Characterization of a prolactin binding protein in rat serum.

To determine the presence of a PRL-binding protein (PRL-BP) in rat serum, female rats were ovariectomized and administered sc estradiol benzoate capsules. Serum was incubated with [125I]rat PRL (rPRL) for 1 h at 37 C with or without different doses of rPRL, ovine PRL (oPRL), or rGH. Separation of the [125I]rPRL-PRL-BP complex from unbound [125I]rPRL was accomplished by Sephadex G100 chromatography or by precipitation with an antibody against rat liver PRL receptor. Results showed that a protein was able to bind specifically to rPRL or oPRL, but not to rGH. Scatchard plots gave an affinity constant of 1.18 +/- 0.7 10(9) M-1 and a capacity of 11.24 +/- 1.4 nM. The complex [125I]rPRL-PRL-BP migrated in the void volume of sephadex G100 column, but with an apparent mol wt of 80K after cross-linking and electrophoresis under reducing conditions. PRL BP was purified from estradiol-treated ovariectomized females by an oPRL sepharose 4B affinity column. Purified protein migrated under reducing conditions with apparent mol wt of 50K and 27K but of 160K under nonreducing conditions. After transfer on nitrocellulose filter, the 160K, 50K and 27K forms were able to bind to monoclonal antibodies directed against rat liver PRL receptor. The 160K and 50K were still able to bind to [125I]oPRL, but not the 27K. These results showed that rat serum contained a PRL-BP, which presented a strong homology to PRL receptor, at least for the immunological and binding characteristics.

Abscissic acid localization by light microscopic immunohistochemistry in Chenopodium polyspermum L. Effect of water stress.

An indirect immunohistochemical technique was developed using a rabbit anti-abscissic acid (ABA) serum and the soluble peroxidase-antiperoxidase (PAP) complex for the localization of endogenous ABA in the aerial parts of Chenopodium. Terminal bud, axillary bud bearing nodes, and adult leaves were prefixed by a soluble carbodiimide to obtain the coupling of ABA on cellular proteins and postfixed by a conventional mixture of aldehydes. They were then embedded in paraffin or in plastic. Numerous controls were carried out on sections and on a model system to test the validity of the technique. Based on the staining patterns observed along the plant, an apico-basal gradient of ABA was revealed. In the older buds, ABA was mainly concentrated in the quiescent meristematic cells of the apex. Phloem cells of the main axis and chloroplasts of the leaves were specifically labeled. No reaction product was visualized in the parenchyma cells or in the cambial zone. Water stress, which is known to increase ABA content, induced an increase of immunoreactivity within the same compartments. This physiological test validates the stain.

Ultrastructural immunolocalization of elastic fibers in rat blood vessels using the protein A-gold technique.

Identification of elastic fibers at the ultrastructural level is accomplished by a post-embedding immunohistochemical technique using the protein A-colloidal gold method. Antisera against elastins from human dermis and rat aorta have been characterized by radioimmunoassay and then applied to thin sections of rat blood vessels. Two fixative solutions and two embedding media have been tested. Both antibodies bind to elastic fibers of normal arteries and veins, indicating crossreactions among organs and species. The high sensitivity of this method is demonstrated by its application to the detection of neo-elastogenesis in the intimal thickening of aortic grafts.

Increased dopaminergic inhibition of prolactin in the hypoprolactinaemic IPL nude rat.

In this study, aimed at investigating whether dopaminergic regulation of prolactin could be implicated in the hypoprolactinaemia observed in the IPL nude rat, dopaminergic inhibition of prolactin was suppressed using a catecholamine synthesis inhibitor alpha-methyltyrosine (MT) and a dopaminergic antagonist sulpiride. Adult male rats (IPL nude and normal) were injected through implanted atrial cannulae with either MT (250 mg/kg) or physiological saline (control). Rats were decapitated 2 h after the injection. Plasma prolactin levels, compared with basal values, increased by 15.6 +/- 1.9 (S.E.M.)- and 5.89 +/- 0.6-fold in IPL nude and normal rats respectively. This difference was highly significant. The pituitary prolactin content was decreased in both groups. In a second experiment, adult male IPL nude or normal rats were injected with either sulpiride (1 mg/kg) or saline and decapitated 2, 4, 8, 12, 14 and 24 h later. Plasma prolactin levels, compared with basal values, were increased in rats injected with sulpiride by 9.2 +/- 1.8 and 3.4 +/- 0.7-fold in IPL nude and normal rats respectively. The pituitary prolactin content was reduced more in IPL nude than in normal sulpiride-injected rats. These data suggest that prolactin secretion, as well as synthesis, is under an increased dopaminergic inhibition in the male IPL nude rat.

