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1.
Neuroscience ; 302: 174-203, 2015 Aug 27.
Article in English | MEDLINE | ID: mdl-25242643

ABSTRACT

Peripheral nerves regenerate following injury due to the effective activation of the intrinsic growth capacity of the neurons and the formation of a permissive pathway for outgrowth due to Wallerian degeneration (WD). WD and subsequent regeneration are significantly influenced by various immune cells and the cytokines they secrete. Although macrophages have long been known to play a vital role in the degenerative process, recent work has pointed to their importance in influencing the regenerative capacity of peripheral neurons. In this review, we focus on the various immune cells, cytokines, and chemokines that make regeneration possible in the peripheral nervous system, with specific attention placed on the role macrophages play in this process.


Subject(s)
Nerve Degeneration/etiology , Nerve Degeneration/immunology , Nerve Regeneration/physiology , Neuroimmunomodulation/physiology , Peripheral Nervous System Diseases/complications , Animals , Cytokines/genetics , Cytokines/metabolism , Humans
2.
Exp Neurol ; 223(2): 516-22, 2010 Jun.
Article in English | MEDLINE | ID: mdl-20144891

ABSTRACT

Sympathetic neurons, like sensory neurons, increase neurite outgrowth after a conditioning lesion. Studies in leukemia inhibitory factor (LIF) knockout animals showed that the conditioning lesion effect in sensory neurons is dependent in part on this cytokine; however, similar studies on sympathetic neurons revealed no such effect. Comparable studies with sensory neurons taken from mice lacking the related cytokine interleukin-6 (IL-6) have yielded conflicting results. LIF and IL-6 belong to a family of cytokines known as the gp130 family because they act on receptors containing the subunit gp130. In sympathetic ganglia, axotomy leads to increases in mRNA for four of these cytokines (LIF, IL-6, IL-11, and oncostatin M). To test the role of this family of cytokines as a whole in the conditioning lesion response in sympathetic neurons, mice in which gp130 was selectively eliminated in noradrenergic neurons were studied. The postganglionic axons of the SCG were transected, and 7days later the ganglia were removed and neurite outgrowth was measured in explant and dissociated cell cultures. In both systems, neurons from wild type animals showed enhanced growth after a conditioning lesion. In contrast, no enhancement occurred in neurons from mutant animals. This lack of stimulation of outgrowth occurred despite an increase in expression of activating transcription factor 3 (ATF3) in the mutant mice. These studies demonstrate that stimulation of enhanced growth of sympathetic neurons after a conditioning lesion is dependent on gp130 cytokine signaling and is blocked in the absence of signaling by these cytokines in spite of an increase in ATF3.


Subject(s)
Cytokine Receptor gp130/genetics , Cytokine Receptor gp130/metabolism , Neurites/physiology , Signal Transduction/physiology , Superior Cervical Ganglion/physiology , Activating Transcription Factor 3/metabolism , Animals , Axotomy , Cells, Cultured , Female , Interleukin-6/metabolism , Leukemia Inhibitory Factor/metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Phosphorylation/physiology , STAT3 Transcription Factor/metabolism , Superior Cervical Ganglion/cytology
3.
Neuroscience ; 150(4): 887-97, 2007 Dec 19.
Article in English | MEDLINE | ID: mdl-18031939

ABSTRACT

Activating transcription factor 3 (ATF3) is induced in a high proportion of axotomized sensory and motor neurons after sciatic nerve transection. In the present study, we looked at the expression of this factor in the superior cervical ganglion (SCG) after axotomy and after other manipulations that induce certain aspects of the cell body response to axotomy. Sympathetic ganglia from intact rats and mice exhibit only a very occasional neuronal nucleus with activating transcription factor 3-like immunoreactivity (ATF3-IR); however, as early as 6 h and as late as 3 weeks postaxotomy, many of the neurons showed intense ATF3-IR. A second population of cells had smaller and generally less intensely stained nuclei, and at least some of these cells were satellite cells. Lesions distal to the SCG induced by administration of 6-hydroxydopamine or unilateral removal of the salivary glands produced increases in ATF3-IR similar to those seen after proximal axotomy, indicating that this response is not strictly dependent on the distance of the lesion from the cell body. Two proposed signals for triggering ATF3 expression were examined: reduction in nerve growth factor (NGF) availability and induction of the cytokine leukemia inhibitory factor (LIF). While administration of an antiserum raised against NGF to intact animals induced ATF3-IR, induction of ATF3-IR after axotomy was not reduced in LIF null mutant mice. Since axotomy, 6-hydroxydopamine, and sialectomy are known to decrease the concentration of NGF in the SCG, our data suggest that these decreases in NGF lead to increases in ATF3-IR. Furthermore, since the number of neurons in the SCG expressing ATF3-IR was greater after axotomy than after antiserum against NGF treatment, this raises the possibility that decreased NGF is not the only process regulating ATF3 expression after axotomy.


