Plunder of Human Blood Leukocytes Containing Ingested Material,by Other Leukocytes: Where Is the Fusagen That Allows Preservation of Membrane Integrity and Motile Function?

In studying phagocytosis of zymosan particles by human blood monocytes in phase-contrast videomicroscopy, we found that monocytes loaded with zymosan particles became chemotactic for polymorphonuclear leukocytes (PMN) which closed on them and purloined their particle content. This despoliation usually occurred in monocytes that had begun to swell—prefiguring their death. The violent seizure of their contents by the aggressing PMN often tore the monocytes apart. However, some apparently healthy monocyte survived the removal of zymosan content by PMN or, more commonly, its removal by another monocyte. PMN—a much hardier cell in slide preparations—that were similarly loaded with zymosan particles, also attracted PMN. The latter could remove zymosan from the target cell without killing it. Thus, leukocytes were sacrificing significant portions of themselves without losing residual membrane integrity and motile function. Their behavior with respect to other particles (e.g., bacteria) will be of interest. We suggest that the membrane fusagen resides in the inner membrane leaflets when they are brought together in an extreme hourglass configuration. This event may be similar to the fragmentation of erythrocytes into intact pieces, the formation of cytokineplasts, the rear extrusion of content by migrating cells on surfaces, and the phagocytic process itself.     

Griseofulvin: Association with tubulin and inhibition of in vitro microtubule assembly

Griseofulvin—shown previously to disrupt the mitotic apparatus $\text{in vivo}$—inhibited the $\text{in vitro}$ microtubule assembly reaction completely at 8 × 10^?4M griseofulvin. In a gel filtration assay, randomly tritiated griseofulvin associated stoichiometrically with purified tubulin, as determined by chromatography on Sephadex G-25. No detectable drug binding was observed when bovine serum albumin was used as a control in an identical column assay. Both gel filtration chromatography and a kinetic analysis of the inhibition of assembly by griseofulvin suggest that the drug interacts directly and stoichimetrically with the tubulin dimer, and that the interaction is both rapid and independent of temperature.     

Phylogeny and systematics of the bee genus Osmia (Hymenoptera: Megachilidae) with emphasis on North American Melanosmia: subgenera,synonymies and nesting biology revisited

The predominantly Holarctic bee genus Osmia Panzer is species‐rich and behaviourally diverse. A robust phylogeny of this genus is important for understanding the evolution of the immense variety of morphological and behavioural traits exhibited by this group. We infer a phylogeny of Osmia using DNA sequence data obtained from three nuclear genes (elongation factor 1‐α, LW‐rhodopsin and CAD) and the mitochondrial gene COI. Our taxon sampling places special attention on North American members of the subgenus Melanosmia Schmiedeknecht; we discuss the novel placement of a number of species traditionally assigned to O. (Melanosmia) and examine the relative support for alternative classifications of this species‐rich subgenus. We use this new phylogeny to guide a reassessment of morphological and behavioural characters within Osmia. Our results provide support for the recognition of Osmia (Hapsidosmia), subgen.n ., a monotypic subgenus containing Osmia iridis Cockerell & Titus. We synonymize Osmia (Mystacosmia) Snelling under O. (Melanosmia), syn.n . We synonymize Osmia (Acanthosmioides) Ashmead under O. (Melanosmia), syn.n ., propose ‘odontogaster species group’ as a replacement for the subgeneric name Acanthosmioides, and refine the morphological characters that serve to diagnose the species group. We additionally propose ‘nigrifrons species group’ for a clade within O. (Melanosmia) containing most species formerly placed in Osmia (Centrosmia) Robertson. We demonstrate more cohesive patterns of nest substrate use in the nigrifrons and odontogaster species groups than was previously believed to occur, reconsider character polarity of aspects of the female mandible, and show that a large number of morphological characters have evolved convergently within the genus. In order to facilitate discussion of relevant taxa, we propose the following 15 new synonymies: O. bakeri Sandhouse under O. melanopleura Cockerell; O. crenulaticornis Michener under O. pinorum Cockerell; O. claremontensis Michener under O. sedula Sandhouse; O. cockerelli Sandhouse under O. dakotensis Michener; O. francisconis White under O. enixa Sandhouse; O. hurdi White under O. austromaritima Michener; O. sladeni Sandhouse under O. nifoata Cockerell; O. titusi Cockerell under O. phenax Cockerell; O. subtrevoris Cockerell, O. physariae Cockerell, and O. erecta Michener under O. giliarum Cockerell; and O. universitatis Cockerell, O. integrella Cockerell, O. amala Cockerell, and O. metitia Cockerell under O. nigrifrons Cresson, syn.n . We remove O. wyomingensis Michener from synonymy with O. nifoata Cockerell, stat.n ., and O. pinorum Cockerell from synonymy with O. physariae Cockerell, stat.n . This published work has been registered in ZooBank, http://zoobank.org/urn:lsid:zoobank.org:pub:A3E7D63B‐5C4C‐4ACF‐BF33‐48E5C5DD1B0D .     

