Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 20 de 50
Filter
Add more filters










Publication year range
1.
J Diabetes Res ; 2022: 8566970, 2022.
Article in English | MEDLINE | ID: mdl-35967127

ABSTRACT

Aims: Mitochondrial dysfunction contributes to many forms of peripheral and central nervous system degeneration. Therapies that protect mitochondrial number and function have the potential to impact the progression of conditions such as diabetic neuropathy. We therefore assessed indices of mitochondrial function in dorsal root ganglia (DRG) and brain cortex of the Zucker diabetic fatty (ZDF) rat model of type 2 diabetes and tested the therapeutic impact of a neurogenic compound, NSI-189, on both mitochondrial function and indices of peripheral and central neurological dysfunction. Materials and Methods: ZDF rats were maintained for 16 weeks of untreated diabetes before the start of oral treatment with NSI-189 for an additional 16 weeks. Nerve conduction velocity, sensitivity to tactile and thermal stimuli, and behavioral assays of cognitive function were assessed monthly. AMP-activated protein kinase (AMPK) phosphorylation, mitochondrial protein levels, and respiratory complex activities were assessed in the DRG and brain cortex after 16 weeks of treatment with NSI-189. Results: Treatment with NSI-189 selectively elevated the expression of protein subunits of complexes III and V and activities of respiratory complexes I and IV in the brain cortex, and this was accompanied by amelioration of impaired memory function and plasticity. In the sensory ganglia of ZDF rats, loss of AMPK activity was ameliorated by NSI-189, and this was accompanied by reversal of multiple indices of peripheral neuropathy. Conclusions: Efficacy of NSI-189 against dysfunction of the CNS and PNS function in type 2 diabetic rats was accompanied by improvement of mitochondrial function. NSI-189 exhibited actions at different levels of mitochondrial regulation in central and peripheral tissues.


Subject(s)
Diabetes Mellitus, Experimental , Diabetes Mellitus, Type 2 , Diabetic Neuropathies , AMP-Activated Protein Kinases/metabolism , Aminopyridines , Animals , Diabetes Mellitus, Experimental/metabolism , Diabetes Mellitus, Type 2/metabolism , Mitochondria/metabolism , Piperazines , Rats , Rats, Zucker
6.
J Peripher Nerv Syst ; 19(2): 77-87, 2014 Jun.
Article in English | MEDLINE | ID: mdl-24934510

ABSTRACT

NIDDK, JDRF, and the Diabetic Neuropathy Study Group of EASD sponsored a meeting to explore the current status of animal models of diabetic peripheral neuropathy. The goal of the workshop was to develop a set of consensus criteria for the phenotyping of rodent models of diabetic neuropathy. The discussion was divided into five areas: (1) status of commonly used rodent models of diabetes, (2) nerve structure, (3) electrophysiological assessments of nerve function, (4) behavioral assessments of nerve function, and (5) the role of biomarkers in disease phenotyping. Participants discussed the current understanding of each area, gold standards (if applicable) for assessments of function, improvements of existing techniques, and utility of known and exploratory biomarkers. The research opportunities in each area were outlined, providing a possible roadmap for future studies. The meeting concluded with a discussion on the merits and limitations of a unified approach to phenotyping rodent models of diabetic neuropathy and a consensus formed on the definition of the minimum criteria required for establishing the presence of the disease. A neuropathy phenotype in rodents was defined as the presence of statistically different values between diabetic and control animals in 2 of 3 assessments (nocifensive behavior, nerve conduction velocities, or nerve structure). The participants propose that this framework would allow different research groups to compare and share data, with an emphasis on data targeted toward the therapeutic efficacy of drug interventions.


