Acetylcholine activates cerebral interneurons and feeding motor program in Limax maximus

The cellular and network effects of acetylcholine (ACh) on the control system for feeding in Limax maximus were measured by intracellular recordings from feeding command-like interneurons and whole nerve recordings from buccal ganglion motor nerve roots that normally innervate the ingestive feeding muscles. The buccal ganglion motor nerve root discharge pattern that causes rhythmic feeding movements, termed the feeding motor program (FMP), was elicited either by attractive taste solutions applied to the lip chemoreceptors or by ACh applied to the cerebral ganglia. The ability of exogenous ACh applied to the cerebral ganglia to trigger FMP was blocked by the cholinergic antagonists curare and atropine. If the strength of the lip-applied taste stimulus was in the range of 1-2 times threshold, cerebral application of the cholinergic antagonists blocked or greatly decreased the ability of lip-applied taste solutions to trigger FMP (5 of 8 trials). The cerebral feeding interneurons, some of which activate FMP when stimulated intracellularly, are excited by small pulses of ACh applied directly to the cell body from an ACh-filled micropipette. A pulse of ACh that activates several of the feeding interneurons simultaneously triggers FMP. The data suggest that under certain stimulus conditions an obligatory set of cholinergic synapses onto the feedininterneurons must be activated for taste inputs to trigger ingestion. The determination of ACh's action within the feeding control system is necessary for understanding how enhanced cholinergic transmission leads to prolonged associative memory retention (Sahley, et al., 1986).     

Role of special stains in the diagnosis of Pneumocystis carinii infection from bronchial washing specimens in patients with the acquired immune deficiency syndrome

Fifty bronchial washing specimens from 36 patients with acquired immune deficiency syndrome (AIDS) were retrospectively reviewed to assess the sensitivity of the various special stains used to diagnose Pneumocystis carinii. In 76% of the cases, the Diff-Quik stain was positive; it was the easiest and most rapid of the special stains used. The sensitivity was increased to 92%, 96% and 100%, respectively, by also doing cresyl echt violet, Grocott's Gomori methenamine silver and both the cresyl violet and Grocott stains in addition to the Diff-Quik stain. We conclude that the Diff-Quik stain is a fairly reliable and rapid screening procedure for making the diagnosis of Pneumocystis infection in bronchial washings from AIDS patients. The routine Papanicolaou stain gave less sensitive results in the smears of the washing specimens, but does give a markedly improved yield in bronchoalveolar lavage specimens.     

Post-ingestive food-aversion learning to amino acid deficient diets by the terrestrial slugLimax maximus

Summary An agar-based artificial diet containing carbohydrates, fats and twenty amino acids was constructed (Fig. 1). This diet is highly palatable and nutritionally complete forLimax maximus as demonstrated by significant ingestion on first encounter, consistent ingestion on subsequent days and good growth of young slugs fed this diet. Removing methionine, an essential amino acid, from the complete diet produces a food which is initially as palatable as the complete diet, but after one day's intake the amount of this deficient diet eaten is greatly reduced (Fig. 2). Removing alanine, a nonessential amino acid, does not produce any decrement in feeding relative to the complete diet (Fig. 5).A single meal can be sufficient for establishing the aversion to the deficient diet (Fig. 5). Following seven days of feeding on the deficient diet the aversion is retained with little or no attenuation for at least 30 days (Fig. 3) and does not generalize to either a known safe food (Fig. 3) or a novel food (Fig. 4). Evidence of a mild neophobia towards the artificial diet which attenuated after one or two meals was seen (Fig. 5).The learned aversion to the deficient diet is reversible if slugs are repeatedly fed the complete diet following feeding on the deficient diet. Also, slugs initially fed the complete diet will develop an aversion to the methionine-deficient diet after sampling it (Fig. 7).Slugs readily ate the artificial diets when these were offered 7 days post-hatch. The methionine-deficient diet however was not eaten in large amounts after the first meals and did not support growth (Fig. 9). Baby slugs fed the methionine-deficient diet for 10 days and then maintained on rat chow ate only small amounts when the deficient diet was presented again 126 days later, while baby slugs fed the complete diet or an alanine deficient diet for 10 days ate large amounts when these diets were presented 126 days later (Fig. 11 b).Supplementing the methionine-deficient diet with an injection of methionine into the haemocoel one hour after the completion of a meal completely blocks the development of a learned aversion while injection ofLimax saline does not (Fig. 8).These results are best explained by the hypothesis that the slugs acquire, post-ingestively, an aversion to the taste and probably the odor of the diet as the result of associative learning. The results of the experiments reported here indicate that there are substantial parallels at the behavioral level between mollusc and mammal with respect to post-ingestive feedback learning.     

