Taste systems of the petrosal ganglion of the rat glossopharyngeal nerve

Single unit recordings were taken from sensory ganglion cellsin the petrosal ganglion (PG) of the glossopharyngeal nerveof the rat. These taste units were examined with respect tospontaneous and evoked discharge patterns and responsivenessto a wide variety of chemical compounds, most of natural occurrence.Spontaneous activity patterns, with few exceptions, tended tobe extremely irregular with both bursting (clusters of 2–3spikes) and grouping (large groups of spikes as in evoked discharges).Most interspike interval histograms of spontaneous activitywere multimodal, similar to rat geniculate ganglion (GG) units.Evoked discharges usually displayed grouping of spikes, andlong latencies of onset and persistence of discharge after rinsewere sometimes seen. Little response was shown to nucleotidesor salts. Units responsive to amino acids tended to show largedischarge to only one or two amino acids; and the most responsiveamino acid usually varied from cell to cell. Units responsiveto alkaloids only responded to a few alkaloids with atropineand quinine being the most stimulatory. Units responsive toacids only discharged to a few of the acids tested and oftenacids of low pH elicited no discharge. Saccharin activated unitsresponsive to both sugar and alkaloids. A few units highly responsiveto both sugar and alkaloids were seen. The units were placedinto four clusters on the basis of chemicals activating themand certain neurophysiological characteristics: PG salt units,PG acid units and, tentatively, amino acid (sugar) units andX (alkaloid and alkaloid plus) units. The PG salt units didnot show the exclusive sensitivity to sodium and lithium compoundsas did the GG salt units. The PG acid units could also be differentiatedfrom the GG acid units. The petrosal amino acid and X units,on the other hand, could not be differentiated from similarunits in the rat GG.     

Stereochemical effects of non-ionic methylphosphonates on nucleotide conformations.

The preferred conformations of deoxyribo and ribonucleoside 3'-methylphosphonates are analysed by minimizing the conformational energy as a function of all the major parameters including the sugar ring for both the S- and R-isomers. The results show that neither the substitution nor the nature of the diastereomer affects significantly the preferred conformations compared to the naturally occurring nucleoside 3'-phosphates. The preferred range of C3'-O3' bond torsions or the phase angles of pseudorotation (P) of the sugar are unaffected. The chiral substitution on the phosphate always adopts a conformation distal to the secondary C3' carbon atom in the minimum energy conformational state. Further, it introduces certain restrictions on the preferred range of P-O3' torsions depending on the methylphosphonate configuration. Methylphosphonate, especially the S-isomer, renders the normal gauche- range of P-O3' bond torsions responsible for the stacked helical duplexes to be energetically unfavourable besides introducing a high energy barrier between trans and gauche conformations. Therefore it is suggested that duplexes with S-methylphosphonate may favour extended phosphodiester conformations. These factors explain the observed lower melting temperature as well as the downfield shifts in the 31P signals in duplexes containing the S-isomer.     

Neurophysiology of geniculate ganglion (facial nerve) taste systems: species comparisons

On the basis of various measures taken from geniculate gangliontaste neurons in four species, it was concluded that withineach species the neurons could be subdivided into distinct functionalgroups. In this report, the neural groups of different specieswere directly compared. Units from all four species were studiedwith a common test series of solutions in addition to otherstimuli. Since these stimuli were presented at the same concentrationsto all species, direct quantitative comparisons across specieswere possible for a wide range of chemical compounds. In addition,the neural groups were compared with respect to spontaneousand evoked activity measures, latency to electrical stimulation,and receptive field characteristics. These neurophysiologicaldata suggest a basic model of four distinct subgroups: acidunits, salt units, amino acid units, and X units.     

Stereochemical studies on nucleic acid analogues. I. Conformations of alpha-nucleosides and alpha-nucleotides: interconversion of sugar puckers via O4'-exo.