Subject(s)
Consensus , Diabetic Neuropathies/physiopathology , Phenotype , Animals , Behavior, Animal/physiology , Biomedical Research/methods , Biomedical Research/standards , Diabetic Neuropathies/pathology , Disease Models, Animal , Humans , Neural Conduction/physiology , Peripheral Nerves/pathology
7.
Exp Neurol ; 241: 169-78, 2013 Mar.
Article in English | MEDLINE | ID: mdl-23159890

ABSTRACT

AIMS/HYPOTHESIS: Dorsal root ganglia (DRG) sensory neurons cultured from 3 to 5 month streptozotocin (STZ)-induced diabetic rats exhibit structural and biochemical changes seen in peripheral nerve fibers in vivo, including axonal swellings, oxidative damage, reduced axonal sprouting, and decreased NF-κB activity. NF-κB is a transcription factor required by DRG neurons for survival and plasticity, and regulates transcription of antioxidant proteins (e.g. MnSOD). We hypothesized that the diabetes-induced decrease in NF-κB activity in DRG contributes to pathological phenomena observed in cultured DRG neurons from diabetic rats. METHODS: NF-κB localization was assessed in intact DRG and neuron cultures using immunostaining. NF-κB activity was manipulated in sensory neuron cultures derived from age-matched normal or 3-5 month STZ-diabetic rats using pharmacological means and lentiviral expression of shRNA. The impact of diabetes and altered NF-κB activity on neuronal phenotype involved analysis of neurite outgrowth, neurite morphology, oxidative stress (lipid peroxidation) and expression of MnSOD. RESULTS: STZ-induced diabetes caused a significant decrease in nuclear localization of NF-κB subunits p50 and c-rel, but no change in p65 in intact DRG. Inhibition of NF-κB in normal neuron cultures significantly increased axonal swellings and oxidative stress, and reduced both neurite outgrowth and expression of MnSOD. These phenomena mimicked markers of pathology in cultured DRG neurons from diabetic rats. Enhancement of NF-κB activity in cultured diabetic DRG neurons ameliorated the sub-optimal neurite outgrowth and MnSOD levels triggered by diabetes. Exogenous insulin enhanced nuclear localization of p50 and c-rel but not p65 in diabetic neuronal cultures. CONCLUSION/INTERPRETATION: The diabetes-induced decrease of nuclear localization of NF-κB subunits p50 and c-rel in DRG contributes to development of in vitro markers of peripheral neuropathy, possibly through impaired mitochondrial ROS scavenging by deficient MnSOD.


Subject(s)
Diabetes Mellitus, Experimental/pathology , Ganglia, Spinal/pathology , Gene Expression Regulation/physiology , NF-kappa B/metabolism , Sensory Receptor Cells/metabolism , Sensory Receptor Cells/pathology , ATPases Associated with Diverse Cellular Activities , Aldehydes/metabolism , Analysis of Variance , Animals , Axons/drug effects , Axons/pathology , Cells, Cultured , DNA Helicases/metabolism , Diabetes Mellitus, Experimental/complications , Disease Models, Animal , GAP-43 Protein/metabolism , Gene Expression Regulation/drug effects , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Hyperglycemia/etiology , Hypoglycemic Agents/pharmacology , Insulin/pharmacology , Male , NF-kappa B/pharmacology , Neoplasm Proteins/metabolism , Neurites/drug effects , Neurites/pathology , Nucleocytoplasmic Transport Proteins/metabolism , Oxidative Stress/physiology , RNA, Small Interfering/genetics , RNA, Small Interfering/metabolism , Rats , Rats, Sprague-Dawley , Sensory Receptor Cells/drug effects , Superoxide Dismutase/metabolism , Time Factors , Transcription Factor RelA/metabolism , Transfection
8.
Exp Neurol ; 194(1): 279-83, 2005 Jul.
Article in English | MEDLINE | ID: mdl-15899264

ABSTRACT

Sensory neurons from streptozotocin (STZ)-diabetic rats exhibit depolarization of mitochondria and the related induction of reactive oxygen species has been proposed to contribute to the etiology of sensory polyneuropathy in diabetes. There is deficient neurotrophin-3 (NT-3)-dependent neurotrophic support of sensory neurons in diabetes and treatment of STZ-diabetic rats with NT-3 prevents neuropathological alterations in peripheral nerve. Therefore, we hypothesized that loss of NT-3 may contribute to mitochondrial dysfunction in sensory neurons in diabetic sensory neuropathy. The specific aim of this study was to determine whether treatment of STZ-diabetic rats with systemic NT-3 could prevent depolarization of the mitochondrial inner membrane potential (Deltapsi(m)). In vitro studies with cultured DRG neurons from control rats revealed that treatment with 50 ng/ml NT-3 for 6 h enhanced the Deltapsi(m), e.g., a higher polarized membrane potential, compared to untreated neurons (P < 0.05). Studies on DRG sensory neurons from control vs. STZ-diabetic rats demonstrated that NT-3 therapy prevented the diabetes-induced depolarization of Deltapsi(m) (P < 0.05) in parallel with normalization of diabetes-dependent deficits in sensory nerve conduction velocity. Furthermore, alterations in mitochondrial function in vitro and in vivo correlated with the level of activation/expression of Akt in DRG neurons.


