Appearance of additional ouabain binding sites on platelet surfaces following activation.

Na,K-ATPase is an integral membrane protein complex involved in the regulation of intracellular Na+ and K+ ion levels. The number of accessible Na,K-ATPase complexes on the surface of human platelets before and after activation by adenosine 5'-diphosphate (ADP) was quantitated using ouabain, a highly specific inhibitor of this enzyme. Studies with [3H]ouabain revealed an increase in the number of accessible ouabain binding sites from approximately 55 per resting platelet to approximately 625 per ADP-activated platelet. Ultrastructural localization of this ion transport complex was also studied in resting human platelets or following activation by ADP. Localization was performed using a novel colloidal gold affinity probe which employed ouabain (ouabain-GAP) as a ligand specific for the Na,K-ATPase. Qualitatively, the number of surface-associated ouabain-GAP labels per platelet was observed to increase following activation by ADP. Fracture-labeling studies of frozen preparations of resting and freshly activated platelets suggested ouabain binding sites were associated with intracellular platelet structures during the early stages of activation. Together these results suggest an increase in the amount of ouabain binding sites at the surface of human platelets as an event of activation and that at least some of these additional sites may originate from intracellular sites.

A PDGF receptor mutation in the mouse (Patch) perturbs the development of a non-neuronal subset of neural crest-derived cells.

The Patch (Ph) mutation in mice is a deletion of the gene encoding the platelet-derived growth factor receptor alpha subunit (PDGFR alpha). Patch is a recessive lethal recognized in heterozygotes by its effect on the pattern of neural crest-derived pigment cells, and in homozygous mutant embryos by visible defects in craniofacial structures. Since both pigment cells and craniofacial structures are derived from the neural crest, we have examined the differentiation of other crest cell-derived structures in Ph/Ph mutants to assess which crest cell populations are adversely affected by this mutation. Defects were found in many structures populated by non-neuronal derivatives of cranial crest cells including the thymus, the outflow tract of the heart, cornea, and teeth. In contrast, crest-derived neurons in both the head and trunk appeared normal. The expression pattern of PDGFR alpha mRNA was determined in normal embryos and was compared with the defects present in Ph/Ph embryos. PDGFR alpha mRNA was expressed at high levels in the non-neuronal derivatives of the cranial neural crest but was not detected in the crest cell neuronal derivatives. These results suggest that functional PDGF alpha is required for the normal development of many non-neuronal crest-derived structures but not for the development of crest-derived neuronal structures. Abnormal development of the non-neuronal crest cells in Ph/Ph embryos was also correlated with an increase in the diameter of the proteoglycan-containing granules within the crest cell migratory spaces. This change in matrix structure was observed both before and after crest cells had entered these spaces. Taken together, these observations suggest that functional PDGFR alpha can affect crest development both directly, by acting as a cell growth and/or survival stimulus for populations of non-neurogenic crest cells, and indirectly, by affecting the structure of the matrix environment through which such cells move.

[The efficacy of dequalinium chloride/benzalkonium chloride as well as medicinal plants on the gingiva]. / Untersuchungen zur Wirksamkeit von Dequaliniumchlorid/Benzalkoniumchlorid sowie von Heilpflanzen auf die Gingiva.

A comparative study involving 50 patients of both genders was conducted in order to determine the effect of dequalinium chloride/benzalconium chloride mouthrinse (Dequonal), and of a preparation of herbal essences (Parodontax) on gingival health. Each of the mouthrinses was used during four weeks by a group of 25 patients who were instructed to abstain from any other oral hygiene measure during this period. Approximal plaque index, sulcus bleeding index and saliva pH were significantly enhanced by both preparations. A slightly better effect shown by dequalinium chloride/benzalconium chloride was not significant.

Association between collagen and glycosaminoglycans is altered in dermal extracellular matrix of fetal Steel (Sld/Sld) mice.

