Effects of water conductivity on electrocommunication in the weak-electric fish Brienomyrus niger (Mormyriformes)

A characteristic electric organ discharge display in social encounters between mormyrid fish is a temporary discharge cessation. Using this response, we have investigated the useful range of electrocommunication under different water conductivity conditions in the mormyrid Brienomyrus niger. An individual fish was confined to a porous ceramic shelter tube and moved from a starting distance of 380 cm toward a similarly confined conspecific until discharge, cessation occurred. The moved fish was subsequently returned to its original, position. Water conductivity affects the peak-to-peak source voltage of the electric organ and the sensitivity of the fish's electroreceptors. Within a range of 10 to 36 000 μS/cm, the peak-to-peak amplitude of the electric organ discharge declined as a power function. At 120 μS/cm, the amplitude was 50%, and at 300μS/cm, 30% of the 10 μS/cm value. The interfish distance at which discharge cessation occurred and the associated electric field gradients were dependent on water conductivity and upon the spatial orientation of the two fish (end-to-end or parallel orientations of their shelter tubes). The respective ranges were from 135 cm and 0.02 mV/cm at 52 μS/cm (parallel orientation) to 22 cm and 0.36 mV/cm at 678 μS/cm (end-to-end orientation). When the data for both tube orientations were combined, the relationship between water conductivity (x) and the distance at which discharge cessation occurred (y) could be expressed by a power function, y=K·x^a (with K=10^2.97 and a=?0.56). When an electrically ‘silent’ fish was moved away from its conspecific, a discharge resumption in the form of a high-frequency rebound occasionally effected changes in the other fish's discharge activity at distances up to 157 cm (with an associated electric, field gradient of 0.01 mV/cm under the lowest conductivity condition).     

Immunochemical studies on blood groups: The internal structure and immunological properties of water-soluble human blood group A substance studied by Smith degradation,liberation, and fractionation of oligosaccharides and reaction with lectins

Carbohydrate structures in the interior of a blood group A active substance (MSS) were exposed by one and by two Smith degradations. Reactivities of the original glycoprotein and its Smith degraded products with 13 different lectins and with anti-I Ma were studied by quantitative precipitin assay. MSS and its first Smith degraded product completely precipitated Ricinus communis hemagglutinin with five times less of the first Smith degraded glycoprotein being required for 50% precipitation. The second Smith degraded material precipitated only 90% of the lectin. MSS did not precipitate peanut lectin, whereas its first and second Smith degraded products completely precipitated the lectin. The first Smith degraded glycoprotein also reacted well with Wistaria floribunda, Maclura pomifera, Bauhinia purpurea alba, and Geodia lectins indicating that its carbohydrate moiety could contain dGalNAc, dGalβ1 → 3dGalNAc, dGalβ1 → 4dGlcNAc, dGalβ1 → 3dGlcNAcβ1 → 3dGal and/or dGalβ1 → 4dGlcNAcβ1 → 6dGal and/or dGalβ1 → 4dGlcNAcβ1 → 6dGalNAc determinants at nonreducing ends. The second Smith degraded material precipitated well with Ricinus communis hemagglutinin, Arachis hypogaea, Geodia cydonium, Maclura pomifera, and Helix pomatia lectins showing that dGalNAc, dGalβ1 → 3dGalNAc, dGalβ1 → 4dGlcNAc residues at terminal nonreducing ends could be involved. Monoclonal anti-I Ma (group 1) serum reacted strongly with the first Smith degraded product indicating large numbers of anti-I Ma determinants, dGalβ1 → 4dGlcNAcβ1 → d 6dGal and/or dGalβ1 → 4dGlcNAcβ1 → 6dGalNAc at nonreducing ends. The comparable activities of the native and Smith degraded products with wheat germ lectin indicate capacity to react with DGlcNAc residues at nonreducing ends and/or at positions in the interior of the chain. The totality of lectin reactivities indicates heterogeneity of the carbohydrate side chains. Oligosaccharides with ³H at their reducing ends released from the protein core of the first and second Smith degraded products were obtained by treatment with 0.05 m NaOH and 1 M NaB³H₄ at 50 °C for 16 h (Carlson degradation). The liberated reduced oligosaccharides were fractionated by dialysis, followed by retardion, Bio-Gel P-2, P-4, and P-6 columns. They were further purified on charcoal-celite columns, and by preparative paper chromatography and high-pressure liquid chromatography. Their distribution by size was estimated by the yields on dialysis, Bio-Gel P-2, and Bio-Gel P-6 chromatography, and from the radioactivity of the reduced sugars. Of the oligosaccharide fractions from the first Smith degraded product, about 77% of the carbohydrate side chain residues contained from 1 to 6 sugars, 13% from 7 to perhaps 12 sugars, and 10% was nondialyzable (polysaccharides and glycopeptide fragments). Of the second Smith degraded product, approximately 82% of carbohydrate residues had from 1 to 6 sugars, 14% from 7 to perhaps 20 sugars and 4% was nondialyzable. The biological activity profile of the two Smith degraded products together with the size distributions of the oligosaccharides indicated that their carbohydrate side chains, comprised a heterogeneous population ranging in size from 1 to about 12 sugars. When most of these chains that are shorter than hexasaccharides are fully characterized it may be possible to reconstruct the overall structure of the carbohydrate moiety of the blood group substances and account for their biological activities.     

