A review of tricaine methanesulfonate for anesthesia of fish

Tricaine methanesulfonate (TMS) is an anesthetic that is approved for provisional use in some jurisdictions such as the United States, Canada, and the United Kingdom (UK). Many hatcheries and research studies use TMS to immobilize fish for marking or transport and to suppress sensory systems during invasive procedures. Improper TMS use can decrease fish viability, distort physiological data, or result in mortalities. Because animals may be anesthetized by junior staff or students who may have little experience in fish anesthesia, training in the proper use of TMS may decrease variability in recovery, experimental results and increase fish survival. This document acts as a primer on the use of TMS for anesthetizing juvenile salmonids, with an emphasis on its use in surgical applications. Within, we briefly describe many aspects of TMS including the legal uses for TMS, and what is currently known about the proper storage and preparation of the anesthetic. We outline methods and precautions for administration and changes in fish behavior during progressively deeper anesthesia and discuss the physiological effects of TMS and its potential for compromising fish health. Despite the challenges of working with TMS, it is currently one of the few legal options available in the USA and in other countries until other anesthetics are approved and is an important tool for the intracoelomic implantation of electronic tags in fish.     

Selectivity is species-dependent: Characterization of standard agonists and antagonists at human,rat, and mouse adenosine receptors

Adenosine receptors (ARs) have emerged as new drug targets. The majority of data on affinity/potency and selectivity of AR ligands described in the literature has been obtained for the human species. However, preclinical studies are mostly performed in mouse or rat, and standard AR agonists and antagonists are frequently used for studies in rodents without knowing their selectivity in the investigated species. In the present study, we selected a set of frequently used standard AR ligands, 8 agonists and 16 antagonists, and investigated them in radioligand binding studies at all four AR subtypes, A₁, A_2A, A_2B, and A₃, of three species, human, rat, and mouse. Recommended, selective agonists include CCPA (for A₁AR of rat and mouse), CGS-21680 (for A_2A AR of rat), and Cl-IB-MECA (for A₃AR of all three species). The functionally selective partial A_2B agonist BAY60-6583 was found to additionally bind to A₁ and A₃AR and act as an antagonist at both receptor subtypes. The antagonists PSB-36 (A₁), preladenant (A_2A), and PSB-603 (A_2B) displayed high selectivity in all three investigated species. MRS-1523 acts as a selective A₃AR antagonist in human and rat, but is only moderately selective in mouse. The comprehensive data presented herein provide a solid basis for selecting suitable AR ligands for biological studies.

Electronic supplementary material

The online version of this article (doi:10.1007/s11302-015-9460-9) contains supplementary material, which is available to authorized users.     

Effects of Karenia brevis diet on RNA:DNA ratios and egg production of Acartia tonsa

Karenia brevis is a harmful alga associated with deleterious effects on zooplankton, but the exact cause (e.g. toxin, nutritional inadequacy or starvation) of these adverse effects is not clear. RNA:DNA ratios, fecundity and fecal pellet production of Acartia tonsa were measured on mono-algal and mixed-algal culture diets of K. brevis and Peridinium foliaceum to examine the usefulness of RNA:DNA ratios as an indicator of nutrition and to determine if adverse effects of K. brevis are due to the presence of toxins, poor nutritional quality or starvation. RNA:DNA ratios and egg production values were significantly higher for 100% P. foliaceum diet compared to 100% K. brevis diet. Significant differences in egg production, but not RNA:DNA ratios, were found between the various mixed diets, suggesting egg production is a more sensitive indicator of nutritional quality than RNA:DNA ratios. Changes in RNA:DNA ratios, fecundity and fecal pellet production of copepods fed two different toxic K. brevis strains were nearly identical, indicating that the presence of brevetoxins has little affect on A. tonsa. The similarity in RNA:DNA ratios, egg production, percent hatching and fecal production between the 100% K. brevis diet and starved copepods suggests that A. tonsa does not consume K. brevis when offered as its sole food source.     

