Recognition of extracellular matrix components by neonatal and adult cardiac myocytes

The histogenesis of renal basement membranes was studied in grafts of avascular, 11-day-old mouse embryonic kidney rudiments grown on chick chorioallantoic membrane (CAM). Vessels of the chick CAM invade the mouse tissue during an incubation period of 7-10 days and eventually hybrid glomeruli composed of mouse epithelium and chick endothelium form. Formation of basement membranes during this development was followed by immunofluorescence and immunoperoxidase stainings using polyclonal and monoclonal antibodies against mouse and chick collagen type IV and against mouse laminin. These antibodies were species-specific as shown in immunochemical and immunohistologic analyses. The glomerular basement membrane contained both mouse and chick collagen type IV, demonstrating its dual cellular origin. All other basement membranes were either exclusively of chick origin (mesangium, vessels) or of mouse origin (tubuli, Bowman's capsule).     

Axon growth from limb motorneurons in the locust embryo: the effect of target limb removal on the pattern of axon branching in the periphery

Metathoracic limb buds have been unilaterally ablated from locust embryos at 25 to 30% of embryonic development and the effect of this operation on the axon morphology of the motorneuron fast extensor tibiae (FETi) observed at later embryonic stages. In control embryos this neuron sends a single axon out the main leg nerve, nerve 5, to the extensor tibiae muscle in the femur. In limb ablated embryos the axon of FETi is found in a wide variety of aberrant peripheral nerve pathways and projects to a wide range of foreign muscles. There is a degree of apparent selectivity, but no rigid hierarchy, in the choice of pathway and muscle made by FETi. A high degree of variability is found between one embryo and another in the extent and pattern of axon branching. The axon of FETi is generally found in pathways that correspond to nerves in control embryos but on occasion grows along novel routes. An anteriorly directed dendritic branch, seldom seen in control FETi neurons, is frequently seen in experimental FETis. These findings are discussed in terms of the rules for specific axon growth in normal development.     

The nutritional ecology of larvae of Alsophila pometaria and Anisota senatoria feeding on early- and late-season oak foliage

The larvae of Alsophila pometaria (Harr.), feeding on the young foliage of oak, has a higher relative growth rate (RGR) and relative nitrogen accumulation rate (RNAR) than the larvae of Anisota senatoria (J. E. Smith), feeding on the mature foliage of oak. Although the young oak foliage is more efficiently digested by A. pometaria (higher AD's), it is not more efficiently assimilated and used for growth (no difference in ECI's). Thus, the higher growth rate of A. pometaria is due entirely to a higher consumption rate (RCR and RNCR). Young foliage is significantly higher in nitrogen and water than mature foliage, but phenol and tannin levels are comparable in young and old foliage. A. pometaria consumes the foliage of different oak species at the same rate, independent of nitrogen content, while A. senatoria increases its consumption rate in response to decreased nitrogen levels. As a result, the growth rate of A. pometaria is directly related to leaf nitrogen content, while the growth rate of A. senatoria is independent of leaf nitrogen. The two species of insects have digestive systems that are very similar biochemically, and that are well-designed for effective protein digestion. Tannins and phenols do not influence the nutrional indices of either species. We suggest that the major benefit of spring feeding is the availability of succulent, high-nitrogen foliage, and not the avoidance of high-tannin foliage. The spring feeder appears to have a feeding strategy that favors rapid growth at the expense of efficiency, while the late summer feeder has a strategy that favors efficiency over rate.