Subject(s)
Activating Transcription Factor 3/metabolism , Axotomy , Ganglia, Sympathetic/cytology , Gene Expression Regulation/physiology , Neurons/metabolism , Activating Transcription Factor 3/genetics , Animals , Antibodies/pharmacology , Gene Expression Regulation/drug effects , Leukemia Inhibitory Factor/metabolism , Male , Mice , Mice, Inbred C57BL , Nerve Growth Factor/immunology , Oxidopamine/pharmacology , Rats , Rats, Sprague-Dawley , Sympatholytics/pharmacology
4.
J Neurobiol ; 66(12): 1322-37, 2006 Oct.
Article in English | MEDLINE | ID: mdl-16967509

ABSTRACT

Axotomized peripheral neurons are capable of regeneration, and the rate of regeneration can be enhanced by a conditioning lesion (i.e., a lesion prior to the lesion after which neurite outgrowth is measured). A possible signal that could trigger the conditioning lesion effect is the reduction in availability of a target-derived factor resulting from the disconnection of a neuron from its target tissue. We tested this hypothesis with respect to nerve growth factor (NGF) and sympathetic neurons by administering an antiserum to NGF to adult mice for 7 days prior to explantation or dissociation of the superior cervical ganglion (SCG) and subsequently measuring neurite outgrowth. The antiserum treatment dramatically lowered the concentration of NGF in the SCG and increased the rate of neurite outgrowth in both explants and cell cultures. The increase in neurite outgrowth was similar in magnitude to that seen after a conditioning lesion. To determine if exogenous NGF could block the effect of a conditioning lesion, mice were injected with NGF or cytochrome C immediately prior to unilateral axotomy of the SCG, and for 7 days thereafter. A conditioning lesion effect of similar magnitude was seen in NGF-treated and control animals. While NGF treatment increased NGF levels in the contralateral control ganglion, it did not significantly elevate levels in the axotomized ganglion. The results suggest that the decreased availability of NGF after axotomy is a sufficient stimulus to induce the conditioning lesion effect in sympathetic neurons. While NGF administration did not prevent the conditioning lesion effect, this may be due to the markedly decreased ability of sympathetic neurons to accumulate the growth factor after axotomy.


Subject(s)
Down-Regulation/physiology , Nerve Growth Factor/metabolism , Nerve Regeneration/physiology , Neurons/metabolism , Sympathetic Nervous System/metabolism , Animals , Antibodies/pharmacology , Axotomy , Cell Differentiation/drug effects , Cell Differentiation/physiology , Cells, Cultured , Down-Regulation/drug effects , Male , Mice , Mice, Inbred C57BL , Nerve Degeneration/metabolism , Nerve Degeneration/physiopathology , Nerve Growth Factor/antagonists & inhibitors , Nerve Regeneration/drug effects , Neurites/drug effects , Neurites/metabolism , Neurons/drug effects , Signal Transduction/drug effects , Signal Transduction/physiology , Superior Cervical Ganglion/cytology , Superior Cervical Ganglion/drug effects , Superior Cervical Ganglion/metabolism , Sympathetic Nervous System/cytology , Sympathetic Nervous System/drug effects
5.
Exp Neurol ; 194(2): 432-43, 2005 Aug.
Article in English | MEDLINE | ID: mdl-16022869

ABSTRACT

Axonal regeneration can be influenced by a conditioning lesion (an axonal injury made prior to a second test lesion). Previously, sympathetic neurons in vivo were shown to respond to a conditioning lesion with decreased neurite outgrowth, in contrast to the enhanced outgrowth observed in all other peripheral neurons examined. The present experiments tested the effects of a conditioning lesion on neurite outgrowth in vitro from the superior cervical ganglion (SCG) and the impact of several factors on that response. Ganglia axotomized 1 week earlier and then explanted in Matrigel or collagen gel responded with a significant increase in neurite extension compared to sham-operated ganglia. A distal axotomy produced by unilateral removal of the salivary glands (sialectomy) caused an increase in neurite outgrowth similar to that of a proximal axotomy. These conditioning lesions induced both an increase in the rate of elongation, and, in the case of the proximally axotomized SCG, a shorter initial delay of outgrowth. The enhanced outgrowth following sialectomy was specific to the nerve containing the majority of axons projecting to the salivary glands, suggesting that the conditioning lesion effect is restricted to previously injured neurons. Deletion of the gene for leukemia inhibitory factor (LIF), a gene induced by axotomy, did not abolish the conditioning lesion effect in SCG explants or dissociated cell cultures. In conclusion, sympathetic neurons are capable of responding to a conditioning lesion with increased neurite outgrowth. The hypothesis that the neuronal cell body response to axotomy plays an important role in the conditioning lesion response is discussed.


