Comparative study of oestrogenic properties of eight phytoestrogens in MCF7 human breast cancer cells

Previous studies have compared the oestrogenic properties of phytoestrogens in a wide variety of disparate assays. Since not all phytoestrogens have been tested in each assay, this makes inter-study comparisons and ranking oestrogenic potency difficult. In this report, we have compared the oestrogen agonist and antagonist activity of eight phytoestrogens (genistein, daidzein, equol, miroestrol, deoxymiroestrol, 8-prenylnaringenin, coumestrol and resveratrol) in a range of assays all based within the same receptor and cellular context of the MCF7 human breast cancer cell line. The relative binding of each phytoestrogen to oestrogen receptor (ER) of MCF7 cytosol was calculated from the molar excess needed for 50% inhibition of 3H]oestradiol binding (IC50), and was in the order coumestrol (35x)/8-prenylnaringenin (45x)/deoxymiroestrol (50x)>miroestrol (260x)>genistein (1000x)>equol (4000x)>daidzein (not achieved: 40% inhibition at 10(4)-fold molar excess)>resveratrol (not achieved: 10% inhibition at 10(5)-fold molar excess). For cell-based assays, the rank order of potency (estimated in terms of the concentration needed to achieve a response equivalent to 50% of that found with 17beta-oestradiol (IC50)) remained very similar for all the assays whether measuring ligand ability to induce a stably transfected oestrogen-responsive ERE-CAT reporter gene, cell growth in terms of proliferation rate after 7 days or cell growth in terms of saturation density after 14 days. The IC50 values for these three assays in order were for 17beta-oestradiol (1 x 10(-11)M, 1 x 10(-11)M, 2 x 10(-11)M), and in rank order of potency for the phytoestrogens, deoxymiroestrol (1 x 10(-10)M, 3 x 10(-11)M, 2 x 10(-11)M)>miroestrol (3 x 10(-10)M, 2 x 10(-10)M, 8 x 10(-11)M)>8-prenylnaringenin (1 x 10(-9)M, 3 x 10(-10)M, 3 x 10(-10)M)>coumestrol (3 x 10(-8)M, 2 x 10(-8)M, 3 x 10(-8)M)>genistein (4 x 10(-8)M, 2 x 10(-8)M, 1 x 10(-8)M)/equol (1 x 10(-7)M, 3 x 10(-8)M, 2 x 10(-8)M)>daidzein (3 x 10(-7)M, 2 x 10(-7)M, 4 x 10(-8)M)>resveratrol (4 x 10(-6)M, not achieved, not achieved). Despite using the same receptor context of the MCF7 cells, this rank order differed from that determined from receptor binding. The most marked difference was for coumestrol and 8-prenylnaringenin which both displayed a relatively potent ability to displace [3H]oestradiol from cytosolic ER compared with their much lower activity in the cell-based assays. Albeit at varying concentrations, seven of the eight phytoestrogens (all except resveratrol) gave similar maximal responses to that given by 17beta-oestradiol in cell-based assays which makes them full oestrogen agonists. We found no evidence for any oestrogen antagonist action of any of these phytoestrogens at concentrations of up to 10(-6)M on either reporter gene induction or on stimulation of cell growth.     

Characterization of changes in the activity of K+-n-nitrophenylphosphatase of erythrocyte ghosts under the influence of ouabain

Variation of K+-p-NPP-ase of human ghosts under the action of ouabain (10(-11)-10(-3) M) has been studied. In human ghosts the activity of ouabain-sensitive K+-p-NPP-ase has been found to make up 65% of the total enzyme activity. The activity of ouabain-sensitive K+-p-NPP-ase reaches a maximum level at pH 7.6-8.0. A decrease in the activity of this enzyme caused by ouabain is of two-phase character. In the range of ouabain concentration from 10(-10) M to 10(-6) M and from 10(-5) M to 10(-3) M the enzyme activity lowers significantly; in the range of 10(-7) M to 10(-5) M it reaches the plato. Two types of the enzyme are assumed to exist differing by 4-5 orders of magnitude in their sensitivity to ouabain, inhibitor affinity constants and Michaelis constants.     

