Diffuse serotoninergic neurohemal systems associated with cerebral and suboesophageal nerves in the head of the Colorado potato beetle Leptinotarsa decemlineata

We analyzed the anatomy of two diffuse neurohemal systems for serotonin in the head of the Colorado potato beetle Leptinotarsa decemlineata by means of immunohistochemistry. One system is formed by axons from two bilateral pairs of neurons in the frontal margin of the suboesophageal ganglion that enter the ipsilateral mandibular nerve, emerge from this nerve at some distance from the suboesophageal ganglion, and cover all branches of the mandibular nerve with a dense plexus of immunoreactive axon swellings. The other system is formed by axons from two large neurons in the frontal ganglion that enter the ipsilateral frontal connectives, emerge from these connectives, and form a network of axon swellings on the labroforntal, pharyngeal, and antennal nerves and on the surface of the frontal ganglion. Immunohistochemical electron microscopy demonstrated that the axon swellings are located outside the neural sheaths of the nerves and hence in close contact with the hemolymph. We therefore suggest that these plexuses represent extensive neurohemal systems for serotonin. Most immunoreactive terminals are in direct contact with the hemolymph, and other terminals are closely associated with the muscles of the mandibles, labrum, and anterior pharynx, as well as with the salivary glands, indicating that these organs are under serotoninergic control.     

In vitro and in vivo properties of an anti-CD5—momordin immunotoxin on normal and neoplastic T lymphocytes

An anti-CD5 monoclonal antibody (mAb) was linked to the plant toxin momordin, a type-1 ribosome-inactivating protein purified fromMomordica charantia. The in vitro cytotoxicity of the immunotoxin was evaluated as the inhibition of protein and/or DNA synthesis on isolated peripheral blood mononuclear cells (PBMC) and on human T cell leukemia Jurkat. The potency of the immunotoxin on PBMC was very high (IC₅₀ = 1–10 pM) and was not affected by blood components. The conjugate was also very efficient in the inhibition of the proliferative response in a mixed lymphocyte reaction (IC₅₀ = 10 pM). Moreover, the in vitro performances of the immunotoxin compared favourably with those reported for other anti-CD5-based immunoconjugates containing ricin A chain. The in vivo activity of the immunotoxin was assessed in the model ofnu/nu mice bearing Jurkat leukemia. A significant inhibition of the tumour development (80%,P <0.01) in the animals treated with immunotoxin was observed. Taken together, the in vitro and in vivo results suggest that the anti-CD5-momordin conjugate may be useful for graft-versus-host disease therapy and potentially in the treatment of CD5-positive leukemias and lymphomas.     

Total DNA 5-methylcytosine level of rice calli derived from protoplasts is independent of regeneration ability

To study the roles of m⁵C in the differentiation of rice calli derived from protoplasts (protoclones), the m⁵C level of the total DNA was analyzed using the³²P post-labeling method. The level of m⁵C in regenerable and nonregenerable protoclones was similar, as was in calli and leaves of plants regenerated from the same protoclones. Treatment with 0.5 mM 5-azacytidine caused significant reduction of the level of m⁵C and of the regeneration frequency of callus. Significantly increased m⁵C levels were observed during prolonged culture.     

A NEW PHYLOGENY FOR CHROMOPHYTE ALGAE USING 16S-LIKE RRNA SEQUENCES FROM MALLOMONAS PAPILLOSA (SYNUROPHYCEAE) AND TRIBONEMA AEQUALE (XANTHOPHYCEAE)1

The 16S-like ribosomal RNA genes from Mallomonas papillosa Harris et Bradley (Synurophyceae) and Tribonema aequale Pascher (Xanthophyceae) were sequenced and compared to those of other eukaryotes. Mallomonas is closely related to Ochromonas (Chrysophyceae) and supports the general hypothesis of a close phylogenetic relationship between the Synurophyceae and Chrysophyceae. Tribonema is specifically related to Costaria costata (C. A. Agardh) Saunders (Phaeophyceae) demonstrating an unexpected phylogenetic relationship between the Xanthophyceae and Phaeophyceae. Distance and parsimony analysis place these four chromophyte genera in a complex evolutionary assemblage that includes the Bacillariophyceae and Oomycetes but excludes the Dinophyceae. The close relationship between the chromophyte algae and the Öomycete fungi supports the hypothesis that protists with tripartite hairs form a natural assemblage.     

Starter culture for the production of ‘soyiru’

Bacillus subtilis or licheniformis facilitated production of soyiru with the best results being given by using both together. Fermentation employing Streptococcus enterococcus was unsuccessful.H.A. Suberu is with the Department of Biological Sciences, Usmanu Danfodiyo University, Sokoto, Nigeria. J.A. Akinyanju is with the Department of Biological Sciences, University of Ilorin, P.M.B. 1515, Ilorin, Nigeria.     

Cloning single-stranded DNA

DNA sequence and expression analyses have greatly benefited from using M13 and pUC derived cloning vectors and their polycloning sites. A chronology of the original concepts and experiments is reviewed.     