Subject(s)
Cell Differentiation/physiology , Nerve Regeneration/physiology , Neurites/physiology , Superior Cervical Ganglion/physiology , Animals , Axotomy/methods , Cells, Cultured , Collagen , Drug Combinations , Growth Cones/physiology , Growth Cones/ultrastructure , Interleukin-6/genetics , Laminin , Leukemia Inhibitory Factor , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Neurites/ultrastructure , Proteoglycans , Rats , Rats, Sprague-Dawley , Salivary Glands/innervation , Superior Cervical Ganglion/cytology
6.
Neuroscience ; 128(4): 741-9, 2004.
Article in English | MEDLINE | ID: mdl-15464282

ABSTRACT

Following axonal damage, sympathetic neurons are capable of regenerating and reinnervating their target tissues. Some years ago exogenous administration of polyamines was shown to enhance this regeneration. Recently, it was found that axonal injury leads to a dramatic up-regulation of the expression of arginase I in sympathetic neurons. This enzyme catalyzes the conversion of arginine to ornithine, which can subsequently be converted to the diamine putrescine and, ultimately, to the polyamines spermidine and spermine. In the present study, using an antiserum that reacts with both spermidine and spermine, we have found an increase in polyamine levels in both neurons and non-neuronal cells in the superior cervical ganglion 2 and 5 days following transection of the ganglion's postganglionic trunks. Using PC12 cells primed with nerve growth factor and then stripped off the culture dish and replated as a model system for axotomized sympathetic neurons, we found that spermidine treatment, with or without nerve growth factor, resulted in an increased percentage of cells with a neurite whose length was at least twice the diameter of the neuron's cell body. These increases could be seen within 48 h and were still evident after 8 days. Together, these data support the possibility that endogenous polyamines are involved in the normal regeneration which occurs following sympathetic axonal damage.


Subject(s)
Nerve Growth Factor/pharmacology , Neuroglia/drug effects , Neurons/drug effects , Polyamines/metabolism , Sympathetic Nervous System/cytology , Animals , Axotomy/methods , Immunohistochemistry/methods , Male , Neurites/drug effects , Neurites/physiology , Neuroglia/metabolism , Neurons/metabolism , PC12 Cells , Rats , Rats, Sprague-Dawley , Time Factors
7.
Trends Neurosci ; 24(9): 494-6; discussion 496, 2001 Sep.
Article in English | MEDLINE | ID: mdl-11506868

ABSTRACT

A recent study has shown that mice containing a loss-of-function mutation in the gene encoding galanin exhibit decreased peripheral nerve regeneration after a lesion. This major advance indicates, for the first time, that the large increases in galanin expression that occur in axotomized peripheral neurons have functional consequences for regeneration. Hopefully, similar functional consequences will soon be found for other peptides induced in these neurons after axotomy, such as vasoactive intestinal peptide and pituitary adenylate cyclase-activating peptide.


Subject(s)
Galanin/physiology , Nerve Growth Factors/physiology , Nerve Regeneration , Peptides/physiology , Animals , Galanin/genetics , Humans , Nerve Growth Factors/genetics , Nerve Regeneration/genetics , Peptides/genetics , Substance P/physiology
8.
Neuroreport ; 12(3): 601-6, 2001 Mar 05.
Article in English | MEDLINE | ID: mdl-11234772

ABSTRACT

EDI-immunoreactive macrophages, absent from the superior cervical ganglia (SCG) of normal rats, appear in these ganglia within 48h after postganglionic axotomy. Further, resident macrophages show changes after axotomy. Since chemokines function as chemoattractants and activators of leukocytes, the effects of axotomy on chemokine expression in the SCG were examined. Within 6 h after nerve transection, increases were seen in mRNA levels for monocyte chemoattractant protein (MCP)-1. MCP-1 mRNA was concentrated in a population of neurons, while MCP-1 protein was localized to endothelial cells. This axotomy-induced neuronal MCP-1 expression may trigger the infiltration and/or activation of macrophages in SCG after injury.


Subject(s)
Chemokine CCL2/genetics , Macrophages/immunology , Superior Cervical Ganglion/injuries , Superior Cervical Ganglion/physiology , Animals , Axotomy , Chemokine CCL5/genetics , Gene Expression/immunology , In Situ Hybridization , Male , Nerve Regeneration/physiology , RNA, Messenger/analysis , Rats , Rats, Sprague-Dawley
9.
J Neurobiol ; 46(3): 178-92, 2001 Feb 15.
Article in English | MEDLINE | ID: mdl-11169504

ABSTRACT

Synaptic transmission in the superior cervical ganglion (SCG) is mediated by nicotinic acetylcholine receptors (nAChR). After transection of the postganglionic nerves of the SCG in the adult rat, the transcript levels of four of the five nAChR subunits present in the ganglion, alpha3, alpha5, alpha7, and beta4, decrease dramatically. In the present study, the effect of axotomy on nAChR subunit expression was examined at the protein level, focusing on the alpha7 and beta4 subunits. Immunohistochemistry with monoclonal antibody mAb306 (for the alpha7 subunit) and polyclonal antibody 4886 (for the beta4 subunit) showed that immunoreactivities for both alpha7 and beta4 subunits were concentrated in neurons in the intact ganglion. Results from double staining with antibodies to these subunits and to tyrosine hydroxylase, the enzyme that catalyzes the rate-limiting step in the biosynthesis of the sympathetic neurotransmitter norepinephrine, demonstrated that most neurons in the SCG express both the alpha7 and beta4 subunits. Three days after axotomy, the number of immunolabeled neurons and the intensity of the immunostaining per labeled neuron were decreased for both subunits. Decreases in subunit levels were also observed by Western blot analysis. Observing changes in these subunits over time after surgery revealed that, while the protein level of the alpha7 subunit recovered substantially within 2 weeks after the lesion, that of the beta4 subunit stayed low. These data demonstrate that decreases in nicotinic receptor subunits are among the changes in proteins that occur in axotomized sympathetic neurons, and suggest that these decreases may contribute to the depression in ganglionic synaptic transmission observed in axotomized ganglia.


