Agonist-induced desensitization of the D-2 dopamine receptor in the intermediate lobe of the rat pituitary gland

The mechanism of agonist-induced desensitization of the D-2 dopamine receptor in the intermediate lobe (IL) of the rat pituitary gland was investigated. Exposure of neurointermediate lobe to 60 microM (-)apomorphine (APO) for 60 min altered the binding of [125I]-N-(p-aminophenethyl)spiperone (NAPS), a D-2 receptor-specific ligand. The capacity of the tissue to bind the ligand (Bmax) was not significantly altered by the exposure to (-)APO but the affinity for [125I]NAPS was decreased 3.6-fold in (-)APO-exposed tissue. The molar potency of YM-09151-2, a D-2 receptor-specific antagonist, showed a minimal difference between in control and (-)-APO-exposed tissue. However, the molar potency of (-)APO towards the D-2 receptor was diminished. The loss of [125I]NAPS binding in (-)APO-exposed tissue was reversed by the addition of guanyl nucleotide. These data suggest that exposure to agonist causes a persistent occupancy of the high affinity state of the receptor. Exposure to (-)APO had no effect on either basal or forskolin-activated adenylate cyclase activity of the intermediate lobe. However, the inhibitory effect of (-)APO upon adenylate cyclase activity of IL homogenates was diminished when the tissue was exposed to (-)APO before homogenization. Furthermore, the ability of GTP but not 5'-guanylyl imidodiphosphate [Gpp(NH)p] to inhibit enzyme activity diminished in the (-)APO-exposed tissue. These data suggest that an agonist-induced desensitization of D-2 receptor in rat IL is thought to occur by uncoupling the receptor from the inhibitory guanyl nucleotide binding protein (Gi) or potentiating the hydrolysis of GTP by Gi.     

Dephosphorylation of the beta 2-adrenergic receptor and rhodopsin by latent phosphatase 2

Recent evidence suggests that the function of receptors coupled to guanine nucleotide regulatory proteins may be controlled by highly specific protein kinases, e.g. rhodopsin kinase and the beta-adrenergic receptor kinase. In order to investigate the nature of the phosphatases which might be involved in controlling the state of receptor phosphorylation we studied the ability of four highly purified well characterized protein phosphatases to dephosphorylate preparations of rhodopsin or beta 2-adrenergic receptor which had been highly phosphorylated by beta-adrenergic receptor kinase. These included: type 1 phosphatase, calcineurin phosphatase, type 2A phosphatase, and the high molecular weight latent phosphatase 2. Under conditions in which all the phosphatases could dephosphorylate such common substrates as [32P]phosphorylase a and [32P]myelin basic protein at similar rates only the latent phosphatase 2 was active on the phosphorylated receptors. Moreover, a latent phosphatase activity was found predominantly in a sequestered membrane fraction of frog erythrocytes. This parallels the distribution of a beta-adrenergic receptor phosphatase activity recently described in these cells (Sibley, D. R., Strasser, R. H., Benovic, J. L., Daniel, K., and Lefkowitz, R. J. (1986) Proc. Natl. Acad. Sci. U. S. A. 83, 9408-9412). These data suggest a potential role for the latent phosphatase 2 as a specific receptor phosphatase.     

Expression of platelet glycoprotein Ib alpha in HEL cells

We have previously shown that platelet glycoprotein Ib is expressed in a minority of cells of the human leukemic cell line HEL (Tabilio, A., Rosa, J. P., Testa, U., Kieffer, N., Nurden, A. T., Del Canizo, M. C., Breton-Gorius, J., and Vainchenker, W. (1984) EMBO J. 3, 453-459). In this report, we have selected a stable HEL subclone with increased expression of glycoprotein (GP) Ib as assessed by 6 different monoclonal antibodies in order to investigate the biochemical characteristics of this glycoprotein. A single polypeptide chain of apparent Mr = 60,000 was precipitated under reducing and nonreducing conditions by a specific polyclonal anti-platelet glycocalicin antibody and two anti-GPIb alpha monoclonal antibodies (AN51 and AP1), both from surface-labeled and metabolically labeled HEL cells. We were unable to demonstrate the presence of a polypeptide corresponding to the beta subunit of GPIb or GPIX which is closely associated with GPIb. Competitive immunoprecipitation performed in the presence of an excess amount of cold platelet glycocalicin completely displaced the Mr = 60,000 polypeptide. Synthesis of N-linked oligosaccharide chains on this Mr = 60,000 polypeptide was inhibited by the antibiotic tunicamycin, and a shift of the apparent Mr from 60,000 to 48,000 was observed. O-Linked oligosaccharide chains identical to platelet GPIb hexasaccharides were deficient or incomplete since no peanut agglutinin binding to the Mr = 60,000 polypeptide was observed after neuraminidase treatment of HEL cells. Thus, our results provide evidence that the Mr = 60,000 polypeptide expressed on the surface membrane of HEL cells is closely related to platelet GPIb and corresponds to an incompletely or abnormally O-glycosylated GPIb alpha subunit.     

Detection of Chlamydia trachomatis in Papanicolaou-stained cervical smears by an indirect immunoperoxidase method

A two-step (indirect) immunoperoxidase method directed against Chlamydia trachomatis was developed. The method was then used to evaluate the specificity of cytologic changes suggestive of C. trachomatis in Papanicolaou smears of cervical specimens from women who were culture-negative for the organism. Positive immunoperoxidase staining was detected in 9 of 21 cases (43%) tested. Technical problems, especially background staining, precluded interpretation in the remainder of the cases. Cervical cytology, as demonstrated by immunoperoxidase staining, may, in some instances, be more sensitive than the culture. However, because the etiology of cytologic changes not specifically identified by immunoperoxidase staining may be due to other organisms or factors, immunoperoxidase procedures, as described, should not replace culture for confirmation of cytologic findings suggestive of C. trachomatis.     