Subject(s)
Diabetes Mellitus, Experimental/drug therapy , Diabetic Neuropathies/prevention & control , Mitochondria/drug effects , Mitochondrial Diseases/prevention & control , Neurons, Afferent/drug effects , Neurotrophin 3/pharmacology , Animals , Cells, Cultured , Diabetes Mellitus, Experimental/metabolism , Diabetic Neuropathies/drug therapy , Diabetic Neuropathies/metabolism , Disease Models, Animal , Ganglia, Spinal/cytology , Male , Mitochondria/metabolism , Mitochondrial Diseases/drug therapy , Mitochondrial Diseases/metabolism , Neurons, Afferent/cytology , Neurons, Afferent/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Protein Serine-Threonine Kinases/metabolism , Proto-Oncogene Proteins/metabolism , Proto-Oncogene Proteins c-akt , Rats , Rats, Wistar , Signal Transduction/drug effects
9.
Brain ; 126(Pt 7): 1671-82, 2003 Jul.
Article in English | MEDLINE | ID: mdl-12805110

ABSTRACT

The objective was to determine whether stress-activated protein kinases (SAPKs) mediated the transfer of diabetes-induced stress signals from the periphery to somata of sensory neurons. Thus, we characterized axonal transport of SAPKs in peripheral nerve, studied any alteration in streptozotocin (STZ)-diabetic rats and examined effects of neurotrophin-3 (NT-3) on diabetes-induced events. We demonstrate that c-jun N-terminal kinase (JNK) and p38 are bidirectionally axonally transported at fast rates in sciatic nerve. In STZ-diabetic rats the relative levels of retrograde axonal transport of phosphorylated (activated) JNK and p38 were raised compared with age-matched controls (all data are in arbitrary units and expressed as fold increase over control: JNK 54-56 kDa isoforms, control 1.0 +/- 0.19, diabetic 2.5 +/- 0.26; p38, control 1.0 +/- 0.09, diabetic 2.9 +/- 0.52; both P < 0.05). Transport of total enzyme levels of JNK and p38 and phosphorylated extracellular signal-regulated kinase (ERK) was not significantly altered and anterograde axonal transport of phosphorylated JNK and p38 was unaffected by diabetes. The transcription factor ATF-2, which is phosphorylated and activated by JNK and p38, also exhibited elevated retrograde axonal transport in STZ-diabetic animals (control 1.0 +/- 0.07, diabetic 3.0 +/- 0.41; P < 0.05). Treatment of STZ-diabetic animals with 5 mg/kg human recombinant NT-3 prevented activation of JNK and p38 in sciatic nerve (phosphorylated JNK, control 1.0 +/- 0.09, diabetic 1.95 +/- 0.35, diabetic + NT-3 1.09 +/- 0.12; P < 0.05 diabetic versus others; phosphorylated p38, control 1.0 +/- 0.16, diabetic 4.7 +/- 0.9, diabetic + NT-3 1.19 +/- 0.18; P < 0.05 diabetic versus others). The results show that JNK and p38 are transported axonally and may mediate the transfer of diabetes-related stress signals, possibly triggered by loss of neurotrophic support, from the periphery to the neuronal soma.


