AsKC11, a Kunitz Peptide from Anemonia sulcata,Is a Novel Activator of G Protein-Coupled Inward-Rectifier Potassium Channels

(1) Background: G protein-coupled inward-rectifier potassium (GIRK) channels, especially neuronal GIRK1/2 channels, have been the focus of intense research interest for developing drugs against brain diseases. In this context, venom peptides that selectively activate GIRK channels can be seen as a new source for drug development. Here, we report on the identification and electrophysiological characterization of a novel activator of GIRK1/2 channels, AsKC11, found in the venom of the sea anemone Anemonia sulcata. (2) Methods: AsKC11 was purified from the sea anemone venom by reverse-phase chromatography and the sequence was identified by mass spectrometry. Using the two-electrode voltage-clamp technique, the activity of AsKC11 on GIRK1/2 channels was studied and its selectivity for other potassium channels was investigated. (3) Results: AsKC11, a Kunitz peptide found in the venom of A. sulcata, is the first peptide shown to directly activate neuronal GIRK1/2 channels independent from Gi/o protein activity, without affecting the inward-rectifier potassium channel (IRK1) and with only a minor effect on K_V1.6 channels. Thus, AsKC11 is a novel activator of GIRK channels resulting in larger K⁺ currents because of an increased chord conductance. (4) Conclusions: These discoveries provide new insights into a novel class of GIRK activators.     

Proteinase, lipase and amylase activities in the small intestine of pigs suffering from colienterotoxaemia.

The activities of proteinases, lipases, amylases and the activities of proteinase inhibitors, as well as the numbers of Escherichia coli in the contents from the small intestine were examined for pigs suffering from colienterotoxaemia and for healthy pigs. Enzyme activities were determined using an agar diffusion test.Compared with healthy animals the activities of proteinases and amylases in diseased animals were reduced while lipases showed increased activity. In pathologically changed contents showing large numbers of E. coli, proteinases could not be demonstrated; however, proteinase inhibitors were found in these contents. In healthy animals, proteinase inhibitors were not demonstrated in ingesta-con-taining contents.In diseased animals, E. coli were found in large numbers in all parts of the small intestine. In healthy animals, E. coli was demonstrated especially in the posterior part of the small intestine and regularly in small numbers.The possible influence of digestive enzymes, especially proteinases and their inhibitors, on enterotoxins from E. coli is discussed.     

The Applicability of Agar-Diffusion Test in Measuring Deoxyribonucleases in Pig Intestinal Content

The suitability of an agar-diffusion test (ADT) using toluidine blue deoxyribonucleic acid agar (TDA) for measuring DNase activity in pig intestinal contents was investigated. The ADT was compared with a spectrophotometrical method. Distinct metachromatic zones around wells in the DNA-containing agar, into which the intestinal content was applied, indicated DNase activity. The DNase activity was determined semiquantitatively by making serial twofold dilutions of the intestinal content. The spectrophotometrical method was optimal at pH 7.2. The ADT proved to be most sensitive at pH 5.6. The ability of the 2 methods employed to measure low concentrations of DNases was equal. However, the ADT was considered more suitable than the spectrophotometrical method because ADT measured reduced amounts of enzyme. DNase activity was demonstrated throughout the small intestine and in the large intestine. By the zymogram technique, at least 3 different DNases could be demonstrated in the lower parts of the small intestine, 1 of which could be of extrapancreatic origin.     

Effects of Oscillatoria agardhii-toxins on blood pressure and isolated organ preparations

Toxic extracts of the blue-green alga Oscillatoria agardhii were tested for their effect on various isolated laboratory animal preparations. The Oscillatoria-toxins did not show hemagglutinative or hemolytical properties. An immediate rise in blood pressure was seen after intravenous injection in rats. As regards isolated organs, effects were most marked on the liver. Addition of toxic extract to isolated liver perfusion fluid led to physiological, biochemical and pathological changes indicating severe damage to the organ. Negative chronotropic and inotropic effects were found on the isolated heart. The toxins did not influence isolated smooth muscle or nerve striated muscle preparations.     

Relationship between production of enterotoxin and various drug resistance patterns in strains of Escherichia coli isolated from piglets suffering from colienterotoxaemia.

