Chemical immobilization of free-ranging dhole (Cuon alpinus), binturong (Arctictis binturong), and yellow-throated marten (Martes flavigula) in Thailand

No published information exists on the chemical immobilization of free-ranging dholes (Cuon alpinus), binturongs (Arctictis binturong), or yellow-throated martens (Martes flavigula). We chemically immobilized these species in Thailand using a mixture of ketamine hydrochloride with xylazine hydrochloride (KH–XH) and tiletamine hydrochloride with zolazepam hydrochloride (TH–ZH). Mean (±SD) dose of KH–XH was 18.1±5.3 and 0.9±0.1 mg/kg for dholes (n=2), 19.7±4.1 and 1.3±0.4 mg/kg for binturongs (n=8), and 28.7±3.3 and 1.1±0.1 mg/kg for yellow-throated martens (n=5), respectively. Mean dose of TH–ZH was 4.1±0.1 mg/kg for dhole (n=2). Induction time, duration of anesthesia, and recovery time were satisfactory for standard field research procedures including radio-collaring, although the effects on yellow-throated martens and binturongs were more variable. Respiration and muscle rigidity were monitored during sedation with no observed adverse physiological effects. Individuals were released after full recovery and monitored via radio telemetry for 4–23 months with no observed detrimental effects due to chemical immobilization. We conclude that KH–XH and TH–ZH are safe and effective immobilization agents for these carnivores; however, we suggest testing different KH–XH ratios and dosages, and other immobilizing agents for these species.     

Dmt1, d-1-Nal3]morphiceptin, a novel opioid peptide analog with high analgesic activity

The morphiceptin-derived peptide [Dmt1, d-1-Nal3]morphiceptin, labeled mu-opioid receptor (MOP) with very high affinity and selectivity in the receptor binding assays. In the mouse hot plate test, [Dmt1, d-1-Nal3]morphiceptin given intracerebroventricularly (i.c.v.) produced profound supraspinal analgesia, being approximately 100-fold more potent than the endogenous MOP receptor ligand, endomorphin-2. The antinociceptive effect of this new analog lasted up to 120min. Thus, [Dmt1, d-1-Nal3]morphiceptin is an interesting and extraordinarily potent analgesic, raising the possibility of novel approaches in the design of clinically useful drugs for pain treatment.     

Effective antagonists of luteinizing hormone releasing hormone modified at position one

Summary The amino acid, D-2-naphthylalanine, has been used by many investigators as a substituent for position one of antagonists of LHRH. We have newly designed substituents for position one in which the carboxy groups of 2-naphthoic acid, 3-quinoline- and 2-quinoxaline-carboxylic acids are linked to the five amino acids, DAla, DThr, DNVal, DSer, and Gly. The substituents in positions 2–10 were DpClPhe², DPal³, Ser⁴, PicLys⁵, DPicLys⁶, Leu⁷, ILys⁸, Pro⁹, DAlaNH₂ ¹⁰.Remarkably, DThr, acylated on the amino group by 3-quinolinecarboxylic acid or by 3-quinoxalinecarboxylic acid, and introduced into position one of a relatively potent antagonist, gave a new class of antagonists of LHRH, which released as little histamine as yet recorded, and yet possessed reasonable antiovulatory activity and greatly improved solubility.These structure-activity results advance the basic knowledge of understanding the structural features of such decapeptides which cause antiovulatory activity and histamine release.Abbreviations ILys N -isopropyllysine; - 1-Nal 3-(1-naphthyl)alanine - 2-Nal 3-(2-naphthyl)alanine - Nap 2-naphthoic acids - NicLys N -nicotinoyllysine; - Pal 3-(3-pyridyl)alanine - pClPhe 3-(4-chlorophenyl)alanine - PicLys N -picolinoyllysine - c-PzACAla cis-3-(4-pyrazinylcarbonylaminocyclohexyl) alanine - 3-Qal 3-(3-quinolyl)alanine - Qui 3-quinolinecarboxylic acid - Qux 2-quinoxalinecarboxylic acid     