Effect of bradykinin on airway neural responses in vitro.

We investigated the effects of bradykinin (BK) on airway excitatory nonadrenergic noncholinergic (e-NANC) and cholinergic nerves in vitro. Neural responses were elicited by electrical field stimulation in guinea pig airways in vitro before and after the addition of BK (10(-10)-10(-7) M). Captopril (10(-5) M) and phosphoramidon (10(-6) M) were added to prevent degradation of BK, and all neural responses were measured in the presence of indomethacin (10(-5) M) and propranolol (10(-6) M). BK potentiated e-NANC responses in bronchi in a concentration-dependent manner (10(-10)-10(-7) M) without changing concentration-response curves to exogenously applied substance P (10(-10)-10(-5) M). BK significantly potentiated e-NANC neural constrictor responses by 22 +/- 7% at 10(-8) M (mean +/- SE, n = 5, P < 0.05) and 32 +/- 7% at 10(-7) M (n = 8, P < 0.01), compared with changes in time-matched control tissues (7 +/- 2%, n = 8). The potentiation of e-NANC responses by BK was abolished by pretreatment with a specific B2-receptor antagonist, HOE 140 (10(-7) M). Cholinergic constrictor responses elicited to electrical field stimulation were not affected by the addition of BK (up to 10(-7) M). These results suggest that BK potentiates e-NANC bronchoconstrictor responses prejunctionally via a B2-receptor.     

Quantitative analysis of the elemental composition and the mass of bacterial polyphosphate bodies using STEM EDX

The quantitative analysis of laboratory grown organisms (Plectonema boryanum and Staphylococcus aureus) revealed that a typical in vivo polyphosphate body (PPB) contains O (4.3 x 10(-8) microg), C (1.2 x 10(-8) microg), P (6.7 x 10(-9) microg), Mg (1.3 x 10(-9) microg), Ca (6.7 x 10(-10) microg), K (6.7 x 10(-10) microg), Fe (6.0 x 10(-10) microg), S (5.4 x 10(-10) microg) and Al (5.9 x 10(-10) microg). Quantitative X-ray analysis of samples from nature showed PPB contain O (1.63 x 10(-8) microg), C (4.75 x 10(-9) microg), P (2.50 x 10(-9) microg), Mg (5.0 x 10(-10) microg), Ca (2.50 x 10(-10) microg), K (2.50 x 10(-10) microg), Fe (2.25 x 10(-10) microg) and S (2.0 x 10(-10) microg). The mass of an average polyphosphate body was 6.7 x 10(-8) microg for P. boryanum, 2.5 x 10(-8) microg for S. aureus and for microbes from the natural environment 6.3 x 10(-8) microg. The results indicate that the PPB may have other unknown functions in addition to essential element storage, acting as a detoxification method by sequestering heavy metals and providing a homeostasis system in the cell.     

Inhibition of small conductance K+ -channels attenuated melatonin-induced relaxation of serotonin-contracted rat gastric fundus

