Exogenous nitric oxide causes collapse of retinal ganglion cell axonal growth cones in vitro

We show that nitric oxide (NO) from applied NO-donating chemicals induces collapse of ganglion cell axonal growth cones extending from explants of tadpole retina in culture. Peroxynitrite, a neurotoxic product of NO and superoxide reaction, did not induce collapse, and oxyhemoglobin, which binds NO, blocked the highly effective collapsing activity of the NO donor S-nitrosocysteine. Membrane-permeable analogs of cyclic guanosine monophosphate had no collapsing activity. Inhibitors of NO synthase did not induce collapse. NO is a potential retrograde messenger through which postsynaptic neurons signal to their inputs to modify synaptic efficacy following NMDA receptor activation. Our results suggest a role for NO as such a messenger during development of the retinotectal projection. © 1996 John Wiley & Sons, Inc.     

Myosin light chain kinase: expression in neurons and upregulation during axon regeneration

Myosin light-chain kinase (MLCK) regulates actin-myosin II interactions in nonskeletal muscle cells, and the use of specific pharmacological inhibitors has implicated MLCK in retinal growth cone motility and neurite outgrowth. To further establish the existence and functions of MLCK in neurons, we isolated cDNAs encoding two forms of goldfish MLCK that were differentially expressed in the brain and gut and we sequenced the form most abundantly expressed in the brain (GFMLCK1). In situ hybridization with a cRNA probe specific to GFMLCK1 revealed widespread expression in CNS neurons, including tectal periventricular neurons and cerebellar and medullary neurons. After optic nerve crush, expression was markedly increased in the retinal ganglion cells. Expression peaked during the phase of axonal outgrowth, which, when taken together with our previous pharmacological studies, further supports a role for MLCK in growth cone motility. © 1996 John Wiley & Sons, Inc.     

Chemical sympathectomy and postganglionic nerve transection produce similar increases in galanin and VIP mRNA but differ in their effects on peptide content

Large changes in neuronal gene expression occur in adult peripheral neurons after axonal transection. In the rat superior cervical ganglion, for example, neurons that do not normally express vasoactive intestinal peptide (VIP) or galanin do so after postganglionic nerve transection. These effects of axotomy could result from a number of aspects of the surgical procedure. To test the idea that the important variable might be the disconnection of axotomized neuronal cell bodies from their target tissues, we examined the effects of producing such a disconnection by means of the compound 6-hydroxydopamine (6-OHDA), a neurotoxin that causes degeneration of sympathetic varicosities and avoids many of the complications of surgery. Two days after 6-OHDA treatment, VIP and galanin immunoreactivities had increased two- and 40-fold, respectively. Nevertheless, these increases were substantially smaller than the 30- and 300-fold changes seen after surgical axotomy. When expression of VIP and galanin was examined at the mRNA level, however, comparable increases were found after either procedure. The results indicate that chemical destruction of sympathetic varicosities produces an equivalent signal for increasing VIP and galanin mRNA as does axonal transection. The differences in the neuropeptide levels achieved suggests that peptide expression after nerve transection is regulated both at the mRNA and protein levels. © 1996 John Wiley & Sons, Inc.     

Screen for mutations affecting development of zebrafish neural crest

The neural crest provides a useful model to learn how cell fate diversification is regulated during vertebrate development. Our approach is to isolate zebrafish mutations in which the development of neural crest derivatives is disrupted, in order to learn about the underlying genetic mechanisms. We describe a screen in which parthenogenetic diploid embryos are examined both for visible phenotypes and for cellular defects in neural crest-derived sensory neurons recognized immunohistochemically. We present preliminary results from this screen and briefly describe a few representative mutations. We also discuss the general utility of our strategy and comment on the future directions of this approach. © 1996 Wiley-Liss, Inc.     

平原就读的藏族与汉族腹型肥胖大学生心肺功能比较研究

目的:探讨腹型肥胖对平原就读藏汉族大学生心肺功能影响的差异研究。方法:选取在校男性大学生96人,依照BMI和WC分类标准,分为藏族腹型肥胖组(TA)、藏族对照组(TC)、汉族腹型肥胖组(HA)、汉族对照组(HC)各24名。使用心脏彩色多普勒超声检查仪检测超声心动图,检测心脏结构和AV、PV、MVE、MVA、TVE、TVA等心功能指标。检测VC、FVC、FEV1、PEF、MMF、MVV等肺功能指标。检测身体形态学,血液和脂肪肝状况等基础指标。结果:腹型肥胖对血压的影响：与TC组相比,TA组SBP显著增加(P<0.01);与HC组相比,HA组SBP和DBP均显著增加(P<0.01,P<0.05)。腹型肥胖对心功能的负性影响：与TC组相比,TA组AV和PV显著减慢(P均<0.01);与HC组相比,HA组IVSA显著增大(P<0.05)。腹型肥胖对肺功能的负性影响：与TC组相比,TA组MMF和肺活量/体重水平显著降低(P<0.05);与HC组相比,HA组FEV1、FEV1%、PEF、MMF、MVV、MVV%和肺活量/体重水平均显著降低(P均<0.05)...     