The differential effects of PGE on the immune response in normal and immunosuppressed mice

The effect of 16,16-dimethyl-PGE₂-methyl ester (di-M-PGE₂) on humoral and cellular immunoresponsiveness has been compared in normal mice and in mice immunosuppressed by splenectomy and thymectomy plus antithymocyte serum (ATS). Splenectomy resulted in immunosuppression manifested by augmentation of B-16 melanoma growth; this stimulatory effect was reversed by di-M-PGE₂. In animals immunosuppressed by thymectomy plus ATS, di-M-PGE₂ augmented the humoral and cellular immune responses; this was manifested by slowing of the growth of B-16 melanoma and by stimulating the number of plaque-forming cells, hemagglutinin titers, and delayed-hypersensitivity reactions to sheep erythrocytes. In contrast, in normal (nonthymectomized) mice, di-M-PGE₂ was mildly immunosuppressive. Finally, adriamycin-immunosuppressed normal mice and this suppression were reversed by the addition of di-M-PGE₂ to the treatment regimen.     

Two Possibly Distinct Prostaglandin E1 Receptors in N1E-115 Clone: One Mediating Inositol Trisphosphate Formation, Cyclic GMP Formation, and Intracellular Calcium Mobilization and the Other Mediating Cyclic AMP Formation

Prostaglandin E1 (PGE1)-mediated transmembrane signal control systems were investigated in intact murine neuroblastoma cells (clone N1E-115). PGE1 increased intracellular levels of total inositol phosphates (IP), cyclic GMP, cyclic AMP, and calcium ([Ca2+]i). PGE1 transiently increased inositol 1,4,5-trisphosphate formation, peaking at 20 s. There was more than a 10-fold difference between the ED50 for PGE1 at cyclic AMP formation (70 nM) and its ED50 values at IP accumulation (1 microM), cyclic GMP formation (2 microM), and [Ca2+]i increase (5 microM). PGE1-mediated IP accumulation, cyclic GMP formation, and [Ca2+]i increase depended on both the concentration of PGE1 and extracellular calcium ions. PGE1 had more potent intrinsic activity in cyclic AMP formation, IP accumulation, and cyclic GMP formation than did PGE2, PGF2 alpha, or PGD2. A protein kinase C activator, 4 beta-phorbol 12 beta-myristate 13 alpha-acetate, had opposite effects on PGE1-mediated IP release and cyclic GMP formation (inhibitory) and cyclic AMP formation (stimulatory). These data suggest that there may be subtypes of the PGE1 receptor in this clone: a high-affinity receptor mediating cyclic AMP formation, and a low-affinity receptor mediating IP accumulation, cyclic GMP formation, and intracellular calcium mobilization.     