The aim of this study was to investigate the effects of melatonin on rat gastric fundus smooth muscle. Melatonin (10(-4) to 10(-3) M) had no effect on the basal tone of gastric smooth muscle. After precontraction with carbachol (10(-6) M) or serotonin (10(-7) M), melatonin caused a concentration dependent inhibitory action. The half maximal effect on serotonin-induced contraction was found with 1.12 +/- 0.86 x 10(-5) M of melatonin. Increasing concentrations of melatonin (10(-5) to 10(-3) M) resulted in a right shift of the serotonin concentration response curve (10(-10) to 10(-5) M). This inhibitory effect of melatonin was partially blocked in the presence of apamin (10(-10) to 10(-7) M), a specific blocker of the small conductance calcium-dependent potassium channel, but not in the presence of other potassium channel blockers like charybdotoxin (10(-8) M), glibenclamide (l0(-5) M), or tetraethylammonium (ODQ, 10(-4) M). The inhibitory effect was not changed in the presence of the neuronal blocker tetrodotoxin (10(-6) M), the selective P2-receptor antagonist pyridoxalphosphate-6-azophenyl-2',4'-disulphonic acid (3 x 10(-5) M), the nitric-oxide synthase inhibitor N-nitro-L-arginine (3 x 10(-4) M), or the guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]-quinoxalin-1-one (10(-4) M), suggesting that neither the purinergic, nitrergic, nor guanylate cyclase pathways were involved. We further investigated inhibitory responses to electrical field stimulation (EFS) at different frequencies under non-adrenergic, non-cholinergic (NANC) conditions on a serotonin-induced contraction in the presence of melatonin (10)-5 to 10(-4) M). Melatonin significantly reduced these inhibitory NANC responses in higher (8-32 Hz), but not lower (05-4 Hz), frequencies (16 Hz without melatonin, 103 +/- 6.3%; melatonin 10(-5) M, 80.4 +/- 7.5%; melatonin 10(-4) M, 39.1 +/- 17.1%). Melatonin had no effect on contractile responses induced by EFS under basal tone. These results demonstrate that the inhibitory effect of melatonin in rat gastric fundus smooth muscle is apamin sensitive, but is not affected by other potassium channel blockers. This suggests that melatonin may be another transmitter candidate for the apamin sensitive responses within the gastrointestinal tract.     

Colinearity between the DNAs of Epstein-Barr virus and herpesvirus papio.

Epstein Barr virus (EBV) and herpesvirus papio (HVPapio) DNAs share a common format and 40% homology. Labeled cloned fragments of EBV DNA were hybridized to blots of XbaI, EcoRI, HindIII, and SalI fragments of HVPapio DNA. EBV fragments which mapped from 2 x 10(6) to 54 x 10(6) and from 59 x 10(6) to 106 x 10(6) daltons hybridized to fragments at identical map positions in HVPapio DNA. Regions of nonhomology were demonstrated at 0 x 10(6) to 2 x 10(6), 54 x 10(6) to 59 10(6), and 106 x 10(6) to 115 x 10(6) daltons.     

Stimulatory effects of LH on release of melatonin and activities of its synthesizing enzymes NAT and HIOMT in organ-cultured pineal glands of female rats.

The pineal hormone melatonin (N-acetyl-5-methoxytryptamine) exerts antigonadotropic effects in some mammalian species. To evaluate the effect of luteinizing hormone (LH) on melatonin release and its synthesizing enzyme activities in pineal glands, pineals of adult female rats undergoing diestrus were organ-cultured in a medium containing 10(-12), 10(-10) or 10(-8) M LH for 6 h. Melatonin release increased significantly in pineals cultured with 10(-12) and 10(-10) M LH, as compared to control values. Similarly, the activity of arylalkylamine N-acetyltransferase (NAT), the key regulatory enzyme in melatonin biosynthesis, was significantly higher in pineals cultured with 10(-12) and 10(-10) M LH for 6 h, while LH at 10(-8) M had no effect. Although LH at 10(-10) M increased pineal hydroxyindole-O-methyltransferase (HIOMT) activity, which catalyzes the final step of melatonin biosynthesis, LH at 10(-12) and 10(-8) M had no effect. These results demonstrate that at relatively low physiological levels, LH stimulates pineal melatonin synthesis and release, mainly by increasing NAT activity.     

Modification of erythrocyte physicochemical properties by millimolar concentrations of glutaraldehyde.

The effects of low levels of glutaraldehyde uptake (less than 120 mumol/10(10) cells) on the physicochemical properties of human red blood cells (RBC) were investigated. Salient effects include: by different measures of cell deformability, the extent of glutaraldehyde uptake required to decrease cellular deformability was shown to range from approximately 8 to 30 mumol/10(10) cells; osmotically stressed red cells exhibit complete hemolysis when the level of glutaraldehyde uptake is less than 28 mumol/10(10) cells and no hemolysis when uptake is less than 70 mumol/10(10) cells with the extent of hemolysis decreasing in an approximately linear manner with glutaraldehyde uptake between these limits; glutaraldehyde uptake of up to 58 mumol/10(10) cells does not change the cells' density, mean cell volume or ability to retain potassium.     

