Biomedical subjects

Patrik Ernfors

Publications and source records attributed to Patrik Ernfors.

At least 19 recordsLinked to original sources

PubMed2006

Engineering the recruitment of phosphotyrosine binding domain-containing adaptor proteins reveals distinct roles for RET receptor-mediated cell survival.

The RET receptor tyrosine kinase is important for several different biological functions during development. The recruitment at the phosphorylated Tyr(1062) site in RET of a number of different phosphotyrosine binding (PTB) domain-containing adaptor proteins, including Shc and Frs2, plays a dominant role for the multiple different biological functions of the RET receptor during development, including stimulation of cell survival. Here, we demonstrate that a competitive recruitment of Shc as opposed to Frs2 mediates the survival signaling arising from RET activation. Based on results from a peptide array, we have genetically engineered the PTB domain binding site of RET to rewire its recruitment of the PTB proteins Shc and Frs2. An engineered RET that has a competitive interaction with Shc at the expense of Frs2, but not a RET receptor that only recruits Frs2, activates cell survival signaling pathways and is protective from cell death in neuronal SK-N-MC cells. Thus, cell type-specific functions involve a competitive recruitment of different PTB adaptor molecules by RET that activate selective signaling pathways.

Adaptor Proteins, Signal Transducing↗

PubMed2006

In vitro and in vivo differentiation of boundary cap neural crest stem cells into mature Schwann cells.

Boundary cap cells can generate neurons as well as peripheral glia during embryonic development (Maro, G.S., Vermeren, M., Voiculescu, O., Melton, L., Cohen, J., Charnay, P., Topilko, P., 2004. Neural crest boundary cap cells constitute a source of neuronal and glial cells of the PNS. Nat Neurosci. 7 (9), 930-938), and, recently, the boundary cap was shown to contain multipotent stem cells (Hjerling-Leffler, J., Marmigère, F., Heglind, M., Cederberg, A., Koltzenburg, M., Enerbäck, S., Ernfors, P., 2005. The boundary cap, a source of neural crest stem cells generating multiple sensory neuron subtypes. Development. 132 (11), 2623-2632). The ability of stem cells to generate mature functional glial phenotypes has not been addressed. In this study, we have explored the competence of boundary neural crest stem cells (bNCSCs) to differentiate into mature functional Schwann cells (SCs) in vitro and in vivo. bNCSCs failed to differentiate into SCs in vitro when cultured in a defined media and in vivo when grafted into adult rat sciatic nerves. However, in the presence of neuregulins, during long-term cultures, the majority of bNCSCs differentiated into SCs. After analysis of the in vivo expression of Sox2, Sox10, S100, GFAP, fibronectin and Krox20 in the glial lineages, we used these markers to characterize differentiation of the bNCSCs. Gliogenesis of bNCSCs proceeded similar to that in vivo by sequentially adopting a SC precursor and immature Schwann cell before maturing into myelinating and non-myelinating SCs. In co-culture with explanted dorsal root ganglia (DRG) as well as in vivo in transplants to the axotomized sciatic nerve, these bNCSC-derived SCs myelinated axons as shown by ensheathing of neuronal processes and expression of myelin basic proteins (MBP). These results show that, under appropriate conditions, bNCSCs can generate mature SCs that are functional and can myelinate axons in regenerating nerves.

Animals↗

PubMed2006

The Runx1/AML1 transcription factor selectively regulates development and survival of TrkA nociceptive sensory neurons.

Neural crest cells (NCCs) can adopt different neuronal fates. In NCCs, neurogenin-2 promotes sensory specification but does not specify different subclasses of sensory neurons. Understanding the gene cascades that direct Trk gene activation may reveal mechanisms generating sensory diversity, because different Trks are expressed in different sensory neuron subpopulations. Here we show in chick and mouse that the Runt transcription factor Runx1 promotes axonal growth, is selectively expressed in neural crest-derived TrkA(+) sensory neurons and mediates TrkA transactivation in migratory NCCs. Inhibition of Runt activity depletes TrkA expression and leads to neuronal death. Moreover, Runx1 overexpression is incompatible with multipotency in the migratory neural crest but does not induce expression of pan-neuronal genes. Instead, Runx1-induced neuronal differentiation depends on an existing neurogenin2 proneural gene program. Our data show that Runx1 directs, in a context-dependent manner, key aspects of the establishment of the TrkA(+) nociceptive subclass of neurons.

