Circulating platelet aggregates and progression of visual field loss in glaucoma

The aim of the study was to assess a relationship between circulating platelet aggregates (CPA) and progression of visual field loss in primary open-angle glaucoma patients. CPA was determined in 27 patients with open-angle glaucoma with nonprogressive visual field loss and 15 patients with open-angle glaucoma and progression of visual field loss. Intraocular pressure (IOP) under topical therapy was < 18 mmHg in all patients. CPA in glaucoma patients with progression of visual field loss was not significantly higher than those without visual field progression (p = 0.59). In conclusion, our study shows that increased platelet aggregability is not solely responsible for progression of visual field loss in glaucoma patients, and indicates the role of IOP in the pathogenesis of visual field loss.     

Transfer of M2 muscarinic acetylcholine receptors to clathrin-derived early endosomes following clathrin-independent endocytosis

Upon agonist stimulation, many G protein-coupled receptors such as beta(2)-adrenergic receptors are internalized via beta-arrestin- and clathrin-dependent mechanisms, whereas others, like M(2) muscarinic acetylcholine receptors (mAChRs), are internalized by clathrin- and arrestin-independent mechanisms. To gain further insight into the mechanisms that regulate M(2) mAChR endocytosis, we investigated the post-endocytic trafficking of M(2) mAChRs in HeLa cells and the role of the ADP-ribosylation factor 6 (Arf6) GTPase in regulating M(2) mAChR internalization. Here, we report that M(2) mAChRs are rapidly internalized by a clathrin-independent pathway that is inhibited up to 50% by expression of either GTPase-defective Arf6 Q67L or an upstream Arf6 activator, Galpha(q) Q209L. In contrast, M(2) mAChR internalization was not affected by expression of dominant-negative dynamin 2 K44A, which is a known inhibitor of clathrin-dependent endocytosis. Nevertheless, M(2) mAChRs, which are initially internalized in structures that lack clathrin-dependent endosomal markers, quickly localize to endosomes that contain the clathrin-dependent, early endosomal markers early endosome autoantigen-1, transferrin receptor, and GTPase-defective Rab5 Q79L, which is known to swell early endosomal compartments. These results suggest that M(2) mAChRs initially internalize via an Arf6-associated, clathrin-independent pathway but then quickly merge with the clathrin endocytic pathway at the level of early endosomes.     

Murine peripherin gene sequences direct Cre recombinase expression to peripheral neurons in transgenic mice

Group B streptococcus (GBS) induced macrophage apoptosis by which it could avoid host defence mechanisms. Macrophages, which constitutively express phosphatidylserine (PtdSer) on the outer leaflet of plasma membrane, increased PtdSer exposure during GBS-induced apoptosis. Induction of apoptosis decreased PtdSer radioactivity of macrophages incubated with [³H]serine. The effect appeared not due to increasing conversion of PtdSer to phosphatidylethanolamine or phosphatidylcholine nor to the release of radioactive membrane vesicles. The radioactivity in lysoPtdSer was also reduced. These results confirm that induction of apoptosis involves a modification of PtdSer metabolism and point out the typical features of the GBS-induced apoptosis with respect to other models of apoptosis.     

