An ultrastructural study of embryonic chick retinal neurons in culture

Summary The differentiation of cells and synapses in explants of 9-day-old chick embryo retina has been studied by light and electron microscopy over a period of 35 days in vitro, and samples of retina from the 9-day chick foetus were directly fixed and prepared for study.At the time of explantation the retinae were poorly differentiated and no lamination was apparent. From day 14 onwards, (i) outer and inner nuclear layers (ONL, INL) separated by a layer of neuropil corresponding to the outer plexiform layer (OPL) and (ii) a layer of scattered large ganglion cells separated from the INL by a zone of neuropil resembling the inner plexiform layer (IPL) were apparent, and (iii) a well-differentiated outer limiting membrane was established close to the surface of the explants. In the oldest cultures some development of photoreceptor outer segments occurred but a distinct optic nerve fibre layer did not form.Although cell identification presented problems even in the oldest cultures, the major retinal cell types described in vivo could be identified. Photoreceptor cells developed pedicles in the OPL which became filled with synaptic vesicles and synaptic ribbons and established ribbon synapses (including triads) with and were commonly invaginated by processes from horizontal and bipolar cells. Processes of bipolar cells in the IPL formed simple and dyad synapses. At least two types of presynaptic amacrine cells were also identified in the INL, one of which contained large numbers of dense-core vesicles. The ganglion cells, though sparse, were large and well differentiated.These findings show that all the major neuronal types of the retina are capable of developing and differentiating in vitro, lagging behind the time-table of development and differentiation in vivo by approximately 7 days, but resulting in a histotypically organised retina with synaptic neuropil showing many similarities to the corresponding neuropil in vivo.     

Vertical community structure and resource utilization in neotropical forest cockroaches

ABSTRACT.

• 1 Patterns of vertical habitat use of ten species of cockroaches are examined. Three assemblages of cockroaches are recognized on the basis of morphology, foraging behaviour, foraging heights, and overlaps and breadths of vertical distributions.
• 2 Three apterous and brachypterous species occur near the ground and comprise one assemblage. They feed mainly on material in the leaf-litter.
• 3 Species that perch higher either migrate into the leaf-litter on a diel basis and feed on both leaf-litter and epiphyllic materials, or some are strictly arboreal and forage on algae, liverworts, lichens, spores, pollen and trichomes on the surfaces of leaves.
• 4 Trophic and behavioural correlations with perch height are described and the functions of perching are examined.
• 5 We conclude that studies of interactions among species are confounded by our lack of understanding of stage- and sex-specific interactions of coexisting species. A simplistic ‘species’approach to such interactions is inadequate because it does not recognize intraspecific variation.

     

Dormancy inDioscorea: Different temperature adaptation of seeds,bulbils and subterranean organs in relation to north-south distribution

Temperature dependencies of sprouting and germination were compared for subterranean perennating organs and seeds of ten closely related species of the genusDioscorea (Dioscoreaceae), a group of monocotyledonous summer perennials which are distributed from the tropics to the subarctic. The species used wereD. nipponica Makino,D. tokoro Makino,D. japonica Thunb.,D. tenuipes Franch. et Savat.,D. septemloba Thunb.,D. quinqueloba Thunb.,D. izuensis Akahori,D. bulbifera L. f.spontanea (Makino) Makino et Nemoto,D. pentaphylla. L. andD. alata L.; they are distributed from cold northern areas to warmer southern areas approximately in this order in and around Japan. Bulbil sprouting was also studied in those forming bulbils. Subterranean organs of the tropical species sprouted faster without any prior temperature treatment, whereas those of species from the more northern areas sprouted after prechilling. Northern species required longer, periods of prechilling for sprouting. On the other hand, with seeds or bulbils, the southern species required longer periods of prior temperature treatment for dormancy breaking. This difference in the length of dormant periods between seeds or bulbils and subterranean organs among the ten species may be related to their size and position of shedding; seeds or bulbils are small and are shed on the ground surface, whereas subterranean organs are large and are located below the surface. It is important to determine in other perennials whether the above relation between dormant features of seeds or bulbils and subterranean organs are common properties or not.     

Studies on the lack of cooperativity in the melting of lipid bilayers

The gel-to-fluid first-order melting transition of lipid bilayers is simulated by the use of a microscopic interaction model which includes a variable number of lipid-chain conformational states. The results suggest that the experimental observation of ‘continuous melting’ in pure wet lipid bilayers, rather than being ascribed to the presence of impurities, may be explained as a result of kinetically caused metastability of intermediate lipid-chain conformations.     

Changes in membrane phospholipid distribution during platelet activation

(1) Exposure of phospholipids at the outer surface of activated and control platelets was studied by incubation with a mixture of phospholipase A₂ from Naja naja and bee venom, solely or in combination with sphingomyelinase from Staphylococcus aureus, using conditions under which cell lysis remained below 10%. (2) Incubation with phospholipase A₂ alone revealed a markedly increased susceptibility of the phospholipids in platelets activated by a mixture of collagen plus thrombin, by the SH-oxydizing compound diamide, or by calcium ionophore A23187, as compared to control platelets or platelets activated separately by collagen or thrombin. (3) Collagen plus thrombin, diamide, and ionophore treated platelets revealed an increased exposure of phosphatidylserine at the outer surface accompanied by a decreased exposure of sphingomyelin, as could be concluded from incubations with a combination of phospholipase A₂ and sphingomyelinase. These alterations were much less apparent in platelets activated either by thrombin or by collagen alone. (4) The increased exposure of phosphatidylserine in activated platelets is accompanied by an increased ability of the platelets to enhance the conversion of prothrombin to thrombin by coagulation factor Xa, in the presence of factor Va and calcium. (5) It is concluded that the altered orientation of the phospholipids in the plasma membrane of platelets activated by collagen plus thrombin, by diamide, or by calcium ionophore, is the result of a transbilayer movement. Moreover, the increased exposure of phosphatidylserine in platelets stimulated by the combined action of collagen and thrombin might be of considerable importance for the hemostatic process.     