Subject(s)
Axonal Transport/drug effects , Diabetes Mellitus, Experimental/enzymology , Diabetic Neuropathies/enzymology , JNK Mitogen-Activated Protein Kinases , Mitogen-Activated Protein Kinase Kinases/metabolism , Neurotrophin 3/pharmacology , Animals , Diabetes Mellitus, Experimental/physiopathology , Diabetic Neuropathies/physiopathology , Enzyme Activation/drug effects , MAP Kinase Kinase 4 , Male , Mitogen-Activated Protein Kinases/metabolism , Phosphorylation , Rats , Rats, Wistar , Recombinant Proteins/pharmacology , Sciatic Nerve/enzymology , p38 Mitogen-Activated Protein Kinases
10.
Cell Calcium ; 32(1): 49-52, 2002 Jul.
Article in English | MEDLINE | ID: mdl-12127062

ABSTRACT

The action of Xestospongin C (XeC) on calcium concentration in the cytosol ([Ca2+]i) and within the lumen of endoplasmic reticulum (ER) ([Ca2+]L) was studied using cultured dorsal root ganglia (DRG) neurones. Application of 2.5 microM of XeC triggered a slow [Ca2+]i transient as measured by Fura-2 video-imaging. The kinetics and amplitude of XeC-induced [Ca2+]i response was similar to that triggered by 1 microM thapsigargin (TG). The [Ca2+]L was monitored in cells loaded with low-affinity Ca2+ indicator Mag-Fura-2. The cytosolic portion of Mag-Fura-2 was removed by permeabilisation of the plasmalemma with saponin. Application of XeC to these permeabilised neurones resulted in a slow depletion of the ER Ca2+ store. XeC, however, failed to inhibit inositol 1,4,5-trisphosphate (InsP3)-induced [Ca2+]L responses. We conclude that XeC is a potent inhibitor of sarco(endo)plasmic reticulum calcium ATPase, and it cannot be regarded as a specific inhibitor of InsP3 receptors in cultured DRG neurones.


Subject(s)
Calcium/metabolism , Endoplasmic Reticulum/metabolism , Enzyme Inhibitors/pharmacology , Inositol 1,4,5-Trisphosphate/metabolism , Neurons, Afferent/metabolism , Oxazoles/pharmacology , Animals , Cells, Cultured , Ganglia, Spinal/cytology , Macrocyclic Compounds , Neurons, Afferent/cytology , Neurons, Afferent/drug effects , Rats , Rats, Sprague-Dawley
11.
Diabetologia ; 45(4): 560-70, 2002 Apr.
Article in English | MEDLINE | ID: mdl-12032634

ABSTRACT

AIMS/HYPOTHESIS: In diabetic sensory polyneuropathy the earliest and most severe pathophysiology occurs in neurones with the longest axons. The aim of this study was to characterise a diabetes-induced neurodegenerative marker that was selective for sensory neurones with the longest axons. We studied alterations in calcium homeostasis since this occurs in other neurodegenerative diseases. METHODS: Sensory neurones were cultured from control and streptozotocin-diabetic rats, treated with or without human recombinant neurotrophin-3 (hrNT-3), and neurones from L4-L6 dorsal root ganglia (DRG) which exhibit the longest axons in vivo were compared with those from C5-L3 DRG. Fluorescent video-imaging was used to measure cytoplasmic calcium dynamics. RESULTS: Streptozotocin diabetes of 8 to 14 weeks, induced an increase in resting internal Ca(2+) concentration ([Ca(2+)](i)), from 67 +/- 7 nmol/l in small neurones and 79 +/- 9 nmol/l in big neurones obtained from control animals to 214 +/- 19 nmol/l in small neurones and 273 +/- 30 nmol/l in big neurones after 14 weeks of diabetes ( p < 0.05) in L4-L6 DRG cultures. Neurones from C5-L3 ganglia and non-neuronal cells were not affected. Treatment of 14-week streptozotocin-diabetic rats with subcutaneous injection of 5 mg/kg NT-3 normalised the increase in resting [Ca(2+)](i). The amplitudes induced by depolarisation, caffeine and ATP [Ca(2+)](i) responses were reduced in small ( < 30 microm diameter) but not big ( > 35 microm diameter) neurones of L4-L6 DRG from streptozotocin-diabetic animals; the C5-L3 DRG were not similarly affected and the changes in the L4-L6 DRG were corrected by NT-3 treatment. CONCLUSIONS/INTERPRETATION: Altered calcium homeostasis could be an early molecular marker linked to the onset of diabetic sensory neuropathy. This neurodegenerative index can be corrected by NT-3 therapy and should encourage further work aimed at understanding the mechanistic basis of these observations.


