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1.
J Dent Res ; 95(10): 1124-31, 2016 09.
Article in English | MEDLINE | ID: mdl-27307048

ABSTRACT

Microglia are the resident immune cells in the spinal cord and brain. Mounting evidence suggests that activation of microglia plays an important role in the pathogenesis of chronic pain, including chronic orofacial pain. In particular, microglia contribute to the transition from acute pain to chronic pain, as inhibition of microglial signaling reduces pathologic pain after inflammation, nerve injury, and cancer but not baseline pain. As compared with inflammation, nerve injury induces much more robust morphologic activation of microglia, termed microgliosis, as shown by increased expression of microglial markers, such as CD11b and IBA1. However, microglial signaling inhibitors effectively reduce inflammatory pain and neuropathic pain, arguing against the importance of morphologic activation of microglia in chronic pain sensitization. Importantly, microglia enhance pain states via secretion of proinflammatory and pronociceptive mediators, such as tumor necrosis factor α, interleukins 1ß and 18, and brain-derived growth factor. Mechanistically, these mediators have been shown to enhance excitatory synaptic transmission and suppress inhibitory synaptic transmission in the pain circuits. While early studies suggested a predominant role of microglia in the induction of chronic pain, further studies have supported a role of microglia in the maintenance of chronic pain. Intriguingly, recent studies show male-dominant microglial signaling in some neuropathic pain and inflammatory pain states, although both sexes show identical morphologic activation of microglia after nerve injury. In this critical review, we provide evidence to show that caspase 6-a secreted protease that is expressed in primary afferent axonal terminals surrounding microglia-is a robust activator of microglia and induces profound release of tumor necrosis factor α from microglia via activation of p38 MAP kinase. The authors also show that microglial caspase 6/p38 signaling is male dominant in some inflammatory and neuropathic pain conditions. Finally, the authors discuss the relevance of microglial signaling in chronic trigeminal and orofacial pain.


Subject(s)
Caspase 6/metabolism , Chronic Pain/enzymology , Facial Pain/enzymology , Microglia/physiology , Signal Transduction/physiology , p38 Mitogen-Activated Protein Kinases/metabolism , Chronic Pain/immunology , Facial Pain/immunology , Female , Humans , Male , Microglia/enzymology , Microglia/immunology , Sex Factors , Signal Transduction/immunology
2.
Neuroscience ; 247: 376-85, 2013 Sep 05.
Article in English | MEDLINE | ID: mdl-23707980

ABSTRACT

Accumulating evidence indicates that activation of spinal cord astrocytes contributes importantly to nerve injury and inflammation-induced persistent pain and chronic opioid-induced antinociceptive tolerance. Phosphorylation of extracellular signal-regulated kinase (pERK) and induction of interleukin-1 beta (IL-1ß) in spinal astrocytes have been implicated in astrocytes-mediated pain. Tissue plasminogen activator (tPA) is a serine protease that has been extensively used to treat stroke. We examined the potential involvement of tPA in chronic opioid-induced antinociceptive tolerance and activation of spinal astrocytes using tPA knockout (tPA(-/-)) mice and astrocyte cultures. tPA(-/-) mice exhibited unaltered nociceptive pain and morphine-induced acute analgesia. However, the antinociceptive tolerance, induced by chronic morphine (10mg/kg/day, s.c.), is abrogated in tPA(-/-) mice. Chronic morphine induces tPA expression in glial fibrillary acidic protein (GFAP)-expressing spinal cord astrocytes. Chronic morphine also increases IL-1ß expression in GFAP-expressing astrocytes, which is abolished in tPA-deficient mice. In cultured astrocytes, morphine treatment increases tPA, IL-1ß, and pERK expression, and the increased IL-1ß and pERK expression is abolished in tPA-deficient astrocytes. tPA is also sufficient to induce IL-1ß and pERK expression in astrocyte cultures. Intrathecal injection of tPA results in up-regulation of GFAP and pERK in spinal astrocytes but not up-regulation of ionized calcium binding adapter molecule 1 in spinal microglia. Finally, intrathecal tPA elicits persistent mechanical allodynia, which is inhibited by the astroglial toxin alpha-amino adipate and the MEK (ERK kinase) inhibitor U0126. Collectively, these data suggest an important role of tPA in regulating astrocytic signaling, pain hypersensitivity, and morphine tolerance.


Subject(s)
Hyperalgesia/metabolism , Interleukin-1beta/biosynthesis , Morphine/administration & dosage , Spinal Cord/metabolism , Tissue Plasminogen Activator/deficiency , Animals , Astrocytes/metabolism , Drug Tolerance/physiology , Hyperalgesia/pathology , Injections, Spinal , MAP Kinase Signaling System/physiology , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Phosphorylation , Signal Transduction/physiology
3.
Br J Anaesth ; 108(4): 662-9, 2012 Apr.
Article in English | MEDLINE | ID: mdl-22307241

