A monoclonal antibody to bovine brain inositol monophosphatase. Immunoaffinity purification of the brain and kidney enzymes and evidence for their structural identity.

A monoclonal IgG2b(K) antibody, G-2A4, has been generated against bovine brain myo-inositol monophosphatase (EC 3.1.3.25). The identity of the antigen recognized by the antibody was established by using e.l.i.s.a. and Western blotting procedures, and by immunoprecipitation of enzyme activity from crude brain supernatant. In addition, the hydrolysis of Ins1P by crude brain extract was inhibited by up to 83% by the pure antibody. Under identical conditions, the hydrolysis of Ins(1,4)P2 was unaffected. An immunoadsorbent column containing monoclonal antibody G-2A4 covalently attached to CNBr-activated Sepharose 4B has been used for rapid purification of the brain enzyme. Elution conditions have been optimized to allow isolation of the enzyme in high yield (54%) with full retention of column-binding capacity. The enzyme was electrophoretically homogeneous, Mr 30,000 and of higher specific activity than that purified conventionally. Chromatography of the pure enzyme on high resolution ion-exchange columns revealed some charge heterogeneity, possibly indicative of some type of post-translational modification. The immunoadsorbent column has also been used to purify the bovine kidney cortex enzyme to homogeneity. Partial proteolytic fragmentation patterns of the brain and kidney enzymes using endoprotease glu-C were identical, suggesting that they are almost certainly products of the same gene.     

The dephosphorylation of inositol 1,4-bisphosphate to inositol in liver and brain involves two distinct Li+-sensitive enzymes and proceeds via inositol 4-phosphate.

1. Hydrolysis of both enantiomers of inositol 1-phosphate and both enantiomers of inositol 4-phosphate to inositol is inhibited by LiCl in liver and brain. 2. The phosphatase activity is predominantly soluble. 3. Inositol 1,4-bisphosphate is also hydrolysed by the soluble fraction of liver and brain. 4. Bisphosphatase activity is inhibited by LiCl, but is less sensitive than monophosphatase activity. 5. The product of bisphosphatase in liver and brain is inositol 4-phosphate.     

Inhibition by Cyclic AMP of Phorbol Ester-Potentiated Norepinephrine Release from Guinea Pig Brain Cortical Synaptosomes

The involvement of Ca2+/phospholipid-dependent protein kinase (protein kinase C, PKC) and cyclic AMP-dependent protein kinase in the K+-evoked release of norepinephrine (NE) was studied using guinea pig brain cortical synaptosomes preloaded with [3H]NE. 12-O-Tetradecanoylphorbol-13-acetate (TPA), a potent activator of PKC, enhanced the K+-evoked release of [3H]NE, in a concentration-dependent manner, but with no effect on the spontaneous outflow and uptake of [3H]NE in the synaptosomes. The apparent affinity of the evoked release for added calcium but not the maximally evoked release was increased by TPA (10(-7) M). Inhibitors of PKC, polymyxin B, and a more potent inhibitor, staurosporine, counteracted the TPA-induced potentiation of the evoked release. Both forskolin and dibutyryl cyclic AMP (DBcAMP) enhanced the evoked release, but reduced the TPA-potentiated NE release. A novel inhibitor of cyclic AMP-dependent protein kinase, KT5720, blocked both the forskolin-induced increase in the evoked release and its inhibition of TPA-induced potentiation in the evoked release, thereby suggesting that forskolin or DBcAMP counteracts the Ca2+-dependent release of NE by activating cyclic AMP-dependent protein kinase. These results suggest that the activation of PKC potentiates the evoked release of NE and that the activation of cyclic AMP-dependent protein kinase acts negatively on the PKC-activated exocytotic neurotransmitter release process in brain synaptosomes of the guinea pig.     

Relationship of serum total calcium and albumin and total protein in owl monkeys (Aotus nancymai)

A positive linear relationship was found between total calcium and albumin and between total calcium and total protein in the serum of 205 owl monkeys. Adjustment formulas for calcium were derived: adjusted calcium (mg/dl) = measured calcium (mg/dl) - 0.84 [albumin (g/dl)] + 3.8 and adjusted calcium (mg/dl) = measured calcium (mg/dl) - 0.82 [total protein (g/dl)] + 6.5. Adjusted calcium calculations for monkeys based on albumin concentration were similar to those for man and dogs, but different from those based on total protein.     