Subject(s)
Calcium Signaling/physiology , Calcium/metabolism , Diabetes Mellitus, Experimental/physiopathology , Ganglia, Spinal/physiopathology , Neurons, Afferent/physiology , Neurotrophin 3/pharmacology , Adenosine Triphosphate/pharmacology , Animals , Caffeine/pharmacology , Calcium Signaling/drug effects , Ganglia, Spinal/drug effects , Homeostasis/drug effects , In Vitro Techniques , Male , Neurons, Afferent/drug effects , Rats , Rats, Wistar , Time Factors
13.
FASEB J ; 15(13): 2508-14, 2001 Nov.
Article in English | MEDLINE | ID: mdl-11689477

ABSTRACT

The onset of diabetic neuropathy, a complication of diabetes mellitus, has been linked to poor glycemic control. We tested the hypothesis that the mitogen-activated protein kinases (MAPK) form transducers for the damaging effects of high glucose. In cultures of adult rat sensory neurons, high glucose activated JNK and p38 MAPK but did not result in cell damage. However, oxidative stress activated ERK and p38 MAPKs and resulted in cellular damage. In the dorsal root ganglia of streptozotocin-induced diabetic rats (a model of type I diabetes), ERK and p38 were activated at 8 wk duration, followed by activation of JNK at 12 wk duration. We report activation of JNK and increases in total levels of p38 and JNK in sural nerve of type I and II diabetic patients. These data implicate MAPKs in the etiology of diabetic neuropathy both via direct effects of glucose and via glucose-induced oxidative stress.


Subject(s)
Diabetic Neuropathies/etiology , JNK Mitogen-Activated Protein Kinases , Mitogen-Activated Protein Kinases/physiology , Animals , Butadienes/pharmacology , Cell Survival/drug effects , Cells, Cultured , Diabetes Mellitus, Experimental/complications , Diabetes Mellitus, Type 1/enzymology , Diabetes Mellitus, Type 2/enzymology , Diabetic Neuropathies/enzymology , Dose-Response Relationship, Drug , Enzyme Activation/drug effects , Enzyme Inhibitors/pharmacology , Ganglia, Spinal/cytology , Ganglia, Spinal/drug effects , Ganglia, Spinal/enzymology , Glucose/pharmacology , Humans , Hydrogen Peroxide/pharmacology , Imidazoles/pharmacology , MAP Kinase Kinase 4 , Male , Mitogen-Activated Protein Kinase Kinases/metabolism , Mitogen-Activated Protein Kinases/antagonists & inhibitors , Mitogen-Activated Protein Kinases/metabolism , Neurons, Afferent/drug effects , Neurons, Afferent/enzymology , Nitriles/pharmacology , Pyridines/pharmacology , Rats , Rats, Wistar , Sural Nerve/enzymology , p38 Mitogen-Activated Protein Kinases
14.
Mol Cell Neurosci ; 18(2): 183-96, 2001 Aug.
Article in English | MEDLINE | ID: mdl-11520179

ABSTRACT

Mature dorsal root ganglion cells respond to neurotrophins, and the intracellular signalling pathways activated by neurotrophins have been characterized in vitro. We have now used immunocytochemistry and Western blots to examine the expression and activation of extracellular signal-regulated protein kinase-1/2 (ERK) in rat dorsal root ganglion cells in vivo, using antisera to total (tERK) and phosphorylated (pERK) forms. This has revealed a number of novel findings. tERK immunoreactivity is present in most dorsal root ganglion cells but is expressed most strongly in small (nociceptive) cells and, surprisingly, is absent in a population of large cells that expressed trkB or trkC but mainly lack p75(NTR) immunoreactivity. In contrast pERK is prominent in a few trkA cells and in satellite glial cells, and is further increased by NGF treatment. tERK and pERK both undergo fast anterograde and retrograde axonal transport, indicated by accumulation at a sciatic nerve ligature, and NGF reduces the level of retrograde pERK transport.


