Anemia in experimental visceral leishmaniasis in hamsters.

Experimental infection of hamsters with Leishmania donovani caused visceral leishmaniasis in which hematological changes occurred. The infected hamsters were anemic and reticulocyte counts were high. No significant change in the serum erythropoietin level was noted. Red cell membrane Na(+)-K(+)-ATPase and acetylcholinesterase activities increased. Osmotic fragility of the erythrocytes from infected animals increased. The level of 2,3-diphosphoglycerate of the red cells increased with the degree of anemia.     

Role of Ca2+ ion on Leishmania-macrophage attachment

Summary Leishmania donovani, the etiological agent for the disease visceral leishmaniasis, attach themselves to the macrophages for initiation of the disease. The attachment process has been found to be regulated by Ca²⁺ ions. Verapamil, a Ca²⁺-channel blocker inhibits Leishmania-macrophage attachment. The inhibitory effect is increased with time. Nifedipine, another Ca²⁺-channel blocker exhibits the same effect. The attachment process is stimulated by Ca²⁺-ionophore alone. The inhibitory effects of the calcium channel blockers are reversed by the ionophore.     

Leishmania donovani: a chemically defined medium suitable for cultivation and cloning of promastigotes and transformation of amastigotes to to promastigotes

A chemically defined medium using commercially available alpha-MEM supplemented with hemin, HEPES, L-glutamine, D-glucose, folic acid, D-biotin and adenine supports the luxuriant growth and propagation of Leishmania donovani promastigotes. A peak parasite population of about 7.0 x 10(7)/ml at stationary phase and a population doubling time of 11.4 h for high-subpassage promastigotes were obtained. The medium was suitable for transformation of isolated amastigotes from infected hamster spleen. Promastigotes could be detected by culturing kala-azar patients' bone-marrow aspirate or spleen puncture material in this medium. Four out of six freshly transformed isolates gradually adapted and grew well in this medium. Macroscopic colonies appeared on agar plates prepared with the medium within 16-20 days after inoculation. The cloning efficiency was increased about five-fold by glycerol supplementation.     

Cyclothiazide Selectively Potentiates AMPA- and Kainate-Induced [3H]Norepinephrine Release from Rat Hippocampal Slices

Abstract: Activation of the α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) subtype of ionotropic glutamate receptors has been shown to result in a rapid desensitization of the receptor in the presence of certain agonists. One effect of AMPA receptor desensitization in the hippocampus may be to decrease the efficacy of AMPA receptor agonists at stimulating the release of norepinephrine from noradrenergic terminals. Recently, cyclothiazide was reported to inhibit AMPA receptor desensitization by acting at a distinct site on AMPA receptors. We have examined the effect of cyclothiazide on AMPA- and kainate (KA)-induced norepinephrine release from rat hippocampal slices to determine whether cyclothiazide would increase the efficacy of AMPA-induced [³H]norepinephrine release by inhibiting AMPA receptor desensitization. Cyclothiazide was observed to potentiate markedly both AMPA- and KA-induced [³H]norepinephrine release. This potentiation is selective for AMPA/KA receptors as cyclothiazide did not potentiate N -methyl- d -aspartate-induced [³H]norepinephrine release or release induced by the nonspecific depolarizing agents veratridine and 4-aminopyridine. These results demonstrate that AMPA receptor-mediated modulation of [³H]norepinephrine release from rat brain slices is a useful approach to studying the cyclothiazide modulatory site on the AMPA receptor complex.     

Enhancement of NMDA-mediated responses by cyanide

The effect of cyanide on NMDA-activated ion current and MK801 binding was studied in cultured rat hippocampal neurons. In microfluorometric analysis using fura-2, removal of extracellular Mg²⁺ resulted in a five-fold increase in NMDA-induced peak of [Ca²⁺]_i. One mM NaCN enhanced the peak NMDA responses in the presence, but not in the absence of extracellular Mg²⁺. Cyanide enhanced the immediate rise in [Ca²⁺]_i produced by NMDA, followed over a 1–5 min period by a gradual increase of [Ca²⁺]_i. Similar results were obtained in whole-cell patch clamp recordings from hippocampal neurons. One mM KCN enhanced the NMDA-activated current in the presence, but not in the absence of extracellular Mg²⁺. This effect was independent of cyanide-mediated metabolic inhibition since the recording pipette contained ATP (2 mM). In binding assays NaCN (1 mM) increased the binding affinity of [³H]MK-801 to rat forebrain membranes in the presence of Mg²⁺, whereas in the absence of Mg²⁺, NaCN did not influence binding. These results indicate that cyanide enhances NMDA-mediated Ca²⁺ influx and inward current by interacting with the Mg²⁺ block of the NMDA receptor. The effect of cyanide can be explained by an initial interaction with the Mg²⁺ block of the NMDA receptor/ionophore which appears to be energy-independent, followed by a gradual increase in Ca²⁺ influx resulting from cellular energy reserve depletion.Abbreviations NMDA N-Methyl-D-Aspartate - EAA excitatory amino acid - MK-801 (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d] cyclohept-5,10-imine maleate     

