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  • 1. Archer, Amena
    et al.
    Lauter, Gilbert
    Södertörn University, School of Life Sciences.
    Hauptmann, Giselbert
    Södertörn University, School of Life Sciences.
    Mode, Agneta
    Gustafsson, Jan-Ake
    Transcriptional activity and developmental expression of liver X receptor (lxr) in zebrafish2008In: Developmental Dynamics, ISSN 1058-8388, E-ISSN 1097-0177, Vol. 237, no 4, p. 1090-1098Article in journal (Refereed)
    Abstract [en]

    Mammalian liver-X-receptors (LXRs) are transcription factors activated by oxysterols. They play an essential role in lipid and glucose metabolism. We have cloned the open reading frame of zebrafish lxr and describe its genomic organization. Zebrafish lxr encodes a 50-kDa protein with high sequence similarity to mammalian LXR alpha. In transfection assays, the encoded protein showed transcriptional activity in response to LXR-ligands. Treatment of adult zebrafish with the synthetic LXR ligand, GW3965, induced expression of genes involved in hepatic cholesterol and lipid pathways. Using qPCR and in situ hybridization, we found ubiquitous expression of lxr mRNA during the first 24 hr of development, followed by more restricted expression, particularly to the liver at 3dpf and the liver and intestine at 4dpf. In adult fish, all examined organs expressed lxr. In addition to a metabolic role of lxr, the temporal expression pattern suggests a developmental role in, e.g., the liver and CNS.

  • 2.
    Bräutigam, Lars
    et al.
    Södertörn University, School of Life Sciences, Molecular biology.
    Hillmer, Janine M.
    Södertörn University, School of Life Sciences, Molecular biology.
    Söll, Iris
    Södertörn University, School of Life Sciences, Molecular biology.
    Hauptmann, Giselbert
    Södertörn University, School of Life Sciences, Molecular biology.
    Localized Expression of Urocortin Genes in the Developing Zebrafish rain2010In: Journal of Comparative Neurology, ISSN 0021-9967, E-ISSN 1096-9861, Vol. 518, no 15, p. 2978-2995Article in journal (Refereed)
    Abstract [en]

    The corticotropin-releasing hormone (CRH) family consists of four aralogous genes, CRH and urocortins (UCNs) 1, 2, and 3. In a previous tudy, we analyzed CRH in the teleost model organism zebrafish and its ranscript distribution in the embryonic brain. Here, we describe ull-length cDNAs encoding urotensin 1 (UTS1), the teleost UCN1 rtholog, and UCN3 of zebrafish. Major expression sites of uts1 in adult ebrafish are the caudal neurosecretory system and brain. By using T-PCR analysis, we show that uts1 mRNA is also present in ovary, aternally contributed to the embryo, and expressed throughout embryonic evelopment. Expression of ucn3 mRNA was detected in a range of adult issues and during developmental stages from 24 hours post fertilization nward. Analysis of spatial transcript distributions by whole-mount in itu hybridization revealed limited forebrain expression of uts1 and cn3 during early development. Small numbers of uts1-synthesizing eurons were found in subpallium, hypothalamus, and posterior iencephalon, whereas ucn3-positive cells were restricted to elencephalon and retina. The brainstem was the main site of uts1 and cn3 synthesis in the embryonic brain. uts1 Expression was confined to he midbrain tegmentum; distinct hindbrain cell groups, including locus oeruleus and Mauthner neurons; and the spinal cord. ucn3 Expression was ocalized to the optic tectum, serotonergic raphe, and distinct hombomeric cell clusters. The prominent expression of uts1 and ucn3 in rainstem is consistent with proposed roles of CRH-related peptides in tress-induced modulation of locomotor activity through monoaminergic rainstem neuromodulatory systems. J. Comp. Neurol. 518:2978-2995, 2010.