Oligonucleotides with novel, cationic backbone substituents: aminoethylphosphonates.

Oligonucleotide (2-aminoethyl)phosphonates in which the backbone consisted of isomerically pure, alternating (2-aminoethyl)-phosphonate and phosphodiester linkages have been prepared and characterized. One of these single isomer oligonucleotides (Rp) formed a more stable duplex with DNA or RNA than its corresponding natural counterpart. Hybrid stability was more pH-dependent, but less salt-dependent than a natural duplex. The specificity of hybridization was examined by hybridization of an oligonucleotide containing one (2-aminoethyl)phosphonate to oligonucleotides possessing mismatches in the region opposite to the aminoethyl group. In contrast to oligonucleotides containing (aminomethyl)-phosphonate linkages, oligonucleotide (2-aminoethyl)phosphonates were completely stable to hydrolysis in aqueous solution. These oligonucleotides were resistant to nuclease activity but did not induce RNase H mediated cleavage of a complementary RNA strand. Incubation in a serum-containing medium resulted in minimal degradation over 24 hours. Studies of cell uptake by flow cytometry and confocal microscopy demonstrated temperature dependent uptake and intracellular localization. (2-Aminoethyl)phosphonates represent a novel approach to the introduction of positive charges into the backbone of oligonucleotides.     

Sequence variation in the outer-surface-protein genes of Borrelia burgdorferi

Borrelia burgdorferi is a spirochete pathogen transmitted among warm- blooded hosts by ixodid ticks. Frequency-dependent selection for variant outer-surface proteins might be expected to arise in this species, since rare variants are more likely to avoid immune surveillance in previously infected hosts. We sequenced the OspA and OspB genes of nine North American strains and compared them with nine strains previously described. For each gene, the mean number of synonymous substitutions per synonymous site and the mean number of nonsynonymous substitutions per nonsynonymous site show only a twofold excess of silent mutations. Synonymous rates vary widely along the OspB protein. Some regions show a significant excess of silent substitutions, while divergence in other regions is constrained by biased base composition or selection. The presence, in antigenically important regions of the protein, of significant variation among strains, as well as evidence for recombination among strains, should be considered in attempts to develop vaccines against this disease.      

Characterisation of a 34 kD protein from potato cyst nematodes, using monoclonal antibodies with potential for species diagnosis

Two monoclonal antibodies, which differentially recognise the two species of potato cyst nematodes (PCN), Globodera pallida and G. rostochiensis, are described. They have been shown to have potential for quantification of these two species, recognising proteins of the same molecular weight (34 kD) in both species. Further investigation showed these proteins to have isoelectric points at pH values of 5.7 in G. pallida and 5.9 in G. rostochiensis, in common with the proteins used by Fleming & Marks (1983) to differentiate the species of PCN. They are likely to be structurally very similar, with the same physiological function (and therefore similar concentrations) in the two species. In cross-reactivity tests with a wide range of soil nematode species, the antibodies reacted strongly only with species of the genus Globodera, and thereby confirmed their potential as the basis of a quantitative immunoassay likely to be useful in management of PCN populations.     

Benzothiadiazole induces disease resistance in Arabidopsis by activation of the systemic acquired resistance signal transduction pathway

Benzothiadiazole (BTH) is a novel chemical activator of disease resistance in tobacco, wheat and other important agricultural plants. In this report, it is shown that BTH works by activating SAR in Arabidopsis thaliana. BTH-treated plants were resistant to infection by turnip crinkle virus, Pseudomonas syringae pv ‘tomato’ DC3000 and Peronospora parasitica. Chemical treatment induced accumulation of mRNAs from the SAR-associated genes, PR-1, PR-2 and PR-5. BTH treatment induced both PR-1 mRNA accumulation and resistance against P. parasitica in the ethylene response mutants, etr1 and ein2, and in the methyl jasmonate-insensitive mutant, jar1, suggesting that BTH action is independent of these plant hormones. BTH treatment also induced both PR-1 mRNA accumulation and P. parasitica resistance in transgenic Arabidopsis plants expressing the nahG gene, suggesting that BTH action does not require salicylic acid accumulation. However, because BTH-treatment failed to induce either PR-1 mRNA accumulation or P. parasitica resistance in the non-inducible immunity mutant, nim1, it appears that BTH activates the SAR signal transduction pathway.