The relationship between production of enterotoxin and drug resistance patterns in 92 E. coli strains isolated from the jejunal content from piglets suffering from colienterotoxaemia was investigated. The drugs included were Oxytetracycline, streptomycin, sulphaisodimidin, neomycin, ampicillin and chloramphenicol. The frequency with which transmission of drug resistance occurred as well as the O-groups of these strains were also recorded.Fifty-one E. coli strains produced enterotoxin. Of these, 42 (approx. 82 %) produced the LT. The same number of strains also produced the ST. Strains simultaneously producing both enterotoxins thus constituted approx. 64 % of the enterotoxin-producing strains. The E. coli strains were divided in 7 groups, each representing different drug resistance patterns. Statistical analysis showed that the various categories of enterotoxin production mentioned above were more frequently found in strains simultaneously resistant to Oxytetracycline, streptomycin and sulphaisodimidin than in strains belonging to the other drug resistance groups in general.Transmission of drug resistance was demonstrated in all resistance pattern groups. Approx. 50–70 % of the enterotoxin-producing strains showed drug transmission. The O-groups 138 and 149 constituted 8 and 54 out of the 62 typable strains, respectively.     

Roe deer (Capreolus capreolus) browsing pressure affects yew (Taxus baccata) recruitment within nature reserves in Norway

Grazing and browsing by large herbivores is known to affect patterns of biodiversity. Still much remains to be done to assess the specific role of browsing on single, red listed plant species. In Norway, the yew (Taxus baccata) is a threatened species and several nature reserves have been established. We show, that within two nature reserves between 1988 and 2003, yew recruitment was strongly dependent on variation in roe deer (Capreolus capreolus) browsing pressure. Our study emphasized that establishing nature reserves for yew is not enough to conserve this species unless control over browsing roe deer population is also taken. We argue that roe deer, being the most abundant large herbivore in Europe, may have a larger effect on conservation than currently realized.     

Drug resistance in strains of Escherichia coli isolated from the intestinal tract of pigs in Norway.

Faecal samples from 95 healthy pigs and samples of jejunal content from 85 piglets suffering from colienterotoxaemia were tested for the presence of drug resistant E. coli strains. Practically all pigs in both groups harboured E. coli strains resistant to one or more of the 6 antibiotics/chemotherapeutic agents tested (Oxytetracycline, streptomycin, sulphaisodimidin, neomycin, ampicillin, chloramphenicol). Almost 100% of healthy and approx. 90% of diseased pigs harboured strains resistant to Oxytetracycline, streptomycin and sulphaisodimidin. Pigs with strains resistant to neomycin, ampicillin and chloramphenicol were less frequently found. The predominant coliform flora consisted of E. coli strains” resistant to Oxytetracycline, streptomycin and sulphaisodimidin in 71% to 81% of diseased pigs and in 47% to 69% of the healthy pigs. In diseased pigs ¾ of the animals had a coliform flora dominated by neomycinresistant E. coli strains.Of the 721 resistant E. coli strains isolated from healthy pigs, 11% were single resistant while the corresponding figure for the 518 resistant strains isolated from diseased pigs was 6%. Thus 89% and 94% of strains showed simultaneous resistance to 2 or more antibiotics. E. coli strains resistant to 3 or more drugs were found in approx. 60% and 70% of the isolates from healthy and diseased animals, respectively. Oxytetracycline/streptomycin/sulphaisodimidin resistance was most commonly found, approx. 22% and 38% of the strains from healthy and diseased pigs, respectively, showing this resistance pattern.Transmission of drug resistance which was examined in E. coli strains originating from the diseased pigs was demonstrated in approx. 76% of the isolates. The incidence of drug resistance transfer in single, double, triple and quadruple resistant strains was 11%, 68%, 97% and 98%, respectively.     

Fimbriae in Escherichia coli isolated from the small intestine of piglets

Ninety E. coli strains, isolated from piglets which had died from neonatal diarrhea, were tested for the presence of K88, K99, 987P and type 1 fimbriae. Two or more types of fimbriae were demonstrated in 14 of the strains, a single fimbria! type in 44 strains while in 32 strains no fimbriae were detected. Of the 14 E. coli strains with more than 1 type of fimbriae, 12;, 10, 8 and 4 strains showed K88, K99, 987P and type 1, respectively.Twelve E. coli strains were isolated from piglets which had died in the neonatal period without showing signs of neonatal diarrhea at necropsy. One strain showed 987P and 3 strains showed type 1 fimbriae, while the remaining 8 strains were unfimbriated.Sixteen fimbriated E. coli strains were subcultured in order to examine the stability of fimbrial expression in the strains. The K88 and the type 1 fimbriae were regularly expressed, while the K99 and 987P were inconsistently demonstrated.