Ecdysone metabolism in the host-parasitoid-system Trichoplusia ni/Chelonus sp

In unparasitized 4th and 5th-instar larvae of Trichoplusia ni and in 4th-instar larvae parasitized by Chelonus sp. 20-hydroxyecdysone, 20,26-dihydroxyec-dysone, and 20-hydroxyecdysonoic acid were the predominant metabolites formed 2 h after injection of [³H]ecdysone. Other unidentified metabolites were seen, but none seemed to be specific for either parasitized or unparasitized larvae. The major difference between parasitized and unparasitized larvae was seen with respect to the quantity of apolar (unidentified) and polar metabolites (20-hydroxyecdysonoic acid and unidentified ones), which were produced to a greater extent in parasitized larvae. Ecdysone was rapidly converted into 20-hydroxyecdysone and the other polar metabolites in all stages investigated, and the parasitoid seemed not to affect the conversion of ecdysone into 20-hydroxyecdysone. When analyzing the fate of [³H]ecdysone in host and parasite separately, at a stage when the parasite drinks hemolymph of its host, we observed that 10–20% of the radioactivity was recovered from the parasitoid. Analysis of the parasitoid's ecdysteroids revealed that ecdysone and 20-hydroxyecdysone represented only a small proportion of the recovered labeled ecdysteroids, the majority being apolar and polar metabolites. Our data suggest that the parasitoid takes up ecdysteroids from its host, converts them, and to some extent releases apolar metabolites into the host.     

Nitrification in terrestrial hot springs of Iceland and Kamchatka

Archaea have been detected recently as a major and often dominant component of the microbial communities performing ammonia oxidation in terrestrial and marine environments. In a molecular survey of archaeal ammonia monooxygenase (AMO) genes in terrestrial hot springs of Iceland and Kamchatka, the amoA gene encoding the alpha-subunit of AMO was detected in a total of 14 hot springs out of the 22 investigated. Most of these amoA-positive hot springs had temperatures between 82 and 97 degrees C and pH range between 2.5 and 7. In phylogenetic analyses, these amoA genes formed three independent lineages within the known sequence clusters of marine or soil origin. Furthermore, in situ gross nitrification rates in Icelandic hot springs were estimated by the pool dilution technique directly on site. At temperatures above 80 degrees C, between 56 and 159 mumol NO(3)(-) L(-1) mud per day was produced. Furthermore, addition of ammonium to the hot spring samples before incubation yielded a more than twofold higher potential nitrification rate, indicating that the process was limited by ammonia supply. Our data provide evidence for an active role of archaea in nitrification of hot springs in a wide range of pH values and at a high temperature.     

The telomeric region of BTA18 containing a potential QTL region for health in cattle exhibits high similarity to the HSA19q region in humans

We have applied a targeted physical mapping approach, based on the isolation of bovine region-specific large-insert clones using homologous human sequences and chromosome microdissection, to enhance the physical gene map of the telomeric region of BTA18 and to prove its evolutionary conservation. The latter is a prerequisite to exploit the dense human gene map for future positional cloning approaches. Partial sequencing and homology search were used to characterize 20 BACs targeted to the BTA18q2.4-q2.6 region. We used fluorescence in situ hybridization (FISH) to create physical maps of 11 BACs containing 15 gene loci; these BACs served as anchor loci. Using these approaches, 12 new gene loci (CKM, STK13, PSCD2, IRF3, VASP, ACTN4, ITPKC, CYP2B6, FOSB, DMPK, MIA, SIX5) were assigned on BTA18 in the bovine cytogenetic map. A resolved physical map of BTA18q2.4-q2.6 was developed, which encompasses 28 marker loci and a comparative cytogenetic map that contains 15 genes. The mapping results demonstrate the high evolutionary conservation between the telomeric region of BTA18q and HSA19q.     

