PubMed Health⌕ Search

Biomedical subjects

M C Fishman

Publications and source records attributed to M C Fishman.

At least 55 records · Page 3Linked to original sources
PubMed1998

Neuronal cell surface molecules mediate specific binding to rabies virus glycoprotein expressed by a recombinant baculovirus on the surfaces of lepidopteran cells.

The existence of specific rabies virus (RV) glycoprotein (G) binding sites on the surfaces of neuroblastoma cells is demonstrated. Spodoptera frugiperda (Sf21) cells expressing G of the RV strain CVS (Gcvs-Sf21 cells) bind specifically to neuroblastoma cells of different species but not to any other cell type (fibroblast, myoblast, epithelial, or glioma). Attachment to mouse neuroblastoma NG108-15 cells is abolished by previous treatment of Gcvs-Sf2 cells with anti-G antibody. Substitutions for lysine at position 330 and for arginine at position 333 in RV G greatly reduce interaction between Gcvs-Sf21 cells and NG108-15 cells. These data are consistent with in vivo results: an avirulent RV mutant bearing the same double mutation is not able to infect sensory neurons or motoneurons (P. Coulon, J.-P. Ternaux, A. Flamand, and C. Tuffereau, J. Virol. 72:273-278, 1998) after intramuscular inoculation into a mouse. Furthermore, infection of NG108-15 cells by RV but not by vesicular stomatitis virus leads to a reduction of the number of binding sites at the neuronal-cell surface. Our data strongly suggest that these specific attachment sites on neuroblastoma cells represent a neuronal receptor(s) used by RV to infect certain types of neurons in vivo.

Animals↗
PubMed1998

"Physiological genomics": mutant screens in zebrafish.

Large-scale mutagenesis screens have proved essential in the search for genes that are important to development in the fly, worm, and yeast. Here we present the power of large-scale screening in a vertebrate, the zebrafish Danio rerio, and propose the use of this genetic system to address fundamental questions of vertebrate developmental physiology. As an example, we focus on zebrafish mutations that reveal single genes essential for normal development of the cardiovascular system. These single gene mutations disrupt specific aspects of rate, rhythm, conduction, or contractility of the developing heart.

Animals↗
PubMed1998

Expression of hypothalamic peptides in mice lacking neuronal nitric oxide synthase: reduced beta-END immunoreactivity in the arcuate nucleus.

The gas nitric oxide (NO) is an important messenger in brain signaling. Along with many other functions, NO is thought to influence the expression and/or release of various hypothalamic hormones (corticotropin-releasing hormone (CRH), gonadotropin-releasing hormone (GnRH) and vasopressin). To learn more about the role of NO in neuroendocrine mechanisms, we studied in mutant mice lacking neuronal isoform of NO synthase (nNOS) the cellular expression of CRH, neurophysin (the carrier protein of vasopressin/oxytocin) and pro-opiomelanocortin (POMC), as well as of the POMC-derived peptides beta-endorphin (beta-END), alpha-melanocyte-stimulating hormone (alpha-MSH) and corticotropin (ACTH) by use of immunohistochemistry and in situ hybridization. Additionally, the remaining NO-generating capacities of the nNOS minus mice were investigated by NADPH-diaphorase histochemistry and citrulline immunohistochemistry as well as by immunohistochemical localization and Western blot analysis of endothelial NOS (eNOS) and nNOS isoforms. Amongst all hypothalamic peptides under investigation, only beta-END was found to be altered in mutant mice. A morphometric analysis of beta-END producing neurons of the arcuate nucleus revealed that significantly less cells were immunoreactive in mutant mice, whereas the expression of the precursor POMC as well as of other POMC-derived peptides was found to be unchanged. In addition to that, fewer beta-END-immunoreactive fibers were found in the paraventricular nucleus of nNOS minus mice in comparison to wild-type animals. Hence, the reduction of hypothalamic beta-END is probably a posttranslational event that might reflect a disturbed endorphinergic innervation of those hypothalamic neurons which normally express nNOS.

Animals↗
PubMed1998

Early development of the zebrafish pronephros and analysis of mutations affecting pronephric function.