Variations of liver prolactin receptors during pregnancy in normal rats and in the genetically hypoprolactinemic IPL nude rat.

Ovine prolactin (oPRL) binding to liver membranes was studied during pregnancy in normal and in genetically hypoprolactinemic rats. Prolactin (PRL) binding was determined using 125I-oPRL in the 100,000 x g pellet. Scatchard plots obtained changed throughout the pregnancy in the normal rat, being almost linear from days 2 to 10, becoming curvilinear (convex) on day 16, and linear again at the end of pregnancy. They were analyzed with reference to the co-operativity and Hill models, which give delta and nH, respectively. During pregnancy, delta values varied and were respectively 2.48 +/- 0.66, 1.84 +/- 0.64, 0.52 +/- 0.06 and 1.69 +/- 0.25 on days 3, 10, 16 and 22, and the delta value on day 16 was significantly different from other days; the nH value estimated on day 16 was 1.10 +/- 0.031. These results suggest the presence of a positive co-operativity on day 16 of pregnancy. Over the same period, a huge increase in the capacity occurred on day 10 and reached a maximum on day 14. It remained elevated until the day before parturition. In the IPL nude rat, the delta value (0.92 +/- 0.45) on day 16 was significantly different from that of normal rats and indicated an absence of positive co-operativity on this day in the IPL nude rat liver. This finding was confirmed by an nH value (0.99 +/- 0.39) close to 1. The PRL-binding capacity was similar to that of normal rats, except on day 14, where it was significantly decreased. These results are discussed in relation to hormonal variations during pregnancy, particularly with regard to serum PRL and placental lactogen values.(ABSTRACT TRUNCATED AT 250 WORDS)

Variations of liver prolactin receptors during the estrous cycle in normal rats and in the genetically hypoprolactinemic IPL nude rat.

Ovine prolactin (oPRL) binding to liver membranes was studied during the estrous cycle in normal and in genetically hypoprolactinemic rats. Serum levels of hormones were measured by radioimmunoassay and prolactin (PRL) binding was determined using 125I-ovine PRL in the 100,000 X g pellet. Scatchard plots obtained were curvilinear throughout the estrous cycle in the normal rat. They were analyzed in reference to the co-operativity model and to the Hill model which give the factor delta and Hill's coefficient (nH), respectively. During the estrous cycle, delta values varied from 3.77 +/- 0.66 on the day of estrous to 13.48 +/- 1.34 on the day of proestrus at 16.00 h. At the same time, nH were 0.97 +/- 0.033 on the day of estrus and 0.72 +/- 0.025 on the day of proestrus at 16.00 h. On the other hand, the number of PRL receptors did not change significantly throughout the estrous cycle. Moreover, the dissociation of 125I-oPRL from its receptor was accelerated by the presence of native ovine oPRL. These results suggest the presence of a negative co-operativity which reached a maximum on the day of proestrus in the normal rat. This co-operativity during the estrous cycle was not found in liver from genetically hypoprolactinemic (IPL nude) rats, which present a total absence of lactation. The delta values did not vary significantly and were 6.52 +/- 1.30 on the day of estrus and 4.41 +/- 0.52 on the day of proestrus at 16.00 h. The difference between the two rat strains was statistically significant on the day of proestrus at 16.00 h for both delta and nH values.(ABSTRACT TRUNCATED AT 250 WORDS)

beta-Endorphin in genetically hypoprolactinemic rat: IPL nude rat.

Beta-endorphin has been reported to regulate not only stress- and suckling-induced but also basal prolactin secretion. In the aim to better evaluate the endogenous beta-endorphin-prolactin interrelation, we measured beta-endorphin levels in a new rat strain, genetically hypoprolactinemic and characterized by a total lack of lactation: IPL nude rat. Beta-endorphin was measured using a specific anti-h-beta endorphin in plasma and extracts of anterior and neurointermediate lobes of the pituitary, hypothalamus and brain. Pituitary extracts were also chromatographed on Sephadex G50 column. Results obtained showed that in IPL nude females on diestrus and males, the beta-endorphin contents of the neurointermediate lobe was significantly lower than in normal rats, while the values found in the other organs and plasma were similar. However, elution pattern of the anterior pituitary extract from male rats showed greater immunoactivity eluting as I125 h-beta-endorphin than in normal rat; this was not the case for the female rat. These results are consistent with a differential regulation of beta-endorphin levels of anterior and neurointermediate lobe by catecholamines. Moreover they suggest that PRL secretion was more related to neurointermediate beta-endorphin.

