Design and synthesis of novel delta opioid receptor agonists and their pharmacologies

We re-examined the accessory site of the 4,5-epoxymorphinan skeleton by camdas conformational analysis in an effort to deign novel δ opioid receptor antagonists. We synthesized three novel compounds (SN-11, 23 and 28) with a 10-methylene bridge and without a 4,5-epoxy ring. Among them, compounds SN-23 (17-isobutyl derivative) and SN-28 (17-methyl derivative) showed very strong agonist activity (over 10 times more than TAN-67). SN-28 also showed high δ selectivity. The δ agonist activity of SN-23 was weaker than that of SN-28, but in terms of the δ selectivity, SN-23 was higher than that of SN-28. These unexpected results indicated that the 4,5-epoxy ring, but not the 10-methylene bridge, was an accessory site in δ opioid receptor agonists.     

Synthesis of pyrrolomorphinan derivatives as κ opioid agonists

We synthesized pyrrolomorphinan derivatives 6, 7, and 9 to examine whether the pyrrole ring would be an accessory site in the κ opioid receptor selective antagonist, nor-binaltorphimine. Derivative 6 had an α,β-unsaturated ketone substituent that strongly bound to the κ receptor. The compound with the highest κ receptor selectivity, 6e, produced a dose-dependent antinociceptive effect in the mouse acetic acid writhing test. However, derivatives 7 and 9, which did not have α,β-unsaturated ketone substituents, showed less κ receptor selectivity than compound 6. Based on structure–activity relationships, we proposed that these compounds adopted active structures for κ selective agonist activity. The pyrrole ring would not function as an accessory site, but the ability of the side chain on the pyrrole ring to localize above the C-ring appeared to confer κ selective agonist activity. These results will promote the design of novel κ agonists.     

Tetranucleotide,trinucleotide, and dinucleotide loci from the bobcat (Lynx rufus)

We describe primers and polymerase chain reaction (PCR) conditions to amplify four dinucleotide, one trinucleotide, and three tetranucleotide microsatellite DNA loci from the bobcat (Lynx rufus). The primers were tested on 22 individuals collected from a population located within southwestern Georgia (USA). The primer pairs developed in this study yielded an average of 7.4 alleles per locus (range four to 10), an average observed heterozygosity of 0.60 (range 0.40 to 0.76), and an average polymorphic information content of 0.70 (range 0.51 to 0.78).     

Quick and reversible inhibition of soybean root nodule growth by nitrate involves a decrease in sucrose supply to nodules

The upper part of a nodulated soybean root hydroponically cultured in a glass bottle was monitored using a computer microscope under controlled environmental conditions, and the diameter of individual nodules was measured from 10-24 d after planting. The diameter of a root nodule attached to the primary root increased from 1 mm to 6 mm for 2 weeks under nitrogen-free conditions. The increase in diameter of the nodules was almost completely stopped after 1 d of supplying 5 mM nitrate, and was due to the cessation of nodule cell expansion. However, nodule growth quickly returned to the normal growth rate following withdrawal of nitrate from the solution. The reversible depression of nodule growth by nitrate was similar to the restriction of photoassimilate supply by continuous dark treatment for 2 d followed by normal light/dark conditions. In addition, the inhibitory effect of nitrate was partially alleviated by the addition of 3% (w/v) sucrose to the medium. Plant leaves were exposed to (11)C or (14)C-labelled carbon dioxide to investigate the effects of 5 mM nitrate on the translocation and distribution of photosynthates to nodules and roots. Supplying 5 mM nitrate stimulated the translocation rate and the distribution of labelled C in nitrate-fed parts of the roots. However, the (14)C partitioning to nodules decreased from 9% to 4% of total (14)C under conditions of 5 mM nitrate supply. These results indicate that the decrease in photoassimilate supply to nodules may be involved in the quick and reversible nitrate inhibition of soybean nodule growth.     

