supernatant of the 3 brain regions. Enzyme activity in hypothalamus and cerebral cortex was maximum at 3 days and decreased with age; in cerebellum it increased through 13 days and decreased thereafter. Thus, the pattern of activity in hypothalamus and cerebral cortex paralleled changes in DNA and RNA synthesis through age 60 days; in cerebellum, it more closely approximated changes in DNA synthesis during early development. Changes inK _m with aging suggest that the brain regions contain more than one form of enzyme. The highest particulate activity was in the microsomal fraction of the cerebellum and hypothalamus at all ages and in the cortex at 35 and 60 days. Relative specific activity for microsomal fractions of the brain regions at 60 days indicate a concentration of the enzyme which may be relevant in the maintenance of RNA activity in adult brain.     

Gonadal steroids modulated hypocretin/orexin type-1 receptor expression in a brain region, sex and daytime specific manner

Orexins A and B (hypocretins A and B) are regulatory peptides that control a variety of neuroendocrine and autonomic functions including feeding and sleep-wakefulness. Previously, we described a clear relationship between the hormonal milieu of the estrous cycle and the mRNA expression of the components of the orexinergic system, in the hypothalamus, pituitary and ovary. Here, we investigate whether steroid hormones are involved in the modulation of the hypocretin/orexin type-1 receptor expression at the protein level, and its time of the day dependence, in hypothalamus and pituitary of castrated male and female rats and castrated receiving hormone replacement.Orchidectomy decreased the hypocretin/orexin type-1 receptor expression in anterior hypothalamus, but not in mediobasal hypothalamus or cortex; in pituitary this treatment resulted in an increase. Testosterone and dihydrotestosterone were able to restore receptor expression and gonadotropins.In females, pituitary and ovarian hormones increased during proestrous afternoon. Hypocretin/orexin type-1 receptor expression was higher at 19:00 of proestrus in hypothalamus and pituitary. Ovariectomized treated with estradiol or oil and sacrificed at 11:00 h showed the receptor expression similar to 11:00 h of proestrus in hypothalamus and pituitary. At 19:00 h, low expression persisted in these areas in oil-treated ovariectomized rats; in contrast, estradiol replacement increased the expression to high levels of normal cycling rats at 19:00 h.Sexual steroids modulate the orexinergic system and the anatomical regions, hormones and times of the day all have to be considered when the roles of orexins, and probably other peptides, are under consideration.     

Effects of fasting on the distribution of cytoplasmic and nuclear oestrogen receptors in rat liver, uterus, pituitary and hypothalamus before and after exogenous oestrogen administration

The accumulation of oestrogen receptors in the liver cell nuclei of intact female rats 45 min after administration of 100 micrograms 17 alpha-ethynyloestradiol-17 beta i.p., decreased progressively during a 72-h fast from 2550 +/- 860 to 257 +/- 67 fmol/mg DNA, a level not significantly different from that in uninjected animals. Cytoplasmic oestrogen receptor concentrations also decreased, but only to about 60% of the original level (from 84.1 +/- 27.5 to 50.3 +/- 2.09 fmol/mg protein during the fast). Similar differences were found when these parameters were examined in normally fed and 72-h-fasted ovariectomized rats. On the other hand these parameters were unaffected in uterus, pituitary and hypothalamus. Uterine cytoplasmic receptor concentrations remained at about 500 fmol/mg protein during the fasting period, those in the pituitary and hypothalamus at about 230 and 30 fmol/mg protein, respectively. Nor was in vivo translocation in these organs affected by fasting. Regardless of nutritional status, the nuclear oestrogen receptor concentrations in uterus rose from about 500 to 2000 fmol/mg DNA after ethynyloestradiol administration, those in the pituitary and hypothalamus from approximately 250 to 2000 and from 250 to 500 fmol/mg DNA respectively.     

Stress-induced changes in brain Met-enkephalin, Leu-enkephalin and dynorphin concentrations.

Methionine-enkephalin (Met-enkephalin), leucine-enkephalin (Leu-enkephalin) and dynorphin A (1-17) (dynorphin A) concentrations in discrete brain areas were determined in the mice showing behavioral changes induced by stress using radioimmunoassay (RIA). In the present experiment, we used environment-induced conditioned suppression of motility and forced swimming-induced immobility. In the environment-induced conditioned suppression of motility, Met-enkephalin concentration in the striatum and hypothalamus significantly decreased. Leu-enkephalin concentration in the hypothalamus also decreased. Dynorphin A concentration in the striatum decreased, but significantly increased in the hypothalamus and pituitary. In the forced swimming-induced immobility, Met-enkephalin concentration in the striatum significantly decreased. Leu-enkephalin concentration in the hypothalamus and pituitary significantly decreased. Dynorphin A concentration in the pituitary decreased, but significantly increased in the hypothalamus. Our results indicated that the concentrations of Met-enkephalin, Leu-enkephalin and dynorphin A in the discrete brain areas changed in two different stressful situations. These findings suggested that these peptides might modulate the behavioral changes induced by stressors.     

