N-Methyl-d-aspartate Receptor- and Calpain-mediated Proteolytic Cleavage of K+-Cl? Cotransporter-2 Impairs Spinal Chloride Homeostasis in Neuropathic Pain

Loss of synaptic inhibition by γ-aminobutyric acid and glycine due to potassium chloride cotransporter-2 (KCC2) down-regulation in the spinal cord is a critical mechanism of synaptic plasticity in neuropathic pain. Here we present novel evidence that peripheral nerve injury diminishes glycine-mediated inhibition and induces a depolarizing shift in the reversal potential of glycine-mediated currents (E_glycine) in spinal dorsal horn neurons. Blocking glutamate N-methyl-d-aspartate (NMDA) receptors normalizes synaptic inhibition, E_glycine, and KCC2 by nerve injury. Strikingly, nerve injury increases calcium-dependent calpain activity in the spinal cord that in turn causes KCC2 cleavage at the C terminus. Inhibiting calpain blocks KCC2 cleavage induced by nerve injury and NMDA, thereby normalizing E_glycine. Furthermore, calpain inhibition or silencing of μ-calpain at the spinal level reduces neuropathic pain. Thus, nerve injury promotes proteolytic cleavage of KCC2 through NMDA receptor-calpain activation, resulting in disruption of chloride homeostasis and diminished synaptic inhibition in the spinal cord. Targeting calpain may represent a new strategy for restoring KCC2 levels and tonic synaptic inhibition and for treating chronic neuropathic pain.     

Echinicola shivajiensis sp. nov., a novel bacterium of the family “Cyclobacteriaceae” isolated from brackish water pond

Strain AK12^T, an orange pigmented Gram-negative, rod shaped, non-motile bacterium, was isolated from a mud sample collected from a brackish water pond at Rampur of West Bengal, India. The strain was positive for oxidase, catalase and phosphatase. The predominant fatty acids were iso-C_15:0 (42.7%), iso-C_17:0 3OH (13.2%), C_16:1ω7c/C_16:1ω6c (summed feature 3) (8.0%), iso-C_17:1 I/anteiso-C_17:1 B (summed feature 4) (6.1%) and iso-C_17:1ω9c/C_16:0 10-methyl (summed feature 9) (9.4%). Strain AK12^T contained MK-7 as the major respiratory quinone and phosphatidylethanolamine, one unidentified aminophospholipid and six unidentified lipids as the polar lipids. The G + C content of DNA of the strain AK12^T was 46.2 mol%. The 16S rRNA gene sequence analysis indicated that strain AK12^T was member of the genus Echinicola and closely related to Echinicola vietnamensis, Echinicola pacifica and Echinicola jeungdonensis with pair-wise sequence similarity of 96.8, 96.3 and 96.0% respectively. Phylogenetic analyses indicated that the strain AK12^T clustered with E. vietnamensis and together with E. pacifica and E. jeungdonensis with a phylogenetic distance of 5.1, 6.3 and 6.6% (94.9, 93.7 and 93.4% similarity) respectively. Based on data from the current polyphasic study, strain AK12^T is proposed as a novel species of the genus Echinicola, for which the name Echinicola shivajiensis sp. nov. is proposed. The type strain of E. shivajiensis is AK12^T (= MTCC 11083^T = JCM 17847^T).     

Biofilm-associated indole acetic acid producing bacteria and their impact in the proliferation of biofilm mats in solar salterns

Biofilm mats appear in salterns distinctively during the monsoon season when the salinity decreases below 12 percentile salinity units and within a short period cover the entire surface area of the saltern. A study was carried out in two salterns viz. Nerul and Curca to find a possible reason for the rapid proliferation of these solar biofilms. Out of the 125 bacteria isolated from these biofilms, 16 produced indole-3-acetic acid (IAA). Rapid in-situ assay with Salkowski reagent and HPLC analysis confirmed the IAA production. Four isolates consistently produced high IAA concentrations ranging from 9.5 to 14.2 μg/mL in the presence of 4 mg/mL tryptophan concentrations in the growth media. The IAA-producing bacteria were Aeromonas aquariorum (N2), Pseudomonas alcaliphila (N3), Vibrio diazotrophicus (N6) and Pseudomonas pachastrellae (C3). These four IAA-producing bacteria also produced other growth promoting factors like ammonia. Three isolates produced siderophores and were phosphate solubilizers. There was enhancement in the growth of Cicer arietinum (length of the shoot and root) under axenic conditions and of biofilm mats (r = 0.9, p < 0.001; r = 0.8, p < 0.05 and r = 0.946, p < 0.01, respectively). This is, according to our knowledge, the first report indicating IAA-producing bacteria isolated from biofilms enhancing the proliferation of these biofilm mats in the solar salterns.     