ABSTRACT

BACKGROUND: There is increasing interest in RNA interference in pain research using the intrathecal route to deliver small-interfering RNA (siRNA). An interferon (IFN) response is a common side-effect of siRNA. However, the IFN response in the spinal cord after intrathecal administration of siRNA remains unknown. We hypothesized that high doses of siRNAs can elicit off-target analgesia via releasing IFN-α. We investigated the IFN response and its role in regulating pain sensitivity in the spinal cords after intrathecal administration of siRNAs. METHODS: Male Sprague-Dawley rats were given intrathecal injections of non-targeting (NT) siRNAs or IFN-α and tested for complete Freund's adjuvant (CFA)-induced mechanical allodynia and heat hyperalgesia. IFN-α in the spinal cord after injection of NT siRNAs was measured by western blotting and immunohistochemical staining. RESULTS: IFN-α was up-regulated in the spinal cord after intrathecal treatment of NT siRNAs. Intrathecal injection of NT siRNAs, at high doses of 10 or 20 µg, reduced CFA-induced inflammatory pain (P<0.05). Intrathecal application of IFN-α inhibited pain hypersensitivity in inflamed rats and produced analgesia in naïve rats (P<0.05). Notably, the anti-nociceptive effects elicited by NT siRNAs and IFN-α were reversed by IFN-α neutralizing antibody and naloxone. CONCLUSIONS: Our data suggest that (i) intrathecal administration of high doses of siRNA (≥ 10 µg) induced up-regulation of IFN-α in the spinal cord and produced analgesic effects through IFN-α, and (ii) IFN-α's analgesic effect is mediated via opioid receptors. Caution must be taken to avoid IFN-α-mediated analgesic effects of siRNAs in pain research.


Subject(s)
Analgesia/methods , Interferon-alpha/pharmacology , Pain/drug therapy , RNA, Small Interfering/pharmacology , Animals , Blotting, Western , Disease Models, Animal , Freund's Adjuvant/administration & dosage , Freund's Adjuvant/pharmacology , Hyperalgesia/drug therapy , Immunologic Factors/pharmacology , Inflammation/complications , Inflammation/drug therapy , Injections, Spinal , Interferon-alpha/administration & dosage , Male , Pain/complications , Pain Management/methods , Pain Threshold/drug effects , RNA, Small Interfering/administration & dosage , Rats , Rats, Sprague-Dawley , Spinal Cord/drug effects , Up-Regulation/drug effects
4.
Anesthesiol Res Pract ; 2011: 124898, 2011.
Article in English | MEDLINE | ID: mdl-21776253

ABSTRACT

Peripheral and neuraxial nerve blockades are widely used in the perioperative period. Their values to diminish acute postoperative pain are established but other important outcomes such as chronic postoperative pain, or newly, cancer recurrence, or infections could also be influenced. The long-term effects of perioperative nerve blockade are still controversial. We will review current knowledge of the effects of blocking peripheral electrical activity in different animal models of pain. We will first go over the mechanisms of pain development and evaluate which types of fibers are activated after an injury. In the light of experimental results, we will propose some hypotheses explaining the mitigated results obtained in clinical studies on chronic postoperative pain. Finally, we will discuss three major disadvantages of the current blockade: the absence of blockade of myelinated fibers, the inappropriate duration of blockade, and the existence of activity-independent mechanisms.

5.
Handb Exp Pharmacol ; (177): 359-89, 2007.
Article in English | MEDLINE | ID: mdl-17087130

ABSTRACT

Pathological pain or clinical pain refers to tissue injury-induced inflammatory pain and nerve injury-induced neuropathic pain and is often chronic. Pathological pain is an expression of neural plasticity that occurs both in the peripheral nervous system (e.g., primary sensory nociceptors), termed peripheral sensitization, and in the central nervous system (e.g., dorsal horn and brain neurons), termed central sensitization. Our insufficient understanding of mechanisms underlying the induction and maintenance of injury-induced neuronal plasticity hinders successful treatment for pathological pain. The human genome encodes 518 protein kinases, representing one of the largest protein families. There is growing interest in developing protein kinase inhibitors for the treatment of a number of diseases. Although protein kinases were not favored as targets for analgesics, studies in the last decade have demonstrated important roles of these kinases in regulating neuronal plasticity and pain sensitization. Multiple protein kinases have been implicated in peripheral and central sensitization following intense noxious stimuli and injuries. In particular, mitogen-activated protein kinases (MAPKs), consisting of extracellular signal-regulated kinase (ERK), p38, and c-Jun N-terminal kinase (JNK), are downstream to many kinases and are activated in primary sensory and dorsal horn neurons by nociceptive activity, growth factors and inflammatory mediators, contributing to the induction and maintenance of pain sensitization via posttranslational, translational, and transcriptional regulation. MAPKs are also activated in spinal glial cells (microglia and astrocytes) after injuries, leading to the synthesis of inflammatory mediators/neuroactive substances that act on nociceptive neurons, enhancing and prolonging pain sensitization. Inhibition of multiple kinases has been shown to attenuate inflammatory and neuropathic pain in different animal models. Development of specific inhibitors for protein kinases to target neurons and glial cells will shed light on the development of new therapies for debilitating chronic pain.