Seasonality of Chesapeake Bay bacterioplankton species

Bacteria, gamma-subclass of Proteobacteria, Vibrio-Photobacterium, Vibrio vulnificus, Vibrio cholerae-Vibrio mimicus, and Vibrio cincinnatiensis in water samples collected from the Choptank River in Chesapeake Bay from 15 April to 16 December 1996 were enumerated using a fluorescent oligonucleotide direct-counting (FODC) procedure. FODC results obtained using a Bacteria taxon-specific probe ranged from one-third the number of to the same number as that obtained by the acridine orange direct count (AODC) procedure. The abundance of individual taxa (per liter) ranged from 0.25 x 10(10) to 2.6 x 10(10) Bacteria, 0.32 x 10(8) to 3.1 x 10(8) gamma-Proteobacteria, 0.2 x 10(8) to 2.1 x 10(8) Vibrio-Photobacterium, 0.5 x 10(7) to 10 x 10(7) V. vulnificus, 0.2 x 10(6) to 6 x 10(6) V. cholerae-V. mimicus, and 0.5 x 10(5) to 8 x 10(5) V. cincinnatiensis. The occurrence of all taxa monitored in this study was higher in summer; however, these taxa made up a larger proportion of the Bacteria when the water temperature was low. Large fluctuations in species abundance as well as in percent composition of Vibrio-Photobacterium occurred from week to week, indicating that localized blooms of these taxa occur. The cross-Choptank River transect sample profile of V. vulnificus and V. cholerae-V. mimicus varied significantly in abundance, and trans-Choptank River transect samples revealed a patchy distribution.     

Fractionation of polyclonal antibody by isoelectric focusing and chromatofocusing: separation of high-affinity rabbit clonotype anti-thyroxine antibody

Immunoglobulin G fractions prepared from conventional rabbit anti-thyroxine (T4) antisera were fractionated by agarose gel isoelectric focusing (IEF) in the range of pH 3 to 10, and by chromatofocusing using a Fast Protein Liquid Chromatography (FPLC) system. The clonotype antibodies were recovered from the fractions and subjected to Scatchard plot analysis. The highest affinity constants of the initial antibody (shown in parentheses) and those of the antibodies recovered were IEF, 1.8 X 10(9) to 8.3 X 10(9) M-1 (2.2 X 10(9) M-1); FPLC, 2.4 X 10(9) to 6.0 X 10(9) M-1 (2.5 X 10(9) M-1). A sensitive radioimmunoassay of T4 was achieved with the isolated high-affinity anti-T4 antibody. The minimum detectable concentration of T4 was 6.3 X 10(-15) to 1.5 X 10(-14) mol/tube, which was three to five times lower than detectable with the initial antibodies.     

The vacuum UV CD spectra of G.G.C triplexes.

Vacuum UV circular dichroism (CD) spectra were measured down to 175 nm for d(C)10, d(G)10, the d(G)10.d(C)10 duplex, and the d(G)10.d(G)10.d(C)10 triplex. A CD difference spectrum was calculated for d(G)10.d(C)10 giving the change in CD induced by forming the duplex from d(G)10 and d(C)10. The d(G)10.d(G)10.d(C)10 CD difference spectrum gave the CD induced by triplex formation from binding of d(G)10 to the d(G)10.d(C)10 duplex. In the near-UV, the d(G)10.d(C)10 and d(G)10.d(G)10.d(C)10 difference spectra resembled the difference spectrum for poly[r(G).r(C)] (Biopolymers 29, 325-333). This similarity may be an indication of similar purine base stacking. The d(G)10.d(G)10.d(C)10 vacuum UV difference spectrum had a negative band at 195 nm and a positive band at 180 nm, making it similar to difference spectra for homopolymer triplexes containing T.A.T and U.A.U triplets (Nucl. Acids Res. 19, 2275-2280). The appearance of these bands in difference spectra should be good indicators of triplex formation. The complementary oligonucleotides c-mycI d(CCCCACCCTCCC) and c-mycII d(GGGAGGGTGGGG) are part of the regulatory sequences of the human c-myc gene. G.G.C rich triplexes formed by binding c-mycII or c-mycIII d(GGGGTGGGTGGG) to the c-mycI.c-mycII duplex had CD difference spectra similar to that of d(G)10.d(G)10.d(C)10 in both the vacuum UV and near UV regions, indicating similar triplet structures.     