Isolation and purification of microbial community DNA from soil naturally enriched for chitin

We made an attempt to isolate and purify metagenomic DNA from chitin enriched soil. In this communication we report a modified direct lysis method for soil DNA extraction including initial pre-lysis washing of sample, followed by a rapid polyvinylpyrrolidone-agarose-based purification and electroelution of DNA using Gene-capsule™ assembly. Rapidity was achieved using low molarity conducting media (sodium-borate buffer) for electrophoresis by reducing run time for both the gel electrophoresis and electroelution. Extracted DNA was sufficiently pure and of high quality, evidenced by amplification of 16S rDNA and chitinase genes by PCR. Metagenomic nature of the DNA was confirmed by running V3 (16S rDNA) region amplicons using denaturing gradient gel electrophoresis. This method requires 30 min for purification, and less than 2 h for complete execution of protocol and becomes the first report on the isolation of metagenomic DNA from soil naturally enriched for chitin.     

A novel device for long bone osteodistraction: description of device and case series

The purpose of this study was to present a novel intramedullary device (M-Bone; Phenix, Paris, France) that contains a mechanism for internal osteodistraction and bone transport in patients with segmental bone defects or limb length discrepancy after limb salvage operations. A total of five patients with primary bone tumors were enrolled in the study. After implantation, daily lengthening was performed in an outpatient setting either by the patient or with the help of a therapist, without the use of anesthesia. This unique device offers a totally new approach for the treatment of segmental bone defects or limb length discrepancy. It is designed to expand the remaining native bone by a magnetically activated drive system and induces new bone formation using osteodistraction and bone transport.     

Biodegradable polymer based encapsulation of neem oil nanoemulsion for controlled release of Aza-A

Azadirachtin a biological compound found in neem have medicinal and pesticidal properties. The present work reports on the encapsulation of neem oil nanoemulsion using sodium alginate (Na-Alg) by cross linking with glutaraldehyde. Starch and polyethylene glycol (PEG) were used as coating agents for smooth surface of beads. The SEM images showed beads exhibited nearly spherical shape. Swelling of the polymeric beads reduced with coating which in turn decreased the rate of release of Aza-A. Starch coated encapsulation of neem oil nanoemulsion was found to be effective when compared to PEG coated encapsulation of neem oil nanoemulsion. The release rate of neem Aza-A from the beads into an aqueous environment was analyzed by UV-visible spectrophotometer (214nm). The encapsulated neem oil nanoemulsion have the potential for controlled release of Aza-A. Neem oil nanoemulsion encapsulated beads coated with PEG was found to be toxic in lymphocyte cells.     

Judgements of agency in schizophrenia: an impairment in autonoetic metacognition

We investigated judgements of agency in participants with schizophrenia and healthy controls. Participants engaged in a computer game in which they attempted to touch downward falling Xs and avoid touching Os. On some trials, participants were objectively in perfect control. On other trials, they were objectively not in complete control because the movement of the cursor on the screen was distorted with respect to the position of the mouse by random noise (turbulence), or it was lagged by 250 or 500 ms. Participants made metacognitive judgements of agency as well as judgements of performance. Control participants' judgements of agency were affected by the turbulence and lag variables-indicating that they knew they were objectively not in control in those conditions, and they were also influenced by their assessments of performance. The patients also used their assessments of performance but neither turbulence nor lag affected their judgements of agency. This indicated an impairment in agency monitoring. The patients, unlike the healthy controls, used only publically available external cues about performance in making judgements of 'agency' and did not rely on any additional access to internal self-relevant cues that were diagnostic in indicating whether or not they were, in fact, in control.     

Changes in sugar levels during slow growth of Dendrocalamus hamiltonii somatic embryos due to liquid paraffin overlay

A liquid paraffin overlay (LPO) was used for storage of rapidly multiplying somatic embryos of Dendrocalamus hamiltonii under slow-growth conditions. Slow growth was associated with changes in sugar metabolism. In rapidly growing embryogenic tissues, a sharp decline in starch and non-reducing sugars indicated rapid utilization of starch. In contrast, under slow-growth conditions in somatic embryos stored under LPO, a gradual decline in starch indicated its slower utilization. As a result, growth of somatic embryos under LPO was suppressed and subculturing was not required. Following retrieval from growth under LPO after 30, 90, 180, 270, and 365 d of storage, the somatic embryos showed high recovery and germination (79.78%, 77.49%, 71.22%, 67.13%, and 59.99%, respectively) and were able to proliferate following transfer to Murashige and Skoog’s (Physiol. Plant. 15:473–497, 1962) medium containing 1 mgl⁻¹ BAP and 2% sucrose. The study provides useful information on in vitro storage of embryogenic tissue of D. hamiltonii.