Subject(s)
Nerve Degeneration/metabolism , Neurons/metabolism , Receptors, Nicotinic/metabolism , Superior Cervical Ganglion/metabolism , Sympathetic Fibers, Postganglionic/injuries , Animals , Axotomy/adverse effects , Immunohistochemistry , Male , Nerve Degeneration/physiopathology , Neurons/cytology , Rats , Rats, Sprague-Dawley , Superior Cervical Ganglion/cytology , Sympathetic Fibers, Postganglionic/metabolism , Sympathetic Fibers, Postganglionic/physiopathology , alpha7 Nicotinic Acetylcholine Receptor
10.
J Neurosci ; 21(2): 363-71, 2001 Jan 15.
Article in English | MEDLINE | ID: mdl-11160417

ABSTRACT

Axonal transection of adult sympathetic and sensory neurons leads to a decrease in their content of target-derived nerve growth factor (NGF) and to dramatic changes in the expression of several neuropeptides and enzymes involved in transmitter biosynthesis. For example, axotomy of sympathetic neurons in the superior cervical ganglion (SCG) dramatically increases levels of galanin, vasoactive intestinal peptide (VIP), and substance P and their respective mRNAs and decreases mRNA levels for neuropeptide Y (NPY) and tyrosine hydroxylase (TH). Axotomy of sensory neurons in lumbar dorsal root ganglia (DRG) increases protein and mRNA levels for galanin and VIP and decreases levels for substance P and calcitonin gene-related peptide (CGRP). To assess whether reduction in the availability of endogenous NGF might play an important role in triggering these changes, we injected nonoperated animals with an antiserum against NGF (alphaNGF). alphaNGF increased levels of peptide and mRNA for galanin and VIP in neurons in both the SCG and DRG. NPY protein and mRNA were decreased in the SCG, but levels of TH protein and mRNA remained unchanged. In sensory neurons the levels of SP and CGRP protein decreased after alphaNGF treatment. These data suggest that the reduction in levels of NGF in sympathetic and sensory neurons after axotomy is partly responsible for the subsequent changes in neuropeptide expression. Thus, the peptide phenotype of these axotomized neurons is regulated both by the induction of an "injury factor," leukemia inhibitory factor, as shown previously, and by the reduction in a target-derived growth factor.


Subject(s)
Immune Sera/pharmacology , Interleukin-6 , Nerve Growth Factor/antagonists & inhibitors , Neurons, Afferent/drug effects , Neuropeptides/metabolism , Sympathetic Nervous System/drug effects , Animals , Axotomy , Brain-Derived Neurotrophic Factor/pharmacology , Calcitonin Gene-Related Peptide/metabolism , Cells, Cultured , Galanin/genetics , Galanin/metabolism , Ganglia, Spinal/cytology , Ganglia, Spinal/drug effects , Ganglia, Spinal/metabolism , Growth Inhibitors/metabolism , In Vitro Techniques , Leukemia Inhibitory Factor , Lymphokines/metabolism , Male , Nerve Growth Factor/pharmacology , Neurons, Afferent/cytology , Neurons, Afferent/metabolism , Neuropeptide Y/genetics , Neuropeptide Y/metabolism , Neuropeptides/genetics , Neurotrophin 3/pharmacology , RNA, Messenger/metabolism , Rats , Rats, Sprague-Dawley , Substance P/metabolism , Superior Cervical Ganglion/cytology , Superior Cervical Ganglion/drug effects , Superior Cervical Ganglion/physiology , Sympathetic Nervous System/cytology , Sympathetic Nervous System/metabolism , Tyrosine 3-Monooxygenase/genetics , Tyrosine 3-Monooxygenase/metabolism , Vasoactive Intestinal Peptide/genetics , Vasoactive Intestinal Peptide/metabolism
11.
J Neurosci ; 20(13): 5076-82, 2000 Jul 01.
Article in English | MEDLINE | ID: mdl-10864965