The zebrafish pronephric kidney provides a simplified model of nephron development and epithelial cell differentiation which is amenable to genetic analysis. The pronephros consists of two nephrons with fused glomeruli and paired pronephric tubules and ducts. Nephron formation occurs after the differentiation of the pronephric duct with both the glomeruli and tubules being derived from a nephron primordium. Fluorescent dextran injection experiments demonstrate that vascularization of the zebrafish pronephros and the onset of glomerular filtration occurs between 40 and 48 hpf. We isolated fifteen recessive mutations that affect development of the pronephros. All have visible cysts in place of the pronephric tubule at 2-2.5 days of development. Mutants were grouped in three classes: (1) a group of twelve mutants with defects in body axis curvature and manifesting the most rapid and severe cyst formation involving the glomerulus, tubule and duct, (2) the fleer mutation with distended glomerular capillary loops and cystic tubules, and (3) the mutation pao pao tang with a normal glomerulus and cysts limited to the pronephric tubules. double bubble was analyzed as a representative of mutations that perturb the entire length of the pronephros and body axis curvature. Cyst formation begins in the glomerulus at 40 hpf at the time when glomerular filtration is established suggesting a defect associated with the onset of pronephric function. Basolateral membrane protein targeting in the pronephric duct epithelial cells is also severely affected, suggesting a failure in terminal epithelial cell differentiation and alterations in electrolyte transport. These studies reveal the similarity of normal pronephric development to kidney organogenesis in all vertebrates and allow for a genetic dissection of genes needed to establish the earliest renal function.

Animals↗
PubMed1998

Regulation in the heart field of zebrafish.

In many vertebrates, removal of early embryonic heart precursors can be repaired, leaving the heart and embryo without visible deficit. One possibility is that this 'regulation' involves a cell fate switch whereby cells, perhaps in regions surrounding normal progenitors, are redirected to the heart cell fate. However, the lineage and spatial relationships between cells that are normal heart progenitors and those that can assume that role after injury are not known, nor are their molecular distinctions. We have adapted a laser-activated technique to label single or small patches of cells in the lateral plate mesoderm of the zebrafish and to track their subsequent lineage. We find that the heart precursor cells are clustered in a region adjacent to the prechordal plate, just anterior to the notochord tip. Complete unilateral ablation of all heart precursors with a laser does not disrupt heart development, if performed before the 18-somite stage. By combining extirpation of the heart precursors with cell labeling, we find that cells anterior to the normal cardiogenic compartments constitute the source of regulatory cells that compensate for the loss of the progenitors. One of the earliest embryonic markers of the premyocardial cells is the divergent homeodomain gene, Nkx2.5. Interestingly, normal cardiogenic progenitors derive from only the anterior half of the Nkx2.5-expressing region in the lateral plate mesoderm. The posterior half, adjacent to the notochord, does not include cardiac progenitors and the posterior Nkx2.5-expressing cells do not contribute to the heart, even after ablation of the normal cardiogenic region. The cells that can acquire a cardiac cell fate after injury to the normal progenitors also reside near the prechordal plate, but anterior to the Nkx2.5-expressing domain. Normally they give rise to head mesenchyme. They share with cardiac progenitors early expression of GATA 4. The location of the different elements of the cardiac field, and their response to injury, suggests that the prechordal plate supports and/or the notochord suppresses the cardiac fate.

Animals↗
PubMed1997

Defective "pacemaker" current (Ih) in a zebrafish mutant with a slow heart rate.

At a cellular level, cardiac pacemaking, which sets the rate and rhythm of the heartbeat, is produced by the slow membrane depolarization that occurs between action potentials. Several ionic currents could account for this pacemaker potential, but their relative prominence is controversial, and it is not known which ones actually play a pacemaking role in vivo. To correlate currents in individual heart cells with the rhythmic properties of the intact heart, we have examined slow mo (smo), a recessive mutation we discovered in the zebrafish Danio rerio. This mutation causes a reduced heart rate in the embryo, a property we can quantitate because the embryo is transparent. We developed methods for culture of cardiocytes from zebrafish embryos and found that, even in culture, cells from smo continue to beat relatively slowly. By patch-clamp analysis, we discovered that a large repertoire of cardiac currents noted in other species are present in these cultured cells, including sodium, T-type, and L-type calcium and several potassium currents, all of which appear normal in the mutant. The only abnormality appears to be in a hyperpolarization-activated inward current with the properties of Ih, a current described previously in the nervous system, pacemaker, and other cardiac tissue. smo cardiomyocytes have a reduction in Ih that appears to result from severe diminution of one kinetic component of the Ih current. This provides strong evidence that Ih is an important contributor to the pacemaking behavior of the intact heart.

Animals↗
PubMed1997

G alpha(o) is necessary for muscarinic regulation of Ca2+ channels in mouse heart.

