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S Korsching

Publications and source records attributed to S Korsching.

At least 19 recordsLinked to original sources
PubMed1997

Functional expression of odorant receptors of the zebrafish Danio rerio and of the nematode C. elegans in HEK293 cells.

Odorant receptors of zebrafish and C elegans were functionally expressed in vertebrate kidney cells (HEK293) using the eucaryotic expression vector pSMyc. Receptor-encoding cDNA cloned into this vector was expressed as a fusion protein with the N-terminal membrane import sequence of the guinea-pig serotonin receptor followed by a myc tag. Immunocytochemical evidence indicates that this strategy directs a protein with the predicted immunoreactivity and approximate molecular weight to the plasma membrane. Fish food extract (TetraMin) evoked a transient increase in intracellular [Ca2+] in HEK293 cells transiently transfected with plasmids containing cDNA for three fish odorant receptors and converted to stable cell lines. The effect of the extract was concentration dependent and limited to the fraction of the extract < 5 kDa. Pretreating the transfected cells with the PLC inhibitor U73122 reduced the odor-evoked signal. Fish food extract also evoked a transient increase in intracellular [Ca2+] in HEK293 cells transiently transfected with plasmids containing cDNA for single fish odorant receptors. Diacetyl evoked a transient increase in intracellular [Ca2+] in HEK293 cells transiently transfected with plasmids encoding the cDNA of ODR10, an odorant receptor of C. elegans suggested in other work to be specific for diacetyl. These results strongly imply that odorant receptors can be functionally expressed in HEK293 cells using this novel expression protocol.

Animals↗
PubMed1997

Tissue-specific and ontogenetic regulation of LIF protein levels determined by quantitative enzyme immunoassay.

To define the physiological role of leukemia inhibitory factor (LIF), it is essential to localize sites of LIF synthesis in vivo. We generated polyclonal antibodies specific for native rat LIF, and developed a two-site immunoassay to detect 10 pg LIF/ml. Using this immunoassay, we determined LIF content of 18 organs, CNS regions, and ganglia throughout postnatal development of rats. High levels of LIF protein (1.0-11.0 ng/g tissue) are present in relatively few tissues: the uterus at late proestrus to estrus and on day 5 of pregnancy, ovary at estrus to early metestrus-1, footpads during early postnatal development and thymus throughout. Intermediate levels (0.5-1.0 ng) are detected in the gut, skin, skeletal muscle, pancreas and lung at one or more postnatal ages. Low levels (0.1-0.5 ng) are observed in most other non-nervous and nervous tissues. LIF protein levels do not completely correspond to reported LIF mRNA levels.

Age Factors↗
PubMed1996

Nested expression domains for odorant receptors in zebrafish olfactory epithelium.

The mapping of high-dimensional olfactory stimuli onto the two-dimensional surface of the nasal sensory epithelium constitutes the first step in the neuronal encoding of olfactory input. We have used zebrafish as a model system to analyze the spatial distribution of odorant receptor molecules in the olfactory epithelium by quantitative in situ hybridization. To this end, we have cloned 10 very divergent zebrafish odorant receptor molecules by PCR. Individual genes are expressed in sparse olfactory receptor neurons. Analysis of the position of labeled cells in a simplified coordinate system revealed three concentric, albeit overlapping, expression domains for the four odorant receptors analyzed in detail. Such regionalized expression should result in a corresponding segregation of functional response properties. This might represent the first step of spatial encoding of olfactory input or be essential for the development of the olfactory system.

Algorithms↗
PubMed1994

Connectional topography in the zebrafish olfactory system: random positions but regular spacing of sensory neurons projecting to an individual glomerulus.