ABSTRACT

Nicotinic acetylcholine receptors (nAChRs) mediate ganglionic transmission in the peripheral autonomic nervous system in mammals. Functional neuronal nAChRs have been shown to assemble from a combination of alpha and beta subunits, including alpha3, alpha5, alpha7, beta2, and beta4 in RNA-injected oocytes, but the subunit composition of functional neuronal nAChRs in vivo in mammals remains unknown. We examined the subunit composition of functional nAChRs in the intracardiac parasympathetic ganglion in a physiologically intact system in vivo. We report here that localized perfusion of the canine intracardiac ganglion in situ with an antagonist specific for nAChRs containing an alpha3/beta2 subunit interface (alpha-conotoxin MII 100-200 nm) resulted in reversible attenuation of the sinus cycle length (SCL) response by approximately 70% to electrical stimulation of the preganglionic vagus nerve. Perfusion with antagonist specific for receptors containing an alpha3/beta4 subunit interface (alpha-conotoxin AuIB 1 micrometer) resulted in attenuation in SCL responses (approximately 20%) compared with baseline when applied by itself, but not in animals pretreated with alpha-conotoxin MII. Perfusion of the ganglion with alpha-bungarotoxin (1 micrometer, which blocks alpha7 receptors) caused a reduction in SCL response by approximately 30% compared with baseline when perfused on its own and when added after blockade with MII and AuIB. Perfusion with hexamethonium bromide resulted in complete blockade of ganglionic transmission, confirming total perfusion of the ganglion and the nicotinic nature of ganglionic transmission at this synapse. Immunohistochemistry using monoclonal antibodies against specific nicotinic subunits confirmed the presence of alpha3, alpha7, beta2, and beta4 subunits. We conclude that functional ganglionic transmission in the canine intracardiac ganglion is mediated primarily by receptors containing an alpha3/beta2 subunit interface, with a smaller contribution by receptors containing alpha7 nAChRs. Despite the presence of beta4 subunits in functional channels, a contribution of a distinct alpha3/beta4 receptor population that does not include an alpha3/beta2 subunit interface was less clear.


Subject(s)
Ganglia, Parasympathetic/physiology , Heart/innervation , Neurons/physiology , Nicotinic Antagonists/pharmacology , Receptors, Nicotinic/physiology , Synaptic Transmission/physiology , Animals , Bungarotoxins/pharmacology , Conotoxins/pharmacology , Dogs , Electric Stimulation , Ganglia, Parasympathetic/drug effects , Heart/drug effects , Hexamethonium/pharmacology , Male , Neurons/drug effects , Synaptic Transmission/drug effects , Vagus Nerve/physiology
12.
J Neurobiol ; 42(1): 14-21, 2000 Jan.
Article in English | MEDLINE | ID: mdl-10623897

ABSTRACT

Cholinergic agonists and certain peptides of the glucagon-secretin family acutely increase tyrosine hydroxylase activity in the superior cervical ganglion in vitro. The present study was designed to investigate possible interactions between these two classes of agonists in regulating catecholamine biosynthesis. Synergistic effects were found between carbachol and either secretin or vasoactive intestinal peptide in the regulation of DOPA (dihydroxyphenylalanine) synthesis. In addition, synergism was found at the level of the accumulation of cyclic adenosine monophosphate, the likely second messenger in the peptidergic regulation of tyrosine hydroxylase activity. The synergism seen with carbachol was blocked by a muscarinic, but not by a nicotinic, antagonist. Synergism was also found between bethanechol, a muscarinic agonist, and secretin, but not between secretin and dimethylphenylpiperazinium, a nicotinic agonist. Since previous immunohistochemical results suggest that vasoactive intestinal peptide and acetylcholine are colocalized in some preganglionic sympathetic neurons, the present data raise the possibility that the two might act synergistically in vivo in regulating catecholamine biosynthesis. Synergistic postsynaptic actions may be a common feature at synapses where peptides of the secretin-glucagon and acetylcholine are colocalized.


Subject(s)
Carbachol/pharmacology , Dihydroxyphenylalanine/drug effects , Muscarinic Agonists/pharmacology , Secretin/pharmacology , Superior Cervical Ganglion/drug effects , Tyrosine 3-Monooxygenase/drug effects , Vasoactive Intestinal Peptide/pharmacology , Animals , Bethanechol/pharmacology , Cyclic AMP/metabolism , Dihydroxyphenylalanine/biosynthesis , Dimethylphenylpiperazinium Iodide/pharmacology , Drug Synergism , Ganglionic Stimulants/pharmacology , Male , Muscarinic Antagonists/pharmacology , Rats , Rats, Sprague-Dawley , Superior Cervical Ganglion/enzymology , Tyrosine 3-Monooxygenase/metabolism
14.
J Neurobiol ; 40(2): 137-48, 1999 Aug.
Article in English | MEDLINE | ID: mdl-10413445