Heterotrimeric G proteins, composed of G alpha and G betagamma subunits, transmit signals from cell surface receptors to cellular effector enzymes and ion channels. The G alpha(o) protein is the most abundant G alpha subtype in the nervous system, but it is also found in the heart. Its function is not completely known, although it is required for regulation of N-type Ca2+ channels in GH3 cells and also interacts with GAP43, a major protein in growth cones, suggesting a role in neuronal pathfinding. To analyze the function of G alpha(o), we have generated mice lacking both isoforms of G alpha(o) by homologous recombination. Surprisingly, the nervous system is grossly intact, despite the fact that G alpha(o) makes up 0.2-0.5% of brain particulate protein and 10% of the growth cone membrane. The G alpha(o)-/- mice do suffer tremors and occasional seizures, but there is no obvious histologic abnormality in the nervous system. In contrast, G alpha(o)-/- mice have a clear and specific defect in ion channel regulation in the heart. Normal muscarinic regulation of L-type calcium channels in ventricular myocytes is absent in the mutant mice. The L-type calcium channel responds normally to isoproterenol, but there is no evident muscarinic inhibition. Muscarinic regulation of atrial K+ channels is normal, as is the electrocardiogram. The levels of other G alpha subunits (G alpha(s), G alpha(q), and G alpha(i)) are unchanged in the hearts of G alpha(o)-/- mice, but the amount of G betagamma is decreased. Whichever subunit, G alpha(o) or G betagamma, carries the signal forward, these studies show that muscarinic inhibition of L-type Ca2+ channels requires coupling of the muscarinic receptor to G alpha(o). Other cardiac G alpha subunits cannot substitute.

Acetylcholine↗
PubMed1997

Expression of a dominant negative mutant of interleukin-1 beta converting enzyme in transgenic mice prevents neuronal cell death induced by trophic factor withdrawal and ischemic brain injury.

To explore the role of the interleukin (IL)-1 beta converting enzyme (ICE) in neuronal apoptosis, we designed a mutant ICE gene (C285G) that acts as a dominant negative ICE inhibitor. Microinjection of the mutant ICE gene into embryonal chicken dorsal root ganglial neurons inhibits trophic factor withdrawal-induced apoptosis. Transgenic mice expressing the fused mutant ICE-lacZ gene under the control of the neuron specific enolase promoter appeared neurologically normal. These mice are deficient in processing pro-IL-1 beta, indicating that mutant ICEC285G blocks ICE function. Dorsal root ganglial neurons isolated from transgenic mice were resistant to trophic factor withdrawal-induced apoptosis. In addition, the neurons isolated from newborn ICE knockout mice are similarly resistant to trophic factor withdrawal-induced apoptosis. After permanent focal ischemia by middle cerebral artery occlusion, the mutant ICEC285G transgenic mice show significantly reduced brain injury as well as less behavioral deficits when compared to the wild-type controls. Since ICE is the only enzyme with IL-1 beta convertase activity in mice, our data indicates that the mutant ICEC285G inhibits ICE, and hence mature IL-1 beta production, and through this mechanism, at least in part, inhibits apoptosis. Our data suggest that genetic manipulation using ICE family dominant negative inhibitors can ameliorate the extent of ischemia-induced brain injury and preserve neurological function.

Animals↗
PubMed1997

Vessel patterning in the embryo of the zebrafish: guidance by notochord.

We have cloned the zebrafish homolog of the receptor tyrosine kinase flk-1 to provide us with a tool to study normal vascular pattern formation in the developing zebrafish embryo and to compare it to mutants in which vascular pattern is perturbed. We find that during normal development the first angioblasts arise laterally in the mesoderm and then migrate medially to form the primordia of the large axial vessels, the dorsal aorta (axial artery) and the axial vein. Lumen formation occurs shortly before onset of circulation at 24 hr postfertilization. We examined the specification of vascular progenitors in the mutant cloche, which fails to form both vessels and blood. cloche lacks all flk-expressing cells and therefore appears to lack angioblasts. The axial vessels of the trunk form in close proximity to notochord and endoderm, which may provide cues for their formation. The dorsal aorta is normally just ventral to the notochord; the axial vein is just below the dorsal aorta and above the endoderm. floating head (flh) and no tail (ntl) mutants both have defects in the formation of notochord. Both are cell-autonomous lesions, flh abolishing notochord and ntl preventing its differentiation. In both mutants the dorsal aorta fails to form, while formation of the axial vein is less affected. Mosaic analysis of mutant embryos shows that transplanted wild-type cells can become notochord in mutant flh embryos. In these mosaic embryos flh cells expressing flk assemble at the midline, beneath the wild-type notochord, and form an aortic primordium. This suggests that signals from the notochord may guide angioblasts in the fashioning of the dorsal aorta. The notochord seems to be less important for the formation of the vein.

