Immunocytochemical applications in neuroanatomy

Summary In the present paper we review immunocytochemical methods for anterograde tracing with the lectin Phaseolus vulgaris-leucoagglutinin (PHA-L), combined PHA-L tracing — neurotransmitter immunocytochemistry, and the immunocytochemical localization of receptor proteins. These methods will be mainly illustrated by examples from tracing- and neurotransmitter studies on the cholinergic basal forebrain system. The morphology of PHA-L labeled neurons strongly resembles that of Golgi impregnated neurons. The complete axonal trajectories and patterns of presynaptic endings of PHA-L labeled neurons are visualized, both for light- and electron microscopic application.PHA-L-tracing can very well be combined with second immunocytochemical labeling procedures. In this way, traced pathways can be studied in their relation to chemically identified fiber systems or target neurons. Application of immunocytochemistry for the localization of the muscarinic acetylcholine receptor, albeit in its early stages, holds great promise for the near future.     

Spatial Analysis of the Canopy of Kiwifruit Vines as it Relates to the Physical, Chemical and Postharvest Attributes of the Fruit

The position of individual fruit on kiwifruit vines (Actinidiadeliciosa var. deliciosa) grown on a horizontal trellis (pergola)and on a T-bar trellis was determined using a theodolite. Thephysical, chemical, and postharvest attributes of the fruitwere related to their position on the vine during development. Fruit from the pergola vines were more numerous, of lesser weight,with lower concentrations of most mineral nutrients, but greaterconcentrations of soluble solids, and similar flesh firmnessafter 12 weeks of storage at 0 °C, than fruit from the T-barvines. The position on the vine accounted for most of the variationin the attributes of the fruit. Differences between fruit ona single lateral accounted for 43-56% of the variation. Variationbetween vines was relatively small (< 4% of the total variance). The heavier fruit were located at the apical ends of the laterals,while greater concentrations of soluble solids were associatedwith fruit located closer to the cordon. The larger fruit fromthe pergola vines developed from the early opening flowers.A similar relationship existed initially for the T-bar vines,but a reduction in growth of fruit from the early opening flowers8 weeks after anthesis resulted in a more even distributionof fruit size at harvest. The strongest relationship between mineral composition and postharvestattributes of the fruit was with soluble solids concentration(29-46% of the variance). The relationship with flesh firmnesswas weak (r = -0·14 to -0·32). Individual elementscould not be considered in isolation but rather in groups ofelements. Nitrogen was grouped strongly with phosphorus, sulphur,potassium, and copper, while calcium was linked with a secondgroup which included manganese and zinc. These two groups werenegatively related to one another. The greatest proportion of fruit with superior characteristicswas located in the denser parts of the canopy. Fruit with lessdesirable attributes were from the extremities of the canopywhere the leaf area index was low.Copyright 1994, 1999 AcademicPress Actinidia deliciosa, kiwifruit, fruit position, fruit quality, within-vine variation     

Seasonal changes in photosynthetic capacity of leaves of kiwifruit (Actinidia deliciosa) vines

A simplified procedure for the assay and purification of an enzyme which activates a galactosyltransferase (EC 2.4.1.96) involved in volume regulation of the unicellular alga Poterioochromonas malhamensis (Peterfi) is described. The enzyme was extracted with water from membranes, followed by chromatography on DEAE-Sephacel, phenyl-Sepharose and fetuin-agarose. Its proteinase activity was demonstrated by cleavage of oxidized insulin A- and B-Chains. The predominant cleavage site of the oxidized A-chain is the peptide bond between ¹³Leu and ¹⁴Tyr whereas ¹⁶Leu-¹⁷Glu is also hydrolyzed with minor activity. Besides this chymotrypsin-like endopeptidase activity some carboxypeptidase activity was also observed.     

Influence of anions on the potassium status and productivity of kiwifruit (Actinidia deliciosa) vines