ABSTRACT

Multiple types of voltage-activated calcium (Ca(2+)) channels are present in all nerve cells examined so far; however, the underlying functional consequences of their presence is often unclear. We have examined the contribution of Ca(2+) influx through N- and L- type voltage-activated Ca(2+) channels in sympathetic neurons to the depolarization-induced activation of tyrosine hydroxylase (TH), the rate-limiting enzyme in norepinephrine (NE) synthesis, and the depolarization-induced release of NE. Superior cervical ganglia (SCG) were decentralized 4 days prior to their use to eliminate the possibility of indirect effects of depolarization via preganglionic nerve terminals. The presence of both omega-conotoxin GVIA (1 microM), a specific blocker of N-type channels, and nimodipine (1 microM), a specific blocker of L-type Ca(2+) channels, was necessary to inhibit completely the stimulation of TH activity by 55 mM K(+), indicating that Ca(2+) influx through both types of channels contributes to enzyme activation. In contrast, K(+) stimulation of TH activity in nerve fibers and terminals in the iris could be inhibited completely by omega-conotoxin GVIA alone and was unaffected by nimodipine as previously shown. K(+) stimulation of NE release from both ganglia and irises was also blocked completely when omega-conotoxin GVIA was included in the medium, while nimodipine had no significant effect in either tissue. These results indicate that particular cellular processes in specific areas of a neuron are differentially dependent on Ca(2+) influx through N- and L-type Ca(2+) channels.


Subject(s)
Action Potentials/physiology , Calcium Channels, L-Type/physiology , Calcium Channels, N-Type/physiology , Calcium Channels , Nerve Tissue Proteins/physiology , Norepinephrine/metabolism , Sympathetic Nervous System/metabolism , Tyrosine 3-Monooxygenase/metabolism , Animals , Calcium/metabolism , Calcium Channel Blockers/pharmacology , Calcium Channels, L-Type/drug effects , Calcium Channels, N-Type/drug effects , Enzyme Activation , Eye Proteins/antagonists & inhibitors , Eye Proteins/physiology , Ion Channel Gating , Ion Transport/drug effects , Iris/drug effects , Iris/innervation , Male , Nerve Endings/drug effects , Nerve Endings/metabolism , Nimodipine/pharmacology , Norepinephrine/biosynthesis , Potassium/antagonists & inhibitors , Potassium/pharmacology , Rats , Rats, Sprague-Dawley , Superior Cervical Ganglion/drug effects , Sympathetic Nervous System/drug effects , omega-Conotoxin GVIA/pharmacology
15.
J Neurosci ; 19(12): 4739-47, 1999 Jun 15.
Article in English | MEDLINE | ID: mdl-10366607

ABSTRACT

Alternative splicing of the avian trkB receptor generates an extracellular deletion (ED) isoform missing 11 amino acids from the neurotrophin-binding domain of the full-length (FL) receptor. When expressed in fibroblasts, the ED isoform exhibited restricted neurotrophin specificity compared with that of the FL receptor. Brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and neurotrophin-4 (NT-4) activated the FL receptor, as determined by tyrosine phosphorylation. However, only BDNF was capable of significant activation of the ED isoform, although to a reduced level. Because positively charged residues in NT-3 are important for binding to trkB, two negatively charged aspartate residues within the 11 amino acid motif of FL trkB were mutated to examine the role of electrostatic interactions on ligand binding. As found for the ED isoform, the FL mutated receptor displayed a similar loss of NT-3- and NT-4-mediated activation, in addition to a diminished responsiveness to BDNF. Because of these profound effects on ligand specificity, reverse transcription-PCR was used to understand the expression of the FL and ED receptor isoforms at the level of single neurons. The predominant expression pattern of either FL or ED isoforms in single embryonic DRG neurons establishes the existence of two subpopulations exhibiting differential responsiveness to trkB ligands, indicating that regulated splicing of the extracellular domain of trkB may serve as a mechanism to restrict neuronal responsiveness to the neurotrophins.


Subject(s)
Nerve Growth Factors/metabolism , Neurons/chemistry , Neuroprotective Agents/metabolism , Receptor Protein-Tyrosine Kinases/genetics , Receptors, Nerve Growth Factor/genetics , Alternative Splicing/physiology , Amino Acid Sequence , Amino Acid Substitution/physiology , Animals , Aspartic Acid , Brain-Derived Neurotrophic Factor/metabolism , Brain-Derived Neurotrophic Factor/pharmacology , Chick Embryo , Fibroblasts/cytology , Fibroblasts/physiology , Ganglia, Spinal/cytology , Gene Deletion , Isomerism , Membrane Proteins/chemistry , Membrane Proteins/genetics , Molecular Sequence Data , Nerve Growth Factors/pharmacology , Neurons/physiology , Neuroprotective Agents/chemistry , Neurotrophin 3 , Protein Binding/drug effects , Protein Binding/genetics , Protein Structure, Tertiary , Receptor Protein-Tyrosine Kinases/chemistry , Receptor, Ciliary Neurotrophic Factor , Receptors, Nerve Growth Factor/chemistry , Reverse Transcriptase Polymerase Chain Reaction
16.
Neuroreport ; 10(6): 1221-4, 1999 Apr 26.
Article in English | MEDLINE | ID: mdl-10363928

ABSTRACT

Galanin expression is co-regulated in peripheral neurons with that of vasoactive intestinal peptide (VIP) under a variety of conditions. For example, the expression of both increase after explantation of adult rat superior cervical ganglia (SCG). Because VIP participates in a positive feedback loop regulating its own expression, we examined whether VIP also increases galanin expression. Galanin mRNA and peptide are nearly undetectable in the SCG in vivo, but increase dramatically after 24-48 h in organ culture. Addition of VIP or forskolin to the culture medium reduced galanin mRNA expression by 75% and 77%, respectively, and reduced galanin peptide expression by 76% and 82%, respectively, compared with ganglia cultured in control medium. In contrast, isoproterenol stimulation did not significantly alter levels of galanin mRNA or peptide, consistent with previous observations that isoproterenol exerts its effect on SCG non-neuronal cells, but not on neurons. The results indicate that galanin and VIP are differentially regulated in sympathetic neurons by cAMP- elevating agents.


