Biomedical subjects

Kenji Okuse

Publications and source records attributed to Kenji Okuse.

12 recordsLinked to original sources

PubMed2006

Pain signalling pathways: from cytokines to ion channels.

Pain is the major reason patients seek medical care. The treatment of pain, particularly chronic pain associated with cancer and damage to the nervous system, is at present inadequate. Lack of effective analgesics is partly due to the fact that pain signalling mechanisms are still not fully understood. Over the recent years, many channels, receptors, and regulatory proteins involved in pain pathways have bee identified, and novel pain signalling mechanisms and pathways at peripheral and spinal levels have been discovered. It is anticipated that increased understanding of the molecular mechanisms of pain would provide a hope for the future development of effective pain killers. This review examines the currently available information on the molecular aspects of pain signalling pathways, and discusses novel and promising therapeutic targets for the treatment of pain in humans.

Adenosine Triphosphate↗

PubMed2006

Deletion of annexin 2 light chain p11 in nociceptors causes deficits in somatosensory coding and pain behavior.

The S100 family protein p11 (S100A10, annexin 2 light chain) is involved in the trafficking of the voltage-gated sodium channel Na(V)1.8, TWIK-related acid-sensitive K+ channel (TASK-1), the ligand-gated ion channels acid-sensing ion channel 1a (ASIC1a) and transient receptor potential vanilloid 5/6 (TRPV5/V6), as well as 5-hydroxytryptamine receptor 1B (5-HT1B), a G-protein-coupled receptor. To evaluate the role of p11 in peripheral pain pathways, we generated a loxP-flanked (floxed) p11 mouse and used the Cre-loxP recombinase system to delete p11 exclusively from nociceptive primary sensory neurons in mice. p11-null neurons showed deficits in the expression of Na(V)1.8, but not of annexin 2. Damage-sensing primary neurons from these animals show a reduced tetrodotoxin-resistant sodium current density, consistent with a loss of membrane-associated Na(V)1.8. Noxious coding in wide-dynamic-range neurons in the dorsal horn was markedly compromised. Acute pain behavior was attenuated in certain models, but no deficits in inflammatory pain were observed. A significant deficit in neuropathic pain behavior was also apparent in the conditional-null mice. These results confirm an important role for p11 in nociceptor function.

Animals↗

PubMed2006

Intense isolectin-B4 binding in rat dorsal root ganglion neurons distinguishes C-fiber nociceptors with broad action potentials and high Nav1.9 expression.

Binding to isolectin-B4 (IB4) and expression of tyrosine kinase A (trkA) (the high-affinity NGF receptor) have been used to define two different subgroups of nociceptive small dorsal root ganglion (DRG) neurons. We previously showed that only nociceptors have high trkA levels. However, information about sensory and electrophysiological properties in vivo of single identified IB4-binding neurons, and about their trkA expression levels, is lacking. IB4-positive (IB4+) and small dark neurons had similar size distributions. We examined IB4-binding levels in >120 dye-injected DRG neurons with sensory and electrophysiological properties recorded in vivo. Relative immunointensities for trkA and two TTX-resistant sodium channels (Nav1.8 and Nav1.9) were also measured in these neurons. IB4+ neurons were classified as strongly or weakly IB4+. All strongly IB4+ neurons were C-nociceptor type (C-fiber nociceptive or unresponsive). Of 32 C-nociceptor-type neurons examined, approximately 50% were strongly IB4+, approximately 20% were weakly IB4+ and approximately 30% were IB4-. Adelta low-threshold mechanoreceptive (LTM) neurons were weakly IB4+ or IB4-. All 33 A-fiber nociceptors and all 44 Aalpha/beta-LTM neurons examined were IB4-. IB4+ compared with IB4- C-nociceptor-type neurons had longer somatic action potential durations and rise times, slower conduction velocities, more negative membrane potentials, and greater immunointensities for Nav1.9 but not Nav1.8. Immunointensities of IB4 binding in C-neurons were positively correlated with those of Nav1.9 but not Nav1.8. Of 23 C-neurons tested for both trkA and IB4, approximately 35% were trkA+/IB4+ but with negatively correlated immunointensities; 26% were IB4+/trkA-, and 35% were IB4-/trkA+. We conclude that strongly IB4+ DRG neurons are exclusively C-nociceptor type and that high Nav1.9 expression may contribute to their distinct membrane properties.