Amygdala Kindling Alters Protein Kinase C Activity in Dentate Gyrus

Kindling is a use-dependent form of synaptic plasticity and a widely used model of epilepsy. Although kindling has been widely studied, the molecular mechanisms underlying induction of this phenomenon are not well understood. We determined the effect of amygdala kindling on protein kinase C (PKC) activity in various regions of rat brain. Kindling stimulation markedly elevated basal (Ca(2+)-independent) and Ca(2+)-stimulated phosphorylation of an endogenous PKC substrate (which we have termed P17) in homogenates of dentate gyrus, assayed 2 h after kindling stimulation. The increase in P17 phosphorylation appeared to be due at least in part to persistent PKC activation, as basal PKC activity assayed in vitro using an exogenous peptide substrate was increased in kindled dentate gyrus 2 h after the last kindling stimulation. A similar increase in basal PKC activity was observed in dentate gyrus 2 h after the first kindling stimulation. These results document a kindling-associated persistent PKC activation and suggest that the increased activity of PKC could play a role in the induction of the kindling effect.     

Low oxygen tension potentiates proliferation and stemness but not multilineage differentiation of caprine male germline stem cells

The milieu of male germline stem cells (mGSCs) is characterized as a low-oxygen (O₂) environment, whereas, their in-vitro expansion is typically performed under normoxia (20–21% O₂). The comparative information about the effects of low and normal O₂ levels on the growth and differentiation of caprine mGSCs (cmGSCs) is lacking. Thus, we aimed to investigate the functional and multilineage differentiation characteristics of enriched cmGSCs, when grown under hypoxia and normoxia. After enrichment of cmGSCs through multiple methods (differential platting and Percoll-density gradient centrifugation), the growth characteristics of cells [population-doubling time (PDT), viability, proliferation, and senescence], and expression of key-markers of adhesion (β-integrin and E-Cadherin) and stemness (OCT-4, THY-1 and UCHL-1) were evaluated under hypoxia (5% O₂) and normoxia (21% O₂). Furthermore, the extent of multilineage differentiation (neurogenic, adipogenic, and chondrogenic differentiation) under different culture conditions was assessed. The survival, viability, and proliferation were significantly (p?<?0.05) improved, thus, yielding a significantly (p?<?0.05) higher number of viable cells with larger colonies under hypoxia. Furthermore, the expression of stemness and adhesion markers were distinctly upregulated under lowered O₂ conditions. Conversely, the differentiated regions and expression of differentiation-specific genes [C/EBPα (adipogenic), nestin and β-tubulin (neurogenic), and COL2A1 (chondrogenic)] were significantly (p?<?0.05) reduced under hypoxia. Overall, the results demonstrate that culturing cmGSCs under hypoxia augments the growth characteristics and stemness but not the multilineage differentiation of cmGSCs, as compared with normoxia. These data are important to develop robust methodologies for ex-vivo expansion and lineage-committed differentiation of cmGSCs for clinical applications.

     

Microbial biofilm ecology,in silico study of quorum sensing receptor-ligand interactions and biofilm mediated bioremediation

Archives of Microbiology - Biofilms are structured microbial communities of single or multiple populations in which microbial cells adhere to a surface and get embedded in extracellular polymeric...     

Identification of Novel Allelic Variants of Integrin Beta 2 (ITGB2) Gene and Screening for Bubaline Leukocyte Adhesion Deficiency Syndrome in Indian Water Buffaloes (Bubalus bubalis)

A fragment of 570 bp corresponding to exon 5 and 6 of integrin beta 2 (ITGB2) gene was amplified for screening D128G mutation in one hundred and fifty two buffaloes (Bubalus bubalis) which causes bovine leukocyte adhesion deficiency syndrome (BLAD) in cattle, as well as to ascertain polymorphism. TaqI PCR-RFLP revealed no such mutation thus indicating the absence of bubaline leukocyte adhesion deficiency (BuLAD) allele in animals under study. However, the polymorphism studies using MspI restriction enzyme revealed two genotypic patterns viz. AA pattern (bands of 293, 141, 105, and 31 bp) and BB pattern (bands of 293, 105, 77, 64, and 31 bp). The sequences of A and B alleles were submitted to the GenBank (EU853307 and AY821799).