Subject(s)
Colforsin/pharmacology , Cyclic AMP/physiology , Galanin/genetics , Gene Expression Regulation/drug effects , Superior Cervical Ganglion/metabolism , Vasoactive Intestinal Peptide/pharmacology , Animals , Blotting, Northern , Culture Media , Feedback , Galanin/biosynthesis , Ganglia, Sympathetic/drug effects , Ganglia, Sympathetic/metabolism , Isoproterenol/pharmacology , Male , Neurons/cytology , Neurons/metabolism , Organ Culture Techniques , RNA, Messenger/genetics , Rats , Rats, Sprague-Dawley , Superior Cervical Ganglion/cytology , Superior Cervical Ganglion/drug effects
17.
J Neurochem ; 72(5): 1871-81, 1999 May.
Article in English | MEDLINE | ID: mdl-10217263

ABSTRACT

Axotomy of sympathetic and sensory neurons leads to changes in their neuropeptide phenotypes. These changes are mediated in part by the induction of leukemia inhibitory factor (LIF) by nonneuronal cells. In the present study, we identified satellite/Schwann cells as a possible source of the injury-induced LIF. Using a Schwann cell line, SC-1 cells, we examined mechanisms of LIF induction. LIF mRNA levels increased rapidly when the cells were treated with 8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate, phorbol 12-myristate 13-acetate (PMA), or A23187. Among these reagents, PMA was the most efficacious. Inhibition of protein kinase C (PKC) by GF-1 09203X significantly reduced the PMA-induced LIF mRNA levels. As PKC is known to activate the extracellular signal-regulated kinase (ERK) signaling pathway, the involvement of this pathway in the PMA-stimulated induction of LIF mRNA was examined. Phosphorylation of ERKs was increased following PMA treatment in SC-1 cells. Moreover, inhibition of ERK kinase activity by PD98059 dramatically reduced PMA-stimulated phosphorylation of ERKs and induction of LIF mRNA. These results indicate that LIF mRNA levels can be regulated by ERK activation via stimulation of PKC in Schwann cells.


Subject(s)
Growth Inhibitors/genetics , Interleukin-6 , Lymphokines/genetics , RNA, Messenger/metabolism , Schwann Cells/metabolism , Second Messenger Systems/physiology , Animals , Animals, Newborn/metabolism , Calcium-Calmodulin-Dependent Protein Kinases/metabolism , Cell Line , Enzyme Activation/physiology , Glial Fibrillary Acidic Protein/metabolism , Leukemia Inhibitory Factor , Protein Kinase C/physiology , Rats , Superior Cervical Ganglion/cytology , Superior Cervical Ganglion/metabolism , Tetradecanoylphorbol Acetate/pharmacology
18.
J Neurosci ; 18(14): 5285-93, 1998 Jul 15.
Article in English | MEDLINE | ID: mdl-9651211

ABSTRACT

Neurons in the adult rat superior cervical sympathetic ganglion (SCG) dramatically increase their content of vasoactive intestinal peptide (VIP) and its mRNA after axotomy in vivo and after explantation. Because the VIP gene contains a functional cAMP response element, the effects of cAMP-elevating agents on VIP expression were examined. VIP, forskolin, or isoproterenol increased cAMP accumulation in explanted ganglia. Secretin, a peptide chemically related to VIP, or forskolin increased VIP levels above those seen in ganglia cultured in control medium, whereas treatment with VIP or secretin increased the level of peptide histidine isoleucine (PHI), a peptide coded for by the same mRNA that encodes VIP. VIP or forskolin also increased VIP-PHI mRNA. In contrast, isoproterenol did not alter levels of VIP, PHI, or VIP-PHI mRNA. Although VIP or forskolin increased cAMP levels in both dissociated neurons and in non-neuronal cells, isoproterenol significantly stimulated cAMP accumulation only in the latter. VIP6-28 was an effective antagonist of the actions of exogenous VIP on cAMP and VIP-PHI mRNA in neuron-enriched cultures. When adult SCG explants were cultured in defined medium, endogenous VIP immunoreactivity was released. When VIP6-28 was added to such cultures, it significantly inhibited the increase in VIP-PHI mRNA that normally occurs. These data indicate that VIP, or a closely related molecule, produced by adult neurons after injury can enhance the expression of VIP. Such a mechanism may prolong the period during which VIP is elevated after axonal damage. The possibility is also discussed that, because VIP is present in preganglionic neurons in normal animals, its release during periods of increased sympathetic nerve activity could alter VIP expression in the SCG.