Animals↗

PubMed2006

Brain-derived neurotrophic factor selectively regulates dendritogenesis of parvalbumin-containing interneurons in the main olfactory bulb through the PLCgamma pathway.

Molecular mechanisms of neurotrophin signaling on dendrite development and dynamics are only partly understood. To address the role of brain-derived neurotrophic factor (BDNF) in the morphogenesis of GABAergic neurons of the main olfactory bulb, we analyzed mice lacking BDNF, mice carrying neurotrophin-3 (NT3) in the place of BDNF, and TrkB signaling mutant mice with a receptor that can activate phospholipase Cgamma (PLCgamma) but is unable to recruit the adaptors Shc/Frs2. BDNF deletion yielded a compressed olfactory bulb with a significant loss of parvalbumin (PV) immunoreactivity in GABAergic interneurons of the external plexiform layer. Dendrite development of PV-positive interneurons was selectively attenuated by BDNF since other Ca2+ -binding protein-containing neuron populations appeared unaffected. The deficit in PV-positive neurons could be rescued by the NT3/NT3 alleles. The degree of PV immunoreactivity was dependent on BDNF and TrkB recruitment of the adaptor proteins Shc/Frs2. In contrast, PLCgamma signaling from the TrkB receptor was sufficient for dendrite growth in vivo and consistently, blocking PLCgamma prevented BDNF-dependent dendrite development in vitro. Collectively, our results provide genetic evidence that BDNF and TrkB signaling selectively regulate PV expression and dendrite growth in a subset of neurochemically-defined GABAergic interneurons via activation of the PLCgamma pathway.

Animals↗

PubMed2006

Vesicular glutamate transporter 3 (VGLUT3) identifies spatially segregated excitatory terminals in the rat substantia nigra.

The excitability of dopaminergic (DA) neurons in the substantia nigra is controlled by the convergent activity of multiple glutamatergic afferents. Here, we show that vesicular glutamate transporter 3 (VGLUT3)-immunoreactive (ir) terminals segregate to the perisomatic region of DA neurons in the substantia nigra pars compacta, and VGLUT3 decorates a synapse population distinct from those marked by vesicular glutamate transporters 1 and 2. VGLUT3-ir nerve endings form asymmetric terminals on DA neurons. Retrograde tracing suggests the superior colliculus as an origin of excitatory VGLUT3-ir afferents. Collectively, our data indicate that VGLUT3 identifies a novel excitatory terminal subset that contributes to the tuning of DA cell excitability in the substantia nigra.

Animals↗

PubMed2005

Endocannabinoids regulate interneuron migration and morphogenesis by transactivating the TrkB receptor.

In utero exposure to Delta(9)-tetrahydrocannabinol (Delta(9)-THC), the active component from marijuana, induces cognitive deficits enduring into adulthood. Although changes in synaptic structure and plasticity may underlie Delta(9)-THC-induced cognitive impairments, the neuronal basis of Delta(9)-THC-related developmental deficits remains unknown. Using a Boyden chamber assay, we show that agonist stimulation of the CB(1) cannabinoid receptor (CB(1)R) on cholecystokinin-expressing interneurons induces chemotaxis that is additive with brain-derived neurotrophic factor (BDNF)-induced interneuron migration. We find that Src kinase-dependent TrkB receptor transactivation mediates endocannabinoid (eCB)-induced chemotaxis in the absence of BDNF. Simultaneously, eCBs suppress the BDNF-dependent morphogenesis of interneurons, and this suppression is abolished by Src kinase inhibition in vitro. Because sustained prenatal Delta(9)-THC stimulation of CB(1)Rs selectively increases the density of cholecystokinin-expressing interneurons in the hippocampus in vivo, we conclude that prenatal CB(1)R activity governs proper interneuron placement and integration during corticogenesis. Moreover, eCBs use TrkB receptor-dependent signaling pathways to regulate subtype-selective interneuron migration and specification.

Animals↗

PubMed2005

Lingual deficits in neurotrophin double knockout mice.