[Anti-inflammatory activity of a copper complex carrying a sugar vector, and observed serum copper levels]. / Activité anti-inflammatoire d'un complexe de cuivre porteur du vecteur sucre et taux de cuivre sérique observés.

(N-2-amino-2-deoxy-alpha, beta-D-glucopyranose salicylaldimino)-Cu (II) shows an anti-inflammatory activity in the cotton wad granuloma on Rats. An increase of the copper level in serum is detected very soon after s.c. drug administration, which is due to the high diffusion of the complex.

Dissociated estradiol (E2) action on the pituitary-testicular axis in a genetically hypoprolactinemic rat (IPL nude rat).

Prolactin (PRL) has been reported to be a possible mediator in the estradiol (E2)-induced inhibition of the pituitary-testicular axis. In order to better characterize the role of PRL, we studied the action of chronic hypoprolactinemia on this E2 inhibitory effect, using a genetically hypoprolactinemic rat (IPL nude). Normal and IPL nude adult male rats were injected either with vehicle or with E2 valerianate (4 mg/rat) once a week for 2 weeks. Rats were decapitated 7 days after the last injection. Results showed that E2 increased, similarly in both strains, pituitary weight and serum PRL levels. Serum testosterone values were reduced by 96% in both strains. However, testis weight was significantly reduced by 30% in normal rat, while in IPL nude rat, no significant decrease was observed. PRL binding sites, expressed as fmol/mg protein, were reduced in normal rat by 40%. No decrease was found in IPL nude rat. The dissociated E2 action observed in IPL nude rat suggested that only testicular growth inhibition could be mediated by PRL and confirm that testosterone level decrease could be due to a direct action of E2 on Leydig cells.

[Radioimmunological determination of the serum IgD]. / Dosage radioimmunologique de l'IgD sérique.

We describe a sensitive liquid phase radioimmunoassay for serum IgD. Extreme values obtained from 85 control patients sera are 0.2 and 121 mg/l with an arithmetic mean of 25 mg/l. In atopic patients (with high serum IgE levels), arithmetic mean is 47 mg/l.

Prolactin receptor regulation by LH in the rat mammary gland.

The aim of this study was to investigate hormonal factors responsible for the huge increase in PRL receptors on the day of estrus in the rat mammary gland. For this purpose, ovariectomized rats were primed with E2 so as to reach a physiological serum concentration of E2 (21.5 +/- 1.2 pg/ml) and high PRL serum values (72.8 +/- 21.9 ng/ml). In these conditions, PRL specific binding and capacity were respectively 22.8 +/- 8.3%/mg protein and 96 +/- 29 fm/mg protein. An injection of either LHRH (500 ng/rat) or LH (60 micrograms LH-RP1/rat) was capable of increasing significantly both PRL specific binding and capacity. Capacity reached the values of 498 +/- 103 and 507 +/- 240 fm/mg protein for LHRH and LH respectively. LHRH action appeared to be mainly mediated through LH secretion, since no difference was found between LHRH and LH. LHRH and LH injections alone were unable to modify PRL binding, suggesting that they only potentiate E2 and PRL action. These results show for the first time that LH is involved in the regulation of PRL receptors in the rat mammary gland.

[Absence of an increase in prolactin receptors on the day of estrus in the mammary gland of the nu IPL rat]. / Absence d'augmentation des récepteurs de la prolactine le jour de l'oestrus dans la glande mammaire de rat nu IPL.

