Effects of sodium ions and monensin on amylase secretion from rat parotid cells

The role of sodium ions in amylase secretion from rat parotid cells was studied using various Na+-free media and monensin. In a sucrose medium, amylase secretion was not stimulated by isoproterenol but was significantly stimulated by dibutyryl cAMP. In choline chloride and LiCl media, both isoproterenol and dibutyryl cAMP clearly evoked amylase release. Monensin itself elicited amylase secretion slightly, but significantly inhibited the secretion stimulated by isoproterenol or dibutyryl cAMP. The inhibitory effect of monensin was detectable even in choline chloride, LiCl and KCl media. These results indicate that sodium ions are not essential for amylase secretion from rat parotid cells and that the inhibitory effect of monensin is independent of influx of sodium ions or efflux of potassium ions.     

Interactions of green or red light with blue light on the dark closure of Albizzia pinnules

The interactions of green or red light with blue light on the dark closing of Albizzia julibrissin Durazz. pinnules have been investigated. Irradiations at 430, 450 and 470 nm progressively delay dark closing with increasing photon fluence rates. Red or green light alone has no effect. However, when the blue fluence rate is low, both red and green light interact with it and increase the delaying effect of the blue light. When the blue fluence rate is high, green light interacts with it to negate some of the effectiveness of the blue light, while red light has no effect. This is similar to results obtained previously with far-red light. It is suggested that the same unidentified photoreceptor is operating in both the far-red and blue regions. The results also indicate the presence of a blue-only absorbing photoreceptor whose action is increased by phytochrome.     

Global bifurcation structure of a Bonhoeffer-van der Pol oscillator driven by periodic pulse trains

The Bonhoeffer-van der Pol (BVP) oscillator is a valuable dynamical system model of pacemaker neurons. Isochrons, phase transition curves (PTC), and two dimensional bifurcation diagrams served to analyze the neuron's response to periodic pulse stimuli. Responses are described and explained in terms of the nonlinear dynamical system theory. An important issue in the generation of spikes by pacemaker neurons is the existence of both slow and fast dynamics in the state point's trajectory in the phase plane. It is this feature in particular that makes the BVP oscillator a faithful model of living pacemaker neurons. Comparison of the model's responses with those of a living pacemaker was based also on return maps of interspike intervals. Analyzed in detail were the complex discharges called stammering which involve interspike intervals that arise unpredictably and exhibit histograms with several modes separated by the equal intervals.Supported by Trent H. Wells Jr. Inc.     

Nitroprusside and Cyclic GMP Stimulate Na+-Ca2+ Exchange Activity in Neuronal Preparations and Cultured Rat Astrocytes

Abstract: The effects of nitric oxide (NO)-generating agents on ⁴⁵Ca²⁺ uptake in rat brain slices and cultured rat astrocytes were studied in the presence of monensin, which is considered to drive the Na⁺-Ca²⁺ exchanger in the reverse mode. Sodium nitroprusside (SNP) at >10 µ M increased monensin-stimulated Ca²⁺ uptake in the slices, although it did not affect high K⁺-stimulated Ca²⁺ uptake. Another NO donor, 3-morpholinosydnonimine, was effective. The effect of SNP was antagonized by hemoglobin (50 µ M ), a NO scavenger, and mimicked by 8-bromo-cyclic GMP (100 µ M ). In rat brain synaptosomes, SNP increased monensin-stimulated Ca²⁺ uptake, but it did not affect high K⁺-stimulated Ca²⁺ uptake. 8-Bromocyclic GMP, but not SNP, increased Na⁺-dependent Ca²⁺ uptake significantly in synaptic membrane vesicles in the absence of monensin. In cultured rat astrocytes, SNP and 8-bromo-cyclic GMP increased Ca²⁺ uptake in the presence of ouabain and monensin, which were required for the Ca²⁺ uptake in the cells. These findings suggest that NO stimulates the Na⁺-Ca²⁺ exchanger in neuronal preparations and astrocytes in a cyclic GMP-dependent mechanism.     

Isolation and Characterization of Hardening-Induced Proteins in Chlorella vulgaris C-27: Identification of Late Embryogenesis Abundant Proteins

