A serotonergic system in the brain of the Antarctic fish, Trematomus bernacchii

The immunohistochemical distribution of serotonin-containing nerve fibres and cells has been described in the brain of the Antarctic fish, Trematomus bernacchii. The largest serotonergic system was associated with the diencephalic and rhombencephalic ventricles. In particular, serotonin-positive cells have been found in the lateral recess and neuropile zone of the diencephalic ventricle, where we have identified the serotonergic portion of the paraventricular organ. Numerous serotonin cells were localized in the dorsal nucleus of the raphe, the dorsal tegmental nucleus and the central gray. Two large cell groups, arranged in a pair of well-defined columns and connecting the central gray with the dorsal reticular formation, were immunostained in the region of the trigeminal nuclei. In addition, few positive cells have been found in the preoptic area and the cerebellar valvula, and few serotonergic nerve fibres, probably belonging to the lateral lemniscus, have been identified. The distribution of serotonin elements in the brain of T. bernacchii has been compared with that described in other fish, where it showed some modifications in the immunoreactive pattern. Finally, the lack of a serotonergic system at the level of the reticular superior formation has been reported; however, it was not possible to rule out a phylogenetic or environmental explanation.     

Neuronal Basis of Innate Olfactory Attraction to Ethanol in Drosophila

The decision to move towards a mating partner or a food source is essential for life. The mechanisms underlying these behaviors are not well understood. Here, we investigated the role of octopamine – the invertebrate analogue of noradrenaline – in innate olfactory attraction to ethanol. We confirmed that preference is caused via an olfactory stimulus by dissecting the function of the olfactory co-receptor Orco (formally known as OR83b). Orco function is not required for ethanol recognition per se, however it plays a role in context dependent recognition of ethanol. Odor-evoked ethanol preference requires the function of Tbh (Tyramine β hydroxalyse), the rate-limiting enzyme of octopamine synthesis. In addition, neuronal activity in a subset of octopaminergic neurons is necessary for olfactory ethanol preference. Notably, a specific neuronal activation pattern of tyraminergic/octopaminergic neurons elicit preference and is therefore sufficient to induce preference. In contrast, dopamine dependent increase in locomotor activity is not sufficient for olfactory ethanol preference. Consistent with the role of noradrenaline in mammalian drug induced rewards, we provide evidence that in adult Drosophila the octopaminergic neurotransmitter functions as a reinforcer and that the molecular dissection of the innate attraction to ethanol uncovers the basic properties of a response selection system.     

Gas exchange studies in two Portuguese grapevine cultivars

Gas exchange characteristics of leaves of Vitis vinifera L. cvs Tinta Amarela and Periquita, two grapevine cultivars grown in distinct climatic regions of Portugal, were studied under natural and controlled conditions. Daily time courses of gas exchange were measured on both a hot, sunny day and a cooler, partly cloudy day. Responses of net photosynthesis to irradiance and internal partial pressure of CO₂, were also obtained. A strong correlation between net photosynthesis (P_N) and leaf conductance (g_s) was found during the diurnal time courses of gas exchange, as well as a relatively constant internal partial pressure of CO₂ (P_i), even under non-steady-state conditions. On the cloudless day, both P_N and g_s were lower in the afternoon than in the morning, despite similar conditions of leaf temperature, air to leaf water vapor deficit and irradiance. The response curves of net photosynthesis to internal CO₂ showed linearity up to p_i values of 50 Pa, possibly indicating a substantial excess of photosynthetic capacity. When measured at low partial pressures of O₂ (1 kPa), P_N became inhibited at high CO₂ levels. Inhibition of P_N at high CO₂ was absent under normal levels of O₂ (21 kPa). Significant differences in gas exchange characteristics were found between the two cultivars, with T. Amarela having higher rates under similar measurement conditions. In particular, the superior performance of T. Amarela at high temperatures may represent adaptation to the warmer conditions at its place of origin.     

Isolation of tomato cell lines with altered response to Fusarium cell wall components

Summary To obtain Tomato cell lines with an altered capacity to respond to heat-released cell wall components (elicitor) of a tomato pathogen (Fusarium oxysporum f. sp. lycopersici), positive and negative selection experiments, using BUdR enrichment techniques, were carried out on suspension cultures of the susceptible, low phytoalexin producer cultivar Red River. Both high and low phytoalexin producing clones were isolated. Further tests demonstrated that not all phytoalexin-producing clones were more susceptible to the elicitor toxic effect, and that they were altered also in the speed of response to fungal cell wall components. Cells selected with Fusarium elicitor showed the same behaviour when challenged by Phytophthora infestans elicitor, thus suggesting in this case lack of specificity. The results are finally discussed with a view to using the technique both as a tool to study the genetics and physiology of hostparasite interactions and as a possible new method for the selection of pathogen resistant genotypes.Paper no. 1224 IPRA-CNR; research supported by an EEC-BAP contract     

Transport of l(-)malic acid and other dicarboxylic acids in the yeast Candida sphaerica