Subject(s)
Axonal Transport/drug effects , Ganglia, Spinal/drug effects , Mitogen-Activated Protein Kinases/drug effects , Nerve Growth Factor/pharmacology , Neurons, Afferent/drug effects , Nociceptors/drug effects , Aging/physiology , Animals , Axonal Transport/physiology , Axons/drug effects , Axons/enzymology , Axons/ultrastructure , Cell Size/physiology , Ganglia, Spinal/cytology , Ganglia, Spinal/enzymology , Gene Expression Regulation, Enzymologic/physiology , Immunohistochemistry , Ligation , Male , Mitogen-Activated Protein Kinases/metabolism , Nerve Growth Factor/metabolism , Neurons, Afferent/cytology , Neurons, Afferent/enzymology , Nociceptors/cytology , Nociceptors/enzymology , Phosphorylation , Rats , Rats, Wistar , Receptor, Nerve Growth Factor , Receptor, trkA/metabolism , Receptor, trkB/metabolism , Receptor, trkC/metabolism , Receptors, Nerve Growth Factor/metabolism , Sciatic Nerve/drug effects , Sciatic Nerve/enzymology , Sciatic Nerve/surgery
16.
Exp Neurol ; 161(2): 597-608, 2000 Feb.
Article in English | MEDLINE | ID: mdl-10686079

ABSTRACT

The expression of brain-derived neurotrophic factor (BDNF) is elevated in the soleus muscle of streptozotocin-diabetic rats. To determine whether this diabetes-induced elevation was associated with or enhanced by muscle activity we have induced high-intensity muscle contraction by electrically stimulating the sciatic nerve. In 6-week diabetic rats, intense contraction of the soleus muscle resulted in a two- to four-fold elevation of BDNF mRNA and increased plasma levels of creatine kinase that were associated with severe focal muscle fiber damage and concomitant satellite cell activation. Focal muscle fiber damage and concomitant satellite cell activation were also observed in the soleus muscle of nonstimulated diabetic rats, but to a much lesser extent. No effects of muscle contraction, i.e., experimentally induced or during normal daily activity, on muscle fiber structure or BDNF mRNA expression were seen in diabetic extensor digitorum longus (EDL) muscle. Using a nonradioactive in situ hybridization technique for electron microscopy, the elevated expression of BDNF mRNA in the diabetic soleus muscle was localized within muscle fibers as well as activated satellite cells. This study shows that diabetic soleus muscle, in contrast to diabetic EDL and to soleus and EDL muscle of normal animals, is highly susceptible to contraction-induced damage. Intense contraction and the associated muscle fiber damage in the diabetic soleus muscle result in an upregulation of BDNF mRNA in muscle fibers and activated satellite cells, which may be involved in the restoration and/or maintenance of nerve/muscle integrity.


Subject(s)
Brain-Derived Neurotrophic Factor/genetics , Diabetes Mellitus, Experimental/physiopathology , Muscle Fibers, Skeletal/metabolism , Muscle, Skeletal/physiopathology , Neuroglia/metabolism , Transcription, Genetic , Animals , Diabetes Mellitus, Experimental/genetics , Diabetes Mellitus, Experimental/pathology , Electric Stimulation , Gene Expression Regulation , Kinetics , Male , Muscle Contraction/physiology , Muscle Fibers, Skeletal/pathology , Muscle, Skeletal/metabolism , Muscle, Skeletal/pathology , Neuroglia/pathology , RNA, Messenger/genetics , Rats , Rats, Wistar , Reference Values , Sciatic Nerve/physiopathology
17.
Brain Res Mol Brain Res ; 71(2): 256-64, 1999 Aug 25.
Article in English | MEDLINE | ID: mdl-10521580