Hardening-induced soluble proteins of Chlorella vulgaris BeijerinkIAM C-27 (formerly Chlorella ellipsoidea Gerneck IAM C-27) wereisolated and purified by two-dimensional high-performance liquidchromatography (2D-HPLC) on an anion-exchange column, with subsequentreversed-phase chromatography. Some of the proteins were resolvedby SDS-PAGE, characterized by amino-terminal sequencing andidentified by searching for homologies in databases. Separationof the soluble proteins during the hardening of Chlorella bya combination of 2D-HPLC and SDS-PAGE revealed that at least31 proteins were induced or increased in abundance. Of particularinterest was the induction after 12 h of a 10-kDa protein withthe amino-terminal amino acid sequence AGNKPITEQISDAVGAAGQKVGand the induction after 6 h of a 14-kDa protein with the amino-terminalsequence ALGEESLGDKAKNAFEDAKDAVKDAAGNVKEAV. The amino-terminalsequences of these proteins indicated that they were homologousto late embryogenesis abundant (LEA) proteins. Furthermore,the level of a 22-kDa protein also increased after 12 h. Theamino-terminal sequence of this protein, AAPLVGGPAPDFTAAAVFD,indicated that it was homologous to thioredoxin peroxidase. (Received June 9, 1995; Accepted September 12, 1995)     

Cell-penetrating peptide-conjugated lipid nanoparticles for siRNA delivery

Lipid nanoparticles (LNP) modified with cell-penetrating peptides (CPP) were prepared for the delivery of small interfering RNA (siRNA) into cells. Lipid derivatives of CPP derived from protamine were newly synthesized and used to prepare CPP-decorated LNP (CPP-LNP). Encapsulation of siRNA into CPP-LNP improved the stability of the siRNA in serum. Fluorescence-labeled siRNA formulated in CPP-LNP was efficiently internalized into B16F10 murine melanoma cells in a time-dependent manner, although that in LNP without CPP was hardly internalized into these cells. In cells transfected with siRNA in CPP-LNP, most of the siRNA was distributed in the cytoplasm of these cells and did not localize in the lysosomes. Analysis of the endocytotic pathway indicated that CPP-LNP were mainly internalized via macropinocytosis and heparan sulfate-mediated endocytosis. CPP-LNP encapsulating siRNA effectively induced RNA interference-mediated silencing of reporter genes in B16F10 cells expressing luciferase and in HT1080 human fibrosarcoma cells expressing enhanced green fluorescent protein. These data suggest that modification of LNP with the protamine-derived CPP was effective to facilitate internalization of siRNA in the cytoplasm and thereby to enhance gene silencing.     

Neutrophil extracellular traps mediate a host defense response to human immunodeficiency virus-1

Neutrophils contribute to pathogen clearance by producing neutrophil extracellular traps (NETs), which are genomic DNA-based net-like structures that capture bacteria and fungi. Although NETs also express antiviral factors, such as myeloperoxidase and α-defensin, the involvement of NETs in antiviral responses remains unclear. We show that NETs capture human immunodeficiency virus (HIV)-1 and promote HIV-1 elimination through myeloperoxidase and α-defensin. Neutrophils detect HIV-1 by Toll-like receptors (TLRs) TLR7 and TLR8, which recognize viral nucleic acids. Engagement of TLR7 and TLR8 induces the generation of reactive oxygen species that trigger NET formation, leading to NET-dependent HIV-1 elimination. However, HIV-1 counteracts this response by inducing C-type lectin CD209-dependent production of interleukin (IL)-10 by dendritic cells to inhibit NET formation. IL-10 suppresses the reactive oxygen species-dependent generation of NETs induced upon TLR7 and TLR8 engagement, resulting in disrupted NET-dependent HIV-1 elimination. Therefore, NET formation is an antiviral response that is counteracted by HIV-1.     

Ancient Mitochondrial DNA Reveals the Origin of Sus scrofa from Rebun Island, Japan

The Kabukai A site (5 to 8C A.D.) of the Okhotsk cultural area is on Rebun Island, a small island near the coast, north–northwest of Hokkaido, Japan. Specimens of Sus scrofa, called the Sakhalin pig, were discovered in five cultural layers at the Kabukai A site. Ancient DNA was extracted from the remains of 42 Sakhalin pig bones. Thirty-nine nucleotide sequences of the 574-bp mitochondrial DNA control region, estimated to have originated from at least 21 individuals, were amplified and analyzed phylogenetically. Nine distinct haplotypes (A1, A2, A3, B1, B2, C1, C2, D1, and D2) from this site were classified into four haplotype groups (A, B, C, and D) by parsimonious network analysis. Phylogenetic analysis of 9 ancient and 55 modern haplotypes indicated that the population of Sakhalin pigs at the Kabukai A site belonged to two distinct clusters; haplotype groups A and B formed a cluster comprised only of themselves, and haplotype groups C and D belonged to the cluster of one of the two genetic groups of Japanese wild boars uniquely distributed in the western part of Japan, including one northeast Mongolian wild boar. Analysis of the haplotype distribution among three archaeological sites and their historical transitions among the five layers reflecting the cultural periods at the Kabukai A site suggests that the Sakhalin pig populations were introduced from Sakhalin island and the Amur River basin in the northeastern Eurasian continent together with some cultural influences. Received: 18 April 2000 / Accepted: 24 November 2000