Electron microscopic determination of phenoloxidase (EC 1.14.18.1) in eosinophilic granulocytes of the small intestine of white rats

It is shown that the light microscopic permanent detectable phenoloxidase activity, using dihydroxyphenylalanine as substrate, of the small intestine of white rats is localized in the most cases in eosinophilic granulocytes. The enzyme has been found as well in the cytoplasma as in the granules. The enzyme proof triggered in the granules the so called reverse effect. The results allow to conclude new aspects for the cell mediated immunological defense reaction.     

Plant–animal interactions between carnivorous plants,sheet‐web spiders,and ground‐running spiders as guild predators in a wet meadow community

1. Plant–animal interactions are diverse and widespread shaping ecology, evolution, and biodiversity of most ecological communities. Carnivorous plants are unusual in that they can be simultaneously engaged with animals in multiple mutualistic and antagonistic interactions including reversed plant–animal interactions where they are the predator. Competition with animals is a potential antagonistic plant–animal interaction unique to carnivorous plants when they and animal predators consume the same prey.
2. The goal of this field study was to test the hypothesis that under natural conditions, sundews and spiders are predators consuming the same prey thus creating an environment where interkingdom competition can occur.
3. Over 12 months, we collected data on 15 dates in the only protected Highland Rim Wet Meadow Ecosystem in Kentucky where sundews, sheet‐web spiders, and ground‐running spiders co‐exist. One each sampling day, we attempted to locate fifteen sites with: (a) both sheet‐web spiders and sundews; (b) sundews only; and (c) where neither occurred. Sticky traps were set at each of these sites to determine prey (springtails) activity–density. Ground‐running spiders were collected on sampling days. DNA extraction was performed on all spiders to determine which individuals had eaten springtails and comparing this to the density of sundews where the spiders were captured.
4. Sundews and spiders consumed springtails. Springtail activity–densities were lower, the higher the density of sundews. Both sheet‐web and ground‐running spiders were found less often where sundew densities were high. Sheet‐web size was smaller where sundew densities were high.
5. The results of this study suggest that asymmetrical exploitative competition occurs between sundews and spiders. Sundews appear to have a greater negative impact on spiders, where spiders probably have little impact on sundews. In this example of interkingdom competition where the asymmetry should be most extreme, amensalism where one competitor experiences no cost of interaction may be occurring.

     

Acyltransferase-catalyzed cleavage of arachidonic acid from phospholipids and transfer to lysophosphatides in lymphocytes and macrophages

The cleavage of fatty acyl moieties from phospholipids was compared in intact cells and homogenates of mouse lymphocytes (thymocytes, spleen cells) and macrophages. Liberation of free arachidonic acid during incubations of intact cells was only detectable in the presence of albumin. Homogenization of prelabeled thymocytes and further incubation of these homogenates at 37 degrees C resulted in a pronounced decrease of phospholipid degradation and cleavage of arachidonoyl residues, while further incubation of homogenates from prelabeled macrophages produced a greatly increased phospholipid degradation. Homogenates of macrophages but not those of thymocytes contain substantial activities of phospholipase A2 detectable using exogenous radiolabeled substrates. These findings indicate that in thymocytes cleavage of arachidonic acid from phosphatidylcholine is an active process that is not catalyzed by phospholipase A2. Addition of CoA and lysophosphatidylethanolamine to prelabeled thymocyte homogenates induced a fast breakdown of phosphatidylcholine and transfer of arachidonic acid to phosphatidylethanolamine, as in seen during incubations of intact thymocytes or macrophages. The transfer is restricted to arachidonic acid and does not require addition of ATP. Sodium cholate, a known inhibitor of the acyl-CoA:lysophosphatide acyltransferase, completely inhibited this transfer reaction. These results suggest that the CoA-mediated, ATP-independent breakdown of phosphatidylcholine and transfer of arachidonic acid is catalyzed by the acyl-CoA:lysophosphatide acyltransferase operating in reverse.     

Evaluation of the silica microbead method for isolation of red beet protoplast plasma membrane sheets

The silica microbead procedure was utilized for the isolation of plasma membrane sheets from protoplasts of a higher plant, the red beet (Beta vulgaris L.). Membrane yields, as determined by recovery of an exogenous membrane marker were approx. 75%. The plasma membrane fraction contained the enzyme marker, pH 6.5, vanadate-sensitive, K+-stimulated, Mg2+-ATPase and small amounts of mitochondria, endoplasmic reticulum, and possibly tonoplast. The silica microbead procedure was also used for the isolation of intact vacuoles from microbead-coated protoplasts.     

Dissection and Culture of Mouse Dopaminergic and Striatal Explants in Three-Dimensional Collagen Matrix Assays

Midbrain dopamine (mdDA) neurons project via the medial forebrain bundle towards several areas in the telencephalon, including the striatum¹. Reciprocally, medium spiny neurons in the striatum that give rise to the striatonigral (direct) pathway innervate the substantia nigra². The development of these axon tracts is dependent upon the combinatorial actions of a plethora of axon growth and guidance cues including molecules that are released by neurites or by (intermediate) target regions^3,4. These soluble factors can be studied in vitro by culturing mdDA and/or striatal explants in a collagen matrix which provides a three-dimensional substrate for the axons mimicking the extracellular environment. In addition, the collagen matrix allows for the formation of relatively stable gradients of proteins released by other explants or cells placed in the vicinity (e.g. see references 5 and 6). Here we describe methods for the purification of rat tail collagen, microdissection of dopaminergic and striatal explants, their culture in collagen gels and subsequent immunohistochemical and quantitative analysis. First, the brains of E14.5 mouse embryos are isolated and dopaminergic and striatal explants are microdissected. These explants are then (co)cultured in collagen gels on coverslips for 48 to 72 hours in vitro. Subsequently, axonal projections are visualized using neuronal markers (e.g. tyrosine hydroxylase, DARPP32, or βIII tubulin) and axon growth and attractive or repulsive axon responses are quantified. This neuronal preparation is a useful tool for in vitro studies of the cellular and molecular mechanisms of mesostriatal and striatonigral axon growth and guidance during development. Using this assay, it is also possible to assess other (intermediate) targets for dopaminergic and striatal axons or to test specific molecular cues.