Subject(s)
Pain/drug therapy , Pain/enzymology , Protein Kinases/drug effects , Animals , Central Nervous System/drug effects , Central Nervous System/enzymology , Humans , Neuronal Plasticity/physiology , Pain/etiology , Pain/physiopathology , Peripheral Nervous System/drug effects , Peripheral Nervous System/enzymology
6.
Neurobiol Dis ; 8(1): 1-10, 2001 Feb.
Article in English | MEDLINE | ID: mdl-11162235

ABSTRACT

Pathological pain, consisting of tissue injury-induced inflammatory and nerve injury-induced neuropathic pain, is an expression of neuronal plasticity. One component of this is that the afferent input generated by injury and intense noxious stimuli triggers an increased excitability of nociceptive neurons in the spinal cord. This central sensitization is an activity-dependent functional plasticity that results from activation of different intracellular kinase cascades leading to the phosphorylation of key membrane receptors and channels, increasing synaptic efficacy. Central sensitization is both induced and maintained in a transcription-independent manner. Several different intracellular signal transduction cascades converge on MAPK (mitogen-activated protein kinase), activation of which appears to be a master switch or gate for the regulation of central sensitization. In addition to posttranslational regulation, the MAPK pathway may also regulate long-term pain hypersensitivity, via transcriptional regulation of key gene products. Pharmacological intervention targeted specifically at the signal transduction pathways in nociceptive neurons may provide, therefore, new therapeutic opportunities for pathological pain.


Subject(s)
Nervous System Diseases/physiopathology , Neuronal Plasticity/physiology , Neurons/physiology , Nociceptors/physiology , Pain/physiopathology , Signal Transduction/physiology , Animals , Humans
7.
Nat Neurosci ; 2(12): 1114-9, 1999 Dec.
Article in English | MEDLINE | ID: mdl-10570489

ABSTRACT

We investigated the involvement of extracellular signal-regulated protein kinases (ERK) within spinal neurons in producing pain hypersensitivity. Within a minute of an intense noxious peripheral or C-fiber electrical stimulus, many phosphoERK-positive neurons were observed, most predominantly in lamina I and IIo of the ipsilateral dorsal horn. This staining was intensity and NMDA receptor dependent. Low-intensity stimuli or A-fiber input had no effect. Inhibition of ERK phosphorylation by a MEK inhibitor reduced the second phase of formalin-induced pain behavior, a measure of spinal neuron sensitization. ERK signaling within the spinal cord is therefore involved in generating pain hypersensitivity. Because of its rapid activation, this effect probably involves regulation of neuronal excitability without changes in transcription.


Subject(s)
Mitogen-Activated Protein Kinases/metabolism , Nociceptors/physiology , Pain Threshold , Pain/physiopathology , Posterior Horn Cells/enzymology , Protein Serine-Threonine Kinases , Animals , Capsaicin/antagonists & inhibitors , Capsaicin/pharmacology , Enzyme Activation/drug effects , Formaldehyde/antagonists & inhibitors , Formaldehyde/pharmacology , Hindlimb , In Vitro Techniques , MAP Kinase Kinase 1 , Male , Mitogen-Activated Protein Kinase Kinases/antagonists & inhibitors , Mitogen-Activated Protein Kinase Kinases/metabolism , Pain/chemically induced , Pain Measurement , Pain Threshold/drug effects , Phosphorylation/drug effects , Rats , Rats, Sprague-Dawley , Receptors, N-Methyl-D-Aspartate/antagonists & inhibitors , Receptors, N-Methyl-D-Aspartate/physiology , Signal Transduction/drug effects , Temperature
8.
Proc Natl Acad Sci U S A ; 96(16): 9385-90, 1999 Aug 03.
Article in English | MEDLINE | ID: mdl-10430952

ABSTRACT

Brain-derived neurotrophic factor (BDNF) is expressed in nociceptive sensory neurons and transported anterogradely to the dorsal horn of the spinal cord where it is located in dense core vesicles in C-fiber terminals. Peripheral inflammation substantially up-regulates BDNF mRNA and protein in the dorsal root ganglion (DRG) in a nerve growth factor-dependent fashion and results in novel expression of BDNF by DRG neurons with myelinated axons. C-fiber electrical activity also increases BDNF expression in the DRG, and both inflammation and activity increase full-length TrkB receptor levels in the dorsal horn. Sequestration of endogenous BDNF/neurotrophin 4 by intraspinal TrkB-Fc fusion protein administration does not, in noninflamed animals, change basal pain sensitivity nor the mechanical hypersensitivity induced by peripheral capsaicin administration, a measure of C fiber-mediated central sensitization. TrkB-Fc administration also does not modify basal inflammatory pain hypersensitivity, but does block the progressive hypersensitivity elicited by low-intensity tactile stimulation of inflamed tissue. BDNF, by virtue of its nerve growth factor regulation in sensory neurons including novel expression in A fibers, has a role as a central modulator of tactile stimulus-induced inflammatory pain hypersensitivity.


Subject(s)
Brain-Derived Neurotrophic Factor/genetics , Gene Expression Regulation , Inflammation/physiopathology , Neurons, Afferent/physiology , Pain/physiopathology , Spinal Cord/physiology , Animals , Axonal Transport , Decerebrate State , Electric Stimulation , Ganglia, Spinal/physiology , Ganglia, Spinal/physiopathology , In Situ Hybridization , Male , Motor Neurons/physiology , Nerve Fibers/physiology , Physical Stimulation , Protein Biosynthesis , RNA, Messenger/genetics , Rats , Rats, Sprague-Dawley , Receptor Protein-Tyrosine Kinases/genetics , Receptor Protein-Tyrosine Kinases/physiology , Receptor, Ciliary Neurotrophic Factor , Receptors, Nerve Growth Factor/genetics , Receptors, Nerve Growth Factor/physiology , Recombinant Fusion Proteins/pharmacology , Spinal Cord/physiopathology , Time Factors , Touch , Transcription, Genetic
9.
Proc Natl Acad Sci U S A ; 95(26): 15635-40, 1998 Dec 22.
Article in English | MEDLINE | ID: mdl-9861022