The effects of K fertiliser (160 kg ha^-1) applied with Cl^- or SO₄ ^2- as the accompanying anion on the K nutrition of kiwifruit (Actinidia deliciosa var. deliciosa) were assessed in a field experiment, using vines with varying degrees of K deficiency. Leaf K concentrations in spring were significantly higher for vines receiving KCl, compared to those receiving K₂SO₄. This effect did not interact significantly with the degree of K deficiency, and persisted for about 6 weeks. Subsequently there was no significant difference between the leaf K concentrations for the vines receiving KCl or K₂SO₄. Applying K as KCl increased the leaf Cl concentration, especially in spring, while applying K as K₂SO₄ had no significant effect on the leaf S concentration at that time. These results implied a greater requirement for organic acid anions for K⁺ uptake from K₂SO₄ than from KCl, and the importance of organic acid anions for K⁺ uptake from different sources of K fertiliser is discussed. This transient effect of the accompanying anion on leaf K status was associated with large effects on flowering, and fruit yields were about 28% higher for plants receiving KCl rather than K₂SO₄.The effects on growth and tissue nutrient composition of varying the concentrations of Cl^-, NO₃ ^-, SO₄ ^2- and H₂PO₄ ^- around the roots of kiwifruit vines were examined in a solution culture experiment. For H₂PO₄ ^-, plant growth was very similar over a wide range of rates of addition. For the other anions, the range between deficiency and toxicity was clearly delineated. For Cl^- and NO₃ ^-, toxicity was associated with high tissue concentrations of Cl and N, respectively, and was consistent with competition for uptake between Cl^- and NO₃ ^-. However, for SO₄ ^2-, toxicity was associated with only a small increase in the tissue S concentration relative to that associated with maximum growth, and appeared to result more from effects on uptake of other anions and cations rather than from direct effects of high tissue S concentrations.It is concluded that the sensitivity of kiwifruit to the anion accompanying K⁺ in fertiliser may be related to the unusually high requirement for Cl previously reported for this species.     

Male Wistar rats are more susceptible to lasting social anxiety than Wild-type Groningen rats following social defeat stress during adolescence

Adolescence is an important period for the development of adult social competences. Social stress during adolescence may contribute not only to an inadequate social development but also to adult vulnerability to social anxiety. There seems to be a clear individual differentiation, however, in the vulnerability to the long-term negative consequences of social stress. The current study further explores this individual vulnerability and is aimed at the influence of social stress during adolescence on adult social anxiety and its context specificity. Rats from different strains (Wistar and Wild-type Groningen rats) were exposed to the resident-intruder paradigm five times during 10 min each in the period between postnatal day 45 and 58. Three and 7 weeks later, the animals were re-exposed to the context in the presence of either a dominant male or an anestrous female behind a wire mesh screen. Wistar rats that were socially defeated spent less time exploring the social stimulus in comparison with socially defeated Wild-type rats and their non-defeated controls. We conclude that the stressed Wistar rat shows signs of generalized social anxiety indicating that the Wistar rat can be considered as a vulnerable phenotype to effects of adolescent social stress.     

Differential long-term effects of social stress during adolescence on anxiety in Wistar and wild-type rats

Severe and chronic stress may interfere with adolescent neuronal plasticity that turns the juvenile brain into an adult brain increasing the vulnerability to develop anxiety disorders. It is well-known from adult stress research that there is a large individual differentiation in stress vulnerability. The current study is aimed at the individual resilience and vulnerability to adolescent social stress. Two strains of rats that differ in social behavioral skills were subjected to social stress during adolescence. In three experiments we studied short and long term effects of adolescent social stress using a water conflict test in different contexts. Wistar rats which had been socially defeated on postnatal days 45 and 46 showed, following water deprivation, a strong decrease in the total amount of water consumed and time spent drinking when tested 2 days and 3 weeks later in the context where they received the defeat experience. Also a strong increase in drinking latency was noticed in the context of the previous defeat. No differences in these parameters were found between defeated and non-defeated wild-type rats. The results of the water conflict test in an environment where no association with the previous defeat experience was present showed that the adolescent social stress did not induce a generalized anxiety.In conclusion, the water conflict test is a useful tool to measure the influence of social defeat on the motivation to obtain resources under conditions with different stimulus properties. In addition, our data suggest the importance of the strain used in adolescent stress experiments. The fact that Wistar rats showed a strong association with the context at adulthood whereas no effect was observed in the wild-type rats shows that victim characteristics are important determining factors for the long term effects of adolescent social stress.     