Action Potentials↗

PubMed2005

Annexin II light chain p11 promotes functional expression of acid-sensing ion channel ASIC1a.

Acid-sensing ion channels (ASICs) have been implicated in a wide variety of physiological functions. We have used a rat dorsal root ganglion cDNA library in a yeast two-hybrid assay to identify sensory neuron proteins that interact with ASICs. We found that annexin II light chain p11 physically interacts with the N terminus of ASIC1a, but not other ASIC isoforms. Immunoprecipitation studies confirmed an interaction between p11 and ASIC1 in rat dorsal root ganglion neurons in vivo. Coexpression of p11 and ASIC1a in CHO-K1 cells led to a 2-fold increase in expression of the ion channel at the cell membrane as determined by membrane-associated immunoreactivity and cell-surface biotinylation. Consistent with these findings, peak ASIC1a currents in transfected CHO-K1 cells were up-regulated 2-fold in the presence of p11, whereas ASIC3-mediated currents were unaffected by p11 expression. Neither the pH dependence of activation nor the rates of desensitization were altered by p11, suggesting that its primary role in regulating ASIC1a activity is to enhance cell-surface expression of ASIC1a. These data demonstrate that p11, already known to traffic members of the voltage-gated sodium and potassium channel families as well as transient receptor potential and chloride channels, also plays a selective role in enhancing ASIC1a functional expression.

Acid Sensing Ion Channels↗

PubMed2005

trkA is expressed in nociceptive neurons and influences electrophysiological properties via Nav1.8 expression in rapidly conducting nociceptors.

To test the hypothesis that trkA (the high-affinity NGF receptor) is selectively expressed in nociceptive dorsal root ganglion (DRG) neurons, we examined the intensity of trkA immunoreactivity in single dye-injected rat DRG neurons, the sensory receptor properties of which were identified in vivo with mechanical and thermal stimuli. We provide the first evidence in single identified neurons that strong trkA expression in DRGs is restricted to nociceptive neurons, probably accounting for the profound influence of NGF on these neurons. Furthermore, we demonstrate that trkA expression is as high in rapidly conducting (Aalpha/beta) as in more slowly conducting (Adelta and C) nociceptors. All Aalpha/beta low-threshold mechanoreceptors (LTMs) are trkA negative, although weak but detectable trkA is present in some C and Adelta LTMs. NGF can influence electrophysiological properties of DRG neurons, probably by binding to trkA. We found positive correlations for single identified Aalpha/beta (but not C or Adelta) nociceptors between trkA immunocytochemical intensity and electrophysiological properties typical of nociceptors, namely long action potential and afterhyperpolarization durations and large action potential amplitudes. Furthermore, for Aalpha/beta (notCorAdelta) nociceptors, trkA intensity is inversely correlated with conduction velocity. Similar relationships, again only in Aalpha/beta nociceptors, between electrophysiological properties and trkA expression exist for sodium channel Nav1.8 but not Nav1.9 immunoreactivities. These findings suggest that in Aalpha/beta nociceptors, influences of NGF on expression levels of Nav1.8 are related to, and perhaps limited by, expression levels of trkA. This view is supported by a positive correlation between immuno-intensities of trkA and Nav1.8 in A-fiber, but not C-fiber, nociceptors.

Action Potentials↗

PubMed2004

Identification of binding domains in the sodium channel Na(V)1.8 intracellular N-terminal region and annexin II light chain p11.

The interaction of p11 (annexin II light chain) with the N-terminal domain of Na(V)1.8, a tetrodotoxin-resistant sodium channel, is essential for the functional expression of the channel. Here we show that p11 binds to Na(V)1.8 but not to sodium channel isoforms Na(V)1.2, 1.5, 1.7 or Na(V)1.9. The binding of amino acids 74-103 of Na(V)1.8 to p11 residues 33-78 occurs in a random coiled region flanked by two EF hand motifs whose crystal structure has been established. As Na(V)1.8 channel expression is associated with pain pathways, drugs that disrupt the Na(V)1.8-p11 interaction and down-regulate channel expression may have analgesic activity.

Amino Acid Sequence↗

PubMed2004

Voltage-gated sodium channels and pain pathways.

