Smooth Pursuit Eye Movements in Children with Strabismus and in Children with Vergence Deficits

Purpose

The objective of our study was to examine horizontal smooth pursuit performance in strabismic children and in children with vergence deficits, and to compare these data with those recorded in a group of control age-matched children.

Methods

Binocular eye movements were recorded by video-oculography in ten strabismic children (mean age: 9.8±0.8) and seven children with vergence deficits (mean age: 10.8±0.6). Data were compared to that of age-matched control children (mean age: 9.8±0.8 years).

Results

Catch-up saccades amplitude in strabismic children and in children with vergence deficits were significantly higher than in control age-matched children. Moreover, in strabismic children the amplitude of catch-up saccades was significantly higher in rightward than in leftward direction. The number of catch-up saccades was also significantly higher in rightward than in leftward direction. The gain value of pursuits in rightward direction was significantly higher in the right eye than in the left one; for the right eye, the gain value was significantly higher in rightward than in leftward direction. Binocular coordination of pursuit was better in control age-matched children than in children with vergence deficits and than in strabismic children.

Conclusions

Binocular coordination of pursuit is abnormal in children with vergence deficits and worse in strabismic children. Binocular vision plays an important role in improving binocular coordination of pursuit.     

Regulated exocytosis in neuroendocrine cells: a role for subplasmalemmal Cdc42/N-WASP-induced actin filaments

In neuroendocrine cells, actin reorganization is a prerequisite for regulated exocytosis. Small GTPases, Rho proteins, represent potential candidates coupling actin dynamics to membrane trafficking events. We previously reported that Cdc42 plays an active role in regulated exocytosis in chromaffin cells. The aim of the present work was to dissect the molecular effector pathway integrating Cdc42 to the actin architecture required for the secretory reaction in neuroendocrine cells. Using PC12 cells as a secretory model, we show that Cdc42 is activated at the plasma membrane during exocytosis. Expression of the constitutively active Cdc42(L61) mutant increases the secretory response, recruits neural Wiskott-Aldrich syndrome protein (N-WASP), and enhances actin polymerization in the subplasmalemmal region. Moreover, expression of N-WASP stimulates secretion by a mechanism dependent on its ability to induce actin polymerization at the cell periphery. Finally, we observed that actin-related protein-2/3 (Arp2/3) is associated with secretory granules and that it accompanies granules to the docking sites at the plasma membrane upon cell activation. Our results demonstrate for the first time that secretagogue-evoked stimulation induces the sequential ordering of Cdc42, N-WASP, and Arp2/3 at the interface between granules and the plasma membrane, thereby providing an actin structure that makes the exocytotic machinery more efficient.     

Calcium-regulated exocytosis of dense-core vesicles requires the activation of ADP-ribosylation factor (ARF)6 by ARF nucleotide binding site opener at the plasma membrane

Vacuole fusion requires a coordinated cascade of priming, docking, and fusion. SNARE proteins have been implicated in the fusion itself, although their precise role in the cascade remains unclear. We now report that the vacuolar SNAP-23 homologue Vam7p is a mobile element of the SNARE complex, which moves from an initial association with the cis-SNARE complex via a soluble intermediate to the docking site. Soluble Vam7p is specifically recruited to vacuoles and can rescue a fusion reaction poisoned with antibodies to Vam7p. Both the recombinant Vam7p PX domain and a FYVE domain construct of human Hrs block the recruitment of Vam7p and vacuole fusion, demonstrating that phosphatidylinositol 3-phosphate is a primary receptor of Vam7p on vacuoles. We propose that the Vam7p cycle is linked to the availability of a lipid domain on yeast vacuoles, which is essential for coordinating the fusion reaction prior to and beyond docking.     

Subjective Visual Vertical and Postural Capability in Children Born Prematurely

Purpose

We compared postural stability and subjective visual vertical performance in a group of very preterm-born children aged 3-4 years and in a group of age-matched full-term children.

Materials and Methods

A platform (from TechnoConcept) was used to measure postural control in children. Perception of subjective visual vertical was also recorded with posture while the child had to adjust the vertical in the dark or with visual perturbation. Two other conditions (control conditions) were also recorded while the child was on the platform: for a fixation of the vertical bar, and in eyes closed condition.