Novel highly potent mu-opioid receptor antagonist based on endomorphin-2 structure

The mu-opioid agonists endomorphin-1 (Tyr-Pro-Trp-Phe-NH(2)) and endomorphin-2 (Tyr-Pro-Phe-Phe-NH(2)) exhibit an extremely high selectivity for the mu-opioid receptor and thus represent a potential framework for modification into mu-antagonists. Here we report on the synthesis and biological evaluation of novel [d-2-Nal(4)]endomorphin-2 analogs, [Sar(2),d-2-Nal(4)]endomorphin-2 and [Dmt(1),Sar(2),d-2-Nal(4)]endomorphin-2 (Dmt=2'6'-dimethyltyrosine; Sar=N-methylglycine, sarcosine; d-2-Nal=3-(2-naphthyl)-d-alanine). [Dmt(1),Sar(2),d-2-Nal(4)]endomorphin-2 possessed very high affinity for the mu-opioid receptor (IC(50)=0.01+/-0.001 nM) and turned out to be a potent and extremely selective mu-opioid receptor antagonist, as judged by the in vitro aequorin luminescence-based calcium assay (pA(2)=9.19). However, in the in vivo hot plate test in mice this analog was less potent than our earlier mu-opioid receptor antagonist, [Dmt(1),d-2-Nal(4)]endomorphin-2 (antanal-2). The exceptional mu-opioid receptor in vitro activity and selectivity of [Dmt(1), Sar(2),d-2-Nal(4)]endomorphin-2 makes this analog a valuable pharmacological tool, but further modifications are needed to improve its in vivo profile.     

Synthesis of novel morphiceptin analogues modified in position 3 and their binding to mu-opioid receptors in experimental mammary adenocarcinoma

Binding of the (125)I-labeled mu-opioid receptor selective ligands, morphiceptin (Tyr-Pro-Phe-Pro-NH(2)) and [D-Phe(3)]morphiceptin, to membranes isolated from experimental mouse mammary adenocarcinoma was examined in vitro using a cross-linking assay followed by a Western blot technique. The radioactive complex had a molecular weight of about 65 kDa and was detectable by anti-mu-opioid receptor antibody, indicating the presence of mu-opioid receptors in tumor membranes. A series of new morphiceptin analogues, modified at the pharmacophoric position 3, was synthesized in order to find the correlation between the lipophilicity, electronic and steric properties of the amino acid in this position and the in vitro affinity of new analogues for mu-opioid receptors on mouse brain and tumor membranes. In in vivo studies the uptake of (131)I-labeled analogues by experimental mammary adenocarcinoma was estimated. The highest affinity for mu-opioid receptors in both, in vitro and in vivo experiments was observed for [D-Phe(3)]morphiceptin and [D-ClPhe(3)]-morphiceptin.     

Binding of endomorphin-2 to mu-opioid receptors in experimental mouse mammary adenocarcinoma.

Endomorphin-2 (Tyr-Pro-Phe-Phe-NH2) binds with high affinity and selectivity to the mu-opioid receptor. In the present study, [125I]endomorphin-2 has been used to characterize mu-opioid-binding sites on transplantable mouse mammary adenocarcinoma cells. Cold saturation experiments performed with [125I]endomorphin-2 (1 nM) show biphasic binding curves in Scatchard coordinates. One component represents high affinity and low capacity (K(d) = 18.79 +/- 1.13 nM, B(max) = 635 +/- 24 fmol/mg protein) and the other shows low affinity and higher capacity (K(d) = 7.67 +/- 0.81 microM, B(max) = 157 +/- 13 pmol/mg protein) binding sites. The rank order of agonists competing for the [125I]endomorphin-2 binding site was [d-1-Nal3]morphiceptin > endomorphin-2 > [d-Phe3]morphiceptin > morphiceptin > [d-1-Nal3]endomorphin-2, indicating binding of these peptides to mu-opioid receptors. The uptake of 131I-labeled peptides administered intraperitoneally to tumor-bearing mice was also investigated. The highest accumulation in the tumor was observed for [d-1-Nal3)morphiceptin, which reached the value of 8.19 +/- 1.14% dose/g tissue.     