---

Résumé Alimentées sur feuillage jeune de chêne, les chenilles d'Alsophila pometaria avaient un taux relatif de croissance (RGR) et un taux relatif d'accumulation d'azote (RNAR) plus élevés que les chenilles d'Anisota senatoria alimentées sur feuillage mûr de chêne. Bien que le jeune feuillage soit plus efficacement digéré par A. pometaria (AD plus élevé), il n'est pas assimilé et utilisé pour la croissance avec de meilleurs rendements (les ECI ne sont pas différents). Ainsi le taux de croissance plus élevé d'A. pometaria est dû entièrement à un taux de consommation plus important (RCR et RNCR). Le feuillage jeune est significativement plus riche en azote et en eau que le feuillage mûr, mais les niveaux de phénol et de tanins sont les mêmes. A pometaria consomme les feuilles de différentes espèces de chênes au même taux, indépendamment de la teneur en azote, tandis que A. senatoria accroît sa consommation en réponse à une diminution de la teneur en azote. Il en résulte que le taux de croissance d'A. pometaria dépend directement de la teneur en azote des feuilles, tandis que celui d'A. senatoria en est indépendant. Les systèmes digestifs des deux insectes sont biochimiquement semblables et sont efficaces pour la digestion des protéines. Les tanins et les phénols n'influent pas sur les indices nutritionnels de ces deux espèces. Nous estimons que le principal intérêt de l'alimentation printanière est la disponibilité en feuillage succulent, riche en azote, et non l'absence de feuilles à haute teneur en tanin. L'alimentation printanière semble correspondre à une strategie alimentaire qui favorise la croissance aux dépens de l'efficacité tandis que l'alimentation en fin d'été est une stratégie qui favorise l'efficacité sur la rapidité.

     

Aphid growth and reproductive rates

Aphid size, although generally a good indicator of fecundity is not a good indicator of aphid performance over a wide range of conditions. In general, the greater the mean relative growth rate (MRGR) of apterous morphs of Rhopalosiphum padi (L.), the greater their fecundity. Intrinsic rate of increase (r_m) is even more closely correlated with mean relative growth rate than fecundity.Once these criteria are quantified for a morph of a particular species of aphid over a range of conditions the morphs intrinsic rate of increase on a particular host can be estimated by a quick measure of its mean relative growth rate.

---

Résumé La taille des pucerons, bien que considérée généralement comme un bon indice de la fécondité, n'est pas un bon indice pour leurs performances dans une grande gamme de conditions. En général, plus le taux moyen de croissance relative est élevé chez les types aptères de Rhopalosiphum padi, plus leur fécondité est importante. Le taux intrinsèque de croissance (r_m) est encore plus étroitement lié au taux moyen de croissance relative que la fécondité.Une fois que ces critères ont été quantifiés pour un type d'une espèce donnée de puceron dans une gamme de conditions, les taux intrinsèques de croissance des différents types sur un hôte particulier peuvent être estimés par une mesure rapide de leur taux moyen de croissance relative.

     

Cryopreservation of Plasmodium chabaudi. II. Cooling and warming rates

Various cooling and warming rates were investigated to determine the optimum conditions for cryopreserving the intraerythrocytic stages of Plasmodium chabaudi. Infected blood, equilibrated in 10% v/v glycerol at 37 degrees C or in 15% v/v Me2SO at 0 degree C for 10 min, was cryopreserved using cooling rates between 1 and 5100 degrees C min-1. After overnight storage in liquid nitrogen the samples were warmed at 12,000 degrees C min-1. Warming rates between 1 and 12,000 degrees C min-1 were investigated using samples previously cooled at 3600 degrees C min-1. After thawing, the glycerol and Me2SO were removed by dilution in 15% v/v glucose-supplemented phosphate-buffered saline. Survival was assayed by inoculation of groups of five mice each with 10(6) infected cells and the time taken to reach a level of 2% parasitemia estimated. The optimum cooling rate was 3600 degrees C min-1 for parasites frozen using either 10% glycerol or 15% Me2SO; the pre-2% patent periods were 0.90 and 1.01 days above control values (representing survival levels of 21 and 17.5%, respectively). The optimum warming rate was 12,000 degrees C min-1; the pre-2% patent periods were 1.01 and 1.32 days above control values, respectively (18 and 10% survival), for glycerol and Me2SO. With ethanediol (5% v/v) and sucrose (15% w/v) as cryoprotectants the optimum warming rates were also 12,000 degrees C min-1 while the optimum cooling rates were 330 and 3600 degrees C min-1, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Reversible inhibition of aggregation-related cohesivity in Dictyostelium discoideum by diffusible metabolites