Results

Postural performance was poor in preterm-born children compared to that of age-matched full-term children: the surface area, the length in medio-lateral direction and the mean speed of the center of pressure (CoP) were significantly larger in the preterm-born children group (p < 0.04, p < 0.01, and p < 0.04, respectively). Dual task in both groups of children significantly affected postural control. The subjective visual vertical (SVV) values were more variable and less precise in preterm-born children.

Discussion-Conclusions

We suggest that poor postural control as well as perception of verticality observed in preterm-born children could be due to immaturity of the cortical processes involved in the motor control and in the treatment of perception and orientation of verticality.     

Molecular characterization and subcellular localization of macrophage infectivity potentiator, a Chlamydia trachomatis lipoprotein

Macrophage infectivity potentiator (MIP) was originally reported to be a chlamydial lipoprotein from experiments showing incorporation of radiolabeled palmitic acid into native and recombinant MIP; inhibition of posttranslational processing of recombinant MIP by globomycin, known to inhibit signal peptidase II; and solubility of native MIP in Triton X-114. However, the detailed structural characterization of the lipid moiety on MIP has never been fully elucidated. In this study, bioinformatics and mass spectrometry analysis, as well as radiolabeling and immunochemical experiments, were conducted to further characterize MIP structure and subcellular localization. In silico analysis showed that the amino acid sequence of MIP is conserved across chlamydial species. A potential signal sequence with a contained lipobox was identified, and a recombinant C20A variant was prepared by replacing the probable lipobox cysteine with an alanine. Both incorporation of U-(14)C-esterified glycerol and [U-(14)C]palmitic acid and posttranslational processing that was inhibitable by globomycin were observed for recombinant wild-type MIP but not for the recombinant C20A MIP variant. The fatty acid contents of native and recombinant MIP were analyzed by gas chromatography-mass spectrometry, and the presence of amide-linked fatty acids in recombinant MIP was investigated by alkaline methanolysis. These results demonstrated a lipid modification in MIP similar to that of other prokaryotic lipoproteins. In addition, MIP was detected in an outer membrane preparation of Chlamydia trachomatis elementary bodies and was shown to be present at the surfaces of elementary bodies by surface biotinylation and surface immunoprecipitation experiments.     

Annexin A2–dependent actin bundling promotes secretory granule docking to the plasma membrane and exocytosis

Annexin A2, a calcium-, actin-, and lipid-binding protein involved in exocytosis, mediates the formation of lipid microdomains required for the structural and spatial organization of fusion sites at the plasma membrane. To understand how annexin A2 promotes this membrane remodeling, the involvement of cortical actin filaments in lipid domain organization was investigated. 3D electron tomography showed that cortical actin bundled by annexin A2 connected docked secretory granules to the plasma membrane and contributed to the formation of GM1-enriched lipid microdomains at the exocytotic sites in chromaffin cells. When an annexin A2 mutant with impaired actin filament–bundling activity was expressed, the formation of plasma membrane lipid microdomains and the number of exocytotic events were decreased and the fusion kinetics were slower, whereas the pharmacological activation of the intrinsic actin-bundling activity of endogenous annexin A2 had the opposite effects. Thus, annexin A2–induced actin bundling is apparently essential for generating active exocytotic sites.     

Annexin 2 promotes the formation of lipid microdomains required for calcium-regulated exocytosis of dense-core vesicles

Annexin 2 is a calcium-dependent phospholipid-binding protein that has been implicated in a number of membrane-related events, including regulated exocytosis. In chromaffin cells, we previously reported that catecholamine secretion requires the translocation and formation of the annexin 2 tetramer near the exocytotic sites. Here, to obtain direct evidence for a role of annexin 2 in exocytosis, we modified its expression level in chromaffin cells by using the Semliki Forest virus expression system. Using a real-time assay for individual cells, we found that the reduction of cytosolic annexin 2, and the consequent decrease of annexin 2 tetramer at the cell periphery, strongly inhibited exocytosis, most likely at an early stage before membrane fusion. Secretion also was severely impaired in cells expressing a chimera that sequestered annexin 2 into cytosolic aggregates. Moreover, we demonstrate that secretagogue-evoked stimulation triggers the formation of lipid rafts in the plasma membrane, essential for exocytosis, and which can be attributed to the annexin 2 tetramer. We propose that annexin 2 acts as a calcium-dependent promoter of lipid microdomains required for structural and spatial organization of the exocytotic machinery.