PubMed Health⌕ Search

Biomedical subjects

Klaus Unsicker

Publications and source records attributed to Klaus Unsicker.

36 records · Page 2Linked to original sources
PubMed2003

Growth differentiation factor-15 prevents low potassium-induced cell death of cerebellar granule neurons by differential regulation of Akt and ERK pathways.

Growth differentiation factor-15 (GDF-15) is a novel member of the transforming growth factor-beta superfamily and has been shown to be induced in neurons subsequent to lesions. We have therefore begun to study its putative role in the regulation of neuron survival and apoptosis. Cultured cerebellar granule neurons (CGN) survive when maintained in high K(+) (25 mm) but undergo apoptosis when switched to low K(+) (5 mm). GDF-15 prevented death of CGN in low K(+). This effect could be blocked by phosphatidylinositol 3-kinase/Akt pathway inhibitors LY294002 or wortmannin. In contrast, mitogen-activated protein kinase (MEK)/extracellular-signal-regulated kinase (ERK) pathway inhibitors U0126 and PD98059 potentiated GDF-15 mediated survival and prevented cell death in low K(+) even without factor treatment. Immunoblots revealed GDF-15-induced phosphorylation of Akt and glycogen synthase kinase-3beta. This activation was suppressed by phosphatidylinositol 3-kinase inhibitors. Low K(+) induced delayed and persistent ERK activation, which was blocked by MEK inhibitors or GDF-15. ERK activation induced c-Jun, a member of the AP-1 transcription factor family. GDF-15 or U0126 prevented c-Jun activation. Furthermore, we show that GDF-15 prevented generation of reactive oxygen species, a known activator of ERK. Together, our data suggest that GDF-15 prevents apoptosis in CGN by activating Akt and inhibiting endogenously active ERK.

Androstadienes↗
PubMed2003

Development of adrenal chromaffin cells is largely normal in mice lacking the receptor tyrosine kinase c-Ret.

c-Ret encodes a receptor tyrosine kinase that is essential for normal development of the kidney as well as enteric and sympathetic neurons. Since sympathetic neurons and neuroendocrine chromaffin cells originate from a common progenitor cell, we have examined the relevance of c-Ret for the development of adrenal chromaffin cells by analyzing mouse mutants lacking c-Ret. Adrenal chromaffin cells express c-Ret mRNA at embryonic day (E) 12.5 and 13.5, yet levels of expression decline at later embryonic and postnatal ages. Adrenal medullae of c-Ret deficient mice show normal numbers of tyrosine hydroxylase (TH)-immunoreactive cells at E13.5 and at birth. Ultrastructurally, adrenal chromaffin cells of c-Ret(-/-) mice appear unaltered: chromaffin cells develop typical secretory chromaffin granules, the morphological hallmark of chromaffin cells, and synaptic terminals appear normal. However, adrenaline levels and numbers of chromaffin cells immunoreactive for the adrenaline synthesizing enzyme phenylethanolamine-N-methyltransferase (PNMT) are reduced by about 30% in c-Ret-deficient mice arguing for a direct or indirect role of c-Ret in the regulation of PNMT. Thus, despite expression of c-Ret, adrenal chromaffin cells develop largely normal in mice lacking c-Ret. We therefore conclude that sympathetic neurons and neuroendocrine chromaffin cells profoundly differ in their requirement for c-Ret signaling during development.

Adrenal Glands↗
PubMed2003

Transforming growth factor beta2 is released from PC12 cells via the regulated pathway of secretion.

Transforming growth factor beta2 (TGF-beta2), a prototypic member of a large superfamily of multifunctional cytokines, is expressed by neurons and glial cells. Its subcellular compartmentalization and release from neurons, however, are largely unknown. Here we show that TGF-beta2 colocalizes with the trans-Golgi network marker TGN38 and a marker molecule for secretory granules, chromogranin B (CgB), in PC12 cells. Similarly, primary hippocampal neurons show colocalization of TGN38 and TGF-beta2. A substantial amount of endogenous as well as transfected TGF-beta2 in PC12 cells comigrates with CgB on an equilibrium gradient, suggesting costorage in secretory granules. TGF-beta biological activity peaks in identical fractions. Depolarization of PC12 cells with high potassium triggers colocalization of CgB and TGF-beta2 at the cell surface, suggesting their regulated corelease from secretory granules. High potassium also liberates biologically active TGF-beta from PC12 cells and primary neurons. Our results indicate that a substantial portion of TGF-beta2 is secreted by the regulated secretory pathway in PC12 cells and hippocampal neurons.