ABSTRACT

Repetitive transcranial magnetic stimulation (rTMS) is a noninvasive technique to induce electric currents in the brain. Although rTMS is being evaluated as a possible alternative to electroconvulsive therapy for the treatment of refractory depression, little is known about the pattern of activation induced in the brain by rTMS. We have compared immediate early gene expression in rat brain after rTMS and electroconvulsive stimulation, a well-established animal model for electroconvulsive therapy. Our result shows that rTMS applied in conditions effective in animal models of depression induces different patterns of immediate-early gene expression than does electroconvulsive stimulation. In particular, rTMS evokes strong neural responses in the paraventricular nucleus of the thalamus (PVT) and in other regions involved in the regulation of circadian rhythms. The response in PVT is independent of the orientation of the stimulation probe relative to the head. Part of this response is likely because of direct activation, as repetitive magnetic stimulation also activates PVT neurons in brain slices.


Subject(s)
Brain/metabolism , Electromagnetic Fields , Genes, fos , Animals , Brain/radiation effects , Circadian Rhythm , Genes, fos/radiation effects , Light , Male , Organ Specificity , Pineal Gland/metabolism , Pineal Gland/radiation effects , Proto-Oncogene Proteins c-fos/genetics , Rats , Rats, Sprague-Dawley , Retina/metabolism , Retina/radiation effects , Transcription, Genetic
10.
Neuropeptides ; 32(5): 447-52, 1998 Oct.
Article in English | MEDLINE | ID: mdl-9845006

ABSTRACT

We examined the effects of intrathecal (i.t.) administration of neuropeptide Y (NPY) on the excitability of the flexor reflex in normal rats and 24 h after inflammation induced by subcutaneous carrageenan. In normal rats, i.t. NPY at low doses (10 and 100 ng) caused a brief facilitation of the flexor reflex with no subsequent depression. At higher doses (1 and 10 microg), the effect of NPY was mainly inhibitory, causing substantial and usually prolonged depression of the flexor reflex. At 24 h after the injection of carrageenan, when inflammation was at its peak, the magnitude of the reflex was increased and discharge duration became prolonged. I.t. NPY produced similar pattern of dose-dependent facilitatory and depressive effects on the flexor reflex. The facilitatory effect of i.t. NPY, particularly for the higher doses, was significantly enhanced in inflamed rats compared to normals. In contrast, the depressive effect of high doses of i.t. NPY was unchanged. These data suggest that the changes in levels of NPY and NPY receptors in the spinal cord known to occur after inflammation, are associated with an increased excitatory effect of this peptide.


Subject(s)
Carrageenan/pharmacology , Inflammation/physiopathology , Neuropeptide Y/pharmacology , Reflex/drug effects , Analysis of Variance , Animals , Carrageenan/administration & dosage , Dose-Response Relationship, Drug , Electric Stimulation , Electromyography , Evoked Potentials, Motor/drug effects , Female , Hindlimb , Inflammation/chemically induced , Injections, Spinal , Injections, Subcutaneous , Male , Neuropeptide Y/administration & dosage , Rats , Rats, Sprague-Dawley
11.
J Neurosci ; 18(23): 9695-702, 1998 Dec 01.
Article in English | MEDLINE | ID: mdl-9822730

ABSTRACT

The synaptic basal lamina protein agrin is essential for the formation of neuromuscular junctions. Agrin mediates the postsynaptic clustering of acetylcholine receptors and regulates transcription in muscles. Agrin expression is not restricted to motor neurons but can be demonstrated throughout the CNS. The functional significance of agrin expression in neurons other than motor neurons is unknown. To test whether agrin triggers responses in neurons that lead to the activation of transcription factors, we have analyzed phosphorylation of the transcriptional regulatory site serine 133 of the transcription factor CREB (cAMP response element binding protein) in primary hippocampal neurons. Our results indicate that the neuronal (Ag4,8), but not the non-neuronal (Ag0,0), isoform of agrin induces CREB phosphorylation in hippocampal neurons. The kinetics of agrin- and BDNF-induced CREB phosphorylation are similar: peak levels are reached in minutes and are strongly reduced 2 hr later. Neuronal responses to agrin require extracellular calcium, and, in contrast to tyrosine kinase inhibitors, the specific inhibition of protein kinase A (PKA) does not affect agrin-evoked CREB phosphorylation. Our results show that hippocampal neurons specifically respond to neuronal agrin in a Ca2+-dependent manner and via the activation of tyrosine kinases.


Subject(s)
Agrin/metabolism , Cyclic AMP/metabolism , Hippocampus/cytology , Neurons/enzymology , Agrin/chemistry , Animals , Brain-Derived Neurotrophic Factor/pharmacology , Calcium/pharmacology , Cells, Cultured , Colforsin/pharmacology , Cyclic AMP/analogs & derivatives , Cyclic AMP/pharmacology , Cyclic AMP Response Element-Binding Protein/metabolism , Cyclic AMP-Dependent Protein Kinases/metabolism , Enzyme Inhibitors/pharmacology , Hippocampus/enzymology , Isomerism , Neurons/chemistry , Neurons/cytology , Phosphorylation , Protein-Tyrosine Kinases/metabolism , Rats , Rats, Sprague-Dawley , Signal Transduction/drug effects , Synapses/chemistry , Synapses/drug effects , Synapses/enzymology , Thionucleotides/pharmacology
12.
J Neurosci ; 17(5): 1776-85, 1997 Mar 01.
Article in English | MEDLINE | ID: mdl-9030636