Amino Acid Sequence↗
PubMed1997

Neuronal nitric oxide synthase mutant mice show smaller infarcts and attenuated apparent diffusion coefficient changes in the peri-infarct zone during focal cerebral ischemia.

Diffusion-weighted MRI at 2 T was used to monitor and assess tissue damage after permanent middle cerebral artery occlusion (MCAO) in wild-type (WT) and mice deficient in nitric oxide synthase gene expression (nNOS-). The ischemic lesion was evaluated 3 h after occlusion and subdivided into the lesion core and peri-infarct zone based on the magnitude of the apparent diffusion coefficient (ADC) change. Infarct volume, measured by using histochemical staining 24 h after MCA occlusion, correlated best with MRI infarct volume as assessed by an ADC threshold of 25% decrease from baseline at 3 h. For ADC thresholds of greater than 25% decrease, lesion size was not significantly different in nNOS- and WT mice. However, brain tissue showing ADC decreases of 10-25% was significantly smaller in the ipsilateral hemisphere of mutants (27 +/- 2% and 21 +/- 2% in WT and nNOS-, respectively; P < 0.05). These findings occurred independently of infarct volume and are consistent with a smaller peri-infarct zone in nNOS- mice. We postulate that the smaller peri-infarct zone is a reflection of less severe metabolic disturbance after ischemia in nNOS- mice, possibly related to diminished production of nitric oxide (NO) or a related product. We conclude that magnetic resonance techniques previously used to assess ischemic damage in larger animals can be extended to the mouse, raising the possibility that the molecular mechanisms leading to ischemic damage can be examined by using genetically engineered mice.

Animals↗
PubMed1997

A novel transmembrane semaphorin can bind c-src.

The semaphorins/collapsins constitute a family of genes unified by the presence of a "semaphorin domain" which has been conserved through metazoan evolution. The semaphorin family comprises both secreted and transmembrane molecules and is thought to be made up of ligands for as yet unidentified receptors. The functions are not known, with the exception of those of sema III (also referred as sem D and collapsin 1), D-sema I, and D-sema II, which have been shown to be involved in axonal pathfinding. Here report the identification of a mouse semaphorin cDNA, termed Sema VIb. Although Sema VIb contains the extracellular semaphorin domain, it lacks the immunoglobulin domain or thrombospondin repeats which are present in other described vertebrate (but not invertebrate) transmembrane semaphorins. During development Sema VIb mRNA is expressed in subregions of the nervous system and is particularly prominent in muscle. In adulthood, Sema VIb mRNA is expressed ubiquitously. The cytoplasmic domain of Sema VIb contains several proline-rich potential SH3 domain binding sites. Using an in vitro binding assay, we show that Sema VIb binds specifically the SH3 domain of the protooncogene c-src. In transfected COS cells Sema VIb coimmunoprecipitates with c-src. These results, along with our evidence that Sema VIb can form dimers, suggests that the semaphorin family not only serves as ligands but may include members, especially those which are transmembrane, which serve as receptors, triggering intracellular signaling via an src-related cascade.

Amino Acid Sequence↗
PubMed1997

MPP+ induced substantia nigra degeneration is attenuated in nNOS knockout mice.

Recent studies showed that neuronal nitric oxide synthase (nNOS) plays a role in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine neurotoxicity. In the present study we examined the effects of striatal injection of 1-methyl-4-phenylpyridinium (MPP+) on substantia nigra degeneration in mutant mice lacking the nNOS gene or the endothelial nitric oxide synthase (eNOS) gene. Both striatal lesion volume and substantia nigra degeneration were significantly attenuated in the nNOS mutant mice but not in the eNOS mutant mice. The mice lacking nNOS showed a significant attenuation of MPP+(-) induced increases of 3-nitrotyrosine concentrations in the striatum. In a separate experiment administration of 7-nitroindazole for 48 h after MPP+ injections significantly attenuated substantia nigra degeneration in rats. Immunohistochemical studies showed apposition of nNOS-positive neuronal processes on tyrosine hydroxylase-positive neurons. These results provide further evidence that neuronally derived NO and peroxynitrite play a role in MPP+ neurotoxicity.