  • 3. Cao, Renhai
    et al.
    Jensen, Lasse Dahl Ejby
    Söll, Iris
    Södertörn University, School of Life Sciences.
    Hauptmann, Giselbert
    Södertörn University, School of Life Sciences.
    Cao, Yihai
    Hypoxia-Induced Retinal Angiogenesis in Zebrafish as a Model to Study Retinopathy2008In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 3, no 7, p. e2748-Article in journal (Refereed)
    Abstract [en]

    Mechanistic understanding and defining novel therapeutic targets of diabetic retinopathy and age-related macular degeneration (AMD) have been hampered by a lack of appropriate adult animal models. Here we describe a simple and highly reproducible adult fli-EGFP transgenic zebrafish model to study retinal angiogenesis. The retinal vasculature in the adult zebrafish is highly organized and hypoxia-induced neovascularization occurs in a predictable area of capillary plexuses. New retinal vessels and vascular sprouts can be accurately measured and quantified. Orally active anti-VEGF agents including sunitinib and ZM323881 effectively block hypoxia-induced retinal neovascularization. Intriguingly, blockage of the Notch signaling pathway by the inhibitor DAPT under hypoxia, results in a high density of arterial sprouting in all optical arteries. The Notch suppression-induced arterial sprouting is dependent on tissue hypoxia. However, in the presence of DAPT substantial endothelial tip cell formation was detected only in optic capillary plexuses under normoxia. These findings suggest that hypoxia shifts the vascular targets of Notch inhibitors. Our findings for the first time show a clinically relevant retinal angiogenesis model in adult zebrafish, which might serve as a platform for studying mechanisms of retinal angiogenesis, for defining novel therapeutic targets, and for screening of novel antiangiogenic drugs.

  • 4.
    Chandrasekar, Gayathri
    et al.
    Södertörn University, School of Life Sciences.
    Lauter, Gilbert
    Södertörn University, School of Life Sciences.
    Hauptmann, Giselbert
    Södertörn University, School of Life Sciences.
    Distribution of corticotropin-releasing hormone in the developing zebrafish brain2007In: Journal of Comparative Neurology, ISSN 0021-9967, E-ISSN 1096-9861, Vol. 505, no 4, p. 337-351Article in journal (Refereed)
    Abstract [en]

    Corticotropin-releasing hormone (CRH) plays a central role in the physiological regulation of the hypothalamus-pituitary-adrenal/interrenal axis mediating endocrine, behavioral, autonomic, and immune responses to stress. Despite the wealth of knowledge about the physiological roles of CRH, the genetic mechanisms by which CRH neurons arise during development are poorly understood. As a first step toward analyzing the molecular and genetic pathways involved in CRH lineage specification, we describe the developmental distribution of CRH neurons in the embryonic zebrafish, a model organism for functional genomics and developmental biology. We searched available zebrafish expressed sequence tag (EST) databases for CRH-like sequences and identified one EST that contained the complete zebrafish CRH open reading frame (ORF). The CRH precursor sequence contained a signal peptide, the CRH peptide, and a cryptic peptide with a conserved sequence motif. RT-PCR analysis showed crh expression in a wide range of adult tissues as well as during embryonic and larval stages. By whole-mount in situ hybridization histochemistry, discrete crh-expressing cell clusters were found in different parts of the embryonic zebrafish brain, including telencephalon, preoptic region, hypothalamus, posterior tuberculum, thalamus, epiphysis, midbrain tegmentum, and rostral hindbrain and in the neural retina. The localization of crh mRNA within the preoptic region is consistent with the central role of CRH in the teleost stress response through activation of the hypothalamic-pituitary-interrenal axis. The widespread distribution of CRH-synthesizing cells outside the preoptic region suggests additional functions of CRH in the embryonic zebrafish brain.

  • 5. Gabriel, J. P.
    et al.
    Mahmood, R.
    Walter, A. M.
    Kyriakatos, A.
    Hauptmann, Giselbert
    Södertörn University, School of Life Sciences. Karolinska Institute.
    Calabrese, R. L.
    El Manira, A.
    Locomotor pattern in the adult zebrafish spinal cord in vitro2008In: Journal of Neurophysiology, ISSN 0022-3077, E-ISSN 1522-1598, Vol. 99, no 1, p. 37-48Article in journal (Refereed)
    Abstract [en]

    The zebrafish is an attractive model system for studying the function of the spinal locomotor network by combining electrophysiological, imaging, and genetic approaches. Thus far, most studies have been focusing on embryonic and larval stages. In this study we have developed an in vitro preparation of the isolated spinal cord from adult zebrafish in which locomotor activity can be induced while the activity of single neurons can be monitored using whole cell recording techniques. Application of NMDA elicited rhythmic locomotor activity that was monitored by recording from muscles or ventral roots in semi-intact or isolated spinal cord preparations, respectively. This rhythmic activity displayed a left-right alternation and a rostrocaudal delay. Blockade of glycinergic synaptic transmission by strychnine switched the alternating activity into synchronous bursting in the left and right sides as well as along the rostrocaudal axis. Whole cell recordings from motoneurons showed that they receive phasic synaptic inputs that were correlated with the locomotor activity recorded in ventral roots. This newly developed in vitro preparation of the adult zebrafish spinal cord will allow examination of the organization of the spinal locomotor network in an adult system to complement studies in zebrafish larvae and new born rodents.

