Normal hepatic glucose production in the absence of GLUT2 reveals an alternative pathway for glucose release from hepatocytes

Glucose production by liver is a major physiological function, which is required to prevent development of hypoglycemia in the postprandial and fasted states. The mechanism of glucose release from hepatocytes has not been studied in detail but was assumed instead to depend on facilitated diffusion through the glucose transporter GLUT2. Here, we demonstrate that in the absence of GLUT2 no other transporter isoforms were overexpressed in liver and only marginally significant facilitated diffusion across the hepatocyte plasma membrane was detectable. However, the rate of hepatic glucose output was normal. This was evidenced by (i) the hyperglycemic response to i.p. glucagon injection; (ii) the in vivo measurement of glucose turnover rate; and (iii) the rate of release of neosynthesized glucose from isolated hepatocytes. These observations therefore indicated the existence of an alternative pathway for hepatic glucose output. Using a [14C]-pyruvate pulse-labeling protocol to quantitate neosynthesis and release of [14C]glucose, we demonstrated that this pathway was sensitive to low temperature (12°C). It was not inhibited by cytochalasin B nor by the intracellular traffic inhibitors brefeldin A and monensin but was blocked by progesterone, an inhibitor of cholesterol and caveolae traffic from the endoplasmic reticulum to the plasma membrane. Our observations thus demonstrate that hepatic glucose release does not require the presence of GLUT2 nor of any plasma membrane glucose facilitative diffusion mechanism. This implies the existence of an as yet unsuspected pathway for glucose release that may be based on a membrane traffic mechanism.

Fluxes of Reserve-Derived and Currently Assimilated Carbon and Nitrogen in Perennial Ryegrass Recovering from Defoliation. The Regrowing Tiller and Its Component Functionally Distinct Zones1

The quantitative significance of reserves and current assimilates in regrowing tillers of severely defoliated plants of perennial ryegrass (Lolium perenne L.) was assessed by a new approach, comprising 13C/12C and 15N/14N steady-state labeling and separation of sink and source zones. The functionally distinct zones showed large differences in the kinetics of currently assimilated C and N. These are interpreted in terms of ”substrate” and ”tissue” flux among zones and C and N turnover within zones. Tillers refoliated rapidly, although C and N supply was initially decreased. Rapid refoliation was associated with (a) transient depletion of water-soluble carbohydrates and dilution of structural biomass in the immature zone of expanding leaves, (b) rapid transition to current assimilation-derived growth, and (c) rapid reestablishment of a balanced C:N ratio in growth substrate. This balance (C:N, approximately 8.9 [w/w] in new biomass) indicated coregulation of growth by C and N supply and resulted from complementary fluxes of reserve- and current assimilation-derived C and N. Reserves were the dominant N source until approximately 3 d after defoliation. Amino-C constituted approximately 60% of the net influx of reserve C during the first 2 d. Carbohydrate reserves were an insignificant source of C for tiller growth after d 1. We discuss the physiological mechanisms contributing to defoliation tolerance.

Oceanic minerals: Their origin, nature of their environment, and significance

The chemical and isotopic compositions of oceanic biogenic and authigenic minerals contain invaluable information on the evolution of seawater, hence on the history of interaction between tectonics, climate, ocean circulation, and the evolution of life. Important advances and greater understanding of (a) key minor and trace element cycles with various residence times, (b) isotopic sources and sinks and fractionation behaviors, and (c) potential diagenetic problems, as well as developments in high-precision instrumentation, recently have been achieved. These advances provided new compelling evidence that neither gradualism nor uniformitarianism can explain many of the new important discoveries obtained from the chemistry and isotopic compositions of oceanic minerals. Presently, the best-developed geochemical proxies in biogenic carbonates are 18O/16O and Sr/Ca ratios (possibly Mg/Ca) for temperature; 87Sr/86Sr for input sources, Cd/Ca and Ba/Ca ratios for phosphate and alkalinity concentrations, respectively, thus also for ocean circulation; 13C/12C for ocean productivity; B isotopes for seawater pH;, U, Th isotopes, and 14C for dating; and Sr and Mn concentrations for diagenesis. The oceanic authigenic minerals most widely used for chemical paleoceanography are barite, evaporite sulfates, and hydrogenous ferromanganese nodules. Marine barite is an effective alternative monitor of seawater 87Sr/86Sr, especially where carbonates are diagenetically altered or absent. It also provides a high-resolution record of seawater sulfate S isotopes, (evaporite sulfates only carry an episodic record), with new insights on factors affecting the S and C cycles and atmospheric oxygen. High-resolution studies of Sr, Nd, and Pb isotopes of well-dated ferromanganese nodules contain invaluable records on climate driven changes in oceanic circulation.

Transport of Sterols to the Plasma Membrane of Leek Seedlings1

To investigate the intracellular transport of sterols in etiolated leek (Allium porrum L.) seedlings, in vivo pulse-chase experiments with [1-14C]acetate were performed. Then, endoplasmic reticulum-, Golgi-, and plasma membrane (PM)-enriched fractions were prepared and analyzed for the radioactivity incorporated into free sterols. In leek seedlings sterols are present as a mixture in which (24R)-24-ethylcholest-5-en-3β-ol is by far the major compound (around 60%). The other sterols are represented by cholest-5-en-3β-ol, 24-methyl-cholest-5-en-3β-ol, (24S)-24-ethylcholesta-5,22E-dien-3β-ol, and stigmasta-5,24(241)Z-dien-3β-ol. These compounds are shown to reside mainly in the PM. Our results clearly indicate that free sterols are actively transported from the endoplasmic reticulum to the PM during the first 60 min of chase, with kinetics very similar to that of phosphatidylserine. Such a transport was found to be decreased at low temperature (12°C) and following treatment with monensin and brefeldin A. These data are consistent with a membrane-mediated process for the intracellular transport of sterols to the PM, which likely involves the Golgi apparatus.

Dynamics of maize endosperm development and DNA endoreduplication.

Endosperm development in Zea mays is characterized by a period of intense mitotic activity followed by a period in which mitosis is essentially eliminated and the cell cycle becomes one of alternating S and G phases, leading to endoreduplication of the nuclear DNA. The endosperm represents a significant contribution to the grain yield of maize; thus, methods that facilitate the study of cellular kinetics may be useful in discerning cellular and molecular components of grain yield. Two mathematical models have been developed to describe the kinetics of endosperm growth. The first describes the kinetics of mitosis during endosperm development; the second describes the kinetics of DNA endoreduplication during endosperm development. The mitotic model is a modification of standard growth curves. The endoreduplication model is composed of six differential equations that represent the progression of nuclei from one DNA content to another during the endoreduplication process. Total nuclei number per endosperm and the number of 3C, 6C, 12C, 24C, 48C, and 96C nuclei per endosperm (C is the haploid DNA content per nucleus) for inbred W64A from 8 to 18 days after pollination were determined by flow cytometry. The results indicate that the change in number of nuclei expressed as a function of the number of days after pollination is the same from one yearly crop to another. These data were used in the model to determine the endosperm growth rate, the maximum nuclei number per endosperm, and transition rates from one C value to the next higher C value. The kinetics of endosperm development are reasonably well represented by the models. Thus, the models provide a means to quantify the complex pattern of endosperm development.