GTPγS restores nucleophosmin (NPM) localization to nucleoli of GTP-depleted HeLa cells

Previous studies showed that localization of nucleophosmin/B23 (NPM) to nucleoli requires adequate cellular GTP levels (Finchet al., J Biol Chem 268, 5823–5827, 1993). In order to study whether hydrolysis of GTP plays a role in NPM localization, we introduced a nonhydrolyzable GTP analog into HeLa cells. Cells were first depleted of GTP with the IMP dehydrogenase inhibitor, mycophenolic acid (MA), to induce translocation of NPM from the nucleoli to the nucleoplasm. Non-hydrolyzable GTP analogs were then introduced into cells by electroporation. We found that introduction of the non-hydrolyzable analog, GTPS, was effective in restoring NPM localization to nucleoli. Cells incubated in medium containing G-nucleotides without electroporation showed no effect. To reduce the possibility that cells use guanine from degraded nucleotide to supplement GTP pools via salvage pathways, experiments were also performed in the presence of (6-mercaptopurine) 6MP, a competitive inhibitor of the salvage enzyme, HGPRT (hypoxanthine guanine phosphoribosyl transferase), in addition to MA. Under these conditions, introduction of GTPS still effectively restored the localization of NPM into nucleoli. This study demonstrates that electroporation can be used effectively to introduce nucleotides into cultured cells without excessive loss of viability. Our results also indicate that the GTP dependent localization of NPM to the nucleoli may not require GTP hydrolysis.     

Theoretical Study of the TiO2 and MgO Surface Acidity and the Adsorption of Acids and Bases

We present ab-initio periodic Hartree–Fock calculations (crystal program) of small molecules on TiO₂ and MgO. The adsorption of the molecules may be molecular or dissociative. This depends on their acid and basic properties in the gas phase. For the molecular adsorption, the molecules are adsorbed as bases on Ti(+IV) sites, the adsorption energies correlate with the proton affinities. The dissociations on the surface correlate with the gas phase cleavages: thus, the dissociation of MeOH leads to a preferential basic cleavage (the fragment HO– is adsorbed on a Ti⁺⁴ ion and the fragment Me⁺ is adsorbed on a O²– ion of the oxide). The opposite result is obtained with MeSH. Another important factor is the adsorbate–adsorbate interaction: favorable cases are a sequence of H-bonds for the hydroxyl groups resulting from the water dissociation and the mode of adsorption for the ammonium ions. Lateral interactions also force the adsorbed CO₂ molecules to bend over the surface so that their mutual orientation resembles the geometry of the CO₂ dimer. With respect to water adsorption, MgO appears to be a basic oxide. As experimentally observed, NH₃ adsorbs preferentially on TiO₂ and CO₂ on MgO. However, this difference of reactivity should not be expressed in terms of acid vs. basic behaviour but in terms of hard and soft acidity. The MgO surface is a 'soft' acidic surface that reacts preferentially with the soft base, CO₂.     

Interaction of force transmission and sarcomere assembly at the muscle-tendon junctions of carp (Cyprinus carpio): ultrastructure and distribution of titin (connectin) and α-actinin

At muscle-tendon junctions of red and of white axial muscle fibres of carp, new sarcomeres are found adjacent to existing sarcomeres along the bundles of actin filaments that connect the myofibrils with the junctional sarcolemma. As the filament bundles that transmit force to the junction originate proximal to new sarcomeres, they probably relieve these new sarcomeres from premature loading. In red fibres, these filament bundles are long (up to 20 m) and dense, permitting light-microscopical immunohistochemistry (double reactions: anti-titin or anti--actinin and phalloidin). New sarcomeres have clear I bands; their A band lengths are similar to those of older sarcomeres and the thick filaments lie in register. T tubules are found at the distal side of new sarcomeres but terminal Z lines are absent. The late addition of -actinin suggests that -actinin mainly has a stabilizing role in sarcomere formation. The presence of titin in the terminal fibre protrusions is in agreement with its supposed role in sarcomere formation, viz. the integration of thin and thick filaments. The absence of a terminal Z line from sarcomeres with well-registered A bands suggests that this structure is not essential for the anchorage of connective (titin) filaments.     

Cortical hypometabolism in injured brain: New correlations with the noradrenergic and serotonergic systems and with behavioral deficits

Changes with time after injury in behavioral deficits, as determined by the Morris swim test, and the in vivo specific binding of HEAT, a selective ₁-adrenoreceptor ligand, were compared with the time-course of development of cortical hypometabolism in rats with focal freezing lesions. In our trauma model, cortical hypometabolism was detectable in the lesioned hemisphere at 4 hr, became maximal (50% of normal) at 3 days and diminished towards normal on days 5 and 10 post-injury. Progressive impairment of acquisition of the Morris water maze task was demonstrated up to day 3 post-lesion with improvement thereafter. On day 3 the latency to reach criterion was 60% longer in lesioned animals than in corresponding sham-operated ones. An increase in the volume of distribution of HEAT, limited to cortical areas of the lesioned hemisphere, was demonstrable at 4 hr post-lesion and reached its maximum on day 3 (200% of normal) with subsequent return toward normal on days 5 and 10. Several types of drugs were shown previously to modify the cortical hypometabolism associated with cerebral injury. The present data indicate that the same drugs also modify the in vivo binding of HEAT and the behavioral deficits induced by brain lesions. Ibuprofen, a non-steroidal anti-inflammatory drug, p-chlorophenylalanine, an inhibitor of serotonin synthesis, ketanserin, a specific 5HT₂-receptor antagonist, and prazosin, an ₁-adrenergic receptor blocker all normalized the in vivo binding of HEAT in the cortical areas of the lesioned hemisphere. All groups of animals treated with these drugs also showed subtle, but statistically highly significant improvements in latency to locate the platform in the Morris water maze. Taken together these results show good correlation between behavioral deficits, changes in ₁-noradrenergic receptor binding and cortical hypometabolism in injured brain. This supports the hypothesis that post-injury cortical hypometabolism is a reflection of cortical functional depression in which both the serotonergic and noradrenergic neurotransmitter systems play a role, compatible with their inhibitory effects in the cortex and their postulated involvement in cortical information processing.Special issue dedicated to Dr. Leon S. Wolfe.