Mammal use of riparian corridors in semi-arid Sonora,Mexico

Wildlife populations in semi-arid regions are increasingly challenged by human activities and dependent on the connectivity of riparian corridors for access to surface water. The Madrean Archipelago is a biodiversity hotspot along the arid United States–Mexico borderlands that support both Neotropical and Nearctic wildlife. Infrastructure development (e.g., the border wall and the expansion of Mexican Federal Highway 2) in this region inhibits wildlife movement along the transnational mountain archipelago by disconnecting habitat. To explore the relationship between habitat variables and mammal use of riparian corridors in northern Sonora, Mexico, we collected data from 19 motion-sensitive cameras between October 2018 and April 2019 and used single-season occupancy models and Royle-Nichols abundance estimation models to analyze our data. We recorded 21 species of mammals, including the first sighting of jaguar (Panthera onca) in this region in 25 years. River characteristics (distance from river, riparian corridor width, water availability), remoteness (distance from highway, productivity, elevation), and topographic variety (vertical elevation difference) influenced patterns of occupancy probability and estimated abundance of mammals >1 kg, but the strength and direction of these relationships varied by species. Additionally, intermittently wet desert washes were comparable in species richness to the perennial system. These results highlight the importance of examining physical and biological aspects of habitat. This is especially true when identifying corridors where mitigation structures should be placed to improve wildlife connectivity in biodiversity hotspots like the Madrean Archipelago and semi-arid ecosystems worldwide.     

Membrane-bound glycoconjugates of fetal mouse erythropoietic cells with special reference to phagocytosis by hepatic macrophages

Using lectin and colloidal iron (CI) stainings in combination with neuraminidase digestion, glycoconjugates on the surface of erythropoietic cells of the yolk sac and liver in fetal mice were examined. Fetal hepatic macrophages were capable of distinguishing between phagocytozed and non-phagocytozed erythroid elements as described in our previous study. Marked differences between these two elements could be ultrahistochemically detected on their cell surface. The phagocytozed elements, such as nuclei expelled from erythroblasts and degenerating primitive erythroblasts, faintly bound neuraminidase-sensitive CI, and neuraminidase digestion imparted a weak peanut agglutinin (PNA) binding. In contrast, erythroblasts at various maturation stages, erythrocytes and normal primitive erythroblasts heavily bound neuraminidase-sensitive CI, and neuraminidase digestion imparted a moderate PNA binding. No differences in binding of either concanavalin agglutinin,Ricinus communis agglutinin-I or PNA were noted between phagocytozed and non-phagocytozed erythroid elements. Desialylation appears to be one of the most important signs for the recognition mechanism of fetal macrophage phagocytosis. During maturation of hepatic erythroblasts, sialic acid changes its affinity forLimax flavus agglutinin from strong to weak, and soybean agglutinin binding sites disappear at the basophilic erythroblast stage. Glycoconjugates on polychromatophilic erythroblasts acquire similar compositions to those of erythrocytes.     

Quantitative studies on brown algal phenols. I. Estimation of absolute polyphenol content of Ascophyllum nodosum (L.) Le Jol. and Fucus vesiculosus (L.)

Methods for estimating the absolute polyphenol content of the brown algae Ascophyllum nodosum (L.) Le Jol. and Fucus vesiculosus (L.) have been developed and tested. Polyphenols were extracted almost quantitatively from Ascophyllum nodosum using aqueous acetone, whereas this procedure was somewhat less efficient with Fucus vesiculosus. Colorimetric methods based on the Folin-Denis reagent, Brentamine Fast Red 2G Salt, and vanillin-H₂SO₄ were applied to acetone-free extracts for determination of polyphenol content relative to suitable reference compounds. Gravimetric methods based on hide powder and on haemoglobin were employed to derive ‘estimation factors’ (EFs) which allow calculation of the absolute polyphenol content from the relative polyphenol content. The values calculated for absolute polyphenol content are considered to be reasonably accurate, despite imprecisions in the methods and despite often large standard deviations, and re-emphasize the potential physiological and ecological significance of brown algal polyphenols. Although the precise EFs calculated here are not valid for other brown algae, the methods are considered to be generally applicable to other Phaeophyceae.