Potentiation of dimethylnitrosamine genotoxicity in rat hepatocytes isolated following ethanol treatment in vivo

Unscheduled DNA synthesis (UDS), following exposure to dimethylnitrosamine (DMN), was potentiated in cultured hepatocytes isolated following treatment of rats for 14 or 28 days with 20% ethanol/5% sucrose solution. Ethanol treatment was associated with increased UDS, a concomitant increase in hepatic microsomal protein concentration and DMN N-demethylase activity. Increased aniline hydroxylase activity of hepatic microsomes from ethanol-treated rats preceded the measured increase in microsomal protein content or DMN metabolism. The increase in metabolism of DMN in vitro and potentiation of DMN-induced UDS associated with ethanol treatment may contribute to a synergistic effect of ethanol on DMN hepatotoxicity and carcinogenicity. In contrast, ethanol pretreatment did not increase the cytotoxicity of DMN as characterized by enzyme release.     

The stimulation of hepatic gluconeogenesis by acetoacetate precursors. A role for the monocarboxylate translocator

The regulation of the gluconeogenic pathway from the 3-carbon precursors pyruvate, lactate, and alanine was investigated in the isolated perfused rat liver. Using pyruvate (less than 1 mM), lactate, or alanine as the gluconeogenic precursor, infusion of the acetoacetate precursors oleate, acetate, or beta-hydroxybutyrate stimulated the rate of glucose production and, in the case of pyruvate (less than 1 mM), the rate of pyruvate decarboxylation. alpha-Cyanocinnamate, an inhibitor of the monocarboxylate transporter, prevented the stimulation of pyruvate decarboxylation and glucose production due to acetate infusion. With lactate as the gluconeogenic precursor, acetate infusion in the presence of L-carnitine stimulated the rate of gluconeogenesis (100%) and ketogenesis (60%) without altering the tissue acetyl-CoA level usually considered a requisite for the stimulation of gluconeogenesis by fatty acids. Hence, our studies suggest that gluconeogenesis from pyruvate or other substrates which are converted to pyruvate prior to glucose synthesis may be limited or controlled by the rate of entry of pyruvate into the mitochondrial compartment on the monocarboxylate translocator.     

Valyl-tRNA synthetase modification-dependent restriction of bacteriophage T4

A strain of Escherichia coli, CP 790302, severely restricts the growth of wild-type bacteriophage T4. In broth culture, most infections of single cells are abortive, although a few infected cells exhibit reduced burst sizes. In contrast, bacteriophage T4 mutants impaired in the ability to modify valyl-tRNA synthetase develop normally on this strain. Biochemical evidence indicates that the phage-modified valyl-tRNA synthetase in CP 790302 is different from that previously described. Although the enzyme is able to support normal protein synthesis, a disproportionate amount of phage structural protein (serum blocking power) fails to mature into particles of the appropriate density. The results with host strain CP 790302 are consistent with either a gratuitous inhibition of phage assembly by faulty modification or abrogation of an unknown role that valyl-tRNA synthetase might normally play in viral assembly.     

Small airway pressure-diameter relationships during nonhomogeneous lung lobe inflation

The pressure-diameter behavior of airways within a collaterally ventilating segment of lung was evaluated radiographically in 12 excised dog lung lobes. The results were compared with the pressure-diameter behavior of airways in a lung region adjacent to the collaterally ventilating segment. Airways in each lung region were dusted with powdered tantalum, and airway diameters were measured during homogeneous and nonhomogeneous lobe inflation. Intrasegmental and extrasegmental airways behaved similarly during homogeneous lobe inflation; airway diameter increased as alveolar pressure increased. The lobe was inflated nonhomogeneously by raising pressure in the collaterally ventilating segment (Ps) while maintaining pressure at the lobar bronchus (Pao) constant at 5, 10, or 15 cmH2O. Increasing Ps at constant Pao reciprocally affected intrasegmental and extrasegmental airways. When Pao was low, intrasegmental airways were expanded, and extrasegmental airways were compressed when Ps was raised. When Pao was high, airway diameter was unaffected by increasing Ps presumably because the airways were already maximally expanded. A comparison of diameters during homogenous and nonhomogenous lobe inflation suggests a very small interdependence effect from the parenchyma surrounding the collaterally ventilating segment. These results demonstrate the combined effects of parenchymal properties and airway pressure-diameter relationships in determining the effect of local lung distortion on airway function.     