Animals↗
PubMed2003

Haploinsufficiency in trkB and/or trkC neurotrophin receptors causes structural alterations in the aged hippocampus and amygdala.

Neurotrophins and their cognate trk receptors regulate key events, most notably survival and differentiation of specific neuron populations, during the development of the nervous system. Their functions in the adult and aged CNS are far less well understood. We have analysed mice aged 21-23 months with haploinsufficiencies of the trkB and/or trkC genes with regard to morphological alterations in the hippocampus and amygdala. Neuronal densities and total numbers of neurons in the dentate gyrus were significantly decreased in trkB+/-, trkC+/-, and trkB+/-/C+/- mutants. In the hippocampal area CA2/3, neuronal density and the total number of neurons were only reduced in double-heterozygous mice. Within the amygdala, neuronal densities were not altered. The lateral, basolateral and basomedial nuclei of the amygdala, as well as the dentate gyrus and area CA3, revealed significant increases in the densities of degenerated axonal fragments; the most pronounced changes were found in the double-heterozygous mice. Thus, partial impairment in trkB and/or trkC receptor expression caused region-specific neuron losses in the hippocampus and increased axonal fragmentation in both hippocampus and amygdala, which may result from degeneration of both intrinsic and extrinsic fibre systems. Together, these data indicate that endogenous ligands to the trkB and trkC receptors are essential to maintain neuronal integrity in the aged hippocampus and amygdala.

Aging↗
PubMed2002

Age-related decline in the tyrosine hydroxylase-immunoreactive innervation of the amygdala and dentate gyrus in mice.

Numbers of catecholaminergic neurons are known to decline with aging. Whether projections of these neurons to the forebrain are similarly affected is not known. High densities of tyrosine hydroxylase-immunoreactive (TH-ir) fibers are found in the hippocampal formation (CA1-3, dentate gyrus) and in the amygdala of normal adult mice. We report here that densities of TH-ir fibers in the amygdala and hippocampus in aged mice (21-26 months) decrease dramatically and in a subregion-specific fashion. There is a reduction of 35% in the dentate gyrus, while hippocampal regions CA1 through CA3 are almost entirely spared. In the amygdala the lateral, basolateral, basomedial, and central nucleus were affected, with fiber reduction ranging from 19% to 34%. These results indicate that the age-related decline of TH-ir catecholaminergic cell bodies in the substantia nigra and the ventral tegmental area induces substantial losses of TH-ir fibers in the amygdala and dentate gyrus, but not in other areas of the hippocampal formation. This suggests that region-specific factors may be implicated in the regulation of maintenance vs. degeneration of TH-ir fibers during aging.

Aging↗
PubMed2002

Neurodegeneration in methylmalonic aciduria involves inhibition of complex II and the tricarboxylic acid cycle, and synergistically acting excitotoxicity.

Methylmalonic acidurias are biochemically characterized by an accumulation of methylmalonate (MMA) and alternative metabolites. There is growing evidence for basal ganglia degeneration in these patients. The pathomechanisms involved are still unknown, a contribution of toxic organic acids, in particular MMA, has been suggested. Here we report that MMA induces neuronal damage in cultures of embryonic rat striatal cells at a concentration range encountered in affected patients. MMA-induced cell damage was reduced by ionotropic glutamate receptor antagonists, antioxidants, and succinate. These results suggest the involvement of secondary excitotoxic mechanisms in MMA-induced cell damage. MMA has been implicated in inhibition of respiratory chain complex II. However, MMA failed to inhibit complex II activity in submitochondrial particles from bovine heart. To unravel the mechanism underlying neuronal MMA toxicity, we investigated the formation of intracellular metabolites in MMA-loaded striatal neurons. There was a time-dependent intracellular increase in malonate, an inhibitor of complex II, and 2-methylcitrate, a compound with multiple inhibitory effects on the tricarboxylic acid cycle, suggesting their putative implication in MMA neurotoxicity. We propose that neuropathogenesis of methylmalonic aciduria may involve an inhibition of complex II and the tricarboxylic acid cycle by accumulating toxic organic acids, and synergistic secondary excitotoxic mechanisms.