Ovine prolactin (o-PRL) binding to rat mammary gland membranes has been shown to vary during the estrous cycle in the normal rat. In this study we report the characteristics of o-PRL binding to mammary gland membranes throughout the estrous cycle in a new rat strain, IPL nude, which presents a total absence of lactation. Prolactin receptors were quantified in the 100 000 g pellet. The mean value of the affinity constant in IPL nude rat was 16.7 X 10(9) M-1 and no variation was observed throughout the estrous cycle. The binding capacity also remained unchanged and the values were situated between 11.6 +/- 1.1. fmoles/mg protein and 38.6 +/- 12.3 fmoles/mg protein, corresponding to the lowest values obtained in normal rat. On the day of estrus, the prolactin binding capacity in the mammary gland of the IPL nude rat was significantly different from that of normal rat. These findings confirm that prolactin certainly plays an important role in the induction or stimulation of the rat mammary gland function.

Steroid regulation of gonadotropins in genetically hypoprolactinemic females (IPL nude rats).

IPL nude females present an absence of lactation with hypoprolactinemia. While males present a slight but significant decrease in serum testosterone and gonadotropins, females show normal values of estradiol, progesterone, LH and FSH during all estrus cycle stages. In this work, we observed that the postovariectomy rise of LH and FSH was significantly lower in the IPL nude females. We studied also the effect of acute (1 injection of 25 micrograms/rat E2Bz) or long-term (E2Bz capsule for 8 days) estradiol benzoate (E2Bz) treatment, with or without progesterone injection (5 mg/rat) in ovariectomized (OVX) IPL and normal females. The sensitivity of gonadotropins to E2 negative feedback is decreased in the IPL nude rats, result in agreement with previous reports and which could be linked to both hypoprolactinemia and decreased beta-endorphin observed in the IPL nude rat. The responsiveness of LH to LHRH was also tested in OVX and OVX + E2Bz or OVX + E2B + P treated. In OVX females responsiveness of LH to LHRH was similar in IPL nude to that of normal females. However, LH responsiveness in acute and long-term steroid-treated OVX IPL nude was significantly depressed. Since the mechanism whereby PRL interacts with steroids to modify gonadotropin secretion is still unexplained, IPL nude rat could be a good model to study it.

A new hypoprolactinemic rat strain. Prolactin, luteinizing hormone, follicle-stimulating hormone, testosterone and corticosterone levels in males and effects of two anesthetics.

A new strain of rat, the IPL nude rat, derived by spontaneous mutation from the Sprague-Dawley strain, is characterized by a lack of hair, delayed puberty and a total lack of lactation. To characterize partially the endocrine system in this rat, serum levels of prolactin (Prl), luteinizing hormone (LH), follicle-stimulating hormone (FSH), testosterone and corticosterone were quantified. The content of gonadotropins and Prl in the pituitary gland was also measured. Blood samples were collected following decapitation or by cardiac puncture after ether or ketamine anesthesia. Serum levels of Prl were decreased by approximately 50% in IPL nude compared to normal Sprague-Dawley rats, regardless of the method used for collection of the samples. There were also significantly reduced levels of testosterone, LH and FSH in the serum collected from IPL nude rats. Levels of corticosterone in serum were similar in IPL nude and normal rats. There was the same relative response to anesthesia in the levels of all the hormones measured when the data from IPL nude and normal rats were compared. The quantities of LH, FSH and Prl contained in the pituitary of the IPL nude rats were not significantly different from those observed in normal rats. These data suggest that the serum hormone deficiencies observed in the nude IPL rat may be due to an altered regulation of hormone production by hypothalamic factors.

Prolactin receptors in the rat mammary gland. Change during the estrous cycle.

Ovine prolactin (o-PRL) binding to mammary gland membranes was studied during the estrous cycle in the rat. Groups of rats were decapitated throughout the 4-day estrous cycle at 10 h00 on the days of diestrus I, diestrus II and estrus and at 10 h00, 12 h00, 16 h00 during the day of proestrus. Daily vaginal smears were taken to determine the stage of the estrous cycle which was also controlled by PRL and LH serum levels. Prolactin receptors were quantified in the 100 000 g pellet. For one Scatchard analysis, mammary gland membranes from 5 animals were pooled. Results given are the mean of 4 or 5 pools. Results obtained showed that the apparent affinity constant (KA) remained unchanged during the days of diestrus II and at all the times studied of proestrus and showed a slight but significant decrease on the days of estrus and diestrus I (or metestrus). The binding capacity did not vary from the day of diestrus II to the proestrus 16h00 (11.3 +/- 2.8 fmoles/mg protein) but sharply increased on the day of estrus (190.4 +/- 35.9 fmoles/mg protein). Binding capacity remained elevated on the day of diestrus I. This increase of PRL receptors on the day of estrous would appear to be an important step in preparing mammary gland for pregnancy and lactation.