ABSTRACT

The involvement of cAMP-responsive element-binding protein (CREB) signaling in tissue injury-induced inflammation and hyperalgesia has been characterized by measuring phosphorylation of CREB at serine-133 (CREB Ser133) using a specific antibody. In the unstimulated state, unphosphorylated CREB was observed in most nuclei of spinal neurons except for motor neurons, where only a small portion of neurons were stained. A few dorsal root ganglion (DRG) neurons were also CREB-positive. After a unilateral injection of formalin into the hindpaw, a strong and bilateral phosphorylation of CREB Ser133 was induced, as assessed by both immunohistochemistry and Western blot. PhosphoCREB (pCREB)-positive neurons were found in laminae I, II, V, and X of spinal cord on both sides. CREB phosphorylation was very rapid and reached peak levels within 10 min of formalin treatment, whereas few pCREB-positive neurons were seen in unstimulated spinal cord. The induction of pCREB was predominantly postsynaptic, because only 5% of DRG neurons were labeled after inflammation. In contrast to CREB phosphorylation, the induction of c-Fos expression reached peak levels 2 hr after formalin treatment and c-Fos induction was mainly ipsilateral. Both formalin-evoked CREB phosphorylation and c-Fos expression in the spinal cord were suppressed by pretreatment with the NMDA receptor antagonist MK-801 (3.5 mg/kg, i.p.) or halothane anesthesia. These results suggest that CREB signaling may play a role in the long-term facilitation of spinal cord neurons after hyperalgesia. Furthermore, our results indicate that CREB phosphorylation may be necessary but not sufficient for c-fos induction.


Subject(s)
Cyclic AMP Response Element-Binding Protein/metabolism , Gene Expression Regulation/drug effects , Genes, fos , Hyperalgesia/metabolism , Protein Processing, Post-Translational/drug effects , Spinal Cord/metabolism , Animals , Dizocilpine Maleate/pharmacology , Excitatory Amino Acid Antagonists/pharmacology , Formaldehyde/toxicity , Halothane/pharmacology , Hyperalgesia/chemically induced , Hyperalgesia/physiopathology , Male , Phosphorylation/drug effects , Protein Kinases/metabolism , Proto-Oncogene Proteins c-fos/biosynthesis , Rats , Rats, Sprague-Dawley , Signal Transduction/physiology , Spinal Cord/physiopathology
13.
Regul Pept ; 66(3): 179-89, 1996 Oct 22.
Article in English | MEDLINE | ID: mdl-8916273

ABSTRACT

Using immunohistochemistry and in situ hybridization the in vivo effects of acidic and basic fibroblast growth factor (aFGF, bFGF), and of nerve growth factor (NGF) on the expression of galanin, neuropeptide Y (NPY) and substance P in axotomized dorsal root ganglia (DRGs) were examined. Self-mutilation (autotomy), a supposed pain-related behavior, was investigated after growth factor treatment. One microgram of aFGF, bFGF or NGF was applied directly to the transected sciatic nerve via a capsule. In normal rats 3.2%, 0% and 17.5% of the neuron profiles in the DRGs contained galanin-, NPY- and substance P-like immunoreactivity (LI), respectively. Sciatic nerve transection induced a distinct increase in galanin- and NPY-LIs, but a downregulation of substance P-LI. Thus three days after axotomy 23.5%, 26.9% and 9.8% of the DRG neuron profiles showed immunoreactivity for galanin-, NPY- and substance P-LI, respectively. In vivo administration of aFGF counteracted the axotomy-induced increase in galanin and NPY, whereas bFGF only suppressed NPY upregulation. NGF reversed in the injury-induced decrease in substance P-LI, but had no significant effect on galanin- and NPY-LIs. These results were confirmed by monitoring the mRNA levels for these neuropeptides. Moreover, aFGF was found to induce autotomy in 60% of the rats 3 days after axotomy. NGF produced autotomy in about 30% of the rats. Taken together, the present results suggest (1) that aFGF, bFGF and NGF differentially regulate neuropeptide expression in vivo; (2) that FGFs can inhibit neuropeptide upregulation of some peptides after nerve injury; and (3) that aFGF and NGF may induce pain-related behavior.


Subject(s)
Fibroblast Growth Factor 1/pharmacology , Fibroblast Growth Factor 2/pharmacology , Galanin/biosynthesis , Ganglia, Spinal/metabolism , Gene Expression Regulation/drug effects , Nerve Growth Factors/pharmacology , Neuropeptide Y/biosynthesis , Substance P/biosynthesis , Animals , Ganglia, Spinal/pathology , Nerve Crush , RNA, Messenger/analysis , Rats , Rats, Sprague-Dawley
14.
Exp Brain Res ; 111(3): 393-404, 1996 Oct.
Article in English | MEDLINE | ID: mdl-8911933