Retention of radioactive substances in the hypothalamus, anterior pituitary, and reproductive organs of male rats after 3H-melatonin administration

Radioactive concentrations were determined in serum, lung, hypothalamus, anterior pituitary, testis, and accessory sex organs of adult male rats at 2, 20, 30 and 60 min after intravenous 3H-melatonin administrations. The retention patterns of 3H-melatonin and other radioactive substance (3H-non-melatonin) in these tissues were compared. The anterior pituitary demonstrated best tissue retention with highest concentration of 3H-melatonin. The testis and prostate gland accumulated 3H-non-melatonin in an increasing manner from 2 to 60 min. The radioactive substances were also preferentially and progressively located in the nuclear fraction of the anterior pituitary, hypothalamus, testis, and prostate gland. This study leads to the following suggestions: the anterior pituitary is another target organ of melatonin; melatonin is converted into other active material which exerts its action in the testis and prostate gland; melatonin and its active derivative exert their action through the nuclei of their respective cells.     

Effects of subchronic alternating cadmium exposure on dopamine turnover and plasma levels of prolactin, GH and ACTH

This study was undertaken to analyze if the effects of subchronic alternating cadmium exposure on pituitary hormone secretion are mediated by changes in dopamine turnover in an age dependent way or are directly correlated to cadmium accumulation at the hypothalamic-pituitary axis. Male rats were treated sc. from day 30 to 60 (prepubertal period) or from day 60 to 90 (adult age) of life, with cadmium chloride (CdCl₂) at a dose of 0.5 and 1.0 mg kg^–1 bw, every 4th day in an alternate schedule, starting with the smaller dose. Dopamine (DA) turnover, expressed as the ratio of acid 3,3-dihidroxifenil acetic (DOPAC)/DA in various hypothalamic areas, the plasma levels of prolactin, growth hormone (GH) and adrenocorticotropic hormone (ACTH), and cadmium accumulation in the hypothalamus and pituitary were studied. Prepubertal cadmium exposure decreased DA content in all hypothalamic areas studied, although its turnover was not modified. A decrease in plasma ACTH levels with no changes in plasma prolactin and GH levels were found. Cadmium did not accumulate in pituitary while it increased in the hypothalamus. Metal exposure during adulthood decreased DA content in mediobasal and posterior hypothalamus, and its turnover in posterior hypothalamus and median eminence. It decreased plasma prolactin and ACTH levels but not those of GH. Cadmium concentration increased in both hypothalamus and pituitary. These results suggest that cadmium exposure produces age dependent changes on the secretory mechanisms of the pituitary hormones studied, related to the selective accumulation of the metal at both hypothalamic and hypophyseal level changes. However the effects of the metal are not mediated by dopamine.     

Possible role of vasoactive intestinal peptide in the hyperprolactinemia induced by ethanol

The effect of the blockade of endogenous VIP by injecting a specific rabbit anti-VIP serum (A-VIP) was studied in rats receiving an acute injection of ethanol. A-VIP administration decreased serum prolactin levels and reduced the hyperprolactinemia induced by ethanol. We also investigated the effect of the acute administration of ethanol on the concentration and release of VIP from the mediobasal hypothalamus. Ethanol decreased VIP concentration in the mediobasal hypothalamus, whereas it stimulated the in vitro K(+)-evoked release of VIP from this tissue. Conversely, ethanol increased VIP concentration in the anterior pituitary gland. The data indicate that VIP may be involved in the pituitary response to ethanol. The increased anterior pituitary VIP after ethanol may be due to an augmented release from the mediobasal hypothalamus.     

Calcium thresholds in the activation of DNA and RNA synthesis in cultured planarian cells: relationship with hormonal and DB cAMP effects