Brain-derived neurotrophic factor (BDNF) and Neurotrophin 3 (NT-3) are members of the neurotrophin family and are expressed in the developing and adult tongue papillae. BDNF null-mutated mice exhibit specific impairments related to innervation and development of the gustatory system while NT-3 null mice have deficits in their lingual somatosensory innervation. To further evaluate the functional specificity of these neurotrophins in the peripheral gustatory system, we generated double BDNF/NT-3 knockout mice and compared the phenotype to BDNF(-/-) and wild-type mice. Taste papillae morphology was severely distorted in BDNF(-/-) xNT-3(-/-) mice compared to single BDNF(-/-) and wild-type mice. The deficits were found throughout the tongue and all gustatory papillae. There was a significant loss of fungiform papillae and the papillae were smaller in size compared to BDNF(-/-) and wild-type mice. Circumvallate papillae in the double knockouts were smaller and did not contain any intraepithelial nerve fibers. BDNF(-/-) xNT-3(-/-) mice exhibited additive losses in both somatosensory and gustatory innervation indicating that BDNF and NT-3 exert specific roles in the innervation of the tongue. However, the additional loss of fungiform papillae and taste buds in BDNF(-/-) xNT-3(-/-) mice compared to single BDNF knockout mice indicate a synergistic functional role for both BDNF-dependent gustatory and NT-3-dependent somatosensory innervations in taste bud and taste papillae innervation and development.

Animals↗

PubMed2005

Increased progenitor proliferation and apoptotic cell death in the sensory lineage of mice overexpressing N-myc.

N-myc, a member of the myc family of bHLH transcription factors, is expressed mainly in the nervous system, including derivatives of neural crest cells in the periphery during development, such as the sensory dorsal root ganglion (DRG). Previous studies suggest that N-myc is involved in the proliferation of progenitor cells in the sensory lineage. To address the role of N-myc in the development of peripheral sensory neurons, we have overexpressed N-myc in sensory progenitor cells. The overexpression of N-myc did not significantly affect the number of multipotent neural crest cells or glial differentiation but caused a brief and marked increase of both proliferation and apoptosis in the DRG at embryonic day 11 (E11), thus coinciding with the stage of cell-cycle exit. At E17, the total number of cells in the lumbar DRG of mice with forced expression of N-myc was significantly reduced compared with that in wild-type mice. Among the different DRG subpopulations examined, the number of parvalbumin-positive neurons representing large-diameter proprioceptive neurons increased significantly. Our results indicate that forced expression of N-myc in the sensory lineage leads to unscheduled cell-cycle re-entry and excessive apoptosis and show that N-myc can affect the composition of different functional subtypes of sensory neurons in the DRG.

Animals↗

PubMed2005

Differential suppression of seizures via Y2 and Y5 neuropeptide Y receptors.

Neuropeptide Y (NPY) prominently inhibits epileptic seizures in different animal models. The NPY receptors mediating this effect remain controversial partially due to lack of highly selective agonists and antagonists. To circumvent this problem, we used various NPY receptor knockout mice with the same genetic background and explored anti-epileptic action of NPY in vitro and in vivo. In Y2 (Y2-/-) and Y5 (Y5-/-) receptor knockouts, NPY partially inhibited 0 Mg2+-induced epileptiform activity in hippocampal slices. In contrast, in double knockouts (Y2Y5-/-), NPY had no effect, suggesting that in the hippocampus in vitro both receptors mediate anti-epileptiform action of NPY in an additive manner. Systemic kainate induced more severe seizures in Y5-/- and Y2Y5-/-, but not in Y2-/- mice, as compared to wild-type mice. Moreover, kainate seizures were aggravated by administration of the Y5 antagonist L-152,804 in wild-type mice. In Y5-/- mice, hippocampal kindling progressed faster, and afterdischarge durations were longer in amygdala, but not in hippocampus, as compared to wild-type controls. Taken together, these data suggest that, in mice, both Y2 and Y5 receptors regulate hippocampal seizures in vitro, while activation of Y5 receptors in extra-hippocampal regions reduces generalized seizures in vivo.

Animals↗

PubMed2005

The boundary cap: a source of neural crest stem cells that generate multiple sensory neuron subtypes.