Animals↗
PubMed2002

Fibroblast growth factor-2 requires glial-cell-line-derived neurotrophic factor for exerting its neuroprotective actions on glutamate-lesioned hippocampal neurons.

FGF-2 is a potent neurotrophic factor for several populations of CNS neurons and has been shown to protect hippocampal neurons from glutamate-induced cell death in vitro and in vivo. Mechanisms underlying the neurotrophic and protective actions of FGF-2 have been resolved only in part. Using glutamate-treated cultured hippocampal neurons we show that FGF-2 shares its neuroprotective capacity with GDNF. Hippocampal neurons express glial-cell-line-derived neurotrophic factor (GDNF), its receptors c-Ret and the lipid-anchored GDNF family receptor-alpha1 (GFRalpha-1), and the FGF receptor 1 (FGFR I). Neutralizing antibodies to GDNF abolish the neuroprotective effect of FGF-2. In support of the notion that GDNF is required to permit the protective effects of FGF-2 we find that FGF-2 up-regulates GDNF and GFRalpha-1 in hippocampal neurons. Furthermore, FGF-2-induced GDNF causes enhanced phosphorylation of c-Ret and the signaling components Akt and Erk. A putative downstream target of FGF-2 and GDNF are bcl-2 gene family members, whose mRNAs are differentially up-regulated by the two factors. Together, these data suggest that GDNF is an important protective factor for glutamate-lesioned hippocampal neurons and an essential mediator of the neuroprotective actions of FGF-2.

Animals↗
PubMed2002

TGF-betas and their roles in the regulation of neuron survival.

Transforming growth factor-betas (TGF-betas) are a still growing superfamily of cytokines with widespread distribution and diverse biological functions. They fall into several subfamilies including the TGF-betas 1, 2, and 3, the bone morphogenetic proteins (BMPs), the growth/differentiation factors (GDFs), activins and inhibins, and the members of the glial cell line-derived neurotrophic factor family. Following a brief description of their general roles and signaling in development, maintenance of homeostasis, and disease, we shall focus on their distribution in the CNS and their involvement in regulating neuron survival and death.

Activins↗
PubMed2002

Expression and developmental regulation of gap junction connexins cx26, cx32, cx43 and cx45 in the rat midbrain-floor.

Connexins (cx) constitute a family of transmembrane proteins that form gap junction channels allowing metabolic and electrical coupling of cellular networks. Initial studies on the expression of cx in the developing brain have suggested that cx may undergo dynamic changes and may possibly be implicated in synchronizing development and differentiation of neural progenitor cells and young neurons. We have investigated expression of cx26, cx32, cx43, and cx45 in the midbrain floor, where nigrostriatal dopaminergic neurons originate and differentiate. This neuron population is of major importance in regulating motor-functions. Semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) revealed low levels of cx26-mRNA in the midbrain floor at E12, which gradually increased during pre- and postnatal development, reaching a maximum in the adult. Cx32-mRNA-levels reached a first peak at E16, and showed highest levels in adulthood. Cx43 was highly expressed at E12, decreased until E18, and subsequently increased again until adulthood. Cx45 mRNA was prominent at all developmental ages, but slightly decreased after the first postnatal week. Double-labeling for the dopaminergic neuronal marker tyrosine hydroxylase (TH), and cx-immunoreactivities (ir) evaluated by quantitative confocal laser microscopy revealed both distinct and similar developmental patterns for the individual cx investigated. Cx26 was highest at E14, decreased towards birth, and subsequently increased again reaching about 50% of the E14 level in the adult. Cx32-ir peaked at E16 and dropped to low levels after birth. Cx43-ir was highest at E12, decreased sharply at E14, reached its lowest levels at birth, but modestly increased again afterwards. Cx45-ir showed a biphasic pattern, with two prominent peaks at E12 and E18, followed by a massive postnatal decrease. Taken together, our results reveal that expression and ir of cx in the midbrain floor and dopaminergic neurons, respectively, follow cx-type specific patterns that temporally coincide with important steps of midbrain morphogenesis, as e.g. progenitor cell formation and migration (E12), early differentiation (E14-16), target encounter (E16-18) and postnatal functional maturation of the nigrostriatal system.