ABSTRACT

Using immunohistochemistry and in situ hybridization, the expression of galanin (GAL)/galanin message associated peptide (GMAP)-, neuropeptide Y (NPY)-, vasoactive intestinal polypeptide (VIP)/peptide histidine isoleucine (PHI)- and nitric oxide synthase (NOS)-like immunoreactivities and mRNAs, and NPY receptor mRNA was studied in normal trigeminal and nodose ganglia and 14 and 42 days after peripheral axotomy. In normal trigeminal ganglia about 11% of the counted neuron profiles contained GAL mRNA, 4% NOS mRNA, 5% NPY mRNA, 7% VIP mRNA, and 19% NPY receptor mRNA. Peptide mRNA- and NPY receptor mRNA-positive neuron profiles were small in size. Fourteen days after axotomy a marked increase in the number of GAL mRNA-(34% of counted neuron profiles), NPY mRNA-(54%) and VIP mRNA-(31%) positive neuron profiles, and a moderate increase in the number of NOS mRNA-(22%) positive neuron profiles were observed in the ipsilateral trigeminal ganglia. The GAL/GMAP, VIP- and NOS-positive profiles were mainly small, the NPY-positive ones mostly large. NPY receptor mRNA was expressed in some large neurons. In normal nodose ganglia, about 3% of the counted neuron profiles contained GAL mRNA, 3% NPY mRNA, 17% NOS mRNA and less than 1% VIP mRNA. Fourteen days after peripheral axotomy, a marked increase in the number of GAL mRNA-(78% of counted neuron profiles), NOS mRNA-(37%) and VIP-(46%) mRNA-positive neuron profiles was seen in the ipsilateral nodose ganglia. The number of NPY-positive (23%) neurons was moderately increased, mainly in small neuron profiles. There were no NPY receptor mRNA-positive neurons, either in normal nodose ganglia or in nodose ganglia ipsilateral to the axotomy. In contralateral nodose ganglia the number of GAL- and NPY-positive neuron profiles was slightly increased, and VIP cells showed a moderate increase. Immunohistochemical analysis revealed parallel changes in expression of peptides and NOS in both trigeminal and nodose ganglia, demonstrating that the changes in mRNA levels are translated into protein. Finally, although not quantified, similar upregulations of peptide and NOS mRNA levels were observed in both ganglia 42 days after nerve injury provided that regeneration was not allowed, suggesting that the changes are long lasting. The present results show that the effect of axotomy on peptide and NOS expression in the trigeminal and nodose ganglia is similar to that previously shown for lumbar dorsal root ganglia. However, no mRNA for the NPY Y1 receptor could be detected in the vagal system. In general the mechanism(s) for and the purpose(s) of the messenger regulation in response to axotomy may be similar in these different sensory systems (dorsal root, trigeminal and nodose ganglia).


Subject(s)
Axons/physiology , Neuropeptides/analysis , Nitric Oxide Synthase/analysis , Nodose Ganglion/chemistry , Receptors, Neuropeptide Y/analysis , Trigeminal Ganglion/chemistry , Animals , Functional Laterality/physiology , Immunohistochemistry , In Situ Hybridization , Male , Rats , Rats, Sprague-Dawley
15.
Brain Res ; 721(1-2): 167-73, 1996 May 20.
Article in English | MEDLINE | ID: mdl-8793097

ABSTRACT

Using the indirect immunofluorescence technique, the distribution of Ca2+/calmodulin-dependent protein kinase IV (CaM kinase IV) was studied in dorsal root ganglia (DRGs) and the sciatic nerve under normal circumstances and after axotomy and nerve ligation. CaM kinase IV-like immunoreactivity (-LI) was observed mainly in small DRG neurons but also in some large ones with the immunoreactivity mainly confined to the cell nuclei and with varying levels in the cytoplasm. CaM kinase IV-LI was present in around 1/4 of all CGRP-positive neurons and in the vast majority of the somatostatin-positive neurons. The enzyme levels decreased markedly after axotomy. The enzyme was also observed in axons in the sciatic nerve and accumulated both proximal and distal to a ligation. The present results suggest that CaM kinase is not of direct importance for upregulation of neuropeptides in DRG neurons after nerve injury. In addition to a nuclear function it may also play a role in the peripheral processes of DRG neurons.


Subject(s)
Axonal Transport/physiology , Axons/physiology , Calcium-Calmodulin-Dependent Protein Kinases/metabolism , Ganglia, Spinal/enzymology , Neuropeptides/metabolism , Animals , Calcitonin Gene-Related Peptide/metabolism , Calcium-Calmodulin-Dependent Protein Kinase Type 4 , Cytoplasm/enzymology , Fluorescent Antibody Technique, Indirect , Male , Neuronal Plasticity/physiology , Rats , Rats, Sprague-Dawley , Sciatic Nerve/cytology , Sciatic Nerve/enzymology , Up-Regulation/physiology
16.
J Neurosci ; 15(12): 8156-66, 1995 Dec.
Article in English | MEDLINE | ID: mdl-8613750