  • 6. Gabriel, Jens Peter
    et al.
    Mahmood, Riyadh
    Kyriakatos, Alexandros
    Söll, Iris
    Södertörn University, School of Life Sciences, Molecular biology.
    Hauptmann, Giselbert
    Södertörn University, School of Life Sciences, Molecular biology.
    Calabrese, Ronald L.
    El Manira, Abdeljabbar
    Serotonergic Modulation of Locomotion in Zebrafish-Endogenous Release and Synaptic Mechanisms2009In: Journal of Neuroscience, ISSN 0270-6474, E-ISSN 1529-2401, Vol. 29, no 33, p. 10387-10395Article in journal (Refereed)
    Abstract [en]

    Serotonin (5-HT) plays an important role in shaping the activity of the spinal networks underlying locomotion in many vertebrate preparations. At larval stages in zebrafish, 5-HT does not change the frequency of spontaneous swimming; and it only decreases the quiescent period between consecutive swimming episodes. However, it is not known whether 5-HT exerts similar actions on the locomotor network at later developmental stages. For this, the effect of 5-HT on the fictive locomotor pattern of juvenile and adult zebrafish was analyzed. Bath-application of 5-HT (1-20 mu M) reduced the frequency of the NMDA-induced locomotor rhythm. Blocking removal from the synaptic cleft with the reuptake inhibitor citalopram had similar effects, suggesting that endogenous serotonin is modulating the locomotor pattern. One target for this modulation was the mid-cycle inhibition during locomotion because the IPSPs recorded in spinal neurons during the hyperpolarized phase were increased both in amplitude and occurrence by 5-HT. Similar results were obtained for IPSCs recorded in spinal neurons clamped at the reversal potential of excitatory currents (0 mV). 5-HT also slows down the rising phase of the excitatory drive recorded in spinal cord neurons when glycinergic inhibition is blocked. These results suggest that the decrease in the locomotor burst frequency induced by 5-HT is mediated by a potentiation of mid-cycle inhibition combined with a delayed onset of the subsequent depolarization.

  • 7. Jensen, Lasse Dahl Ejby
    et al.
    Cao, Renhai
    Hedlund, Eva-Maria
    Söll, Iris
    Södertörn University, School of Life Sciences, Molecular biology.
    Lundberg, Jon O.
    Hauptmann, Giselbert
    Södertörn University, School of Life Sciences, Molecular biology.
    Steffensen, John Fleng
    Cao, Yihai
    Nitric oxide permits hypoxia-induced lymphatic perfusion by controlling arterial-lymphatic conduits in zebrafish and glass catfish2009In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 106, no 43, p. 18408-18413Article in journal (Refereed)
    Abstract [en]

    The blood and lymphatic vasculatures are structurally and functionally coupled in controlling tissue perfusion, extracellular interstitial fluids, and immune surveillance. Little is known, however, about the molecular mechanisms that underlie the regulation of bloodlymphatic vessel connections and lymphatic perfusion. Here we show in the adult zebrafish and glass catfish (Kryptopterus bicirrhis) that blood-lymphatic conduits directly connect arterial vessels to the lymphatic system. Under hypoxic conditions, arterial-lymphatic conduits (ALCs) became highly dilated and linearized by NO-induced vascular relaxation, which led to blood perfusion into the lymphatic system. NO blockage almost completely abrogated hypoxia-induced ALC relaxation and lymphatic perfusion. These findings uncover mechanisms underlying hypoxia-induced oxygen compensation by perfusion of existing lymphatics in fish. Our results might also imply that the hypoxia-induced NO pathway contributes to development of progression of pathologies, including promotion of lymphatic metastasis by modulating arterial-lymphatic conduits, in the mammalian system.