Subject(s)
Neurons/drug effects , Peptide Fragments/pharmacology , Superior Cervical Ganglion/drug effects , Vasoactive Intestinal Peptide/pharmacology , Animals , Axotomy , Cells, Cultured , Colforsin/pharmacology , Cyclic AMP/metabolism , Feedback , Isoproterenol/pharmacology , Male , Neurons/metabolism , Neurons/pathology , Organ Culture Techniques , RNA, Messenger/biosynthesis , Radioimmunoassay , Rats , Rats, Sprague-Dawley , Stimulation, Chemical , Superior Cervical Ganglion/metabolism , Superior Cervical Ganglion/pathology , Vasoactive Intestinal Peptide/biosynthesis
19.
Proc Natl Acad Sci U S A ; 95(13): 7727-30, 1998 Jun 23.
Article in English | MEDLINE | ID: mdl-9636218

ABSTRACT

Axonal damage to adult peripheral neurons causes changes in neuronal gene expression. For example, axotomized sympathetic, sensory, and motor neurons begin to express galanin mRNA and protein, and recent evidence suggests that galanin plays a role in peripheral nerve regeneration. Previous studies in sympathetic and sensory neurons have established that galanin expression is triggered by two consequences of nerve transection: the induction of leukemia inhibitory factor (LIF) and the reduction in the availability of the target-derived factor, nerve growth factor. It is shown in the present study that no stimulation of galanin expression occurs following direct application of LIF to intact neurons in the superior cervical sympathetic ganglion. Injection of animals with an antiserum to nerve growth factor concomitant with the application of LIF, on the other hand, does stimulate galanin expression. The data suggest that the response of neurons to an injury factor, LIF, is affected by whether the neurons still receive trophic signals from their targets.


Subject(s)
Growth Inhibitors/physiology , Interleukin-6 , Lymphokines/physiology , Nerve Growth Factors/physiology , Neurons/physiology , Superior Cervical Ganglion/physiology , Animals , Galanin/biosynthesis , Galanin/genetics , Growth Inhibitors/antagonists & inhibitors , Growth Inhibitors/genetics , Immune Sera , Leukemia Inhibitory Factor , Lymphokines/antagonists & inhibitors , Lymphokines/genetics , Male , Nerve Growth Factors/immunology , Nerve Regeneration , Peripheral Nervous System/injuries , Peripheral Nervous System/physiology , RNA, Messenger/metabolism , Rats , Rats, Sprague-Dawley , Superior Cervical Ganglion/drug effects
20.
J Neurobiol ; 34(2): 164-78, 1998 Feb 05.
Article in English | MEDLINE | ID: mdl-9468387

ABSTRACT

Axotomy of adult peripheral neurons produces decreases in the levels of transcripts for a number of proteins involved in synaptic transmission. For example, tyrosine hydroxylase and neuropeptide Y mRNA decrease in axotomized sympathetic neurons in the superior cervical ganglion (SCG). In the present study, the effects of axotomy on the expression of nicotinic receptor subunit transcripts were examined in the SCG and the results were compared to those produced by deafferentation and explantation. Normally, neurons in the SCG express five different nicotinic subunits: alpha3, alpha5, alpha7, beta2, and beta4. Forty-eight hours after axotomy in vivo or explantation, dramatic decreases in these transcripts were seen, except for beta2, which increased. In contrast, deafferentation of the SCG had negligible effects on any of these transcripts. Both leukemia inhibitory factor (LIF) and nerve growth factor (NGF) have been shown to play a role in the decrease in neuropeptide Y mRNA expression after axotomy. In the cases of these nicotinic receptor transcripts, however, similar decreases were seen in wild-type and LIF knockout animals. Furthermore, administration of an antiserum to NGF in intact animals produced no changes in transcript levels. On the other hand, providing exogenous NGF to axotomized SCG in vivo or in explant cultures partially prevented the decreases in the transcripts for alpha3, alpha5, alpha7, and beta4. These data indicate that axotomy produces dramatic decreases in the expression of several nicotinic receptor subunit transcripts, and that the molecular signals underlying these changes differ from those previously shown to mediate the decrease in neuropeptide Y expression.


Subject(s)
Gene Expression Regulation , Interleukin-6 , Neurons/physiology , Receptors, Nicotinic/metabolism , Sympathetic Nervous System/physiology , Animals , Axotomy , Gene Expression Regulation/drug effects , Growth Inhibitors/genetics , Growth Inhibitors/physiology , Leukemia Inhibitory Factor , Lymphokines/genetics , Lymphokines/physiology , Male , Mice , Mice, Knockout , Nerve Growth Factors/pharmacology , Neurons/cytology , Neurons/drug effects , Rats , Rats, Sprague-Dawley , Receptors, Nicotinic/genetics , Superior Cervical Ganglion/drug effects , Superior Cervical Ganglion/physiology , Sympathetic Nervous System/metabolism , Transcription, Genetic
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