ABSTRACT

Recently, antisera that recognize unique epitopes of the cloned mu-, delta-, and kappa-opioid, receptors (MOR, DOR, KOR, respectively) have been developed. In the present study MOR-, DOR-, and KOR-like immunoreactivities (LIs) were examined in rat dorsal root ganglia (DRGs, L4-5) after injection of carrageenan (CAR) into the hindpaw. In normal control rats 20.9%, 13.5%, and 9% of the DRG neurons contained MOR-, DOR-, KOR-LI, respectively. A marked upregulation in MOR-LI was observed in DRG neurons 1 and 3 d after inflammation. In contrast, CAR induced a distinct downregulation in DOR- and KOR-LIs. MOR-, DOR-, and KOR-LIs were preferentially localized in small DRG neurons. MOR-LI was often located in patches in the cytoplasm, and in some cells close to the somatic plasmalemma. However, DOR- and KOR-LIs mainly showed a diffuse staining pattern within cytoplasm. Two or even all three receptors could sometimes be found to coexist in DRG neurons. In the spinal cord, these receptors were mainly confined to the superficial dorsal horn, with a somewhat diffuse staining which was strong for MOR-LI, and weak for KOR-LI. DOR-LI had distinctly punctate, varicose distribution. CAG induced-alterations in opioid receptor staining in spinal cord were much less pronounced than those in the DRGs with a small increase in MOR-LI and a slight decrease in DOR-LI ipsilaterally. There was an accumulation of all three types of receptors in the sciatic nerve both proximal and distal to the ligation site as early as 2 hr, indicating both antero- and retrograde transport of multiple opioid receptors. However, DOR-LI accumulation was stronger than that of MOR- and KOR-LIs. Taken together, these results suggest that all three opioid receptors are involved in the response to inflammation and that they may play different roles in this pathological state. The coexistence of MOR, DOR, and KOR in at least some primary sensory neurons provides a substrate for functional interactions between these receptors.


Subject(s)
Ganglia, Spinal/metabolism , Neuritis/metabolism , Receptors, Opioid, delta/metabolism , Receptors, Opioid, kappa/metabolism , Receptors, Opioid, mu/metabolism , Animals , Axonal Transport , Carrageenan , Immunohistochemistry , Male , Neuritis/chemically induced , Rats , Rats, Sprague-Dawley , Spinal Cord/metabolism , Tissue Distribution
17.
Eur J Neurosci ; 7(12): 2458-68, 1995 Dec 01.
Article in English | MEDLINE | ID: mdl-8845951

ABSTRACT

Using quantitative in situ hybridization and immunohistochemistry the expression of acidic and basic fibroblast growth factors (aFGF, bFGF) in dorsal root ganglia (DRGs) was examined. Around 5% of the small neurons expressed bFGF mRNA in normal DRGs. Nerve injury induced a very dramatic and rapid up-regulation in bFGF mRNA levels, and around 80% of all DRG neurons expressed bFGF mRNA 3 days after axotomy. A distinct increase in bFGF-like immunoreactivity (LI) was also detected as early as 15 h after axotomy. The elevation of bFGF mRNA and protein levels declined after 1 week. bFGF mRNA was also up-regulated in non-neuronal cells following axotomy. Normally bFGF-LI was mainly localized in the nuclei of DRG neurons and in some non-neuronal cells. After nerve section, bFGF-LI was in addition found in the cytoplasm, and many more bFGF-positive non-neuronal cells were observed. By means of confocal microscopy analysis of axotomized DRGs, some bFGF-LI could be detected in vesicle-like structures in the cytoplasm as well as in the nucleoli, in addition to the nuclear location. Application of leukaemia inhibitory factor to the transected sciatic nerve significantly increased the number of bFGF-positive neurons, whereas the bFGF-LI in non-neuronal cells was strongly suppressed. About 70% of the normal DRG neurons expressed aFGF mRNA and aFGF-LI. Axotomy produced a moderate increase in aFGF mRNA levels, but no detectable effect on protein levels. Taken together, the results show that bFGF may be involved in the neuronal response to injury and suggest a role in neuronal survival and regeneration in axotomized DRG neurons.


Subject(s)
Axons/physiology , Fibroblast Growth Factor 2/metabolism , Ganglia, Spinal/metabolism , Animals , Fibroblast Growth Factor 2/physiology , Immunohistochemistry , In Situ Hybridization , Male , Rats , Rats, Sprague-Dawley , Time Factors
18.
Brain Res ; 705(1-2): 149-58, 1995 Dec 24.
Article in English | MEDLINE | ID: mdl-8821745

ABSTRACT

Pituitary adenylate cyclase-activating polypeptide (PACAP) has recently been demonstrated in sensory neurons. In the present study on rat 17.5% of all neurons, mainly of small size, contained PACAP in normal dorsal root ganglia (DRGs). Transection of the sciatic nerve induced a rapid and strong upregulation in PACAP peptide and mRNA levels which could be seen already after 15 h. After 3 days more than 51.5% of neurons of different sizes expressed PACAP. However, the intensity of PACAP-LI in the DRG neurons declined after 10 days. Thirty days after axotomy, 56.7% of the DRG neurons still expressed PACAP, but with a low intensity, in fact even lower than in normal controls. No VIP- or NPY-positive neurons were observed in normal or axotomized DRGs at 15 h. However a distinct increase in VIP and NPY levels were seen 3 days after the lesion, and their levels were considerably higher after 30 days. PACAP was often present in neurons expressing VIP, NPY and/or galanin. Thus, 3 days after injury, PACAP was present in 84.4%, 95.7%, and 76.8% of the VIP-, NPY-, and galanin-positive neurons, respectively. PACAP was also found in nerve fibers in control sciatic nerves. After nerve ligation, accumulation of PACAP was seen mainly proximal to the injury but also distally, suggesting both anterograde and retrograde transport of the peptide. Also a moderate increase (about 20%) in PACAP levels was found in the superficial spinal dorsal horn 3 days after nerve transection. Taken together, our results suggest that PACAP is involved in the response to nerve injury. The very high levels of expression in different populations of DRG neurons after axotomy, and its different time course as compared to galanin, NPY and VIP indicate that it may play a complementary and/or different role than these peptides in the adaptation to nerve injury, especially in its early phase.