Evidence presented elsewhere (G.B. Williams, E.M. Elder, and M. Sussman 1984, Dev. Biol. 105, 377-388) indicates that NH3 and certain carboxylic acids including propionate, succinate, and acetate modulate the cAMP relay in Dictyostelium discoideum. The former appears to act as a cAMP accumulation inhibitor, the latter as cAMP release inhibitors. The cohesive properties of aggregation competent cells have been assayed quantitatively in the presence of these modulators. The following results were obtained: (1) At pH 7.5, EDTA-resistant cohesivity was greatly inhibited by NH4C within the concentration range tested (30-3.8 mM). Even at the higher concentrations the effect was not immediate but required ca. 10 min for full expression. At the lower concentrations, the inhibitory level was only slightly reduced but the time for full expression progressively increased. At pH 6.5, the level of inhibition was marginal, indicating that NH3 is the active molecular species. By themselves, neither ambient pH nor ionic strength appeared to affect cohesive performance within the ranges employed. The inhibition was immediately and completely reversed upon removal of NH4Cl or a shift of ambient pH from 7.5 to 6.5. The presence of cycloheximide did not affect the recovery of cohesivity after NH4Cl removal. (2) The presence of 15 mM succinate, propionate, or acetate also reduced cell cohesivity. The timing and extent of the inhibition were identical at pH 7.5 and 6.5. The inhibition was expressed immediately and was reversible. Each of the acids acted synergistically with NH4Cl. The relative potencies of these metabolites acting singly or in combination as inhibitors of cohesivity corresponded roughly to their potencies as modulators of the cAMP relay (Williams et al., 1984). (3) The sensitivity to the metabolites was stage specific, being maximal during and shortly after aggregation and disappearing abruptly at 11-12 hr. This corresponds to the time at which this cohesive system, responsible for the end-to-end cell associations evident during aggregation (H. Beug, G. Gerisch, S. Kempff, V. Riedel, and G. Cremer, 1970, Exp. Cell. Res. 63, 147-158) is supplanted by a newly arisen, serologically and genetically distinct system which thereafter maintains the integrity of the aggregate (C. Steinemann and R.W. Parish, 1980, Nature (London) 286, 721-724; D.K. Wilcox and M. Sussman, 1981, Dev. Biol. 82, 102-112, and Proc. Natl. Acad. Sci. USA 78, 358-362; C.L. Saxe III and M. Sussman, 1982, Cell 29, 755-759). The activities of the metabolites, detailed above, are discussed in relation to their previously demonstrated activities as morphogens.     

Stage-specific expression of three cell surface carbohydrate antigens during murine spermatogenesis detected with monoclonal antibodies

We have identified three germ cell surface carbohydrate antigens that exhibit a common, stage-specific pattern of expression during spermatogenesis in the mouse. IgM-class monoclonal antibodies designated "J1," "C6," and "A5" were absorbed by adult testis, but not by any adult somatic tissue tested. In indirect immunofluorescence assays using collagenase-dissociated prepuberal and adult testicular cells, these antibodies labeled the surfaces of early and late pachytene spermatocytes and round spermatids. Gonocytes from fetal and neonatal testes were not labeled. In paraffin sections of prepuberal and adult testes, sialidase treatment exposed antigens recognized by antibodies C6 and A5 on preleptotene, leptotene, and zygotene spermatocytes located near the perimeter of seminiferous tubules. The determinants recognized by antibodies J1, C6, and A5 were characterized partially using a sugar hapten inhibition assay. The binding of J1 to adult testicular cells was inhibited specifically by N-acetylglucosamine and the binding of both C6 and A5 was inhibited by N-acetyllactosamine. The glycoconjugates recognized by J1, C6, and A5 eluted from gel filtration columns with an apparent molecular weight greater than 1 X 10(6) and were sensitive to endo-beta-galactosidase (keratanase) treatment. The apparent high molecular weight of these glycoconjugates was confirmed by immunolabeling Western blots of testis extracts separated by SDS-polyacrylamide gel electrophoresis. The results suggest that polylactosamine (keratan) glycoconjugates of high molecular weight are associated with the plasma membranes of meiotic and haploid male germ cells. The effects of sialidase on antibody labeling patterns suggest that changes in cell surface sialylation accompany the transition of early meiotic germ cells to pachytene spermatocytes during spermatogenesis.     