DNA and RNA syntheses were reduced to a basal level in dissociated planarian cells grown for 48 h in a Ca2+-free medium. These syntheses could be triggered anew by raising the Ca2+ concentration in the medium. Serotonin could be substituted for Ca2+ in stimulating DNA synthesis by Ca2+-depleted cells, while dopamine greatly enhanced RNA synthesis in these cells. When Ca2+ concentration was raised in hormone-treated cultures, DNA synthesis was again slightly increased but RNA synthesis was depressed. Both hormonal effects were completely inhibited by the anticalmodulin drug trifluoperazine. As serotonin and dopamine are both known to stimulate the adenylate cyclase system, it was further investigated whether the hormonal effects were mediated by cAMP. Indeed, a DB cAMP concentration of 1 microM increased DNA labelling when applied for 8 h to Ca2+-depleted cultures. However, when Ca2+ was present, the 8-h treatment with 1 microM DB cAMP was inhibiting. A 4-h pulse with 1 microM DB cAMP just after Ca2+ addition was a condition for a high stimulation of DNA labelling. The other DB cAMP concentrations used, 0.1 and 10 microM, reduced DNA labelling. In the absence of Ca2+, RNA labelling was only slightly increased by 0.1 microM DB cAMP, but was highly stimulated by a 4-h treatment of 1 microM DB cAMP in the presence of Ca2+. The noted effects with 1 or 0.1 microM DB cAMP on DNA or RNA labelling corresponded to true changes in synthesis rather than alterations of the specific activity of the nucleotide pool by DB cAMP. Besides, it was precluded that these effects were due to butyrate issued from DB cAMP degradation. It was further shown that DB cAMP at 1 microM increased Ca2+ uptake in planarian cells, whereas the other concentration reduced it. This observation might explain the stimulating effect on nucleic acid synthesis of 1 microM DB cAMP applied at the appropriate moment. Based on these results it seems that, for triggering RNA synthesis, the threshold value of Ca2+ was lower than for DNA synthesis. These Ca2+ thresholds might be reached, in the absence of Ca2+ in the medium, by treatments with DB cAMP or hormones at the appropriate doses and periods. This interpretation is in agreement with the succession of biochemical events described in regenerating planarians and suggests that these events might be causally related.     

Effect of testosterone and progesterone on carboxypeptidase H activity in the hypothalamo-hypophyseal-gonadal axis and the adrenal gland in mice

Testosterone propionate decreased activity of carboxypeptidase H in the male mongrel mice pituitary gland, activity of the CP H in testicles, and increased these parameters in 4 hours after treatment. Progesterone decreased the CP H activity in pituitary gland and increased it in testicles. Progesterone increased the CP H activity in female pituitary gland and decreased it in adrenal gland and hypothalamus. The role of the CP H in the effects of the sex steroids on the function of peptidergic systems is discussed.     

Pituitary luteinizing hormone level in the male house musk shrew, Suncus murinus L., following castration and testosterone treatment

The effect of castration and replacement therapy with testosterone propionate (TP) on the pituitary LH concentration and contents in the house musk shrew was investigated by using an in vitro bioassay for LH, the Rat Interstitial Cell Testosterone assay. The concentration and contents of LH increased slightly 10 days after castration, but decreased progressively thereafter to about a half of the pre-operation level by 90 days after the operation. The replacement with TP (100 micrograms/day) for 7 days significantly depressed LH contents when it was begun 10 days after castration, while the same treatment started immediately after or 30 days after the operation did not significantly affect the pituitary LH level. The feedback mechanism between the gonad and the pituitary may be slightly different in the shrew from that in other mammals. TP replacement, started immediately after castration, completely inhibited the decrease in the weight of male accessory sex organs in castrated shrews. In castrated animals when more than 10 days had elapsed after the operation, however, the decreased weight of the organs could not be fully restored by the TP replacement for 7 days.     

The effect of luteinizing hormone releasing hormone (LH-RH) and oestrogen on RNA synthesis in anterior pituitary and different brain regions of rats.

Oestradiol-17beta caused a marked reduction of RNA content in the cortex, hippocampus, brain stem, hypothalamus and RNA synthesis in the pituitary. LH-RH had facilitatory effect on the cortical and inhibitory influence on the hypothalamic RNA synthesis in vitro, and suppressed the pituitary RNA synthesis both in vivo and in vitro. The possible regulatory role of the LH-RH in the nucleic acid metabolism in discussed.     

Effects of lipopolysaccharide on neurokinin A content and release in the hypothalamic-pituitary axis

The administration of bacterial lipopolysaccharide (LPS) markedly affects pituitary secretion, and its effects are probably mediated by cytokines produced by immune cells or by the hypothalamo-pituitary axis itself. Since neurokinin A (NKA) plays a role in inflammatory responses and is involved in the control of prolactin secretion, we examined the in vivo effect of LPS on the concentration of NKA in hypothalamus and pituitary (assessed by RIA) and serum prolactin levels in male rats. One hour after the intraperitoneal administration of LPS (250 microg/rat), NKA content was decreased in the posterior pituitary but not in the hypothalamus or anterior pituitary. Three hours after injection, LPS decreased NKA concentration in the hypothalamus and anterior and posterior pituitary. In all the conditions tested, LPS significantly decreased serum prolactin. We also examined the in vitro effects of LPS (10 microg/ml), interleukin-6 (IL-6, 10 ng/ml) and tumor necrosis factor alpha (TNF-alpha, 50 ng/ml) on hypothalamic NKA release. Interleukin-6 increased NKA release without modifying hypothalamic NKA concentration, whereas neither LPS nor TNF-alpha affected them. Our results suggest that IL-6 may be involved in the increase of hypothalamic NKA release induced by LPS. NKA could participate in neuroendocrine responses to endotoxin challenge.     