It is unknown how neuronal connections are specified in the olfactory system. To define rules of connectivity in this system, we investigated whether the projection of sensory neurons from the olfactory epithelium to the olfactory bulb is topographically ordered. By backtracking with 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI), we find that neurons projecting into a single identified glomerulus are widely dispersed over the olfactory epithelium. Their positions in the sensory surface do not predict their glomerulus specificity and are probably random. A statistical analysis reveals that neurons connected to the same glomerulus are spaced at distances of several cell diameters from each other. The convergence of projections to one point in the target area from neurons that are widely and evenly distributed in the sensory surface constitutes an unusual type of connectional topography that contrasts with the precise topological (neighborhood-preserving) maps found in other sensory systems. It may maximize the probability to detect odorants that activate a single glomerular unit.

Animals↗
PubMed1994

Olfactory glomeruli in the zebrafish form an invariant pattern and are identifiable across animals.

Glomeruli are anatomical and possibly functional modules in the vertebrate olfactory bulb. We investigated the spatial arrangement of glomeruli in the olfactory bulbs of adult zebrafish (Brachydanio rerio). A solution of the lipophilic tracer Dil was injected into the nasal cavities. Axons of sensory neurons projecting from the olfactory epithelium into the bulb were traced anterogradely, thus labeling the whole population of glomeruli. The glomerular distribution was analyzed in detail by confocal laser-scanning microscopy. We find that a typical olfactory bulb contains a small number of about 80 glomeruli that have a stereotyped configuration in all animals investigated. All glomeruli exhibit bilateral symmetry. Twenty-two single glomeruli could be identified from animal to animal by their characteristic position and morphology. The remaining glomeruli either are embedded in glomerular plexus and therefore cannot be delineated reliably, or belong to a densely clustered subpopulation of on average 49 glomeruli in the dorsal olfactory bulb. No sexually dimorphic glomeruli were identified. To test whether glomerular constancy is specific for the zebrafish, we performed similar tracing experiments in the goldfish and found several indications for a similar invariance of glomeruli in this species. The remarkable stereotypy of this pattern is reminiscent of the insect olfactory system and has been demonstrated here for the first time in a vertebrate. It will now be possible to examine whether these identifiable glomeruli are functionally specialized in terms of odor processing. If so, zebrafish may emerge as a tractable model system for studies on olfactory coding.

Animals↗
PubMed1993

The neurotrophic factor concept: a reexamination.

The neurotrophic factor concept in its basic form envisages that innervated tissues produce a signal for the innervating neurons for the selective limitation of neuronal death occurring during development (Purves, 1986; Oppenheim, 1991). This concept arose several decades ago on the basis of the observation that experimental manipulation of the amount of target tissue could modulate the size of neuronal populations. By making the survival of neurons dependent on their target, nature would provide a means to match neuron and target cell populations. NGF, discovered in the 1950s, represents the first known molecular realization of the neurotrophic factor concept. NGF was found to regulate survival, neurite growth, and neurotransmitter production of a particular neuronal type, the sympathetic neurons of the PNS. NGF produced by target cells is specifically bound and internalized by sympathetic neurons, followed by retrograde axonal transport of NGF to the cell soma, where NGF exerts its effects via the cotransported receptor molecule (Levi-Montalcini, 1987; Thoenen et al., 1987). Strictly speaking, increased neurite growth and neurotransmitter production are not trophic effects; however, I will use the term "neurotrophic" in the extended meaning of enhancing neuronal differentiation as well as neuronal survival. It was expected that these results could be generalized to a model of multiple, mutually independent, retrograde trophic messengers, which are synthesized in distinct target areas and act on restricted neuronal types (Fig. 1). This assumption leads to a conceptually simple way to arrange and maintain a variety of neuronal subsystems. One might call this a modular approach to the construction of the nervous system. The hypothesis of multiple retrograde signals has gained widespread experimental support in recent years. Originally proposed for the PNS, the model could be extended to the CNS, in which target neurons synthesize trophic factors for their afferent neurons (Ernfors et al., 1990b). In addition to NGF, a family of NGF-related molecules (now commonly called neurotrophins), which are thought to exert retrograde trophic influences (DiStefano et al., 1992), has been identified.