Evidence that hypoxic pulmonary vascular remodeling in rats is polyamine dependent

This study tested the hypothesis that the polyamines, a family of low-molecular-weight organic cations with documented regulatory roles in cell growth and differentiation, are mediators of chronic hypoxia-induced pulmonary vascular remodeling. Relative to room air controls, chronically hypoxic animals (inspired O2 fraction = 0.1; 21 days) exhibited higher pulmonary arterial pressures (measured in room air), thicker medial layers in pulmonary arteries of 50-100 microns diam, increased hematocrits, and right ventricular hypertrophy. In addition, lung contents of the polyamines, putrescine, spermidine, and spermine were greater in hypoxic animals than in controls. alpha-Difluoromethylornithine (DFMO), an inhibitor of polyamine synthesis, attenuated the hypoxia-induced elevations in lung putrescine and spermidine content and blunted the increases in pulmonary arterial pressure and medial thickness. Neither the increased hematocrit nor right ventricular hypertrophy associated with chronic hypoxia were abrogated by DFMO. In addition, DFMO failed to influence vasoconstrictor responses provoked by acute hypoxic ventilation in isolated, buffer-perfused rat lungs. These observations suggest that depression of polyamine biosynthesis with DFMO blunts the sustained increase in pulmonary arterial pressure by attenuating hypoxia-induced medial thickening.     

The effect of monensin on gametogenesis and zoosporogenesis in the aquatic fungus,Allomyces macrogynus

Summary Cytoplasmic cleavage in the gametangia and zoosporangia ofA. macrogynus was studied using monensin, an ionophore known to disrupt several endomembrane functions in plant and animal cells. Monensin interfered with normal gamete and zoospore formation in a dose dependent manner such that at a 20 M concentration very abnormal cells were released from the reproductive structures. It was evident that monensin's effect was most pronounced during the first 25 minutes of gametogenesis and parallels in time the onset and continuation of the cytoplasmic cleavage events. Observations using fluorescence and differential interference contrast microscopy indicated that the ionophore inhibited normal cytoplasmic cleavage resulting in the production of multinucleate cells, many of which had either no flagella or multiple flagella. Transmission electron microscopy showed that the monensin-treated gametangia had many large vacuoles which contained amorphous electron-opaque material. X-ray microprobe analysis demonstrated that the elemental composition of the large vacuoles was identical to that of the dense globular inclusions seen in untreated gametangia, and morphological analysis confirmed the relationship between these endomembrane structures. Thus this swollen endomembrane component probably is not involved in the cleavage process. Single endomembrane cisternae which were very common in untreated gametangia were seldom seen in monensin-treated preparations. Instead, many smaller electron-transparent vacuoles were observed. These swollen cisternae may both represent monensin-modified Golgi apparatus equivalents and/or play a critical role during the process of gametogenesis and zoosporogenesis inA. macrogynus.     

Effects of light and nitrogen limitation on the cell cycle of the dinoflagellate Amphidinium carteri

Cell cycle phase durations of cultures of Amphidinium carteriin light- or nitrogen-limited balanced growth were determinedusing flow cytometry. For both types of growth rate limitation,the increases in generation time caused by increasing degreesof limitation were due solely to expansion of the G₁ phase ofthe cell cycle. The durations of the S and G₂ + M phases wereindependent of growth rate. Furthermore, when cells were deprivedcompletely of light and nitrogen, they arrested in the G₁ phaseof the cell cycle. The results indicate that light- and nitrogen-dependentprocesses are heavily concentrated in the early part of thecell cycle, while DNA replication and cell division, once initiated,are independent of light or nitrogen supply.     