Inhibitory Modulation by Sodium Ions of the N-Methyl-D-Aspartate Recognition Site in Brain Synaptic Membranes

Specific binding of radiolabeled L-glutamic acid (Glu) was examined using rat brain synaptic membranes treated with a low concentration of Triton X-100. The binding drastically increased in proportion to increasing concentrations of the detergent used up to 0.1%. Addition of 100 mM sodium acetate significantly potentiated the binding in membranes not treated with Triton X-100, whereas it markedly inhibited the binding in Triton-treated membranes. The binding in Triton-treated membranes was inversely dependent on incubation temperature and reached a plateau within 10 min after the initiation of incubation at 2 degrees C, whereas the time required to attain equilibrium at 30 degrees C was less than 1 min. Sodium acetate invariably inhibited the binding detected at both temperatures independently of the incubation time via decreasing the affinity for the ligand. The binding was significantly displaced by agonists and antagonists for an N-methyl-D-aspartate (NMDA)-sensitive subclass of brain excitatory amino acid receptors, but not by those for the other subclasses. Inclusion of sodium acetate reduced the potencies of NMDA agonists to displace the binding without virtually affecting those of NMDA antagonists. Moreover, sodium ions inhibited the ability of Glu to potentiate the binding of N-[3H] [1-(2-thienyl)cyclohexyl]piperidine to open NMDA channels in Triton-treated membranes. These results suggest that sodium ions may play an additional modulatory role in the termination process of neurotransmission mediated by excitatory amino acids via facilitating a transformation of the NMDA recognition site from a state with high affinity for agonists to a state with low affinity.     

Enhanced Rate of Expression and Biosynthesis of Neuropeptide Y After Kainic Acid-Induced Seizures

Recent studies have shown marked increases in brain content of neuropeptide Y (NPY) after seizures induced by intraperitoneal injection of kainic acid and after pentylenetetrazole kindling in the rat. We have now investigated possible changes in the rate of biosynthesis of NPY after kainic acid treatment, by using pulse-labeling of the peptide and by determining prepro-NPY mRNA concentrations. For pulse labeling experiments, [3H]tyrosine was injected into the frontal cortex, and the incorporation of the amino acid into NPY was determined after purifying the peptide by gel filtration chromatography, antibody affinity chromatography, and reversed-phase HPLC. At 2 and 30 days after kainic acid treatment, the rate of tyrosine incorporation was enhanced by approximately 380% in the cortex. In addition, concentrations of pre-pro-NPY mRNA were determined in four different brain areas by hybridization of Northern blots with a complementary 32P-labeled RNA probe 2, 10, 30, and 60 days after kainic acid treatment. Marked increases were observed in the frontal cortex (by up to 350% of controls), in the dorsal hippocampus (by 750%), and in the amygdala/pyriform cortex (by 280%) at all intervals investigated. In the striatum only a small, transient increase was observed. The data demonstrate increased expression of prepro-NPY mRNA and an enhanced rate of in vivo synthesis of NPY as a result of seizures induced by the neurotoxin kainic acid.     

Classical Conditioning-Induced Changes in Low-Molecular-Weight GTP-Binding Proteins in Rabbit Hippocampus

Classical conditioning of Hermissenda, involving paired light-rotation events, results in a 30-35% decrease in the levels of a 20-kDa G protein (cp20). To test whether a similar protein exists in vertebrates, rabbits were trained to associate a tone with periorbital electrical stimulation and G proteins were analyzed by photoaffinity labeling with [alpha-32P]GTP-azidoanilide. A 20-kDa G protein similar to cp20 decreased by 36% in the hippocampus of rabbits subjected to paired tone and electrical stimulation, but not in unpaired controls. Learning-specific decreases were also found in the amount of ras protein.     