The preferred conformations of ribo and deoxyribo alpha-nucleosides and alpha-nucleotides, the stereoisomers of the naturally occurring beta-isomers, are worked out by minimizing the conformational energy as a function of all the major parameters including the sugar ring conformations along the pseudorotation path. The results of the studies bring out certain distinct conformational features that are at variance with their beta counterparts. The range of glycosyl conformations are found to be not only quite restricted here but favor predominantly the anti conformation. The syn glycosyl conformation for the entire region of P values are found to be energetically less favorable, with the barrier to anti in equilibrium with syn interconversion being higher especially in alpha-ribonucleosides. The energetically preferred range of pseudorotation phase angles (P) is also considerably restricted and P values corresponding to the C1'-exo range of sugars are highly unfavorable for alpha-nucleosides, in sharp contrast to the broad range of sugar ring conformations favored by beta-isomers. While both trans congruent to 180 degrees and skew congruent to 270 degrees conformations around the C3'-O3' (phi') bond are favored for alpha-3'-nucleotides with deoxyribose sugars, ribose sugars seem to favor only the skew values of phi'. Most interestingly and in sharp contrast to beta-stereoisomers, an energy barrier is encountered at P values corresponding to O4'-endo sugars. This suggests that the possible sugar pucker interconversion between C2'-endo/C3'-exo and C3'-endo/C2'-exo in alpha-anomers could take place only through the O4'-exo region. Likewise the possible path of anti in equilibrium with syn interconversion in alpha-nucleosides is not via high anti, in sharp contrast to beta-nucleosides. These observations should be borne in mind while assigning the sugar ring conformers in alpha-nucleosides and those containing them from nmr investigations. Comparison of the results with beta-anomers seem to suggest on the whole a lack of conformational variability or the restricted nature of alpha-stereoisomers. This could be one of the reasons for its nonselection in the naturally occurring nucleic acids.     

Foliar regeneration of the endangered Red Vanda,Renanthera imschootiana Rolfe (Orchidaceae)

An in vitro method was developed to regenerate large numbers of phenotypically uniform plants from the basal parts of the leaves of flowering plants of Renanthera imschootiana Rolfe. Differentiation of up to 10 shoot buds free of callus and protocorm-like bodies occurred in 10–12 weeks from the base of a single leaf implanted in Mitra et al. (1976) medium supplemented with 2% sucrose, 2 g l^-1 peptone, 44.4 M benzyladenine (BA) and 10.7 M naphthaleneacetic acid (NAA). Subculture of the tissues in medium enriched with 10% coconut water and 35 g l^-1 ripe banana pulp resulted in the production of highest average number of 40 shoots in 12 weeks. No difference in the regeneration potential was observed among the three young leaves while mature leaves did not respond. All the leaves of the regenerated shoots were easily recultured to increase shoot multiplication. Shoots readily formed roots on transfer to a medium containing 4.4 M BA, 10.7 M NAA and 1% activated charcoal. All regenerated plants examined were normal diploids with 2n=38. Foliar meristem culture appears to have great potential for ex situ conservation and propagation of this extremely endangered orchid.Abbreviations BA benzyladenine - NAA naphthaleneacetic acid     

Molecular docking analysis of stachydrine and sakuranetin with IL-6 and TNF-α in the context of inflammation

Inflammation is a process triggered by pro-inflammatory cytokines and anti-inflammatory molecules. Therefore, it is of interest to document the anti-inflammatory activity of Stachydrine and Sakuranetin against the inflammatory target proteins IL-6 and TNF-α by using molecular docking analysis. Both compounds showed good binding features with the selected target proteins. Compared to Sakuranetin, the Stachydrine have low binding energy and good hydrogen bond interactions. Hence, data show that Stachydrine possessed high and specific inhibitory activity on tumor necrosis factor-α and interleukin-6.     