The boundary cap (BC) is a transient neural crest-derived group of cells located at the dorsal root entry zone (DREZ) that have been shown to differentiate into sensory neurons and glia in vivo. We find that when placed in culture, BC cells self-renew, show multipotency in clonal cultures and express neural crest stem cell (NCSCs) markers. Unlike sciatic nerve NCSCs, the BC-NCSC (bNCSCs) generates sensory neurons upon differentiation. The bNCSCs constitute a common source of cells for functionally diverse types of neurons, as a single bNCSC can give rise to several types of nociceptive and thermoreceptive sensory neurons. Our data suggests that BC cells comprise a source of multipotent sensory specified stem cells that persist throughout embryogenesis.

Animals↗

PubMed2005

Brain-derived neurotrophic factor and antidepressant drugs have different but coordinated effects on neuronal turnover, proliferation, and survival in the adult dentate gyrus.

Antidepressants increase proliferation of neuronal progenitor cells and expression of brain-derived neurotrophic factor (BDNF) in the hippocampus. We investigated the role of BDNF signaling in antidepressant-induced neurogenesis by using transgenic mice with either reduced BDNF levels (BDNF+/-) or impaired trkB activation (trkB.T1-overexpressing mice). In both transgenic strains, chronic (21 d) imipramine treatment increased the number of bromodeoxyuridine (BrdU)-positive cells to degree similar to that seen in wild-type mice 24 h after BrdU administration, although the basal proliferation rate was increased in both transgenic strains. Three weeks after BrdU administration and the last antidepressant injection, the amount of newborn (BrdU- or TUC-4-positive) cells was significantly reduced in both BDNF+/- and trkB.T1-overexpressing mice, which suggests that normal BDNF signaling is required for the long-term survival of newborn hippocampal neurons. Moreover, the antidepressant-induced increase in the surviving BrdU-positive neurons seen in wild-type mice 3 weeks after treatment was essentially lost in mice with reduced BDNF signaling. Furthermore, we observed that chronic treatment with imipramine or fluoxetine produced a temporally similar increase in both BrdU-positive and terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end-labeled neurons in the dentate gyrus, indicating that these drugs simultaneously increase both neurogenesis and neuronal elimination. These data suggest that antidepressants increase turnover of hippocampal neurons rather than neurogenesis per se and that BDNF signaling is required for the long-term survival of newborn neurons in mouse hippocampus.

Animals↗

PubMed2005

Genetic evidence for selective neurotrophin 3 signalling through TrkC but not TrkB in vivo.

Neurotrophins control neuronal survival in a target-derived manner during the period of naturally occurring cell death in development. The specificity of this mechanism has been attributed to a restricted spatio-temporal expression of neurotrophin ligands in target tissues, as well as a selective expression of their cognate tyrosine kinase (Trk) receptors in different neuronal subpopulations. However, several in vitro and in vivo studies of null mutant mice have suggested that neurotrophin 3 (NT 3) also signals through the non-preferred TrkB receptor. In this study, we have directly addressed the in vivo preference of NT 3 to signal through TrkB or TrkC, by crossing the NT 3 knock-in mice (BDNF(NT 3/NT 3) mice) with the TrkB- or TrkC-null mutant mice. We find that TrkB is dispensable, whereas TrkC is required for the neuronal rescue by the NT 3 allele in the brain-derived neurotrophic factor- and NT 3-dependent cochleovestibular system. Our results show that NT 3 maintains survival of cells as well as target innervation only through interactions with TrkC in vivo. TrkB and TrkC receptors are thus not functionally redundant for NT 3, even when coexpressed in neurons of the cochleovestibular system.

Adaptor Proteins, Signal Transducing↗

PubMed2005

Brain-derived neurotrophic factor modulates dopaminergic deficits in a transgenic mouse model of Huntington's disease.

