Induction of endoreduplication in Chinese hamsters V79 cells by cytosine arabinoside

Endoreduplication (ER) could be induced very effectively in Chinese hamster V79 cells exposed to cytosine arabinoside (1-β-D-arabinofuranosylcytosine; Ara-C). Cells were cultured for 48 hours in Ara-C containing medium. ER frequency increases rapidly after Ara-C release. About 60% of metaphase cells were endoreduplicated at 8–10 hours after release from Ara-C (5 μg/ml). Induction of ER also depends on Ara-C concentrations.     

Catecholamine Secretion by Chemically Skinned Cultured Chromaffm Cells

The secretory system of intact chromaffin cells is not accessible to direct chemical manipulation because of the selective permeability of the plasmalemma. We have devised a simple procedure for chemically "skinning" (permeabilizing) cultured adrenal medullary chromaffin cells by brief exposure to the detergent saponin. This procedure disrupts the continuity of the plasmalemma, thus allowing us to bypass those aspects of the secretory process controlled by the cell membrane and giving direct access to exogenous substances to the cellular secretory machinery. We report here that the skinned cells retain a fully competent secretory mechanism dependent only on exogenous calcium and MgATP. Saponin treatment had no significant effect on the total catecholamine content of the cells. Secretion could be initiated by either MgATP or calcium as long as the other was present in the medium. Catecholamine and dopamine-beta-hydroxylase release by the skinned cells was dependent on the calcium concentration of the medium. The ratio of secreted catecholamine and enzyme was similar to that of the cells, indicating that secretion occurred by an exocytotic mechanism. About half the total cellular content of the cytoplasmic enzyme lactic dehydrogenase was released during the permeabilization process and subsequent incubations, indicating plasmalemma permeability to molecules as large as protein. Calcium-induced secretion was unaffected by several drugs known to affect catecholamines and granule function. Saponin treatment of chromaffin cells in culture appears to be a simple means for allowing access to exogenous substances to the cells' secretory machinery. Therefore, it offers the opportunity to use chemical treatments, and perhaps specific antibodies to cellular components, to determine the role of these elements in the secretory process. These techniques should also be applicable to other cells known to secrete by an exocytotic mechanism.     

Respiratory mechanics during halothane anesthesia and anesthesia-paralysis in humans

In six spontaneously breathing anesthetized subjects [halothane approximately 1 maximum anesthetic concentration (MAC), 70% N2O-30% O2], we measured flow (V), volume (V), and tracheal pressure (Ptr). With airway occluded at end-inspiration tidal volume (VT), we measured Ptr when the subjects relaxed the respiratory muscles. Dividing relaxed Ptr by VT, total respiratory system elastance (Ers) was obtained. With the subject still relaxed, the occlusion was released to obtain the V-V relationship during the ensuing relaxed expiration. Under these conditions, the expiratory driving pressure is V X Ers, and thus the pressure-flow relationship of the system can be obtained. By subtracting the flow resistance of equipment, the intrinsic respiratory flow resistance (Rrs) is obtained. Similar measurements were repeated during anesthesia-paralysis (succinylcholine). Ers averaged 23.9 +/- 4 (+/- SD) during anesthesia and 21 +/- 1.8 cmH2O X 1(-1) during anesthesia-paralysis. The corresponding values of intrinsic Rrs were 1.6 +/- 0.7 and 1.9 +/- 0.9 cmH2O X 1(-1) X s, respectively. These results indicate that Ers increases substantially during anesthesia, whereas Rrs remains within the normal limits. Muscle paralysis has no significant effect on Ers and Rrs. We also provide the first measurements of inspiratory muscle activity and related negative work during spontaneous expiration in anesthetized humans. These show that 36-74% of the elastic energy stored during inspiration is wasted in terms of negative inspiratory muscle work.     