Animals↗
PubMed1989

The cholinergic neuronal differentiation factor from heart cells is identical to leukemia inhibitory factor.

A protein secreted by cultured rat heart cells can direct the choice of neurotransmitter phenotype made by cultured rat sympathetic neurons. Structural analysis and biological assays demonstrated that this protein is identical to a protein that regulates the growth and differentiation of embryonic stem cells and myeloid cells, and that stimulates bone remodeling and acute-phase protein synthesis in hepatocytes. This protein has been termed D factor, DIA, DIF, DRF, HSFIII, and LIF. Thus, this cytokine, like IL-6 and TGF beta, regulates growth and differentiation in the embryo and in the adult in many tissues, now including the nervous system.

Amino Acid Sequence↗
PubMed1988

Developmental changes of nerve growth factor levels in sympathetic ganglia and their target organs.

The predominant source of nerve growth factor (NGF) used by mature sympathetic neurons originates in their target organs (Heumann, R., Korsching, S., Scott, J., and Thoenen, H. (1984), EMBO J. 3, 3183-3189; Korsching, S., and Thoenen, H. (1985), J. Neurosci. 5, 1058-1061). We have determined the NGF content of two sympathetically innervated mouse organs, submandibular gland and heart ventricle, and of sympathetic ganglia from mouse and rat between embryonic Day 12 (E12) and adulthood. NGF levels were measured by a two-site enzyme immunassay (Korsching, S., and Thoenen, H. (1983), Proc. Natl. Acad. Sci. USA 80, 3513-3516). In heart ventricle and submandibular gland, NGF first became detectable around the time of initial innervation by sympathetic neurons (E12 and E13, respectively) and increased respectively 14- and 7-fold in the following 2 days, to reach adult levels already at E14 for heart ventricle (1.4 +/- 0.2 ng NGF/g wet wt). NGF in the superior cervical ganglion (SCG) was first detected at the same time as in its target organ, the submandibular gland. NGF content in the SCG then increased 6-fold during the next 2 days and continued to increase until the end of the third postnatal week, when adult levels were reached. Although the levels of NGF in the adult mouse submandibular gland are sexually dimorphic and six orders of magnitude higher than those in other sympathetic target organs, no sex difference in the NGF content was found in either developing submandibular gland or SCG until the end of the third postnatal week. Moreover, the steep NGF increase observed in the male submandibular gland after postnatal Day 18 (250-fold within the following 3 days and up to the 55,000-fold in the next 7 days) was not reflected in a corresponding increase in the NGF content of the male SCG. These data indicate that, in accordance with earlier findings (see Levi-Montalcini, R., and Angeletti, P. U. (1968), Physiol. Rev. 48, 534-569), SCG neurons do not have access to the large amounts of NGF synthesized during and after adolescence in the mouse submandibular gland. Our results support the concept that initial fiber outgrowth of sympathetic neurons is neither dependent on NGF nor mediated by it. The time course of NGF levels in the SCG is consistent with the concept that sympathetic neurons are provided with NGF by means of retrograde axonal transport from the innervated organs already early in development.

Aging↗
PubMed1988

Nerve growth factor synthesis in cultured rat iris: modulation by endogenous transmitter substances.

Organ cultures of rat iris show a characteristic change in the levels of both nerve growth factor (NGF) and its mRNA: a rapid but transient initial increase is followed by a smaller but persistently elevated NGF synthesis. This time course may be influenced by release of a factor(s) from degenerating nerve terminals and/or by the lack of some factor(s) repressing NGF synthesis in vivo. We therefore analyzed the influence of biogenic amine transmitter substances and putative neuropeptides on this elevation of NGF synthesis in cultured iris. The marked increase of NGF synthesis seen initially in culture was not completely mimicked by any of the substances tested. A specific increase in NGF production up to 150% of control was observed only with cGMP. We also obtained some evidence that reaction to trauma following the culture procedure could enhance NGF production: cutting of irides into small pieces increased NGF production in culture up to 250% of control and, vice versa, treatment with 1 microM dexamethasone decreased NGF production to about 60% of control. However, the sympathetic neurotransmitter norepinephrine (NE) decreased both NGF and its mRNA levels specifically in a dose-dependent manner (0.01-1 mM) to a minimum of about 25% of control. In situ hybridization with mRNA(NGF)-specific probes showed that in cultures of dissociated iris cells all cells were capable of expressing mRNA(NGF), but that 0.1 mM NE preferentially decreased expression of mRNA(NGF) in smooth muscle cells. Thus, our results indicate that the sympathetic transmitter NE is capable of downregulating NGF synthesis in the target cells of sympathetic neurons.