Estradiol benzoate induced changes in protein synthesis and lysosomal hydrolases in the pituitary of male rats

In vitro protein synthesis, lysosomal hydrolases activity and peroxidase activity in the anterior pituitary were estimated in adult male rats treated with 50 micrograms of estradiol benzoate (EB) for 1 day or 7 days. Pituitary protein synthesis, protein and RNA content increased after 7 days. A significant increase in total and membrane-bound acid phosphatase was noted after 1 day or 7 days of EB treatment whereas total beta-glucuronidase activity decreased in both 1 and 7 day group. Cathepsin activity increased after 7 days and pituitary peroxidase system did not change by EB treatment. These findings suggest that immediate change in the enzyme milieu may be one of the first reactions by which EB expresses its feedback control.     

Pituitary concentration of GH messenger ribonucleic acid in rats with anterolateral hypothalamic deafferentation

To study neuronal mechanism through which the hypothalamus exerts its influence on growth hormone (GH) synthesis, anterolateral hypothalamic knife cuts (ALHD) were used. After ALHD, GH messenger RNA (mRNA) content in the pituitary was reduced to 47% of the control value. Neither the prolactin mRNA level nor the total DNA content showed any significant change. In such animals, the serum GH level was significantly higher (184%) and pituitary GH content was lower (43%) than the control values. These results suggest that neuronal factor(s) outside the mediobasal hypothalamus plays an important role in the regulation of GH synthesis.     

Galanin-like immunoreactivity is increased in the brain of estradiol- and methyltestosterone-treated eels.

A galanin-like peptidergic system was demonstrated in the brain of Anguilla. A group of immunoreactive perikarya was located in the nucleus preopticus periventricularis close to the recessus preopticus. Galaninergic fibers occurred in various brain areas. Galanin identified in mammalian pituitary cells was undetectable in fish adenohypophysial cells. Estradiol increased the immunostaining of the rostral perikarya and brain fibers in both male and female European eels kept in fresh water and in female American eels in sea water. Methyltestosterone, an aromatizable androgen, increased galanin immunoreactivity in rostral perikarya and brain fibers of male European eels and female American eels. The cross-sectional area of these perikarya increased significantly after both treatments whereas cell bodies of the posteroventral hypothalamus were slightly affected. Dihydrotestosterone showed no clear effect. Fibers close to the corticotropes were sometime increased, but galanin synthesis was not induced in pituitary cells. In contrast, estradiol induced galanin synthesis in rat pituitary cells, but had a still controversed effect on hypothalamic galanin. A putative influence of galanin on the pituitary-gonadal axis is discussed as gonadal hormones diversely affect gonadotropes and gonosomatic indices in Anguilla.     

Effect of dopaminergic drugs on hypothalamic and pituitary immunoreactive beta-endorphin concentrations in the rat

Beta-endorphin concentrations have been evaluated in the hypothalamus, pituitary lobes and plasma after 1-and 3-week treatment with 2-Br-alpha-ergocriptine or lisuride, two potent dopaminergic drugs. Hypothalamic beta-endorphin concentrations were significantly decreased after the administration of the dopaminergic agents for 1 or 3 weeks. Similarly, beta-endorphin concentrations decreased in the neurointermediate lobe and plasma. After gel chromatography, it appeared that in the anterior pituitary, beta-lipotropin concentrations were unchanged or lightly increased concomitantly with a decrease of beta-endorphin. Our data indicate that, both in the hypothalamus and the neurointermediate pituitary lobe, beta-endorphin is under an inhibitory dopaminergic tone. The latter may also play a role in inhibiting beta-endorphin cleavage from beta-lipotropin in the anterior pituitary.     

Effects of cycloheximide on protein, RNA and DNA synthesis in cultures of CHO cells and human diploid fibroblasts]

In CHO cell line and primary human diploid fibroblasts culture an incorporation of protein, RNA and DNA biosyntheses precursors was investigated under different conditions of inhibition of translation by cycloheximide (CHM). Both CHO and human fibroblasts transitory treatment by CHM in the serumfree medium resulted in inhibition of protein and DNA syntheses during S-period while RNA synthesis increased up to 130% (CHM concentration from 0.003 to 2 Mg/ml), as well as in Go--an incorporation of 3H-U increased to 200% (CHM concentration-100 Mg/ml). Long-term treatment (48 hours) in the serum-free medium resulted in decreased uptake of 3H-T and 3H-L during first 6 hours of experiment, while incorporation of 3H-U increased to 160%. By 16-th hour of treatment characters of protein, RNA and DNA syntheses came back to control levels.