Characteristics and determinants of osmotic lysis in chromaffin granules

(1) Using isolated bovine chromaffin granules, we demonstrate that osmotic lysis is not a random process and establish the osmotic pressure dependence of osmotic lysis in chromaffin granules, the so-called osmotic fragility curve. (2) We show by measuring the release of constituents of the granule core and correlating these with changes in spectroscopic parameters (turbidity and endogenous catecholamine fluorescence), that the latter can be safely used to measure lysis. (3) Within a particular granule population, noradrenaline granules lyse at higher osmolarities than adrenaline granules, suggesting a higher core osmolarity of the noradrenaline granules. (4) The size distribution of chromaffin granules as a function of lysis was determined by the use of whole mount electron microscopy. It is shown that the mean size of chromaffin granules decreases as a function of lysis. (5) On the basis of theoretical considerations three alternative models of the sequence of osmotic lysis in chromaffin granules are proposed. The experimental results best support a model which postulates that during partial osmotic lysis, granule membranes reseal into smaller vesicles after graded release of contents. The osmotic fragility would represent several cycles of lysis and resealing and would not be a reflection of the distribution of osmotic pressures in the granule population.     

Cell Surface Sialoglycoproteins of Cultured Rat Cerebellar Interneurons

Abstract: The sialoglycoproteins of cultures of relatively pure rat cerebellar interneurons were labelled by NaIO₄ oxidation/NaB ³H₄ reduction. The labelled molecules were analysed by polyacrylamide gel electrophoresis in sodium dodecyl sulphate followed by fluorography. Faint labelling could be detected in three components if cells were labelled without any oxidation. In young cultures, oxidation by galactose oxidase alone failed to reveal any additional bands. After oxidation by NaIO₄ or galactose oxidase in the presence of neuraminidase, many more components were labelled. After NaIO₄ oxidation, about 80% of the cell-associated radioactivity could be removed by treating the cells with neuraminidase, which left the cells more than 95% viable. The majority of the bands seen after neuraminidase treatment were substantially reduced when compared with untreated controls, supporting a surface localisation of these molecules. Reproducible developmental changes were seen in the profiles of bands labelled by NaIO₄/NaB ³H₄ in time course studies of cultures up to 8 days in vitro . Some bands became more prominent, and others disappeared. The gel profiles of the neuron cultures were quite distinct from those of cerebellar astrocyte cultures, which contain all the cell types likely to be contaminants of the neuron cultures.     

Root growth and litter decomposition in a coffee plantation under shade trees

In a non-fertilized coffee plantation under shade trees the root biomass was excavated to estimate its distribution in the soil profile. One third of total fine (less than 1 mm) roots was found in the first 10 cm of soil; the cumulative total to 30 cm reached 73%. A highly variable and transient amount of fine roots colonized the litter layer. Root production both in the litter and in the first 7.5 cm of mineral soil was estimated from sequential samplings and was 10 g m^–2 yr^–1 and 660 g m^–2 yr^–1 respectively. The decomposition rate of weighed averages of litter fractions in the coffee plantation, calculated as the ratio of litter fall rate to the amount found in the soil was k=4.8. Shade tree leaves, the major component of litter descomposed slower than coffee leaves and these slower than flowers and fruits. Litter bag experiments showed considerable slower rates when mesh was 0.03 mm than 0.5 mm. Nitrogen and phosphorous showed increases in concentrations as decomposition progressed while potassium, calcium and magnesium followed a decrease in concentration that paralleled that of dry weight loss. In comparing the decomposition rate for litter with or without coffee roots growing in the bags, a tendency to show faster decomposition rates was found for the treatment with roots. These differences were however, only significant for one month for shade tree leaves litter. Nitrogen amounts remaining in shade tree leaves litter was lower in the treatment with roots that without roots. Potassium concentration in roots was positively correlated with potassium concentration in decomposing leaf litter where roots were growing. These results suggest that while roots growing attached to decomposing litter had little or no effect in speeding the decomposition process, the superficial roots seem to play an important role in absorbing very efficiently the mineralized nutrients from litter. The anatomical study of roots showed that the plantation is intensely infected with V-A mycorrhiza. External mycorrhizal hyphae did not to play a role in attachment of roots to decomposing litter while root hairs were found to grow in profusion on root surfaces oriented toward litter.     

Statistical treatment of VAM infection data

Summary The amount of vesicular-arbuscular mycorrhizal (VAM) infection, when expressed as length of infected roots, is commonly quite variable among replicate pots within an experimental treatment. In this paper we show that frequency distributions of VAM infection parameters are often non-normal in form and may follow the negative binomial, a distribution commonly associated with aggregated organisms in nature. The lack of normality means that statistical procedures should either be non-parametric or should include data transformations.