  • 8.
    Kitambi, Satish Srinivas
    et al.
    Södertörn University, School of Life Sciences. Karolinska Institutet.
    Hauptmann, Giselbert
    Södertörn University, School of Life Sciences. Karolinska Institutet.
    The zebrafish orphan nuclear receptor genes nr2e1 and nr2e3 are expressed in developing eye and forebrain2007In: Gene Expression Patterns, ISSN 1567-133X, E-ISSN 1872-7298, Vol. 7, no 4, p. 521-528Article in journal (Refereed)
    Abstract [en]

    Mammalian Nr2e1 (Tailless, Mtll or Tlx) and Nr2e3 (photoreceptor-specific nuclear receptor, Pnr) are highly related orphan nuclear receptors, that are expressed in eye and forebrain-derived structures. In this study, we analyzed the developmental expression patterns of zebrafish nr2e1 and nr2e3. RT-PCR analysis showed that nr2e1 and nr2e3 are both expressed during embryonic and post-embryonic development. To examine the spatial distribution of nr2e1 and nr2e3 during development whole-mount in situ hybridization was performed. At tailbud stage, initial nr2e1 expression was localized to the rostral brain rudiment anterior to pax2.1 and eng2 expression at the prospective midbrain-hindbrain boundary. During Subsequent stages, nr2e1 became widely expressed in fore- and midbrain primordia, eye and olfactory placodes. At 24 hpf, strong nr2e1 expression was detected in telencephalon, hypothalamus, dorsal thalamus, pretectum, midbrain tectum, and retina. At 2 dpf, the initially widespread nr2e1 expression became more restricted to distinct regions within the fore- and midbrain and to the retinal ciliary margin, the germinal zone which gives rise to retina and presumptive iris. Express on of nr2e3 was exclusively found in the developing retina and epiphysis. In both structures, nr2e3 expression was found in photoreceptor cells. The developmental expression profile of zebrafish nr2e1 and nr2e3 is consistent with evolutionary conserved functions in eye and rostral brain structures.

  • 9.
    Lauter, Gilbert
    et al.
    Södertörn University, School of Life Sciences. Karolinska Institutet.
    Söll, Iris
    Södertörn University, School of Life Sciences. Karolinska Institutet.
    Hauptmann, Giselbert
    Södertörn University, School of Life Sciences. Karolinska Institutet.
    PACAP in zebrafish neurodevelopment2009In: Mechanisms of Development, ISSN 0925-4773, E-ISSN 1872-6356, Vol. 126, p. S235-S235Article in journal (Refereed)
  • 10. Lee, Samantha Lin Chiou
    et al.
    Rouhi, Pegah
    Jensen, Lasse Dahl
    Zhang, Danfang
    Ji, Hong
    Hauptmann, Giselbert
    Södertörn University, School of Life Sciences, Molecular biology.
    Ingham, Philip
    Cao, Yihai
    Hypoxia-induced pathological angiogenesis mediates tumor cell dissemination, invasion, and metastasis in a zebrafish tumor model2009In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 106, no 46, p. 19485-19490Article in journal (Refereed)
    Abstract [en]

    Mechanisms underlying pathological angiogenesis in relation to hypoxia in tumor invasion and metastasis remain elusive. Here, we have developed a zebrafish tumor model that allows us to study the role of pathological angiogenesis under normoxia and hypoxia in arbitrating early events of the metastatic cascade at the single cell level. Under normoxia, implantation of a murine T241 fibrosarcoma into the perivitelline cavity of developing embryos of transgenic fli1:EGFP zebrafish did not result in significant dissemination, invasion, and metastasis. In marked contrast, under hypoxia substantial tumor cells disseminated from primary sites, invaded into neighboring tissues, and metastasized to distal parts of the fish body. Similarly, expression of the hypoxia-regulated angiogenic factor, vascular endothelial growth factor (VEGF) to a high level resulted in tumor cell dissemination and metastasis, which correlated with increased tumor neovascularization. Inhibition of VEGF receptor signaling pathways by sunitinib or VEGFR2 morpholinos virtually completely ablated VEGF-induced tumor cell dissemination and metastasis. To the best of our knowledge, hypoxia- and VEGF-induced pathological angiogenesis in promoting tumor dissemination, invasion, and metastasis has not been described perviously at the single cell level. Our findings also shed light on molecular mechanisms of beneficial effects of clinically available anti-VEGF drugs for cancer therapy.

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