1-Methyl-4-phenylpyridinium↗
PubMed1997

Cyclooxygenase-2 mediates increased renal renin content induced by low-sodium diet.

We hypothesized that neuronal nitric oxide synthase and cyclooxygenase-2, which both exist in the renal cortex, predominantly in the macula densa, play a role in the control of renal renin tissue content. We studied the possible role of neuronal nitric oxide synthase in regulating renal renin content by using mice in which the neuronal nitric oxide synthase gene has been disrupted (nNOS-/-) compared with its two progenitor strains, the 129/SvEv and the C57BL/6, to determine if the absence of neuronal nitric oxide synthase would result in decreased renal renin content or blunt the increase observed during low sodium intake. Renal renin content from cortical slices was determined in adult mice from all three strains maintained on a normal sodium diet. Renal renin content was significantly reduced in the nNOS-/- mice compared with the 129/SvEv and the C57BL/6 mice (3.11 +/- 0.23 versus 5.66 +/- 0.50 and 7.55 +/- 1.17 micrograms angiotensin l/mg dry weight, respectively; P < .005), suggesting that neuronal nitric oxide synthase may stimulate renal renin content under basal conditions. Neither selective pharmacological inhibition of neuronal nitric oxide synthase using 7-nitroindazole or disruption of the neuronal nitric oxide synthase gene affected the increase in renal content observed during dietary sodium restriction. The influence of cyclooxygenase-2 on renal renin content through a macula densa-mediated pathway was studied using a selective cyclooxygenase-2 inhibitor, NS398, in 129/SvEv mice. A low-sodium diet increased renal renin content from 6.97 +/- 0.52 to 11.59 +/- 0.79 micrograms angiotensin l/mg dry weight (P < .005); but this increase was blocked by NS398. In addition, treatment with NS398 reduced renin mRNA in response to a low-sodium diet. Thus, increased renal renin content in response to dietary sodium restriction appears to require the induction of cyclooxygenase-2, while neuronal nitric oxide synthase appears to affect basal but not stimulated renal renin content.

Animals↗
PubMed1997

Pulmonary vasoconstriction and hypertension in mice with targeted disruption of the endothelial nitric oxide synthase (NOS 3) gene.

NO, synthesized in endothelial cells by endothelial NO synthase (NOS 3), is believed to be an important endogenous pulmonary vasodilator substance that contributes to the normal low pulmonary vascular resistance. To selectively investigate the role of NOS 3 in the pulmonary circulation, mice with targeted disruption of the NOS 3 gene were studied. Pulmonary hemodynamics were studied by measuring pulmonary artery pressure, left ventricular end-diastolic pressure, and lower thoracic aortic flow by using a novel open-chest technique. Transient partial occlusion of the inferior vena cava was used to assess the pulmonary artery pressure-flow relationship. Tension developed by isolated pulmonary artery segments after acetylcholine stimulation was measured in vitro. The histological appearance of NOS 3-deficient and wild-type murine lungs was compared. NOS 3-deficient mice (n = 27), when compared with wild-type mice (n = 32), had pulmonary hypertension (pulmonary artery pressure, 19.0 +/- 0.8 versus 16.4 +/- 0.6 mm Hg [mean +/- SE]; P < .05) that was due to an increased total pulmonary resistance (62 +/- 6 versus 33 +/- 2 mm Hg.min.g.mL-1; P < .001). In vitro, acetylcholine induced vasodilation in the main pulmonary arteries of wild-type but not NOS 3-deficient mice. The morphology of the lungs of NOS 3-deficient mice did not differ from that of wild-type mice. We conclude that NOS 3 is a key enzyme responsible for providing basal pulmonary NO release. Congenital NOS 3 deficiency produces mild pulmonary hypertension in mice.

Acetylcholine↗
PubMed1997

Fashioning the vertebrate heart: earliest embryonic decisions.

Our goal here is to set out the types of unitary decisions made by heart progenitor cells, from their appearance in the heart field until they form the simple heart tube. This provides a context to evaluate cell fate, lineage and, finally, morphogenetic decisions that configure global heart form and function. Some paradigms for cellular differentiation and for pattern generation may be borrowed from invertebrates, but neither Drosophila nor Caenorhabditis elegans suffice to unravel higher order decisions. Genetic analyses in mouse and zebrafish may provide one entrance to these pathways.

Animals↗