The solution structure of a gallium-substituted putidaredoxin mutant: GaPdx C85S

The Fe₂S₂ cluster of the ferredoxin putidaredoxin (Pdx) can be replaced by a single gallium ion, giving rise to a colorless, diamagnetic protein in which, apart from the metal binding site, the major structural features of the native ferredoxin are conserved. The solution structure of the C85S variant of gallium putidaredoxin (C85S GaPdx), in which a non-ligand cysteine is replaced by a serine, has been determined via multidimensional NMR methods using uniformly ¹⁵N,¹³C labeled samples of C85S GaPdx. Stereospecific assignments of leucine and valine methyl resonances were made using ¹³C,¹H HSQC spectra obtained with fractionally ¹³C-labeled samples, and backbone dihedral angle restraints were obtained using a combination of two-dimensional J-modulated ¹⁵N,¹H HSQC and three-dimensional (HN)CO(CO)NH experiments. A total of 1117 NOE-derived distance restraints were used in the calculations, including 454 short range ($i - j 3$), 456 long range (i - j 4) interresidue restraints and 207 non-trivial intraresidue restraints. 97 and 55 ₁ angular restraints were also included in the calculation of a family of 20 structures using a combined distance geometry-simulated annealing protocol. Most regions of the protein are well defined in the calculations, with an RMSD of 0.525 Å for backbone atoms excluding the metal binding loop (residues 34–48) and the last three C-terminal residues (residues 103–106). Where comparison is possible, these regions show an increase in dynamic behavior over the native protein, as does the loop containing residues 74–76. Structural and dynamic differences between native Pdx and GaPdx are discussed in relation to charge and packing of the metal binding site.     

Molecules and cognition: The latterday lessons of levels,language, and iac. Evolutionary overview of brain structure and function in some vertebrates and invertebrates

The characteristics of the nervous systems of a number of organisms in different phyla are examined at the recombinant DNA, protein, neuroanatomic, neurophysiological, and cognitive levels. Among the invertebrates, special attention is paid to the advantages as well as the shortcomings of the fly Drosophila melanogaster, the worm Caenorhabditis elegans, the honey bee Apis mellifera, the sea hare Aplysia californica, the octopus Octopus vulgaris, and the squid Loligo pealei. Among vertebrates, the focus is on Homo sapiens, the mouse Mus musculus, the rat Rattus norvegicus, the cat Felis catus, the macaque monkey Macaca fascicularis, the barn owl Tyto alba, and the zebrafish Brachydanio rerio. Vertebrate nervous systems have also been compared in fossil vs. extant organism. I conclude that complex nervous systems arose in the Early Cambrian via a big bang that was underpinned by a modular method of construction involving massive pleiotropy of gene circuits. This rapidity of construction had enormous implications for the degrees of freedom that were subsequently available to evolving nervous systems. I also conclude that at the level of neuronal populations and interactions of neuropiles there is no model system between phyla except at the basic macromolecular level. Further, I argue that to achieve a significant understanding of the functions of extant nervous systems we need to concentrate on fewer organisms in greater depth and manipulate genomes via transgenic technologies to understand the behavioral outputs that are possible from an organism. Finally, I analyze the concepts of “perceptual categorization” and “information processing” and the difficulties involved in the extrapolation of computer analogies to sophisticated nervous systems. © 1993 John Wiley & Sons, Inc.     

An aminomethylpyrimidine DPP-IV inhibitor with improved properties

A recently identified DPP-IV inhibitor (1) was found to induce phospholipidosis and to inhibit CYP3A4. A small series of less lipophilic and less amphiphilic analogues was synthesized in an effort to overcome these issues. One compound from this series was equipotent to 1, did not induce phospholipidosis and showed a reduced CYP3A4 inhibition.     

Functional conservation of RNA polymerase II in fission and budding yeasts

The complementary DNAs of the 12 subunits of fission yeast (Schizosaccharomyces pombe) RNA polymerase II were expressed from strong promoters in Saccharomyces cerevisiae and tested for heterospecific complementation by monitoring their ability to replace in vivo the null mutants of the corresponding host genes. Rpb1 and Rpb2, the two largest subunits and Rpb8, a small subunit shared by all three polymerases, failed to support growth in S. cerevisiae. The remaining nine subunits were all proficient for heterospecific complementation and led in most cases to a wild-type level of growth. The two alpha-like subunits (Rpb3 and Rpb11), however, did not support growth at high (37 degrees C) or low (25 degrees C) temperatures. In the case of Rpb3, growth was restored by increasing the gene dosage of the host Rpb11 or Rpb10 subunits, confirming previous evidence of a close genetic interaction between these three subunits.