Low density lipoprotein receptor activity in homozygous familial hypercholesterolemia fibroblasts

We have identified specific low affinity low density lipoprotein (LDL) receptors in skin fibroblasts from two patients previously classified as having LDL receptor-negative homozygous familial hypercholesterolemia (FHC). Km and maximum capacity for cell-associated and degraded 125I-LDL were determined by two independent methods, a traditional technique in which increasing amounts of 125I-LDL were added until receptor saturation was achieved and a new technique in which the displacement of a small amount of 125I-LDL tracer was observed during the addition of variable amounts of unlabeled LDL. The Km for specific cell-associated 125I-LDL in FHC cells was 3.5-7.3 times that of normal cells and the maximum specific capacity was reduced to 11% of normal. Thus, some FHC cells have reduced affinity as well as reduced capacity for LDL. The FHC cell receptors share many but not all properties of the normal skin fibroblast LDL receptor. Specific degradation of bound 125I-LDL occurred concomitantly with LDL binding and was greatly reduced by the addition of chloroquine, an inhibitor of lysosomal function. Preincubation of FHC cells with cholesterol or LDL resulted in significant suppression of receptor function. Modification of lysine residues of LDL abolished receptor activity in both normal and FHC cells. Treatment of FHC cells with compactin, a cholesterol synthesis inhibitor, resulted in significant increases in specific 125I-LDL binding and degradation compared to FHC cells without compactin treatment. Normal cells also showed increases in 125I-LDL binding and degradation with compactin treatment, but the mean percentage increase in specific 125I-LDL degradation was significantly greater in FHC cells (strain GM 2000, 160 +/- 18%) than in normal cells (29 +/- 8%).     

Synthesis of quinolinomorphinan-4-ol derivatives as δ opioid receptor agonists

The previously reported morphinan derivative SN-28 showed high selectivity and agonist activity for the δ opioid receptor. In the course of examining the structure-activity relationship of SN-28 derivatives, the derivatives with the 4-hydroxy group (SN-24, 26, 27) showed higher selectivities for the δ receptor over the μ receptor than the corresponding SN-28 derivatives with the 3-hydroxy group (SN-11, 23, 28). Derivatives with the 4-hydroxy group showed potent agonist activities for the δ receptor in the [(35)S]GTPγS binding assay. Although the 17-cyclopropylmethyl derivative (SN-11) with a 3-hydroxy group showed the lowest selectivity for the δ receptor among the morphinan derivatives, the agonist activity toward the δ receptor was the most potent for candidates with the 3-hydroxy group.     

Lifetable demography and population growth of the rotifer Brachionus angularis in Kenya: influence of temperature and food density

Lifetable demography and reproductive traits of a Kenyan strain of the rotifer Brachionus angularis were investigated using individual and small batch culture approaches. The rotifer was identified morphologically before conducting studies at 20, 25 and 30 °C, using Chlorella vulgaris at 2.5 × 10⁵ to 2.5 × 10⁷ cells ml^–1. The rotifers were highly fecund, producing 2.11 ± 0.07 offspring female^–1 day^–1 and reproductive, producing 8.43 ± 0.24 offspring female^–1 at 25 °C with 2.5 × 10⁶ algal cells ml^–1. The highest intrinsic rate of natural increase (0.74 ± 0.02 d^–1), specific population growth rate (0.49 ± 0.01), longest life expectancy at hatching (12.41 ± 0.28 d) and shortest generation time (2.87 ± 0.03 d) also occurred at 25 °C with 2.5 × 10⁶ algal cells ml^–1. The duration of hatching to first spawning was shortest (2.86 ± 0.21 h) at 30 °C with 2.5 × 10⁷ algal cells ml^–1 and longest (8.83 ± 0.39 h) at 20 °C with 2.5 × 10⁵ algal cells ml^–1. The highest population density (255.7 ± 12.6 ind. ml^–1) was realised at 25 °C with 2.5 × 10⁶ cells ml^–1 on Day 8, whereas the lowest population density (122.0 ± 3.6 ind. ml^–1) was realised at 20 °C with 2.5 × 10⁵ cells ml^–1 on Day 8. The lorica length and width of the Kenyan strain of B. angularis are 85.6 ± 3.1 µm and 75.4 ± 3.6 µm, respectively. The rotifer optimally reproduces at 25 °C when fed with 2.5 × 10⁶ algal cells ml^–1.     