Early Detection of Erlotinib Treatment Response in NSCLC by 3′-Deoxy-3′-[18F]-Fluoro-L-Thymidine ([18F]FLT) Positron Emission Tomography (PET)

Background

Inhibition of the epidermal growth factor receptor (EGFR) has shown clinical success in patients with advanced non-small cell lung cancer (NSCLC). Somatic mutations of EGFR were found in lung adenocarcinoma that lead to exquisite dependency on EGFR signaling; thus patients with EGFR-mutant tumors are at high chance of response to EGFR inhibitors. However, imaging approaches affording early identification of tumor response in EGFR-dependent carcinomas have so far been lacking.

Methodology/Principal Findings

We performed a systematic comparison of 3′-Deoxy-3′-[¹⁸F]-fluoro-L-thymidine ([¹⁸F]FLT) and 2-[¹⁸F]-fluoro-2-deoxy-D-glucose ([¹⁸F]FDG) positron emission tomography (PET) for their potential to identify response to EGFR inhibitors in a model of EGFR-dependent lung cancer early after treatment initiation. While erlotinib-sensitive tumors exhibited a striking and reproducible decrease in [¹⁸F]FLT uptake after only two days of treatment, [¹⁸F]FDG PET based imaging revealed no consistent reduction in tumor glucose uptake. In sensitive tumors, a decrease in [¹⁸F]FLT PET but not [¹⁸F]FDG PET uptake correlated with cell cycle arrest and induction of apoptosis. The reduction in [¹⁸F]FLT PET signal at day 2 translated into dramatic tumor shrinkage four days later. Furthermore, the specificity of our results is confirmed by the complete lack of [¹⁸F]FLT PET response of tumors expressing the T790M erlotinib resistance mutation of EGFR.

Conclusions

[¹⁸F]FLT PET enables robust identification of erlotinib response in EGFR-dependent tumors at a very early stage. [¹⁸F]FLT PET imaging may represent an appropriate method for early prediction of response to EGFR TKI treatment in patients with NSCLC.     

Molecular analysis of PKU haplotypes in the population of southern Poland

Summary Out of a population of 138598 infants born in southern Poland between 1987 and 1989, and screened for phenylketonuria (PKU), 28 cases were ascertained giving an incidence of 1 in 5000. DNA from 22 of these probands and their parents was isolated and eight polymorphic restriction sites were analyzed within the phenylalanine hydroxylase gene region. Twenty-one different haplotypes (HT) were revealed, five of them representing new categories. The most common haplotypes among those carrying normal alleles were: HT1 (27.3%) and HT4 (11.4%). Within the group of haplotypes with mutant alleles the most frequent was HT2 (56.8%), whereas the frequency of this haplotype in other European populations, such as French, Danish and German, ranged from 12% to 24%. HT3, being the most common in Danish (38%), and relatively frequent in the other western European (13–14%) populations, appeared to be very rare in our sample (2.3%). The mutation of codon 408 (exon 12, CT, ArgTrp), which has been described to be tightly linked to HT2, was tested on amplified DNA by dot-blot hybridization. This mutation was found in 25 out of 44 proband chromosomes. In one case it was linked to HT5, in the remaining 24 to HT2. Our results confirm molecular heterogeneity of PKU haplotypes, as well as their significant interpopulation variation.     

Nitrogenase activity in composting horse bedding and leaves

To date little or no nitrogen fixation has been found in composting materials. However, some materials have high C:N ratios and thus might be expected to support nitrogen fixation. We examined windrows of composting horse bedding and leaves for nitrogenase activity via acetylene reduction and used carbon monoxide controls to differentiate ethylene evolved by nitrogenase from that evolved by endogenous processes. In both piles temperatures were substantially elevated and partial pressures of oxygen greatly reduced thus indicating vigorous microbial activity. In the horse bedding, there were only low rates of ethylene evolution and little of this was due to nitrogen fixation. In contrast, much higher rates of ethylene evolution were measured in the leaf pile and 94% of this was due to nitrogenase activity. We estimate that the leaf pile fixed approximately 0.062 mg N/g leaves during a 5-week period.