Altered extracellular matrix produced by Steel mutant fetuses affects the pigmentation of neural crest cells in vitro (K. Morrison-Graham, L. West-Johnsrud, and J.A. Weston, 1990, Dev. Biol. 139). Here, we demonstrate that collagen bundle morphology and hyaluronidase sensitivity of the glycosaminoglycans associated with the collagen fibrils differ between normal and mutant dermis. Although no differences were detected in the amounts of collagen or glycosaminoglycans produced in vitro or present in vivo, hyaluronic acid was more readily extracted from Sld/Sld than from normal skin. We suggest that the Steel mutation alters the organization of collagen bundles and associated hyaluronic acid within the extracellular matrix.

Neuronal development in the Drosophila compound eye: rap gene function is required in photoreceptor cell R8 for ommatidial pattern formation.

In the compound eye of Drosophila, cell-cell interactions are thought to play an important role in the determination of neuronal cell fate and pattern morphogenesis. Recent work on the bride of sevenless (boss) gene has demonstrated an inductive role for photoreceptor R8 in the differentiation of photoreceptor R7. These studies have shown that while R8 differentiates early in the scheme of ommatidial assembly, it continues to play an active role in subsequent patterning events. We describe studies on a new genetic locus rap (retina aberrant in pattern), whose functions are critical for normal pattern formation in the developing eye. Mutations in the rap gene perturb the early stages of pattern formation and lead to a variable number of photoreceptor cells (R cells) in each ommatidium. Experiments with a temperature-sensitive allele have shown that rap gene function is required during the period of development when pattern formation occurs. In addition, a somatic mosaic analysis of rap has shown that its function is required only in photoreceptor cell R8 for normal ommatidial patterning. These studies suggest an important role for rap in the initial events leading to pattern formation and are consistent with R8 playing a central role in directing ommatidial pattern formation.

Molecular architecture of Escherichia coli F1 adenosinetriphosphatase.

The structure of the E. coli F1 ATPase (ECF1) has been studied by a novel combination of two specimen preparation and image analysis techniques. The molecular outline of the ECF1 was determined by three-dimensional reconstruction of images of negatively stained two-dimensional crystals of ECF1. Internal features were revealed by analysis of single particles of ECF1, preserved in their native state in a thin layer of amorphous ice, and examined by cryoelectron microscopy. Various projections of the unstained ECF1 were interpreted consistently with the three-dimensional structure in negative stain, yielding a more informative description of the enzyme than otherwise possible. Results show that the ECF1 is a roughly spherical complex approximately 90-100 A in diameter. Six elongated protein densities (the alpha and beta subunits, each approximately 90 A X approximately 30 A in size) comprise its hexagonally modulated periphery. At the center of the ECF1 is an aqueous cavity which extends nearly or entirely through the length of the complex. A compact protein density, located at one end of the hexagonal barrel and closely associated with one of the peripheral subunits, partially obstructs the central cavity.

Factors guiding optic fibers in developing Xenopus retina.

We have characterized, by electron microscopy, the growth of pioneering axons from the retina into the visual pathway during early development of Xenopus laevis. The subsequent development of following fibers from the growing retinal margin as they accumulated in the ganglion cell fiber layer (GCFL) of the retina was also studied. Extracellular channels bordered by neuroepithelial cells appear in the developing retina in a dorsal to ventral gradient before any pioneering axons are seen. Pioneering axons are subsequently observed in these channels, usually surrounded by neuroepithelial cell processes. Ruthenium red treatment of embryonic retinas reveals extracellular matrix (ECM) within these retinal channels, while extracellular spaces in the proximal optic stalk, just beyond the optic disc, lack this material. ECM is also seen in optic tectum wherever ingrowing retinal and nonretinal axons are found. The channels and the ECM contained within them may provide guidance cues for pioneering retinal axons. The early association of pioneering retinal axons with neuroepithelial cell processes (putative glia) appears to be important in further development of the GCFL. The so-called following fibers of ganglion cells, arising later in development, fasciculate with pioneer axons in extracellular spaces and form fiber bundles of the GCFL on top of the layer of glial cell endfeet. It is not clear whether pioneering axons, glial cell surfaces, or both serve as guidance cues for following fiber migration.