Endogenous chloride channels of insect sf9 cells. Evidence for coordinated activity of small elementary channel units

The endogenous Cl- conductance of Spodoptera frugiperda (Sf9) cells was studied 20-35 h after plating out of either uninfected cells or cells infected by a baculovirus vector carrying the cloned beta-galactosidase gene (beta-Gal cells). With the cation Tris+ in the pipette and Na+ in the bath, the reversal potential of whole-cell currents was governed by the prevailing Cl- equilibrium potential and could be fitted by the Goldman-Hodgkin-Katz equation with similar permeabilities for uninfected and beta-Gal cells. In the frequency range 0.12 < f < 300 Hz, the power density spectrum of whole-cell Cl- currents could be fitted by three Lorentzians. Independent of membrane potential, >50% of the total variance of whole-cell current fluctuations was accounted for by the low frequency Lorentzian (fc = 0.40 +/- 0.03 Hz, n = 6). Single-Cl- channels showed complex gating kinetics with long lasting (seconds) openings interrupted by similar long closures. In the open state, channels exhibited fast burst-like closures. Since the patches normally contained more than a single channel, it was not possible to measure open and closed dwell-time distributions for comparing single-Cl- channel activity with the kinetic features of whole-cell currents. However, the power density spectrum of Cl- currents of cell-attached and excised outside-out patches contained both high and low frequency Lorentzian components, with the corner frequency of the slow component (fc = 0.40 +/- 0.02 Hz, n = 4) similar to that of whole-cell current fluctuations. Chloride channels exhibited multiple conductance states with similar Goldman-Hodgkin-Katz-type rectification. Single-channel permeabilities covered the range from approximately 0.6.10(-14) cm5/s to approximately 6.10(-14) cm3/s, corresponding to a limiting conductance (gamma 150/150) of approximately 3.5 pS and approximately 35 pS, respectively. All states reversed near the same membrane potential, and they exhibited similar halide ion selectivity, P1 > PCl approximately PBr. Accordingly, Cl- current amplitudes larger than current flow through the smallest channel unit resolved seem to result from simultaneous open/shut events of two or more channel units.     

THE MELANOCYTE MODEL : Colchicine-like Effects of Other Antimitotic Agents

The effect of various agents that cause metaphase arrest in dividing cells was studied on the rapid reversible darkening of frog skin under the influence of melanocyte-stimulating hormone (MSH). Darkening is due to dispersion of melanin granules in melanocytes and is thought to be accompanied by a gel-to-sol cytoplasmic transformation. After subsequent washing the skin lightens, with aggregation of melanin granules and cytoplasmic gelation. As previously shown with colchicine, preincubation of frog skin with vinblastine, vincristine, or colcemid produced an increase in darkening induced by MSH, as compared to control skins, and a dosage-dependent inhibition of subsequent lightening. Preincubation with each drug, without subsequent MSH, produced a gradual, irreversible, dosage-dependent darkening over several hours. On a molar basis, the relative strength of the various agents was vinblastine > vincristine > colcemid > colchicine; vinblastine was about 100 times stronger than colchicine. Preincubation of frog skin with griseofulvin, followed by washing, had no subsequent effects on darkening or lightening. However, effects similar to those of the Colchicum and Vinca alkaloids were seen if griseofulvin was kept in the ambient media. These effects were rapidly reversible on removal of the drug from the media. These findings support the melanocyte model originally proposed for the action of colchicine, and emphasize certain facts that models of melanin granule movement will have to accommodate.     