Dysfunction of dopaminergic neurons may contribute to motor impairment in Huntington's disease. Here, we study the role of brain-derived neurotrophic factor (BDNF) in alterations of the nigrostriatal system associated with transgenics carrying mutant huntingtin. Using huntingtin-BDNF+/- double-mutant mice, we analyzed the effects of reducing the levels of BDNF expression in a model of Huntington's disease (R6/1). When compared with R6/1 mice, these mice exhibit an increased number of aggregates in the substantia nigra pars compacta. In addition, reduction of BDNF expression exacerbates the dopaminergic neuronal dysfunction seen in mutant huntingtin mice, such as the decrease in retrograde labelling of dopaminergic neurons and striatal dopamine content. However, mutant huntingtin mice with normal or lowered BDNF expression show the same decrease in the anterograde transport, number of dopaminergic neurons and nigral volume. In addition, reduced BDNF expression causes decreased dopamine receptor expression in mutant huntingtin mice. Examination of changes in locomotor activity induced by dopamine receptor agonists revealed that, in comparison with R6/1 mice, the double mutant mice exhibit lower activity in response to amphetamine, but not to apomorphine. In conclusion, these findings demonstrate that the decreased BDNF expression observed in Huntington's disease exacerbates dopaminergic neuronal dysfunction, which may participate in the motor disturbances associated with this neurodegenerative disorder.

Amphetamine↗

PubMed2005

Lack of the central nervous system- and neural crest-expressed forkhead gene Foxs1 affects motor function and body weight.

To gain insight into the expression pattern and functional importance of the forkhead transcription factor Foxs1, we constructed a Foxs1-beta-galactosidase reporter gene "knock-in" (Foxs1beta-gal/beta-gal) mouse, in which the wild-type (wt) Foxs1 allele has been inactivated and replaced by a beta-galactosidase reporter gene. Staining for beta-galactosidase activity reveals an expression pattern encompassing neural crest-derived cells, e.g., cranial and dorsal root ganglia as well as several other cell populations in the central nervous system (CNS), most prominently the internal granule layer of cerebellum. Other sites of expression include the lachrymal gland, outer nuclear layer of retina, enteric ganglion neurons, and a subset of thalamic and hypothalamic nuclei. In the CNS, blood vessel-associated smooth muscle cells and pericytes stain positive for Foxs1. Foxs1beta-gal/beta-gal mice perform significantly better (P < 0.01) on a rotating rod than do wt littermates. We have also noted a lower body weight gain (P < 0.05) in Foxs1beta-gal/lbeta-gal males on a high-fat diet, and we speculate that dorsomedial hypothalamic neurons, expressing Foxs1, could play a role in regulating body weight via regulation of sympathetic outflow. In support of this, we observed increased levels of uncoupling protein 1 mRNA in Foxs1beta-gal/beta-gal mice. This points toward a role for Foxs1 in the integration and processing of neuronal signals of importance for energy turnover and motor function.

Animals↗

PubMed2004

Attenuation of acute experimental colitis by preventing NPY Y1 receptor signaling.

Neuropeptide Y (NPY), a 36-amino acid peptide, is widely expressed in the central and peripheral nervous system. NPY is involved in the regulation of several physiological processes, including energy balance, food intake, and nociception. Recently, we showed that activation of the NPY Y1 receptor is required for cutaneous neurogenic inflammation. Because neurogenic inflammation could participate in colitis, the aim of this study was to investigate the role of the NPY Y1 receptor in acute colitis using mice genetically deficient of NPY Y1 receptor. In addition, the Y1 receptor antagonist H409/22, was also investigated. Animals received 5% dextran sulfate sodium (DSS) in drinking water for 7 days. One group of animals also received the Y1 receptor antagonist, administered intraperitoneally twice daily. Disease activity was assessed daily for 7 days in all groups. DSS induced colitis in all animals resulting in weight loss, diarrhea, epithelial damage, crypt shortening, and inflammatory infiltration. However, clinical manifestation of the disease was markedly attenuated in Y1 null mutant mice as well as in mice receiving the Y1 antagonist. Histological analysis showed that tissue damage and ulceration were less severe in Y1-deficient animals. Consistent with the clinical and histological data, capsaicin-induced plasma extravasation was significantly reduced in the gut of Y1 null mutant animals compared with treated wild-type animals. These data indicate that NPY and Y1 receptor are involved in intestinal inflammation and suggest that inhibition of NPY Y1 receptor signaling may provide a novel therapeutic approach in the treatment of colonic inflammation.

Acute Disease↗

PubMed2004

Whole-genome expression profiling through fragment display and combinatorial gene identification.