Prolactin receptors in the hypoprolactinemic male rat (IPL nude rat): measurement in testis and induction in liver.

In order to evaluate the importance of PRL in the regulation of its own receptors, characteristics of specific binding for PRL were studied in membrane preparations from liver and testis of a new hypoprolactinemic male rat, the IPL nude male rat, and this was compared to those found for normal male rats. Under basal conditions, hepatic specific binding of PRL in IPL nude rats, as in normal rats was not detectable. Following castration, it became detectable in both groups, and was 6.99 +/- 0.78% and 6.34 +/- 0.87% for IPL nude and normal rats respectively. Under such conditions, the apparent affinity constant (Ka) and the binding capacity (Nmax) obtained were also similar for both groups (Ka) = 1.36 +/- 0.14.10(9) M-1, Nmax = 102 +/- 14 fmol/mg protein in IPL and Ka = 1.34 +/- 0.28.10(9) M-1, Nmax = 97 +/- 11 fmol/mg protein in normal rats) although a decrease in serum levels of PRL was observed in both groups. This decrease was greater for IPL nude rats. As already reported, estradiol injection following castration was able to further increase the percentage of PRL hepatic specific binding (4 times). Furthermore, our results demonstrated that the affinity constant was significantly increased by estradiol injection in both groups. On the other hand, for testicular PRL binding characteristics, a statistically significant difference was found between IPL nude and normal rats. The PRL specific binding percentage was 7.01 +/- 0.85% for the IPL nude rat and 10.07 +/- 0.64% for the normal rat. By Scatchard analysis, the Ka of testicular membranes for labelled oPRL was similar in both groups, while the capacity differed (Nmax = 9.82 +/- 1.25 fmol/mg protein for IPL nude rat and Nmax = 26.06 +/- 4.39 fmol/mg protein for normal rats). These data established the fact that IPL nude male rats presented characteristics of hepatic PRL receptors similar to those of normal rats, while their testicular oPRL binding significantly differed. These findings therefore suggest that in genetic hypoprolactinemic rats (IPL nude rats), PRL might be more involved in the regulation of testicular PRL receptors than in that of hepatic receptors.

[Utero-ovarian implantation. Experimental study and critical evaluation of its use in women]. / L'implantation ovaro-utérine. Etude expérimentale et étude critique de son application humaine.

The authors studied experimentally ovario-uterine implantation in rats, an operation first performed many years ago yielding very rare and debatable results. This procedure is ineffective in the treatment of sterility due to endometrial proliferation over the ovary, which closes off rapidly the endometrial cavity and separates it from the implanted ovary. Much uncertainty surrounds the majority of previously reported successes, the few resultant pregnancies from ovario-uterine implantation considered due to hazard and completely unreliable. This operation has no place in the treatment of sterility.

[Casein-like immunoreactivity in the male rat]. / Immunoactivité semblable à la caséine chez le rat mâle.

A substance presenting a beta-casein-like immunoreactivity (beta-cas-LI) was found in male serum and tissues. This beta-cas-LI was measured by a specific radioimmunoassay using a purified beta-casein and an anti beta-casein kindly supplied by L. M. Houdebine. Serial dilutions of serum or tissues samples showed a good parallelism to the standard curve. In male rat, beta-cas-Li was found in all sera and in some tissues = mammary gland, prostate and testis which contained respectively 6,200, 408 and 38 ng per organ. beta-cas-LI value in the serum was regulatable. Hypophysectomy and long-term castration strongly reduced basal levels. Insertion of testosterone silastic capsules (25 mg/capsule) was able to restore control values, while estradiol (E2) (4 mg/kg E2 valerianate) to castrated or intact rats increased serum beta-cas-LI above basal values. In conclusion, these results showed, for the first time, the presence in male rat serum and tissues of a beta-casein immunoreactive-like substance. Moreover, this beta-cas-LI seemed to be under pituitary and gonad regulation.

[Anti-inflammatory activity of 5 copper complexes]. / Activité anti-inflammatoire de cinq complexes du cuivre.

The anti-inflammatory activity of five copper complexes is shown in the cotton wad granuloma test in Rats. The activity due to copper seems to be only modulated by the ligands in the complexes studied.