Spantide III,a superior tachykinin antagonist with high potency and negligible neurotoxicity

Summary Five new antagonists of Substance P were designed and synthesized toward increasing potency and safety. One of them was more effective than Spantide II, which was the basis for the design. It was named Spantide III and has the structure: D-NicLys,Pro,Pal,Pro,D-Cl₂Phe,Asn,D-Trp,Phe,D-Pal,Leu, NleNH₂.     

Production and characteristics of human tissue plasminogen-activator antibodies with region-oriented synthetic peptides.

We selected six peptide sequences as belonging to potential epitopes of tissue plasminogen activator (tPA) using, as the main criterion for their choice, the location of the peptide sequences on the surface of the protein molecule. The six peptides (corresponding to amino acids 4-8, 11-16, 96-101, 272-277, 371-376 and 514-519) were synthesized, coupled to carrier proteins and injected into rabbits. All of these peptides elicited antibodies and 15-75% binding of the corresponding iodinated peptide was obtained with a 1:100 dilution of antiserum. Only two anti-(peptide) sera [anti-(tPA96-101) and anti-(tPA272-277)] reacted with intact tPA and its heavy chain in Western immunoblotting analysis. These two peptides sequences and fragment tPA11-16 appear to be involved in the structure of native antigenic epitopes of tPA, since they were recognized and antibodies present in antisera raised against native tPA. There was no interaction between anti-(tPA4-8) and anti-(tPA371-376) sera with intact one-chain or two-chain tPA. In the case of anti-(tPA4-8) cleavage of one-chain tPA to two-chain tPA and reduction of disulfide bonds exposed this epitope.     

A method of the rapid preparation of adenosine 5'-gamma-[32P] triphosphate by chemical synthesis.

A new chemical method for the synthesis of adenosine 5'-gamma-[32P] triphosphate has been developed based on the reaction of adenosine 5'-diphosphate with ethyl chloroformate. The resulting active mixed anhydride was able to react with [32P]-triethylammonium orthophosphate to give gamma-[32P]ATP.     

Genetic differences in the response to landscape fragmentation by a habitat generalist,the bobcat,and a habitat specialist,the ocelot

The ecology of a species strongly influences genetic variation and population structure. This interaction has important conservation implications because taxa with low dispersal capability and inability to use different habitats are more susceptible to anthropogenic stressors. Ocelots (Leopardus pardalis albescens) and bobcats (Lynx rufus texensis) are sympatric in Texas and northeastern Mexico; however, their ecology and conservation status are markedly different. We used 10 microsatellite loci and a 397-bp segment of the mitochondrial control region to examine how historical and ecological differences in these two species have influenced current patterns of genetic diversity in a landscape heavily altered by anthropogenic activities. Substantially higher genetic diversity (heterozygosity and haplotype diversity) and population connectivity was observed for bobcats in comparison to ocelots. The level of divergence among proximate ocelot populations (<30 km) was greater than between bobcat populations separated by >100 km. Ocelot populations in the US have never recovered from reductions experienced during the twentieth century, and their low genetic variation and substantial isolation are exacerbated by strong preference for dense native thornshrub and avoidance of open habitat. In contrast, despite continued legal harvesting and frequent road-related mortality, bobcats have maintained wide distribution, high abundance, and population connectivity. Our study illustrates that sympatric species with a similar niche can still have sufficient ecological differences to alter their response to anthropogenic change. Sensitive species, such as the ocelot, require additional conservation actions to sustain populations. Ecological differences among species occupying a similar guild are important to consider when developing conservation plans.