Light microscopy of glial cells in turtles and birds

Summary Four classes of glial cells can be recognized in the central nervous system of turtles and birds on the basis of nuclear characteristics (methylene blue) and external morphology (Golgi technique). It seems likely that astrocytes and ependymal cells have a similar origin and function, but no evidence has been seen to indicate that transitional forms exist between astrocytes and oligodendrocytes or microgliacytes. Ependymal cells in the tectum and forebrain are covered by lamellate excrescences which are absent on cells in the spinal cord. Protoplasmic astrocytes are restricted to the gray matter. In the turtle they have an elongate shape characteristic of primitive elements, but stellate forms typical of mammals predominate in the bird. Fibrous astrocytes are abundant in the white matter. Endfeet are lacking in the turtle except on cells located near the pia; they are common for all elements in the bird and can sometimes be observed to outline the course of capillaries. Oligodendrocytes are identical to mammalian and amphibian forms. Small, round somata and long, thin processes are typical of types I and II while a tubular reticulum or membranous sheath characterizes type IV. The lack of a well defined somata and absence of transitional forms (type III) are compatible with the possibility that type IV is not a true cell type but corresponds to the inner cytoplasmic tongue of myelin. Microgliacytes are present in gray and white matter; they have a smaller overall size in the turtle and young chicken than in adult birds.Supported by a postdoctoral fellowship from the United States National Institutes of Health, NB 28,013-01Al.     

Nesting behavior ofHalictus scabiosæ in Switzerland (Hymenoptera,Halictidæ)

Summary The structure and populations of seven nests ofHalictus scabiosæ were studied in Geneva during July and August, 1964. Cells, containing brood of various ages, were radially arranged along the branching burrows. The one to six females in each nest did not belong to distinct worker and queen castes. This is in contrast to reports of distinct castes in what appears to be the same species from France. Most pollen collectors in the Geneva population were inseminated and some probably laid eggs.

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Zusammenfassung Die Struktur und die Bewohner von sieben Nestern vonHalictus scabiosæ, die im Juli und August 1964 in der Gegend um Genf gesammelt wurden, werden beschrieben. Zellen mit Brut verschiedenen Alters sind entlang verzweigter Gänge radiär angeordnet. Bis zu sechs Weibchen befanden sich in einem einzelnen Nest. Sie waren nicht in Königinnen und Arbeiterinnen differenziert, wie dies von Bienen aus Frankreich berichtet ist, die wahrscheinlich zur selben Art gehören. Die meisten Pollen-Sammlerinnen der Genfer Population waren befruchtet und einige von ihnen legten auch wahrscheinlich Eier.

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Résumé La structure et la population de sept nids deHalictus scabiosæ ont été étudiées à Genève durant les mois de juillet et d'août 1964. Des cellules contenant du couvain d'âge différent sont disposées en rayons le long de terriers qui se ramifient. Chaque nid était occupé par une à six femelles qui n'appartenaient pas à une caste distincte d'ouvrières ou de reine. La plupart des abeilles qui collectent le pollen furent inséminées et quelques-unes probablement pondèrent des ufs.

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Contribution No. 1303 from the Department of Entomology, The University of Kansas, Lawrence.     

A practical method for rescuing desired hybridomas during monoclonal antibody production

Summary We present a practical method for the rescue of previosly stable hybridoma clones which increases the proportion of desired cells in the population before cloning by limiting dilution. When the antibody activity of a culture supernatant was lower than that previously obtained, a precloning distribution at a density of 10 cells per microtiter well greatly improved the chances of obtaining a single active clone by subsequent limiting dilution. The Poisson distribution model was used to evaluate the method. Probabilities calculated clearly demonstrate the advantage of this precloning distribution step when attempting to isolate a hydridoma cell line that is relatively rare in a population. This work was supported in part by grants EY 06225 and EY 06226 from the National Eye Institute of the National Institutes of Health, Bethesda, MD and by an unrestricted departmental award from Research to Prevent Blindness, Inc.