Animals↗
PubMed2002

Morphological alterations in the amygdala and hippocampus of mice during ageing.

Declines in memory function and behavioural dysfunction accompany normal ageing in mammals. However, the cellular and morphological basis of this decline remains largely unknown. It was assumed for a long time that cell losses in the hippocampus accompany ageing. However, recent stereological studies have questioned this finding. In addition, the effect of ageing is largely unknown in another key structure of the memory system, the amygdala. In the present study, we have estimated neuronal density and total neuronal numbers as well as density of fragments of degenerated axons in different hippocampal subfields and amygdaloid nuclei. Comparisons were made among aged (21-26 months old) mice and normal adult littermates (8 months old). No significant volume loss occurs in the hippocampus of aged mice. Small but insignificant reductions in total neuronal numbers were found in the hippocampus and in the amygdaloid nuclei. In contrast to the mild effects of ageing upon neuronal numbers, fragments of degenerated axons were increased in both hippocampus and amygdala of aged mice. These data suggest that ageing does not induce prominent cell loss in the hippocampus or amygdala, but leads to degeneration of axons that innervate these forebrain structures. Thus, mechanisms underlying age-related dysfunction depend on parameters other than neuronal numbers, at least in the hippocampal formation and the amygdala.

Afferent Pathways↗
PubMed2002

Generation of neuroendocrine chromaffin cells from sympathoadrenal progenitors: beyond the glucocorticoid hypothesis.

The developmental diversification of neural crest-derived sympathoadrenal (SA) progenitor cells into neuroendocrine adrenal chromaffin cells and sympathetic neurons has been thought to be largely understood. Based on two decades of in vitro studies with isolated SA progenitor and chromaffin cells, it was widely assumed that chromaffin cell development crucially depends on glucocorticoid hormones provided by adrenal cortical cells. However, analysis of mice lacking the glucocorticoid receptor has revealed that the chromaffin cell phenotype develops largely normally in these mice, except for the induction of the adrenaline synthesizing enzyme phenylethylamine N-methyl transferase. In a search for novel candidate genes that might be involved in triggering the sympathetic neuron/chromaffin cell decision, we have studied putative contributions of transforming growth factor (TGF)-alpha, BMP-4, and the transcription factor MASH-1, molecules with distinct expressions in SA progenitor cells, in their migratory pathways and final destinations. TGF-beta2 and -beta3 and BMP-4 are highly expressed in the wall of the dorsal aorta and in the adrenal anlagen during and after immigration of SA progenitors but expressed at much lower levels in sympathetic ganglia. We found that neutralizing antibodies against all three TGF-beta isoforms applied to the chorionic-allantoic membrane (CAM) of quail embryos interfere with proliferation of immigrated adrenal chromaffin cells but do not affect their specific neuroendocrine ultrastructural phenotype. Grafting of noggin-producing cells to the CAM, which scavenges BMPs, interferes with visceral arch and limb development but does not overtly affect the chromaffin phenotype. The transcription factor MASH-1 promotes early differentiation of SA progenitors. Mice deficient for MASH-1 lack sympathetic ganglia, whereas the adrenal medulla previously has been reported to be present. We show here that most adrenal medullary cells in MASH-1(-/-) mice identified by Phox2b immunoreactivity lack the catecholaminergic marker tyrosine hydroxylase. More surprisingly, most cells do not contain chromaffin granules and display a neuroblast-like ultrastructure and show strongly enhanced expression of c-RET comparable to that observed in sympathetic ganglia. Together, our data suggest that TGF-betas and BMP-4 do not seem to be essential for chromaffin cell differentiation. In contrast with previous reports, however, MASH-1 apparently plays a crucial role in chromaffin cell development.