Synergistic Regulation of Cytosolic Ca2+ Concentration in Mouse Astrocytes by NK1 Tachykinin and Adenosine Agonists

The effects on cytosolic Ca2+ concentration of 2-chloroadenosine and [L-Pro9]-substance P, a selective agonist of NK1 receptors, were investigated on astrocytes from embryonic mice in primary culture. Cells responded to [L-Pro9]-substance P with a transitory increase in cytosolic Ca2+ which was of shorter duration when external Ca2+ was removed. A transient response to 2-chloroadenosine alone occurred. When simultaneously applied, [L-Pro9]-substance P and 2-chloroadenosine evoked a prolonged elevation of cytosolic Ca2+ (up to 30 min). This phenomenon was dependent on the presence of extracellular Ca2+, but insensitive to dihydropyridines, La3+, and Co2+, excluding the implication of voltage-operated Ca2+ channels. Arachidonic acid also induced a sustained elevation of cytosolic Ca2+, but did not increase further the response evoked by [L-Pro9]-substance P and 2-chloroadenosine. The activation of protein kinase C by a diacylglycerol analogue mimicked the effect of [L-Pro9]-substance P in potentiating the 2-chloroadenosine-evoked response. Like 2-chloroadenosine, pinacidil, which hyperpolarizes the cells by opening K+ channels, prolonged the elevation of cytosolic Ca2+ concentration induced by [L-Pro9]-substance P. Conversely, depolarization with 50 mM KCl canceled the effects of either pinacidil or 2-chloroadenosine applied with [L-Pro9]-substance P. Pertussis toxin pretreatment suppressed all the effects induced by 2-chloroadenosine.     

Developmental Expression of HNK-1-Reactive Antigens in Rat Cerebral Cortex and Molecular Heterogeneity of Sulfoglucuronylneolactotetraosylceramide in CNS Versus PNS

Monoclonal antibody HNK-1 reacts with a carbohydrate epitope present in proteins, proteoglycans, and sulfoglucuronylglycolipids (SGGLs). On high-performance TLC plates, SGGLs of the CNS from several species migrated consistently slower than those from the PNS, a result indicating possible differences in the structures. The structural characteristics of the major SGGL, sulfoglucuronylneolactotetraosylceramide (SGGL-1), from CNS was compared with those of SGGL-1 from PNS. Although the composition, sequence, and linkages of the carbohydrate moiety of the SGGL-1 species were identical, SGGL-1 from CNS contained mainly short-chain fatty acids, 16:0, 18:0, and 18:1, amounting to 85% of the total fatty acids, whereas SGGL-1 from PNS contained large proportions (59%) of long-chain fatty acids (greater than 18:0). These differences in the fatty acid composition accounted for the different migration pattern observed. The developmental expression of SGGLs and HNK-1-reactive proteins was studied in rat cerebral cortex between embryonic day (ED) 15 to adulthood. SGGLs in the rat cortex were maximally expressed around ED 19 and almost completely disappeared by postnatal day (PD) 20. This expression was contrary to their increasing expression in the cerebellum and sciatic nerve with postnatal development. Six to eight protein bands with a molecular mass of greater than 160 kDa were HNK-1 reactive in the rat cerebral cortex at different ages. The major HNK-1 reactivity to the 160-kDa protein band seen in ED 19 to PD 10 cortex decreased and completely disappeared from the adult cortex, whereas several other proteins remained HNK-1 reactive even in the adult. Western blot analyses of the neural cell adhesion molecules (N-CAMs) during development of the rat cortex with a polyclonal anti-N-CAM antibody showed that the major HNK-1-reactive protein bands were not N-CAMs. Between PD 1 and 10, 190-200-kDa N-CAM was the major N-CAM, and between PD 15 to adulthood, 180-kDa N-CAM was the only N-CAM present in the rat cortex.