Cloning and characterization of zRICH, a 2',3'-cyclic-nucleotide 3'-phosphodiesterase induced during zebrafish optic nerve regeneration

We previously reported cloning of cDNAs encoding both components of a protein doublet induced during goldfish optic nerve regeneration. The predicted protein sequences showed significant homology with the mammalian 2',3'-cyclic-nucleotide 3'-phosphodiesterases (CNPases). CNPases are well-established markers of mammalian myelin; hence, the cDNAs were designated gRICH68 and gRICH70 (for goldfish Regeneration-Induced CNPase Homologues of 68 and 70 kDa). Homologous cDNAs have now been isolated from zebrafish encoding a highly related protein, which we have termed zRICH. RNase protection assays show that zRICH mRNA is induced significantly (fivefold) in optic nerve regenerating zebrafish retinas 7 days following nerve crush. Western blots show a single band in zebrafish brain and retina extracts, with immunoreactivity increasing three-fold in regenerating retinas 21 days postcrush. Immunohistochemical analysis indicated that this increase in zRICH protein expression is localized to the retinal ganglion cell layer in regenerating retina. We have characterized and evaluated the relevance of a conserved beta-ketoacyl synthase motif in zRICH to CNPase activity by means of site-directed mutagenesis. Two residues within the motif, H334 and T336, are critical for enzymatic activity. A cysteine residue within the motif, which corresponds to a critical residue for beta-ketoacyl synthase, does not appear to participate in the phosphodiesterase activity.     

The levels of leukemia inhibitory factor mRNA in a Schwann cell line are regulated by multiple second messenger pathways

Axotomy of sympathetic and sensory neurons leads to changes in their neuropeptide phenotypes. These changes are mediated in part by the induction of leukemia inhibitory factor (LIF) by nonneuronal cells. In the present study, we identified satellite/Schwann cells as a possible source of the injury-induced LIF. Using a Schwann cell line, SC-1 cells, we examined mechanisms of LIF induction. LIF mRNA levels increased rapidly when the cells were treated with 8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate, phorbol 12-myristate 13-acetate (PMA), or A23187. Among these reagents, PMA was the most efficacious. Inhibition of protein kinase C (PKC) by GF-1 09203X significantly reduced the PMA-induced LIF mRNA levels. As PKC is known to activate the extracellular signal-regulated kinase (ERK) signaling pathway, the involvement of this pathway in the PMA-stimulated induction of LIF mRNA was examined. Phosphorylation of ERKs was increased following PMA treatment in SC-1 cells. Moreover, inhibition of ERK kinase activity by PD98059 dramatically reduced PMA-stimulated phosphorylation of ERKs and induction of LIF mRNA. These results indicate that LIF mRNA levels can be regulated by ERK activation via stimulation of PKC in Schwann cells.     

Morphology and ultrastructure of spiracles in phlebotomine sandfly larvae

The morphology and ultrastructure of the larval spiracle system of three phlebotomine sandfly species, Phlebotomus perniciosus, P. perfiliewi and P. papatasi, were examined by scanning (SEM) and transmission (TEM) electron microscopy and by confocal scanning laser microscopy (CSLM). During larval development, thoracic and abdominal spiracles show considerable modifications. In fourth instar larvae, the spiracles consist of a plate with a sclerotized central portion and a peripheral circle of papillae. The latter is distinctive in the larvae of P. papatasi, which are readily distinguished from the other species. Opening clefts across the papillae communicate with an internal chamber that encircles an electrondense plug. Many cylindrical projections cross the chamber, uniting the central plug with the larval body, forming an air filter. Spiracular development in successive larval instars has both a taxonomic and adaptive value.     

Identification of an abdominal ganglion neuron antagonizing L7-driven gill contraction in Aplysia

Gill motor neuron L7-induced longitudinal shortening of the gill in Aplysia kurodai and A. juliana was suppressed when extracellular stimuli were applied to a restricted dorsal central region of the abdominal ganglion. We found a neuron there which antagonized the L7-driven contraction. Since the contraction was suppressed when the identified neuron was activated simultaneously with L7, we refer to the newly found neuron as “Anti-L7”. Anti-L7 did not change the L7 impulse generation in the abdominal ganglion. No direct synaptic connection from L7 to Anti-L7 was detected. A fluorescent dye injected into the soma of Anti-L7 revealed that the neuron sent axonal branches to the branchial nerve. These results may show that Anti-L7 antagonizes L7 at the periphery inside the gill, rather than in the abdominal ganglion. EJPs induced by L7 were unaffected by Anti-L7. Activation of Anti-L7 alone did not induce any change in tone or membrane potential of the gill musculature. The suppressive effect of Anti-L7 lasts many seconds after the cessation of a train of Anti-L7 impulses. The results may suggest that the suppression is mediated through an inhibitory neuromodulatory mechanism without inhibition of L7 itself. Accepted: 1 April 1999