Adrenal Glands↗
PubMed2002

Development of chromaffin cells depends on MASH1 function.

The sympathoadrenal (SA) cell lineage is a derivative of the neural crest (NC), which gives rise to sympathetic neurons and neuroendocrine chromaffin cells. Signals that are important for specification of these two types of cells are largely unknown. MASH1 plays an important role for neuronal as well as catecholaminergic differentiation. Mash1 knockout mice display severe deficits in sympathetic ganglia, yet their adrenal medulla has been reported to be largely normal suggesting that MASH1 is essential for neuronal but not for neuroendocrine differentiation. We show now that MASH1 function is necessary for the development of the vast majority of chromaffin cells. Most adrenal medullary cells in Mash1(-/-) mice identified by Phox2b immunoreactivity, lack the catecholaminergic marker tyrosine hydroxylase. Mash1 mutant and wild-type mice have almost identical numbers of Phox2b-positive cells in their adrenal glands at embryonic day (E) 13.5; however, only one-third of the Phox2b-positive adrenal cell population seen in Mash1(+/+) mice is maintained in Mash1(-/-) mice at birth. Similar to Phox2b, cells expressing Phox2a and Hand2 (dHand) clearly outnumber TH-positive cells. Most cells in the adrenal medulla of Mash1(-/-) mice do not contain chromaffin granules, display a very immature, neuroblast-like phenotype, and, unlike wild-type adrenal chromaffin cells, show prolonged expression of neurofilament and Ret comparable with that observed in wild-type sympathetic ganglia. However, few chromaffin cells in Mash1(-/-) mice become PNMT positive and downregulate neurofilament and Ret expression. Together, these findings suggest that the development of chromaffin cells does depend on MASH1 function not only for catecholaminergic differentiation but also for general chromaffin cell differentiation.

Adrenal Medulla↗
PubMed1990

Maintenance of Intermediolateral Spinal Cord Neurons by Fibroblast Growth Factor Administered to the Medullectomized Rat Adrenal Gland: Dependence on Intact Adrenal Innervation and Cellular Organization of Implants.

We have previously shown that basic fibroblast growth factor (bFGF; 2 microg), when administered in gelfoams implanted to medullectomized adult rat adrenal glands, maintains target-deprived preganglionic neurons located in the intermediolateral column (IML) of the spinal cord. We show now that administration of 600 or 60 ng/implant/organ of the biologically active recombinant protein also maintains IML-neurons. The effects depend on an intact organ nerve supply since administration of bFGF-implants to the medullectomized and denervated (splanchnicotomized) organ abolishes the maintenance effects for IML-neurons. A possible regeneration of cholinergic nerve fibres was assessed by quantitative determination of choline acetyltransferase (ChAT)-activity and ultrastructurally under various treatments. When compared to a cytochrome-(Cyto)-C treatment, bFGF significantly increased ChAT-activity in the innervated adrenal gland after 4 weeks, suggesting that sprouting of cholinergic nerve fibres and/or recovery of enzyme activity had occurred. Electron microscopically, unmyelinated axons associated with Schwann cells were only found in bFGF, but not in Cyto-C-implants. We conclude that exogenous bFGF or a yet unidentified trophic molecule possibly induced by bFGF in the adrenal gland acts as a retrogradely transported messenger serving for the maintenance of spinal cord IML-neurons in vivo. By stimulating axonal sprouting, bFGF might facilitate access of trophic material to nerve fibres.

Journal Article↗
PubMed1989

Basic Fibroblast Growth Factor in the Adrenal Gland.