Photoaffinity labeling of mammalian alpha 1-adrenergic receptors. Identification of the ligand binding subunit with a high affinity radioiodinated probe

We have synthesized and characterized a novel high affinity radioiodinated alpha 1-adrenergic receptor photoaffinity probe, 4-amino-6,7-dimethoxy-2-[4-[5-(4-azido - 3 - [125I]iodophenyl) pentanoyl] - 1 - piperazinyl] quinazoline. In the absence of light, this ligand binds with high affinity (KD = 130 pM) in a reversible and saturable manner to sites in rat hepatic plasma membranes. The binding is stereoselective and competitively inhibited by adrenergic agonists and antagonists with an alpha 1-adrenergic specificity. Upon photolysis, this ligand incorporates irreversibly into plasma membranes prepared from several mammalian tissues including rat liver, rat, guinea pig, and rabbit spleen, rabbit lung, and rabbit aorta vascular smooth muscle cells, also with typical alpha 1-adrenergic specificity. Autoradiograms of such membrane samples subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis reveal a major specifically labeled polypeptide at Mr = 78,000-85,000, depending on the tissue used, in addition to some lower molecular weight peptides. Protease inhibitors, in particular EDTA, a metalloprotease inhibitor, dramatically increases the predominance of the Mr = 78,000-85,000 polypeptide while attenuating the labeling of the lower molecular weight bands. This new high affinity radioiodinated photoaffinity probe should be of great value for the molecular characterization of the alpha 1-adrenergic receptor.     

A revision of the Scapholeberinae (Crustacea: Cladocera)

The group of hyponeustonic daphniid cladocera previously known under the generic name Scapholeberis is raised to the rank of a subfamily (Scapholeberinae) and contains two genera, Megafenestra n.gen. (2 species), and Scapholeberis s.s. (7 species and one subspecies). The characters upon which this revision is based are: structure of the rostrum, structure of the first antennae, structure of trunk limbs 1, 2, and 5, presence and nature of headpore(s), structure of the ventral rim of the valves, armature of the distal rim of the valves, structure of the postabdomen and its end-claws. Males were examined in all but two species, and proved to be more primitive than females and much less diagnostic than in the Chydoridae. It also appeared that the shape and armature of the postabdomen are less important taxonomical tools than heretofore supposed. In more than half of the species, it is of generalised shape and hardly usable beyond the species-group level. The same, although less drastically, holds true for the P1 of males. Of greatest diagnostic value at the species level are the shape of the rostrum and the armature of the valve margins. A phylogeny of the subfamily is proposed, with Megafenestra as the more primitive genus, closely allied to Ceriodaphnia, and within Scapholeberis, S. mucronata as the most apomorphic species. In biogeographical respect, no species is cosmopolitan, although their ranges may be of continental dimensions. Species and populations appear to concentrate in the temperate and continental climate belts of the northern hemisphere, but more research in the southern hemisphere is needed. A key for the identification of all species is given.     

Flore levuriforme intestinale du babouin,Papio papio (Desm.)

Résumé Chez 44 Babouins,Papio papio (Desm.), appartenant à la collection du Parc zoologique de Paris, distingués en trois groupes, en fonction de l'âge, nous avons pratiqué 101 prélèvements aux niveaux suivants du tube digestif: intestin grêle gros intestin et rectum.C'est chez les animaux les plus âgés que nous avons obtenu les valeurs les plus élevées pour les deux critères étudiés, le pourcentage de positivité et la richesse spécifique des prélèvements et ce, aussi bien en ce qui concerne l'ensemble des levures identifiées que pour les seuls constituants de la flore probable. La fréquence de ces champignons aux divers étages du tube digestif, ainsi que l'incidence saisonnière sur les isolements ont été abordés.

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In 44 Baboons,Papio papio (Desm.), of the Zoological Park of Paris, 101 samples have been taken at three levels of the alimentary tract: small intestine, large intestine and rectum. Three groups of subjects have been recognized according to age. The yeasts or the yeast-like flora, have been found with maximum of frequency and diversity in the oldest animals. The variations of the isolation in the three levels of this Mammal's gut and the seasonal influence have been discussed.

     

Calorimetric measurement of free energy utilization by Nitrosomonas and Nitrobacter

Summary Calorimetric estimates of the utilization efficiency of the free-energy derived from substrate oxidation by cell suspensions of two nitrifying bacteria, Nitrosomonas and Nitrobacter, provided two ranges of values: 11 to 27% and 15 to 51%, respectively. About 15 to 30% of the utilized free-energy is used for driving endergonic reactions other than CO₂ fixation, probably the synthesis of polyphosphates.The molar heat of substrate oxidation does not seem to be influenced by the age of cells harvested during growth or by the length of the incubation period during which cells have been kept in a buffer suspension in a starved condition. The loss of respiratory activity measured either by oxygen uptake or heat evolution in the presence of the specific substrate, nitrite or ammonium, decreases according to kinetics which are influenced by the aerobiosis of the suspension. The viability of the starved cells decreases in a way which is similar to that of the respiratory activity. It seemed impossible to obtain cells which had lost their viability but kept the ability to oxidize their substrate.Two inhibitors of the respiratory chain, quinacrine and cyanide, are without effect on the molar heat of substrate oxidation and consequently on the free-energy utilization efficiency. 2.4 dinitrophenol did decrease the rate of heat evolution during substrate oxidation at concentrations at which the rate of oxygen uptake was not depressed, with the consequences that free-energy efficiency was apparently increased.