Subject(s)
Ganglia, Spinal/physiology , Neuropeptides/genetics , Neurotransmitter Agents/genetics , Animals , Axonal Transport/physiology , Denervation , Galanin/analysis , Galanin/genetics , Ganglia, Spinal/cytology , Gene Expression/physiology , Immunohistochemistry , In Situ Hybridization , Male , Neuronal Plasticity/physiology , Neurons, Afferent/physiology , Neurons, Afferent/ultrastructure , Neuropeptide Y/analysis , Neuropeptide Y/genetics , Neuropeptides/analysis , Pituitary Adenylate Cyclase-Activating Polypeptide , RNA, Messenger/analysis , Rats , Rats, Sprague-Dawley , Sciatic Nerve/cytology , Sciatic Nerve/metabolism , Sciatic Nerve/surgery , Time Factors , Vasoactive Intestinal Peptide/analysis , Vasoactive Intestinal Peptide/genetics
19.
Neuroscience ; 68(4): 1219-27, 1995 Oct.
Article in English | MEDLINE | ID: mdl-8544995

ABSTRACT

The effects of moderate and high intrathecal doses of clonidine, an alpha 2 adrenoceptor agonist, or a high dose of morphine on sciatic nerve section-induced expression of c-Fos-like immunoreactivity was studied in laminae I and II of the dorsal horn and laminae VIII and IX of the ventral horn of rat lumbar spinal cord. c-Fos-like immunoreactivity was examined by immunohistochemistry in normal rats (group 1), rats implanted with an intrathecal catheter with its tip on the lumbar spinal cord (group 2), injected with 10 micrograms (group 3) or 50 micrograms (group 4) clonidine intrathecally 3 h before being killed. In other groups, saline, 10 or 50 micrograms clonidine or 30 micrograms morphine was injected 1 h before unilateral nerve section, and the expression of c-Fos-like immunoreactivity was examined 2 h after axotomy. Few labeled neurons were found in normal controls. The intrathecal catheter itself caused a significant increase in bilateral c-Fos-like immunoreactivity in spinal dorsal and ventral horn compared to normals. The level of c-Fos-like immunoreactivity after 10 or 50 micrograms intrathecal clonidine was similar as in the intrathecal catheter group. Sciatic nerve section caused a significant ipsilateral increase in c-Fos-like immunoreactivity in the dorsal horn compared to the intact side in rats injected with saline. Pretreatment with 10 or 10 micrograms clonidine did not reduce sciatic nerve section-induced expression of c-Fos-like immunoreactivity, but instead caused a significant bilateral increase in c-Fos-like immunoreactivity.(ABSTRACT TRUNCATED AT 250 WORDS)


Subject(s)
Adrenergic alpha-Agonists/pharmacology , Clonidine/pharmacology , Morphine/pharmacology , Narcotics/pharmacology , Proto-Oncogene Proteins c-fos/biosynthesis , Sciatic Nerve/physiology , Spinal Cord/metabolism , Adrenergic alpha-Agonists/administration & dosage , Animals , Clonidine/administration & dosage , Immunohistochemistry , Injections, Spinal , Male , Morphine/administration & dosage , Narcotics/administration & dosage , Rats , Rats, Sprague-Dawley , Spinal Cord/drug effects
20.
Neuroscience ; 68(2): 563-76, 1995 Sep.
Article in English | MEDLINE | ID: mdl-7477966

ABSTRACT

Using in situ hybridization, immunohistochemistry and receptor binding methodology, the galanin messenger RNA levels, galanin binding and galanin-like immunoreactivity were examined in rats injected with carrageenan into the left hindpaw. Three days after injection, a distinct increase (63%) in galanin messenger RNA-positive neurons was observed in the medial laminae I and II of the ipsilateral dorsal horn (lumbar 4 and 5) as compared to the contralateral side. However, no alteration was found in galanin binding and galanin-like immunoreactivity in the dorsal horn. In dorsal root ganglia (lumbar 5), inflammation induced a significant decrease in galanin messenger RNA (39%) and galanin peptide (47%) on the ipsilateral side. Galanin binding was not detected in dorsal root ganglia, neither on the inflammatory nor on the control side. Increased levels of galanin-like immunoreactivity and galanin messenger RNA were seen in cells in the inflamed dermis and epidermis, especially in stratum granulosum. Most of the galanin-immunoreactive cells contained ED1-like immunoreactivity, a marker for macrophages. A strong galanin binding was seen in the inflamed dermis. Such binding sites may be targets for galanin released from local cells in inflamed dermis. Taken together, our results suggest that both neuronal and non-neuronal galanin or a galanin-like peptide is involved in the response to inflammation.


Subject(s)
Central Nervous System/metabolism , Galanin/biosynthesis , Inflammation/metabolism , Peripheral Nerves/metabolism , Animals , Autoradiography , Carrageenan , Ganglia, Spinal/metabolism , Immunohistochemistry , In Situ Hybridization , Inflammation/chemically induced , Male , Microscopy, Confocal , RNA, Messenger/biosynthesis , Rats , Rats, Sprague-Dawley , Skin/metabolism , Spinal Cord/cytology , Spinal Cord/drug effects , Spinal Cord/metabolism
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