Stimulation of glycogenolysis by adenine nucleotides in the perfused rat liver.

Infusion of adenine nucleotides and adenosine into perfused rat livers resulted in stimulation of hepatic glycogenolysis, transient increases in the effluent perfusate [3-hydroxybutyrate]/[acetoacetate] ratio, and increased portal vein pressure. In livers perfused with buffer containing 50 microM-Ca2+, transient efflux of Ca2+ was seen on stimulation of the liver with adenine nucleotides or adenosine. ADP was the most potent of the nucleotides, stimulating glucose output at concentrations as low as 0.15 microM, with half-maximal stimulation at approx. 1 microM, and ATP was slightly less potent, half-maximal stimulation requiring 4 microM-ATP. AMP and adenosine were much less effective, doses giving half-maximal stimulation being 40 and 20 microM respectively. Non-hydrolysed ATP analogues were much less effective than ATP in promoting changes in hepatic metabolism. ITP, GTP and GDP caused similar changes in hepatic metabolism to ATP, but were 10-20 times less potent than ATP. In livers perfused at low (7 microM) Ca2+, infusion of phenylephrine before ATP desensitized hepatic responses to ATP. Repeated infusions of ATP in such low-Ca2+-perfused livers caused homologous desensitization of ATP responses, and also desensitized subsequent Ca2+-dependent responses to phenylephrine. A short infusion of Ca2+ (1.25 mM) after phenylephrine infusion restored subsequent responses to ATP, indicating that, during perfusion with buffer containing 7 microM-Ca2+, ATP and phenylephrine deplete the same pool of intracellular Ca2+, which can be rapidly replenished in the presence of extracellular Ca2+. Measurement of cyclic AMP in freeze-clamped liver tissue demonstrated that adenosine (150 microM) significantly increased hepatic cyclic AMP, whereas ATP (15 microM) was without effect. It is concluded that ATP and ADP stimulate hepatic glycogenolysis via P2-purinergic receptors, through a Ca2+-dependent mechanism similar to that in alpha-adrenergic stimulation of hepatic tissue. However, adenosine stimulates glycogenolysis via P1-purinoreceptors and/or uptake into the cell, at least partially through a mechanism involving increase in cyclic AMP. Further, the hepatic response to adenine nucleotides may be significant in regulating hepatic glucose output in physiological and pathophysiological states.     

The level of nerve growth factor (NGF) as a function of innervation : A correlative radio-immunoassay and bioassay study of the rat iris

A two-site radio-immunoassay for βNGF demonstrated 5–10 pg of NGF in the normal, adult rat iris. Ciliarectomy or sympathectomy did not significantly alter the amount of NGF after 10 days. However, denervation including all sensory axons (stereotactic lesion distal to the trigeminal ganglion) increased the level to about 100 pg of NGF. Total denervation resulting from homologous transplantation of the iris gave a similar increase after only 2 days. Fibre outgrowth responses evoked by corresponding iris explants in an NGF bioassay supported the results and suggested in addition that sympathetic denervation may cause a moderate transient increase in NGF after 3 days. It seems that sensory nerves in particular influence the level of NGF in a terminal field, either by a high capacity for uptake and removal of NGF or by exerting a negative feed-back on the production or processing of this growth factor.     

Mutagen sensitivity of Neurospora meiotic mutants

The Neurospora meiotic mutants, mei-1, mei-2, mei-3, and mei-4 were tested for cross sensitivity to mutagens. Mei-2 and mei-3 are sensitive to MMS, gamma-irradiation and histidine. Mei-2 is not sensitive to ultraviolet (UV) at 20 degrees C. Tests with recombinants with crossovers to either side of mei-2 or mei-3 show that these traits are the pleiotropic properties of a single gene which also determines meiotic behavior. The mei-2 gene maps to the right of al-3 on linkage group V. It is not allelic to mus-11. Upon backcrossing, the originally dominant meiotic effect of mei-2 becomes recessive to partially dominant. The histidine sensitivity is recessive.