Effects of cropping, nutrition and water supply on accumulation and distribution of biomass and nutrients for apple trees on'M9'root systems

The effects of cropping, nutrition and water supply on accumulation and distribution of biomass and nutrients for apple trees on'M9'root systems were examined using trees growing in lysimeters. For 3-year-old 'Golden Delicious' trees, cropping significantly increased total biomass, although leaf and root biomass were reduced. For 3-year-old'Golden Delicious', 'Coxs Orange' and 'Gloster' trees, cropping only in the third year reduced root biomass by 39 to 45%. Cropping did not affect water use by trees of any cultivar early in the season, but increased water use later in the season for 'Gloster' trees. For trees with fruit, total non-structural carbohydrate (TNC) contents at the end of the season were highest in fruit (up to 1 400 g plant ⁻¹). For trees without fruit, TNC contents were generally highest in the roots (up to 110 g planr ⁻¹). Supplying trees with nutrient solution diluted to 10% of the standard composition increased the root biomass for 'Golden Delicious' trees only, but decreased biomass of above-ground tissues for all cultivars. Reducing the nutrient supply decreased water and nutrient uptake, and reduced nutrient contents within vegetative parts of the tree more than within roots. For 'Golden Delicious' trees, restricting the water supply to 50% or 25% of that consumed by control trees significantly reduced above-ground biomass, but root biomass was not significantly affected. The N, P and K. contents for the trees were also reduced by water stress, due to reduced contents within above-ground organs. Water stress reduced the TNC and starch contents of all tissues, except the roots. These results are discussed in relation to the efficiency of root systems for water and nutrient uptake.     

Temporal expression of effects of varying nitrogen supply on canopy growth, photosynthesis and fruit production for Actinidia deliciosa vines in the field

The effects of varying nitrogen supply on canopy leaf area, response of leaf net photosynthesis (A_n) to quantum flux density (Q), and fruit yields of kiwifruit vines (Actinidia deliciosa var. deliciosa) were examined in a two-year field experiment. Vines were grown with 0, 250 or 750 kg N ha^?1 year^?1. The responses to nitrogen supply were compared with responses to shade, to examine the impact of reduced carbon assimilation on canopy leaf area and fruit yields. Nitrogen supply did not affect significantly any of the measured variables during the first season of the experiment. In the second season, canopy leaf area was reduced significantly where nitrogen supply was limited. The quantum efficiency of photosynthesis (φ_q) increased from 0. 03 mol CO₂ mol^?1 Q soon after leaf emergence to more than 0. 05 mol CO₂ mol^?1 Q during the middle of the growing season. The quantum saturated rate of A_n (A_sat) also increased during the season, from 7–10 μmol CO₂ m^?2 s^?1 soon after leaf emergence, to 15–20 (μmol CO₂ m^?2 s^?1 during the middle of the growing season. φ_q and A_sat increased significantly with nitrogen supply at all measurement times during the second season. For vines with high nitrogen, fruit yields in both seasons were similar, averaging 3. 05 kg m^?2. Fruit yields in the second season were reduced significantly where nitrogen supply was limited, due to reduced fruit numbers. The relative effects of reduced leaf area and reduced leaf photosynthesis for carbon assimilation by nitrogen deficient vines were examined using a mathematical model of canopy photosynthesis for kiwifruit vines. Simulations of canopy photosynthesis indicated that effects on leaf area and on leaf photosynthesis were of similar importance in the overall effects of nitrogen deficiency on carbon assimilation. The effects of nitrogen supply on fruit numbers (i. e. flower development) preceded the measured effects on carbon assimilation, indicating that the nitrogen supply affected carbon partitioning to reserves in the first season.     

Immunocytochemical localization of muscarinic acetylcholine receptors in the rat endocrine pancreas

Summary Immunocytochemical application of the antimuscarinic acetylcholine receptor antibody M35 to pancreas tissue revealed the target areas for the parasympathetic nervous system. Immunoreactivity in the endocrine pancreas was much higher than that in the exocrine part. Moreover, the endocrine cells at the periphery of the islets of Langerhans displayed the highest level of immunoreactivity. Based on these findings in the mantle of the islets, two types of islets have been distinguished: type-I islets with intensely stained mantle cells, and type-II islets with a much lower concentration of these cells. On average, type-I islets were larger (244.8 m±6.1 SEM) than type-II islets (121.5 m±3.8 SEM). M35-immunoreactivity was present on the majority of D cells, which were characterized by their immunoreactivity to somatostatin [of 446 D cells 356 (79.8%) were M35-immunopositive]. However, only a small proportion of the intensely stained mantle cells belonged to the D cell population. Therefore, it is concluded that the majority of the intensely stained mantle cells represent glucagon-secreting A and/or pancreatic polypeptide-secreting F cells. The intensity of M35-immunoreactivity at the periphery and central core of the islets paralleled the density of cholinergic innervation, suggesting a positive correlation between the intensity of cholinergic transmission and the number of muscarinic acetylcholine receptors at the target structures. The present study further revealed some striking parallels for the muscarinic acetylcholine receptor characteristics between the (endocrine) pancreas and the central nervous system.