Molecular and structural characterization of the biosurfactant produced by Pseudomonas aeruginosa DAUPE 614

Pseudomonas aeruginosa DAUPE 614 produced rhamnolipids (3.9gL(-1)) when cultivated on a medium containing glycerol and ammonium nitrate. These rhamnolipids reduced the surface tension of water to 27.3mNm(-1), with a critical micelle concentration of 13.9mgL(-1). The maximum emulsification index against toluene was 86.4%. The structure of the carbohydrate moiety of the glycolipid was determined by gas chromatography-mass spectroscopy (GC-MS) analysis allied to electrospray ionization mass spectrometry and nuclear magnetic resonance (NMR) 1D, 2D (13)C, (1)H spectroscopy. The hydroxyl fatty acids were analyzed by GC-MS as hydroxy-acetylated fatty acid methyl ester derivatives. The positions of the fatty acids in the lipid moiety were variable, with 6 mono-rhamnolipid homologues (Rha-C(10)-C(10); Rha-C(10)-C(8); Rha-C(8)-C(10); Rha-C(10)-C(12:1); Rha-C(12)-C(10); Rha-C(10)-C(12)) and 6 di-rhamnolipid homologues (Rha(2)-C(10)-C(10); Rha(2)-C(10)-C(8); Rha(2)-C(8)-C(10); Rha(2)-C(10)-C(12:1); Rha(2)-C(12)-C(10); Rha(2)-C(10)-C(12)). The ratio of Rha(2)-C(10)-C(10) to Rha-C(10)-C(10) was higher than has been reported in previous studies. Our methodology allowed us to distinguish between the isomeric pairs Rha-C(10)-C(8)/Rha-C(8)-C(10), Rha-C(10)-C(12)/Rha-C(12)-C(10), Rha(2)-C(10)-C(8)/Rha(2)-C(8)-C(10) and Rha(2)-C(12)-C(10)/Rha(2)-C(10)-C(12). For each isomeric pair, the congener with the shorter chain adjacent to the sugar was always more abundant than the congener with longer chain.     

Induction of metabolic shocks in source and sink of two cucurbits by molybdenum stress

Different concentrations of ammonium molybdate (10(-7) to 10(-4)M) affected the levels of metabolites in the source and sink organs of the seedlings of C. melo and C. vulgaris and data were recorded at 7, 14 and 21 days after treatment (DAT) of molybdenum (Mo). Reducing and non reducing sugars declined with an increase in concentration of ammonium molybdate from 10(-7) to 10(-4) M. Soluble protein and dry weight of seedlings increased in source at lower concentration (10(-7) M) and gradually decreased in all other concentrations (10(-6), 10(-5) and 10(-4) M). Starch was slightly accumulated in hypocotyl and fresh weight constantly declined with an increase in ammonium molybdate concentration from 10(-7) to 10(-4) M in all the parts of seedlings viz. cotyledon, hypocotyl and roots. Thus molybdenum at higher concentration induced decline in the metabolite levels in source and sink as well as in transporting organs.     

Sources of spermatozoa loss during collection and artificial insemination of horses

During artificial insemination of horses, it is important to accurately estimate the number of spermatozoa in each insemination dose. However, little research exists regarding sources of spermatozoa loss during collection and artificial insemination. Therefore, spermatozoal losses were quantified in the dismount loss (187.6×10(6)±62.5×10(6)spermatozoa), gel fraction (179.8×10(6)±61.7×10(6)spermatozoa), and the collection receptacle (136.1×10(6)±26.9×10(6)spermatozoa). Spermatozoal losses were examined in the centrifuge tube (25.8×10(6)±2.1×10(6)spermatozoa), AI pipette during the air removal (90.9×10(6)±8.5×10(6)spermatozoa), and spermatozoa remaining in the AI pipette after insemination (342.9×10(6)±21.4×10(6)spermatozoa). The average cumulative loss was 14.2±2.9% of the total spermatozoa ejaculated with approximately half of the loss due to the process of semen collection and half due to the process of artificial insemination. Spermatozoa retained in the AI pipette, after insemination with extended semen, represented the greatest source of loss.     