Induction of synthesis of bovine adrenocortical cytochromes P-450scc, P-45011 beta, P-450C21, and adrenodoxin by prostaglandins E2 and F2 alpha and cholera toxin

To further elucidate the mechanisms by which ACTH (adrenocorticotropin) exerts its long-term action to maintain normal levels of adrenocortical cytochromes P-450 and related enzymes, the abilities of cholera toxin and prostaglandins E2 and F2 alpha to induce the synthesis of cytochromes P-450scc, P-45011 beta, and P-450C21 and adrenodoxin have been examined. These effectors stimulate the production of cyclic AMP and thus steroidogenesis in the adrenal cortex. Using bovine adrenocortical cells in primary monolayer culture, we have shown that treatment with cholera toxin results in increased synthesis of cytochromes P-450scc and P-45011 beta and adrenodoxin, similar to the effect observed upon ACTH treatment. Prostaglandins E2 and F2 alpha are less effective at inducing the synthesis of the mitochondrial cytochromes P-450, and do not seem to induce the synthesis of adrenodoxin. Furthermore, cholera toxin was found to be less effective at inducing the synthesis of microsomal cytochrome P-450C21 than ACTH, and no more effective than the prostaglandins. Thus, while it appears that elevation of cyclic AMP levels is a necessary step leading to increased synthesis of adrenocortical forms of cytochrome P-450, the detailed mechanism of this induction will be found to be different for each of the different enzymes.     

1H NMR of glycosaminoglycans and hyaluronic acid oligosaccharides in aqueous solution: the amide proton environment

The exchangeable amide protons of hyaluronic acid (HA) oligosaccharides and a higher-molecular-weight segment dissolved in H2O at pH 2.5 or 5.5 were examined by H NMR spectroscopy at 250 MHz. The HA segment preparation showed a single amide resonance, near the chemical shift for the amide proton of the monosaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose (beta-GlcNAc). Smaller HA oligosaccharides showed two or three separate amide proton resonances, corresponding in relative peak area to interior or end GlcNAc residues. The interior GlcNAc amide resonance occurred at the same chemical shift as the single resonance of the HA segment. For the end GlcNAc residues, linkage to D-glucuronopyranose (GlcUA) through C1 resulted in an upfield shift relative to the beta-anomer of GlcNAc, whereas linkage through C3 resulted in a downfield shift relative to the corresponding anomer of GlcNAc. These chemical-shift perturbations appeared to be approximately offsetting in the case of linkage at both positions. The amide proton vicinal coupling constant (ca. 9 Hz) was found to be essentially independent of chain length, residue position, or solution pH. These data favor a nearly perpendicular orientation for the acetamido group with respect to the sugar ring, little affected by linkage of GlcNAc to GlcUA. No evidence for the existence of a stable hydrogen bond linking the amide proton with the carboxyl(ate) oxygen of the adjacent uronic acid residue was found. The amide proton resonances for chondroitin, chondroitin 4-sulfate, and dermatan sulfate were compared to that of HA. The chemical shifts of these resonances deviated no more than 0.1 ppm from that of HA. A small dependence on the identity of the adjacent uronic acid residue was noted, based on the observation of two resonances for dermatan sulfate.     

Disposition of polar and nonpolar residues on outer surfaces of transmembrane helical segments of proteins involved in proton translocation

"Helical wheel" projections of transmembrane helical segments of membrane proteins involved in proton translocation were constructed. The particular proteins studied were the uncF protein subunit of the Escherichia coli proton-ATPase, the uncE protein subunit of the E. coli proton-ATPase, and cytochrome oxidase subunit III. Clear demarcation of polar and nonpolar regions on surfaces of transmembrane helical segments was seen in the uncF protein and in uncE protein helical segment two, but not in uncE protein helical segment one. The transmembrane segment of cytochrome oxidase subunit III which includes the dicyclohexylcarbodiimide (DCCD)-reactive residue was very similar to E. coli uncE protein helical segment two. The DCCD-reactive residue in both was clearly located on a nonpolar surface.