Cation–π Interactions in β-Lactamases: The Role in Structural Stability

β-lactam group of antibiotics is the most widely used therapeutic molecules for treating bacterial infections. The main mode of bacterial resistance to β-lactams is by β-lactamases. In the present study, we report our results on the role of cation–π interactions in β-lactamases and their environmental preferences. The number of interactions formed by arginine is higher than lysine in the cationic group, while tyrosine is comparatively higher than phenylalanine and tryptophan in the π group. Our results indicate that cation–π interactions might play an important role in the global conformational stability of β-lactamases.     

Optimization of a Method for Chromatin Immunoprecipitation Assays in the Marine Invertebrate Chordate Ciona

Chromatin immunoprecipitation (ChIP) assays allow the efficient characterization of the in vivo occupancy of genomic regions by DNA-binding proteins and thus facilitate the prediction of cis-regulatory sequences in silico and guide their validation in vivo. For these reasons, these assays and their permutations (e.g., ChIP-on-chip and ChIP-sequencing) are currently being extended to several non-mainstream model organisms, as the availability of specific antibodies increases. Here, we describe the development of a polyclonal antibody against the Brachyury protein of the marine invertebrate chordate Ciona intestinalis and provide a detailed ChIP protocol that should be easily adaptable to other marine organisms.     

Reversal of Succinylcholine Induced Apnea with an Organophosphate Scavenging Recombinant Butyrylcholinesterase

Background

Concerns about the safety of paralytics such as succinylcholine to facilitate endotracheal intubation limit their use in prehospital and emergency department settings. The ability to rapidly reverse paralysis and restore respiratory drive would increase the safety margin of an agent, thus permitting the pursuit of alternative intubation strategies. In particular, patients who carry genetic or acquired deficiency of butyrylcholinesterase, the serum enzyme responsible for succinylcholine hydrolysis, are susceptible to succinylcholine-induced apnea, which manifests as paralysis, lasting hours beyond the normally brief half-life of succinylcholine. We hypothesized that intravenous administration of plant-derived recombinant BChE, which also prevents mortality in nerve agent poisoning, would rapidly reverse the effects of succinylcholine.

Methods

Recombinant butyrylcholinesterase was produced in transgenic plants and purified. Further analysis involved murine and guinea pig models of succinylcholine toxicity. Animals were treated with lethal and sublethal doses of succinylcholine followed by administration of butyrylcholinesterase or vehicle. In both animal models vital signs and overall survival at specified intervals post succinylcholine administration were assessed.

Results

Purified plant-derived recombinant human butyrylcholinesterase can hydrolyze succinylcholine in vitro. Challenge of mice with an LD₁₀₀ of succinylcholine followed by BChE administration resulted in complete prevention of respiratory inhibition and concomitant mortality. Furthermore, experiments in symptomatic guinea pigs demonstrated extremely rapid succinylcholine detoxification with complete amelioration of symptoms and no apparent complications.

Conclusions

Recombinant plant-derived butyrylcholinesterase was capable of counteracting and reversing apnea in two complementary models of lethal succinylcholine toxicity, completely preventing mortality. This study of a protein antidote validates the feasibility of protection and treatment of overdose from succinylcholine as well as other biologically active butyrylcholinesterase substrates.     

Abundant Intracellular IgG in Enterocytes and Endoderm Lacking FcRn

FcRn, a non-classical MHCI molecule, transports IgG from mother to young and regulates the rate of IgG degradation throughout life. Brambell proposed a mechanism that unified these two functions, saying that IgG was pinocytosed nonspecifically by the cell into an FcRn-expressing endosome, where, at low pH, it bound to FcRn and was exocytosed. This theory was immediately challenged by claims that FcRn specificity for ligand could be conferred at the cell surface in neonatal jejunum. Assessing Brambell''s hypothesis we found abundant nonspecifically endocytosed IgG present in the cytoplasm of FcRn^−/− enterocytes. Further, IgG was present in the intercellular clefts and the cores of FcRn^+/+ but not FcRn^−/− jejunum. FcRn specificity for ligand could be determined within the cell.