Effects of oestradiol and progesterone on the metabolism of [U-14C]glucose by mouse morulae and early blastocysts in vitro

The addition of progesterone (10(-7) to 10(-5) M) and/or oestradiol (10(-10) M) during 24-h chase culture of pulse-labelled morulae-early blastocysts did not affect the degradation of radiolabelled glycogen or other biochemical fractions. The presence of a high concentration of progesterone (10(-5) M) during 5-h pulse culture significantly inhibited incorporation of substrate carbon from [U-14C]glucose into both the acid-soluble and acid-insoluble glycogen fractions, but had no effect on non-glycogen fractions. Catabolic utilization of glucose as estimated by the rate of carbon dioxide and lactate production was not affected by the presence of progesterone (10(-7) to 10(-5) M), oestradiol (10(-10) to 10(-8) M) or a combination of both. The results indicate that ovarian steroids at expected physiological concentrations do not directly influence embryonic energy metabolism.     

Immunochemical analysis and protective properties of components of Pseudomonas aeruginosa mucus with different molecular weights

P. aeruginosa slime has been separated into fractions XM-300 (3 X 10(5) daltons and more), XM-100 (1 X 10(5) = 3 X 10(5) daltons), PM-30 (3 X 10(4) = 1 X 10(5) daltons) and PM-10, (1 X 10(4) = 3 X 10(4) daltons) by ultrafiltration. The high-molecular slime components (3 X 10(4) daltons and more) have been found to be serologically more active than the low-molecular components (1 X 10(4) = 3 X 10(4) daltons). As shown in experiments on mice, both high-molecular toxic and low-molecular nontoxic slime components have protective activity, but the high-molecular components are more active than the low-molecular ones. The slime components stimulate the formation of immunity to homologous and partially heterologous P. aeruginosa strains in mice. Antigenic relationship between the slime components (especially the high-molecular ones) and the corresponding lipopolysaccharides has been noted.     

Effects of dried bonito (katsuobushi) and captopril, an angiotensin I-converting enzyme inhibitor, on rat isolated aorta: a possible mechanism of antihypertensive action

In order to elucidate the mechanism of the antihypertensive action of dried bonito (katsuobushi), we compared the effects of dried bonito extracts with those of captopril, an angiotensin I-converting enzyme (ACE) inhibitor, on aorta preparations isolated from rats. Dried bonito extracts (3 x 10(-4) to 3 x 10(-3) g/ml) more potently relaxed contractions induced by norepinephrine (10(-7) M) than contractions induced by KCl (55.9 mM). Dried bonito extracts (3 x 10(-3) g/ml) slightly inhibited 10(-7) M angiotensin I-induced contractions. In contrast, captopril (10(-8) to 10(-7) M) did not affect 10(-7) M norepinephrine- or 55.9 mM KCl-induced contractions, but a higher concentration of captopril (10(-6) M) very slightly relaxed it. Captopril (10(-8) to 10(-6) M) markedly inhibited 10(-7) M angiotensin I-induced contractions in a concentration-dependent manner. These results suggest that antihypertensive mechanism of action induced by dried bonito involves direct action on vascular smooth muscle in addition to ACE-inhibitory activity.     