Animals↗
PubMed1988

Antibodies against mouse nerve growth factor interfere in vivo with the development of avian sensory and sympathetic neurones.

The monoclonal antibody 27/21 directed against mouse nerve growth factor (NGF) interferes in vivo with the survival of sensory dorsal root ganglion (DRG) neurones during the development of the quail embryo: the number of DRG neurones at embryonic day 11 (E11) was reduced by about 30% in embryos treated with the antibody between E3 and E11. Neurone numbers in the nodose ganglion were not affected. The effect of NGF antibodies on sympathetic neurones was assessed by determining the levels of the adrenergic marker enzyme tyrosine hydroxylase. Both total tyrosine hydroxylase activity and protein levels in sympathetic chains were reduced by about 30% in embryos treated with 27/21 antibody but not in embryos treated with a control antibody. The 27/21 antibody cross-reacts with chick NGF-like activity as shown in vitro by the ability of the antibody to partially block the survival activity of chick-embryo-fibroblast-conditioned medium for E9 chick DRG neurones.

Animals↗
PubMed1987

Developmental changes of nerve growth factor and its mRNA in the rat hippocampus: comparison with choline acetyltransferase.

Previous experiments have demonstrated that in the septo-hippocampal system choline acetyltransferase (ChAT) is induced by nerve growth factor (NGF) (Gnahn et al. (1983) Dev. Brain Res. 9, 45-52) and that hippocampal NGF and mRNANGF levels are correlated with the density of cholinergic innervation (Korsching et al. (1985) EMBO J. 4, 1389-1393). In the present investigation we have compared the developmental changes of ChAT, NGF, and mRNANGF levels in this system. During the postnatal development of the hippocampus the time courses of NGF and ChAT were well correlated including the most rapid increase between P12 and P14. This increase in hippocampal NGF was preceded by a corresponding increase in mRNANGF. The developmental changes in hippocampal NGF levels were also closely reflected by corresponding changes in the septum. This, together with previous observations (Korsching et al., 1985) that the adult septum, in spite of relatively high NGF levels, does not contain measurable quantities of mRNANGF, suggests that the NGF levels in the septum are determined by the quantity of NGF transported retrogradely from the field of innervation rather than by local synthesis. During the prenatal period hippocampal NGF levels were relatively high, whereas the mRNANGF was below the level of detection. Since the ingrowth of septal fibers, and with that also the removal of NGF by retrograde transport, begins around birth, the relatively high prenatal NGF levels probably result from an accumulation produced by a small copy number of mRNANGF prior to the removal of NGF by retrograde axonal transport. It is concluded that the correlation of the developmental changes in NGF and mRNANGF with the ChAT activity in the hippocampus further supports the concept of a physiological role of NGF in the central nervous system.

Animals↗
PubMed1987

Changes of nerve growth factor synthesis in nonneuronal cells in response to sciatic nerve transection.