Electrical and adaptive properties of rod photoreceptors in bufo marinus. I. Effects of altered extracellular Ca(2+) levels

The effects of altering extracellular Ca(2+) levels on the electrical and adaptive properties of toad rods have been examined. The retina was continually superfused in control (1.6 mM Ca(2+)) or test ringer’s solutions, and rod electrical activity was recorded intracellularly. Low-calcium ringer’s (10(-9)M Ca(2+)) superfused for up to 6 min caused a substantial depolarization of the resting membrane potential, an increase in light-evoked response amplitudes, and a change in the waveform of the light-evoked responses. High Ca(2+) ringer’s (3.2 mM) hyperpolarized the cell membrane and decreased response amplitudes. However, under conditions of either low or high Ca(2+) superfusion for up to 6 min, in both dark-adapted and partially light-adapted states, receptor sensitivity was virtually unaffected; i.e., the V-log I curve for the receptor potential was always located on the intensity scale at a position predicted by the prevailing light level, not by Ca(2+) concentration. Thus, we speculate that cytosol Ca(2+) concentration is capable of regulating membrane potential levels and light-evoked response amplitudes, but not the major component of rod sensitivity. Low Ca(2+) ringer’s also shortened the period of receptor response saturation after a bright but nonbleaching light flash, hence accelerating the onset of both membrane potential and sensitivity recovery during dark adaptation.

Exposure of the retina to low Ca(2+) (10(-9)M) ringer’s for long periods (7-15 min) caused dark-adapted rods to lose responsiveness. Response amplitudes gradually decreased, and the rods became desensitized. These severe conditions of low Ca(2+) caused changes in the dark-adapted rod that mimic those observed in rods during light adaptation. We suggest that loss of receptor sensitivity during prolonged exposure to low Ca(2+) ringer’s results from a decrease of intracellular (intradisk) stores of Ca(2+); i.e., less Ca(2+) is thereby released per quantum catch.

     

Interleukin 1 (IL 1) as a mediator of crystal arthritis. Stimulation of T cell and synovial fibroblast mitogenesis by urate crystal-induced IL 1

We reported before that monosodium urate (MSU) crystals were potent stimulators of endogenous pyrogen (EP) production from human and rabbit mononuclear phagocytes, and proposed that this property of MSU crystals may be important in the pathogenesis of gout. EP activity is now attributed to interleukin 1 (IL 1) peptides but IL 1 is not the only pyrogenic monocyte-derived cytokine, since both interferon-alpha (alpha-IFN) and tumor necrosis factor (TNF) are also pyrogenic in rabbits. Using a T cell comitogenic assay based on a murine helper T cell clone that does not respond to IFN or TNF, we now report the release of IL 1 activity from human blood monocytes and synovial fluid mononuclear cells (MNC), following stimulation with MSU crystals. MSU-induced supernatants with IL 1 activity were neutralized with rabbit antiserum to human IL 1 and also stimulated the growth ([3H]thymidine incorporation) of long-term fibroblast-like cell lines derived from human synovial rheumatoid exudate. Two other crystals associated with articular inflammation were tested: hydroxyapatite was a much less potent stimulus compared with MSU crystals, and calcium pyrophosphate dihydrate did not stimulate IL 1 release from human monocytes or synovial fluid MNC. As a model for the inflammatory consequences of acute and chronic overproduction of IL 1, gout is the only sterile inflammatory disease where the local and systemic pathology is compatible with such overproduction; raised IL 1 levels have been found at the site of inflammation, and a necessary etiologic agent, crystalline urate, has been shown unequivocally to be a direct activator of mononuclear IL 1 release.