Effects of carbon monoxide and heme oxygenase inhibitors in cerebral vessels of rats and mice

Carbon monoxide (CO) has been postulated to be a signaling molecule in many tissues, including the vasculature. We examined vasomotor responses of adult rat and mouse cerebral arteries to both exogenously applied and endogenously produced CO. The diameter of isolated, pressurized, and perfused rat middle cerebral arteries (MCAs) was not altered by authentic CO (10(-6) to 10(-4) M). Mouse MCAs, however, dilated by 21 +/- 10% at 10(-4) M CO. Authentic nitric oxide (NO., 10(-10) to 10(-7) M) dilated both rat and mouse MCAs. At 10(-8) M NO., rat vessels dilated by 84 +/- 4%, and at 10(-7) M NO., mouse vessels dilated by 59 +/- 9%. Stimulation of endogenous CO production through heme oxygenase (HO) with the heme precursor delta-aminolevulinic acid (10(-10) to 10(-4) M) did not dilate the MCAs of either species. The metalloporphyrin HO inhibitor chromium mesoporphyrin IX (CrMP) caused profound constriction of the rat MCA (44 +/- 2% at 3 x 10(-5) M). Importantly, this constriction was unaltered by exogenous CO (10(-4) M) or CO plus 10(-5) M biliverdine (both HO products). In contrast, exogenous CO (10(-4) M) reversed CrMP-induced constriction in rat gracilis arterioles. Control mouse MCAs constricted by only 3 +/- 1% in response to 10(-5) M CrMP. Magnesium protoporphyrin IX (10(-5) M), a weak HO inhibitor used to control for nonspecific effects of metalloporphyrins, also constricted the rat MCA to a similar extent as CrMP. We conclude that, at physiological concentrations, CO is not a dilator of adult rodent cerebral arteries and that metalloporphyrin HO inhibitors have nonspecific constrictor effects in rat cerebral arteries.     

Structure of Marek's disease virus DNA: detailed restriction enzyme map.

Purified virion DNA (120 X 10(6) molecular weight [MW]) of Marek's disease virus strain GA was cleaved with BamHI restriction endonuclease, and 27 out of the 29 fragments were cloned into bacterial plasmids. Restriction maps for BamHI, BglI, and SmaI endonucleases were constructed. The genomic structure of Marek's disease virus DNA was found to be similar to that of herpes simplex virus types 1 and 2. A long unique region (75 X 10(6) MW, located at 10 X 10(6) to 85 X 10(6) MW [10-85] from the left end of the genome), which was subdivided into segment 1 (22 X 10(6) MW, located at 10-32) and segment 2 (51 X 10(6) MW, located at 34-85) by direct repeats (32-34), was flanked by a long terminal region (10 X 10(6) MW, located at 0-10) and a long inverted region (10 X 10(6) MW, located at 85-95). A short unique region (8 X 10(6) MW, located at 103-111) was flanked by a short terminal region (8 X 10(6) MW, located at 111-119) and a short inverted region (8 X 10(6) MW, located at 95-103). The direct repeat fragments (0.9 X 10(6) could be isolated by cleavage with SmaI. The right terminal end was found to be heterogenous .     

Existence of two types of postjunctional alpha-adrenoceptors in the isolated canine internal carotid artery

The pharmacological characteristics of postjunctional alpha-adrenoceptors in isolated canine internal carotid arteries were investigated by the use of selective agonists and antagonists for alpha 1- and alpha 2-adrenoceptors. Norepinephrine, phenylephrine, and xylazine caused concentration-dependent contractions in the helical strips. The contraction induced by 10(-4)M xylazine was significantly smaller than that produced by 10(-4)M norepinephrine or 10(-4)M phenylephrine. The contraction induced by 10(-4)M phenylephrine was almost the same value as that induced by 10(-4)M norepinephrine. Phentolamine (10(-8) and 10(-7)M) caused a parallel shift to the right of the concentration-response curve to norepinephrine. The contractile responses to low concentrations of norepinephrine were significantly suppressed by pretreatment with an alpha 2-antagonist such as yohimbine (10(-9) and 10(-8)M) or DG5128 (10(-7) and 10(-6)M). On the other hand, the responses to higher concentrations of norepinephrine were mainly reduced by low concentrations of an alpha 1-antagonist, prazosin (3 x 10(-10) and 3 x 10(-9)M). These results suggest that both alpha 1- and alpha 2-adrenoceptors are located on the plasma membrane of smooth muscle cells in canine internal carotid arteries and that the norepinephrine-induced contractions at low and high concentrations are mainly mediated by activation of alpha 2- and alpha 1-adrenoceptors, respectively.