There is a growing demand for highly parallel gene expression analysis with whole genome coverage, high sensitivity and high accuracy. Open systems such as differential display are capable of analyzing most of the expressed genome but are not quantitative and generally require manual identification of differentially expressed genes by sequencing. Closed systems such as microarrays use gene-specific probes and are, therefore, limited to studying specific genes in well-characterized species. Here, we describe Tangerine, a PCR-based system that combines the scope and generality of open systems with a robust and immediate identification algorithm using publicly available sequence information. By combinatorial analysis of three independent and complete DNA indexing profiles, each displaying the complete set of expressed transcripts on capillary electrophoresis, the method allows transcripts to be simultaneously quantified and identified. The method is sensitive, accurate and reproducible, and is amenable to high-throughput automated operation.

Animals↗

PubMed2004

Brain-derived neurotrophic factor regulates the onset and severity of motor dysfunction associated with enkephalinergic neuronal degeneration in Huntington's disease.

The mechanism that controls the selective vulnerability of striatal neurons in Huntington's disease is unclear. Brain-derived neurotrophic factor (BDNF) protects striatal neurons and is regulated by Huntingtin through the interaction with the neuron-restrictive silencer factor. Here, we demonstrate that the downregulation of BDNF by mutant Huntingtin depends on the length and levels of expression of the CAG repeats in cell cultures. To analyze the functional effects of these changes in BDNF in Huntington's disease, we disrupted the expression of bdnf in a transgenic mouse model by cross-mating bdnf(+/ -) mice with R6/1 mice. Thus, we compared transgenic mice for mutant Huntingtin with different levels of BDNF. Using this double mutant mouse line, we show that the deficit of endogenous BDNF modulates the pathology of Huntington's disease. The decreased levels of this neurotrophin advance the onset of motor dysfunctions and produce more severe uncoordinated movements. This behavioral pathology correlates with the loss of striatal dopamine and cAMP-regulated phosphoprotein-32-positive projection neurons. In particular, the insufficient levels of BDNF cause specific degeneration of the enkephalinergic striatal projection neurons, which are the most affected cells in Huntington's disease. This neuronal dysfunction can specifically be restored by administration of exogenous BDNF. Therefore, the decrease in BDNF levels plays a key role in the specific pathology observed in Huntington's disease by inducing dysfunction of striatal enkephalinergic neurons that produce severe motor dysfunctions. Hence, administration of exogenous BDNF may delay or stop illness progression.

Age of Onset↗

PubMed2004

Endocannabinoid-independent retrograde signaling at inhibitory synapses in layer 2/3 of neocortex: involvement of vesicular glutamate transporter 3.

Recent studies implicate dendritic endocannabinoid release from subsynaptic dendrites and subsequent inhibition of neurotransmitter release from nerve terminals as a means of retrograde signaling in multiple brain regions. Here we show that type 1 cannabinoid receptor-mediated endocannabinoid signaling is not involved in the retrograde control of synaptic efficacy at inhibitory synapses between fast-spiking interneurons and pyramidal cells in layer 2/3 of the neocortex. Vesicular neurotransmitter transporters, such as vesicular glutamate transporters (VGLUTs) 1 and 2, are localized to presynaptic terminals and accumulate neurotransmitters into synaptic vesicles. A third subtype of VGLUTs (VGLUT3) was recently identified and found localized to dendrites of various cell types. We demonstrate, using multiple immunofluorescence labeling and confocal laser-scanning microscopy, that VGLUT3-like immunoreactivity is present in dendrites of layer 2/3 pyramidal neurons in the rat neocortex. Electron microscopy analysis confirmed that VGLUT3-like labeling is localized to vesicular structures, which show a tendency to accumulate in close proximity to postsynaptic specializations in dendritic shafts of pyramidal cells. Dual whole-cell recordings revealed that retrograde signaling between fast-spiking interneurons and pyramidal cells was enhanced under conditions of maximal efficacy of VGLUT3-mediated glutamate uptake, whereas it was reduced when glutamate uptake was inhibited by incrementing concentrations of the nonselective VGLUT inhibitor Evans blue (0.5-5.0 microm) or intracellular Cl- concentrations (4-145 mm). Our results present further evidence that dendritic vesicular glutamate release, controlled by novel VGLUT isoforms, provides fast negative feedback at inhibitory neocortical synapses, and demonstrate that glutamate can act as a retrograde messenger in the CNS.

Amino Acid Transport Systems, Acidic↗