Summary dl-Malic acid grown cells of Candida sphaerica (anamorph of Kluyveromyces marxianus) formed a saturable transport system that mediated accumulative transport of l(-)malic acid with the following kinetic parameters at pH 5.0: V _max, 0.44 nmol l(-)malate·s^-1 per milligram dry weight; K _m ,0.1 mM l(-)malate. Initial uptake of the acid was accompanied by disappearance of extracellular protons, the rates of which followed Michaelis-Menten kinetics as a function of the acid concentration. Variation with extracellular pH of the K _m values, calculated either as the concentrations of anions or of undissociated acid, pointed to anions as the transported form. Furthermore, accumulated free acid suffered rapid efflux after the addition of the protonophore carbonylcyanide-M-chlorophenyl-hydrazone (CCCP). These results suggested that the transport system was a dicarboxylate-proton symporter. The system was inducible and was subject to glucose repression. Succinic, fumaric, -ketoglutaric, oxaloacetic and d-malic acid, but not maleic, malonic, oxalic nor l(+)-tartaric acid, apparently used the same transport system since they acted as competitive inhibitors of l(-)malic acid transport and induced proton movements that followed Michaelis-Menten kinetics. Experiments with glucose-repressed cells showed that undissociated dicarboxylic acid (measured with labelled succinic acid) entered the cells slowly by simple diffusion. The permeability of the cells for undissociated acid increased exponentially with pH, the diffusion constant increasing 100-fold between pH 3.5 and 6.0.     

Fine structure of the receptors at the myotendinous junction of human extraocular muscles

The myotendinous junction of the human extraocular muscles was studied by electron microscopy. Some peculiar receptorial structures have been found in the majority of the samples examined. These structures are very small and consist of 1) the terminal portion of one muscle fibre, 2) the tendon into which it inserts and 3), within the tendon, a rich nerve arborization, whose branches are always very close to the muscle component. Only one discontinuous layer, made up of flat cells, which lack a basal lamina and often show pinocytotic vesicles, encapsules every musculo-tendinous complex. The tendinous component consists of amorphous ground substance of different electron density, of collagen and elastic fibres and is divided in compartments by ramified cells, which make an inner capsular-like covering to the nerve fibres. Three types of afferent nerve endings can be identified. One type is usually more frequent than the others, possesses a large number of neurotubules and neurofilaments and few mitochondria and is always surrounded by a Schwann cell which forms finger-like processes penetrating into the axoplasm. The second type is only partially enveloped by the Schwann cell. The axoplasm is devoid of neurotubules and contains few neurofilaments, several mitochondria and groups of small clear vesicles placed in the areas uncovered by the glial sheath. The third one is completely surrounded by the Schwann cell, but is devoid of neurotubules and neurofilaments and full of mitochondria. These morphological features correspond well with the probable role of these receptorial structures, which is to ensure very exact and precise ocular movements.     

Linkage data suggesting allelic heterogeneity for paramyotonia congenita and hyperkalemic periodic paralysis on chromosome 17

Summary Paramyotonia congenita (PC), an autosomal dominant non-progressive muscle disorder, is characterised by cold-induced stiffness followed by muscle weakness. The weakness is caused by a dysfunction of the sodium channel in muscle fibre. Parts of the gene coding for the -subunit of the sodium channel of the adult human skeletal muscle (SCN4A) have been localised on chromosome 17. To investigate the role of this gene in the etiology of PC, a linkage analysis in 17 well-defined families was carried out. The results (z=20.61, =0.001) show that the mutant gene responsible for the disorder is indeed tightly linked to the SCN4A gene. The mutation causing hyperkalemic periodic paralysis (HyperPP) with myotonia has previously been mapped to this gene locus by the same candidate gene approach. Thus, our data suggest that PC and HyperPP are caused by allelic mutations at a single locus on chromosome 17.Dedicated to Professor P. E. Becker on the occasion of his 83rd birthday.     

Isolation and characterization of three protein proteinase isoinhibitors from the granular fraction of horse neutrophilic granulocytes

Three cathodically migrating protein protease isoinhibitors were isolated from the granule-rich fraction of equine neutrophilic granulocytes by means of FPLC chromatography, in addition to two previously described anodically migrating inhibitors. The three isoinhibitors had an identical enzyme specificity which was equal to the two previously described isoinhibitors; they inhibited exclusively proteinase K and subtilisin. The inhibitors retained their activity between pH 1 and 12. They also were heat stable at 100 degrees C for 20 min. Neither the biological function of isoinhibitors nor the fundamental role of granular protease inhibitors of such narrow and peculiar enzyme specificity are known.     

Isolation and characterization of two new low-molecular-weight protein proteinase inhibitors from the granule-rich fraction of equine neutrophilic granulocytes

A new species of protein proteinase inhibitors was detected in the granule-rich fraction of equine neutrophilic granulocytes. Five isoinhibitors were identified with a narrow enzyme specificity towards two microbial proteinases, e.g., proteinase K and subtilisin. Two isoinhibitors were purified and partially characterized. They had an Mr of 11,300 and 7400, respectively, and were resistant to perchloric acid and heat treatment at 100 degrees C for 20 min. The inhibitors retained their activity over a broad range of pH (1-9 and 1-12, respectively). The possible biological function of this species of protein proteinase inhibitors as defensins (= endogenous antibiotics) is tentatively discussed.     

Different structure of polytene chromosomes ofPhaseolus coccineus suspensors during early embryogenesis

Summary The pattern of DNA and RNA puffs in pair VII of polytene chromosomes has been investigated in the suspensor ofPhaseolus coccineus during early embryo development. The pattern of³H-TdR and³H-U incorporation has been also detected. Collected data indicate that: 1. both heterochromatic regions, p₁₁ and q_(111+112), of chromosome pair VII, organize large DNA puffs; 2. DNA puffs of both regions are specific of different embryo differentiation steps; 3. a seasonal influence on the DNA puffing seems also to be present, as demonstrated by the comparison of the results collected in two different crops; 4. the incorporation experiment by³H-TdR evidences that not all DNA puffs show clustered labeling; 5. the RNA puffing of the two regions seems also to be specific of determined embryo stages.