The importance of trophic agents for the development and maintenance of neurons and their presence in mesenchyme-derived neuronal target organs such as muscle is well exemplified by the protein nerve growth factor (NGF) and its synthesis in target areas of sympathetic and sensory nerves. Stringent conceptualization of target organ-regulated neuronal maintenance would imply that neurons were able to provide trophic support to their presynaptic counterparts. We present data suggesting that basic fibroblast growth factor (bFGF), a mitogen and trophic factor for several neuron populations in vitro, may be such a protein involved in retrograde trophic neuron - neuron interaction. Basic FGF or a closely related protein is present in the adrenal medulla and its sympathetic neuron-like chromaffin cells. A polyclonal antibody specific for bFGF recognizes an 18 kD band in Western blots of bFGF-enriched bovine adrenal medulla extracts and immunostains isolated bovine chromaffin cells. This antibody also blocks the bFGF-like activity present in adrenal medullary extracts and chromaffin granule extracts that both promote in vitro survival of embryonic chick ciliary ganglionic neurons. Furthermore, like bFGF, the soluble proteins of bovine chromaffin granules are mitogenic for cultured bovine aorta endothelial cells. Electrothermal unilateral destruction of the adrenal medulla causes the disappearance of 25% of Nissl-stained neurons in the ipsilateral intermediolateral column (IML) of the spinal cord between levels Th7 and L1, which contains the preganglionic neurons projecting to the adrenal medulla. Substitution of the adrenal medulla by gel foams soaked with bFGF prevents neuron losses in the IML. The effects are specific in that NGF and cytochrome C are ineffective. Our results suggest that bFGF is located in chromaffin cells and maintains target-deprived autonomic spinal cord neurons, thus possibly acting as an interneuronal trophic messenger in vivo.

Journal Article↗
PubMedDate not supplied

The chromaffin cell and its development.

This article summarizes some of the recent progress in understanding the development of chromaffin cells. These cells are derivatives of the neural crest and are intimately associated with the sympathetic nervous system. Although a common sympathoadrenal (SA) progenitor cell for chromaffin cells and sympathetic neurons has been postulated, there is evidence to suggest that chromaffin progenitors are already distinct, at least in part, from neuronal SA progenitors prior to invading the adrenal gland. The concept of an essential role of glucocorticoid signalling for chromaffin cell development has been shaken by the observation that chromaffin cells in mice lacking the glucocorticoid receptor develop largely normal. Distinct developmental requirements of chromaffin cells and sympathetic neurons must also be assumed based on the analyses of mice carrying targeted mutations of the genes for two transcription factors, MASH1 and Phox2B. Both genes are expressed by SA progenitors, but are distinctly required for the development of chromaffin cells and sympathetic neurons. There is an ongoing search for molecules selectively operating at the sites, where chromaffin cells develop. Such molecules may be candidates for triggering the distinct developmental pathway of chromaffin cells, as opposed to sympathetic neurons.

Adrenal Glands↗
PubMedDate not supplied

TGF-beta and the regulation of neuron survival and death.

Transforming growth factor-betas (TGF-betas) constitute a superfamily of multifunctional cytokines with important implications in morphogenesis, cell differentiation, and tissue remodeling. In the developing nervous system, TGF-beta2 and -beta3 occur in radial and astroglial cells as well as in many populations of postmitotic, differentiating neurons. TGF-beta1 is restricted to the choroid plexus and meninges. In addition to functions related to glial cell maturation and performances, TGF-beta2 and -beta3 are important regulators of neuron survival. In contrast to neurotrophic factors, as for example, neurotrophins, TGF-betas are most likely not neurotrophic by themselves. However, they can dramatically increase the potency of select neurotrophins, fibroblast growth factor-2, ciliary neurotrophic factor, and glial cell line-derived neurotrophic factor (GDNF). In the case of GDNF, we have shown that GDNF fails to promote the survival of highly purified neuron populations in vitro unless it is supplemented with TGF-beta. This also applies to the in vivo situation, where antibodies to all three TGF-beta isoforms fully prevent the trophic effect of GDNF on axotomized, target-deprived neurons. In addition to the TGF-beta isoforms -beta2 and -beta3, other members of the TGF-beta superfamily are expressed in the nervous system having important roles in embryonic patterning, cell migration, and neuronal transmitter determination. We have cloned and expressed a novel TGF-beta, named growth/differentiation factor-15 (GDF-15). GDF-15 is synthesized in the choroid plexus and released into the CSF, but also occurs in all regions investigated of the developing and adult brain. GDF-15 is a potent trophic factor for developing and 6-OHDA-lesioned midbrain dopaminergic neurons in vitro and in vivo, matching the potency of GDNF.

Animals↗