ABSTRACT

The aim of the study was to determine which factors regulated the expression of neurotrophin-3 (NT-3) mRNA in cultured primary Schwann cells derived from sciatic nerve of neonatal rats. Treatment of primary Schwann cells with the adenylate cyclase activator, forskolin, or the cAMP agonist, 8-Br-cAMP, induced a significant reduction in NT-3 transcript levels. Transforming growth factor-beta1 (TGF-beta1) and glial growth factor 2 (GGF(2)) also reduced the levels of NT-3 mRNA in a dose and time-dependent manner. Treatment with nerve growth factor, brain-derived neurotrophic factor, NT-3, ciliary neurotrophic factor or interleukin-1beta was without effect. The TGF-beta1, GGF(2) and forskolin dependent reduction in NT-3 mRNA levels involved a destabilization of transcripts which was antagonised by co-treatment with cycloheximide. The cAMP-dependent protein kinase A (PKA) inhibitor, H-89, blocked the reduction in levels of NT-3 mRNA induced by TGF-beta1, GGF(2) and forskolin. The data show that the effects of TGF-beta1, GGF(2) and forskolin on the downregulation of NT-3 mRNA, at least in part, were due to a post-transcriptional event involving a labile protein intermediate under the control of PKA. The results suggest that the down-regulation of NT-3 mRNA in Schwann cells at a site of peripheral nerve damage may be mediated via a cAMP-dependent pathway and possibly involve neuroma-related elevations in TGF-beta1 and GGF(2).


Subject(s)
Cyclic AMP/metabolism , Nerve Tissue Proteins , Neuregulin-1/pharmacology , Neurotrophin 3/genetics , RNA, Messenger/biosynthesis , Schwann Cells/metabolism , Sulfonamides , Transforming Growth Factor beta/pharmacology , 8-Bromo Cyclic Adenosine Monophosphate/pharmacology , Animals , Cells, Cultured , Colforsin/pharmacology , Cyclic AMP-Dependent Protein Kinases/metabolism , Dactinomycin/pharmacology , Enzyme Inhibitors/pharmacology , Humans , Isoquinolines/pharmacology , Protein Synthesis Inhibitors/pharmacology , Rats , Rats, Wistar , Schwann Cells/drug effects
18.
J Neurosci ; 19(18): RC24, 1999 Sep 15.
Article in English | MEDLINE | ID: mdl-10479717

ABSTRACT

The aim of this study was to determine whether axonal transport of activating transcription factor-2 (ATF2) occurs in adult sensory neurons, and whether this process is under neurotrophin control. Antisera to both total ATF2 and to the activated (i.e., phosphorylated) form were used for immunocytochemistry and Western blotting. ATF2 was localized to predominantly nociceptive dorsal root ganglion cells in adult rats and shown to accumulate proximal and distal to a sciatic nerve ligature as a result of axonal transport. Subcutaneous injection of nerve growth factor (NGF) decreased the levels of fast retrograde axonal transport of activated ATF2 by 97% (p < 0.05) and elevated levels of retrograde axonal transport of total ATF2 by twofold (p < 0.02). In contrast, blocking endogenous NGF using an anti-NGF antibody induced an elevation in retrograde axonal transport of activated ATF2 of 4. 5-fold (p < 0.05) and decreased retrograde axonal transport of total ATF2 by 72% (p < 0.05). NGF or anti-NGF treatment had no effect on the anterograde transport levels of total or activated ATF2. This study shows that signaling by target-derived NGF to the cell bodies of sensory neurons consists, in part, of the modulation of levels and activation status of a retrogradely transported transcription factor, ATF2.


Subject(s)
Axonal Transport , Cyclic AMP Response Element-Binding Protein/metabolism , Nerve Growth Factor/physiology , Neurons, Afferent/physiology , Nociceptors/physiology , Transcription Factors/metabolism , Activating Transcription Factor 2 , Animals , Fluorescent Antibody Technique, Indirect , Ganglia, Spinal/metabolism , Lumbosacral Region , Male , Nerve Growth Factor/pharmacology , Neurons, Afferent/metabolism , Nociceptors/cytology , Rats , Rats, Wistar , Sciatic Nerve/metabolism
19.
Neuroscience ; 91(1): 265-71, 1999.
Article in English | MEDLINE | ID: mdl-10336076