The intact sciatic nerve contains levels of nerve growth factor (NGF) that are comparable to those of densely innervated peripheral target tissues of NGF-responsive (sympathetic and sensory) neurons. There, the high NGF levels are reflected by correspondingly high mRNANGF levels. In the intact sciatic nerve, mRNANGF levels were very low, thus indicating that the contribution of locally synthesized NGF by nonneuronal cells is small. However, after transection an increase of up to 15-fold in mRNANGF was measured in 4-mm segments collected both proximally and distally to the transection site. Distally to the transection site, augmented mRNANGF levels occurred in all three 4-mm segments from 6 h to 2 wk after transection, the longest time period investigated. The augmented local NGF synthesis after transection was accompanied by a reexpression of NGF receptors by Schwann cells (NGF receptors normally disappear shortly after birth). Proximal to the transection site, the augmented NGF synthesis was restricted to the very end of the nerve stump that acts as a "substitute target organ" for the regenerating NGF-responsive nerve fibers. While the mRNANGF levels in the nerve stump correspond to those of a densely innervated peripheral organ, the volume is too small to fully replace the lacking supply from the periphery. This is reflected by the fact that in the more proximal part of the transected sciatic nerve, where mRNANGF remained unchanged, the NGF levels reached only 40% of control values. In situ hybridization experiments demonstrated that after transection all nonneuronal cells express mRNANGF and not only those ensheathing the nerve fibers of NGF-responsive neurons.

Animals↗
PubMed1986

Cholinergic denervation of the rat hippocampus by fimbrial transection leads to a transient accumulation of nerve growth factor (NGF) without change in mRNANGF content.

Unilateral stereotaxic fimbrial transection was used to disrupt the cholinergic pathway from the medial septum to the hippocampus in adult rats. The resulting alterations in hippocampal nerve growth factor (NGF) and mRNANGF levels were then analysed by two-site enzyme immunoassay and quantitative Northern blots, respectively. NGF levels did not change up to one week after lesion but had increased to 145% of contralateral control values by two weeks. In contrast, mRNANGF levels were not changed at this time point. Neither bilateral fimbrial transection, nor removal of the sympathetic innervation of hippocampal vasculature by excision of the superior cervical ganglia, led to more pronounced increases in NGF levels. These results are consistent with the concept that the magnocellular cholinergic neurons in the medial septum are constantly reducing hippocampal NGF levels by retrograde axonal transport.

Animals↗
PubMed1985

Nerve growth factor supply for sensory neurons: site of origin and competition with the sympathetic nervous system.

Nerve growth factor (NGF) levels in the sensory nervous system were measured by a highly sensitive two-site enzyme immunoassay for NGF. Dorsal root ganglia and the adjacent spinal nerves contained 2.8 +/- 0.3 and 1.7 +/- 0.4 ng NGF/g wet wt., respectively, whereas no NGF was detectable in dorsal roots and spinal cord (less than 0.05 ng NGF/g wet wt.). It is concluded that sensory neurons are supplied with NGF exclusively from their peripheral and not from their central field of projection. Two days after treatment with 6-hydroxydopamine, which destroys sympathetic nerve terminals and thereby prevents the removal of NGF by sympathetic neurons, the NGF content of dorsal root ganglia and trigeminal ganglia increased to 285% and 161% of control, respectively. This indicates that in peripheral target tissues sensory and sympathetic neurons compete for NGF.

Animals↗
PubMed1985

Nerve growth factor.

In contrast to all other molecules which are labelled 'growth factor', NGF is not a mitogen. It is a neurotrophic molecule essential for the development and maintenance of function of specific populations of peripheral and possibly also central neurons. The availability of NGF in large quantities from exocrine glands (e.g. male mouse submandibular gland), where NGF does not play a neurotrophic role, has allowed the purification of NGF, the production of specific antibodies, the determination of its amino acid sequence and finally the molecular cloning of NGF leading to the elucidation of its precursor structure and its genomic organization. Comparison of the biological activities and the immunological properties of NGF isolated from different sources demonstrated that the active centre of the molecule has been highly conserved during evolution, whereas other parts of the molecule determining immunological properties have undergone considerable changes. After a survey of the essential biological actions of NGF, this paper concentrates on two actual questions of NGF research, namely the regulation of NGF synthesis in the target tissues of NGF-responsive neurons, and the molecular mechanism(s) of action of NGF on these neurons.

Animals↗