Fluorine-containing aryloxyethyl thiocyanate derivatives are potent inhibitors of Trypanosoma cruzi and Toxoplasma gondii proliferation

As a part of our project aimed at developing new safe chemotherapeutic and chemoprophylactic agents against tropical diseases, fluorine-containing drugs structurally related to 4-phenoxyphenoxyethyl thiocyanate (1) were designed, synthesized, and evaluated as antiproliferative agents against Trypanosoma cruzi, the parasite responsible of American trypanosomiasis (Chagas' disease), and Toxoplasma gondii, the etiological agent of toxoplasmosis. This thiocyanate derivative had previously proven to be an effective agent against T. cruzi proliferation. Fluorine-containing thiocyanate derivatives 2 and 3 were threefold more potent than our lead drug 1 against intracellular T. cruzi. The biological evaluation against T. gondii was also very promising. The IC(50) values corresponding to 2 and 3 were at the very low micromolar level against tachyzoites of T. gondii. Both of these drugs are interesting examples of effective antiparasitic agents that have outstanding potential not only as lead drugs but also to be used for further in vivo studies.     

Water level fluctuations in Lake Baringo,Kenya, during the 19th and 20th centuries: Evidence from lake sediments

This article presents a fossil diatom-based, semi-quantitative reconstruction of water level fluctuations for Lake Baringo over the past 200 years as a consequence of climatic variations. A 285 cm long sediment core sample was collected using a Rod-Operated Single-drive Stationary Piston corer. Lake level was inferred using indices based on the proportion of planktonic to benthic diatom taxa (P/B ratio). The sediment archive presented distinct zones dominated by planktonic and benthic diatom flora. An initial transgression in the early 19th century was characterised as a shallow water environment dominated by planktonic Aulacoseira spp. This was a response to extreme drought during the late 18th to early 19th century. Mid-19th century was defined by a high lake stand. The late 19th to early 20th centuries experienced low water level following the widely documented aridity at the time. The mid-20th century was marked by a spectacular rise in water level that coincided with remarkably wet years during the early 1960s and late 1970s. The first decade of the 21st century witnessed widespread changes in water level. The proxy records show that lake ramping and drawdown over the years follow approximately 50-year climatic cycles.     

Kinetic model for isometric contraction in smooth muscle on the basis of myosin phosphorylation hypothesis.

A kinetic model was proposed to simulate an isometric contraction curve in smooth muscle on the basis of the myosin phosphorylation hypothesis. The Ca2+-calmodulin-dependent activation of myosin light-chain kinase and the phosphorylation-dephosphorylation reaction of myosin were mathematically treated. Solving the kinetic equations at a steady state, we could calculate the relationship between the Ca2+ concentration and the myosin phosphorylation. Assuming that two-head-phosphorylated myosin has an actin-activated Mg2+-ATPase activity and that this state corresponds to an active state, we computed the time courses of the myosin phosphorylation and the active state for various Ca2+ transients. The time course of the active state was converted into that of isometric tension by use of Sandow's model composed of a contractile element and a series elastic component. The model could simulate not only the isometric contraction curves for any given Ca2+ transient but also the following experimental results: the calmodulin-dependent shift of the Ca2+ sensitivity of isometric tension observed in skinned muscle fibers, the disagreement between the Ca2+ sensitivity of myosin phosphorylation and that of isometric tension at a steady state, and the disagreement between the time course of myosin phosphorylation and that of isometric tension development.