ABSTRACT

Effects of delivery of nerve growth factor, from a catheterized osmotic mini-pump to the proximal stump of a transected sciatic nerve, were compared with the effects of normal saline. A pilot measured retrograde axonal transport of nerve growth factor to determine a pump concentration which raised axonal transport ipsilaterally, but not contralaterally. The effects of this delivery over 12 days were then determined on expression of growth-associated protein-43, trkA, p75NTR and preprotachykinin A ipsilateral and contralateral to the pump in dorsal root ganglia at L4 and L5 (pooled). Ganglionic expression was measured both as messenger RNA and protein. Axotomy (saline pumps) increased growth-associated protein-43 messenger RNA (318 +/- 14%: all changes are percent of contralateral, non-axotomized ganglia with saline pumps) and immunoreactivity (431 +/- 43%). The increase was significantly less (P < 0.001) ipsilateral to nerve growth factor pumps (191 +/- 45%). Axotomy reduced expression of p75NTR (messenger RNA: 52 +/- 17%, P < 0.01; immunoreactivity: 74 +/- 3%, P < 0.05). These decreases were converted to increases by nerve growth factor delivery (respectively 143 +/- 40% and 281 +/- 67%; both P < 0.01). With trkA, axotomy decreased the expression of the messenger RNA (68 +/- 40%, P < 0.01) and of the primary translation product--110,000 mol. wt protein (55 +/- 12%, P < 0.01)--but not the fully glycosylated trkA protein (mol. wt 145,000). Nerve growth factor delivery did not affect trkA expression. Axotomy reduced messenger RNA for the substance P precursor, preprotachykinin A, to 42 +/- 17% (P < 0.01) and this reduction was prevented by nerve growth factor treatment. We suggest that the primary effect of nerve growth factor on axotomized C-fibres is not to promote regeneration, although that may be its secondary effect via an action on Schwann cells. It is possible that reduced neuronal sensitivity to nerve growth factor during regeneration is advantageous in suppressing nociception.


Subject(s)
Nerve Growth Factors/pharmacology , Nerve Regeneration/drug effects , Neurons, Afferent/drug effects , Peripheral Nerves/growth & development , Animals , Axotomy , Blotting, Western , GAP-43 Protein/biosynthesis , Ganglia, Spinal/cytology , Ganglia, Spinal/drug effects , Ganglia, Spinal/metabolism , Male , Nerve Tissue Proteins/biosynthesis , Nerve Tissue Proteins/genetics , Peripheral Nerves/cytology , Peripheral Nerves/drug effects , RNA, Messenger/biosynthesis , Rats , Rats, Wistar
20.
Diabet Med ; 16(2): 113-8, 1999 Feb.
Article in English | MEDLINE | ID: mdl-10229303

ABSTRACT

AIMS: This study set out to establish a novel procedure for the measurement of human nerve growth factor (NGF) messenger ribonucleic acid (mRNA) and to use this method to measure NGF expression in skin biopsies from control subjects and from patients with early neuropathies. NGF mRNA levels were related to functional measures of the competence of NGF-responsive nerves. METHODS: mRNA levels were measured by competitive reverse transcription with polymerase chain reaction amplification (cRT-PCR). Functional correlates of this observation were assessed by indices of thermal sensitivity--mediated by C-fibres, whose phenotype is regulated by NGF. RESULTS: NGF mRNA was increased in skin biopsies from 19 diabetic patients (5.12+/-3.88 (SD)) compared with samples from eight controls (1.57+/-0.95; P=0.001). Diabetic patients showed significantly (P < 0.001) diminished detection of cool and warm stimuli compared to age matched control group (n=24), but there were no differences in detection of heat as pain, or correlation with NGF mRNA levels. CONCLUSIONS: These findings suggest abnormally increased expression of NGF in diabetic neuropathy, which may represent a compensatory mechanism for impaired phenotype in NGF-responsive neurones.


Subject(s)
Diabetic Neuropathies/metabolism , Nerve Growth Factors/genetics , RNA, Messenger/biosynthesis , Skin/pathology , Adult , Aged , Case-Control Studies , Diabetic Neuropathies/pathology , Humans , Leg , Middle Aged , Reverse Transcriptase Polymerase Chain Reaction
SELECTION OF CITATIONS
SEARCH DETAIL
...