Acute, inflammatory, and neuropathic pain can all be attenuated or abolished by local treatment with sodium channel blockers such as lidocaine. The peripheral input that drives pain perception thus depends on the presence of functional voltage-gated sodium channels. Remarkably, two voltage-gated sodium channel genes (Nav1.8 and Nav1.9) are expressed selectively in damage-sensing peripheral neurons, while a third channel (Nav1.7) is found predominantly in sensory and sympathetic neurons. An embryonic channel (Nav1.3) is also upregulated in damaged peripheral nerves and associated with increased electrical excitability in neuropathic pain states. A combination of antisense and knock-out studies support a specialized role for these sodium channels in pain pathways, and pharmacological studies with conotoxins suggest that isotype-specific antagonists should be feasible. Taken together, these data suggest that isotype-specific sodium channel blockers could be useful analgesics.

Amino Acid Sequence↗

PubMed2004

Ion channel activities implicated in pathological pain.

Altered expression of voltage-gated sodium, calcium and potassium channels has been associated with neuropathic pain conditions. In addition, roles for the ligand-gated P2X3 and NMDA receptors, as well as pacemaker HCN channels have also been invoked in the pathogenesis of neuropathic pain. In this chapter, evidence of an important role for post-translational regulation of Nav1.9 in setting pain thresholds is presented. Despite the importance of tactile allodynia and mechanical hyperalgesia in chronic pain, we remain ignorant of the molecular nature of mechanosensors present in sensory neurons. A number of candidate mechanosensor genes, identified because of their structural similarity with mechanosensors in Caenorbabditis elegans and Drosophila melanogaster have been identified. Acid-sensing ion channels (ASICs) are structurally related to putative mechanosensors in C. elegans, whilst transient receptor potential channels (TRPs) have been implicated in mechanosensation in the Drosophila acoustic system. Evidence against a role for ASICs as primary transducers of mechanosensation is provided here, and recent evidence implicating TRP channels is reviewed. Finally, the use of sensory neuron-specific gene deletion approaches to unravel the significance of individual ion channels in the regulation of sensory neuron excitability and the induction of pain will be described.

Animals↗

PubMed2003

The TTX-resistant sodium channel Nav1.8 (SNS/PN3): expression and correlation with membrane properties in rat nociceptive primary afferent neurons.

We have examined the distribution of the sensory neuron-specific Na+ channel Nav1.8 (SNS/PN3) in nociceptive and non-nociceptive dorsal root ganglion (DRG) neurons and whether its distribution is related to neuronal membrane properties. Nav1.8-like immunoreactivity (Nav1.8-LI) was examined with an affinity purified polyclonal antiserum (SNS11) in rat DRG neurons that were classified according to sensory receptive properties and by conduction velocity (CV) as C-, Adelta- or Aalpha/beta. A significantly higher proportion of nociceptive than low threshold mechanoreceptive (LTM) neurons showed Nav1.8-LI, and nociceptive neurons had significantly more intense immunoreactivity in their somata than LTM neurons. Results showed that 89, 93 and 60% of C-, Adelta- and Aalpha/beta-fibre nociceptive units respectively and 88% of C-unresponsive units were positive. C-unresponsive units had electrical membrane properties similar to C-nociceptors and were considered to be nociceptive-type neurons. Weak positive Nav1.8-LI was also present in some LTM units including a C LTM, all Adelta LTM units (D hair), about 10% of cutaneous LTM Aalpha/beta-units, but no muscle spindle afferent units. Nav1.8-LI intensity was negatively correlated with soma size (all neurons) and with dorsal root CVs in A- but not C-fibre neurons. Nav1.8-LI intensity was positively correlated with action potential (AP) duration (both rise and fall time) in A-fibre neurons and with AP rise time only in positive C-fibre neurons. It was also positively correlated with AP overshoot in positive neurons. Thus high levels of Nav1.8 protein may contribute to the longer AP durations (especially in A-fibre neurons) and larger AP overshoots that are typical of nociceptors.

Action Potentials↗

PubMed2003

Sensory neuron proteins interact with the intracellular domains of sodium channel NaV1.8.

Voltage-gated sodium channels initiate and propagate action potentials in excitable cells. The tetrodotoxin-resistant Na(+) channel (Na(V)1.8/SNS) is expressed in damage-sensing neurons (nociceptors) and plays an important role in pain pathways. Expression of high levels of functional Na(V)1.8 in heterologous cells has proved problematic, even in the presence of known sodium channel accessory beta-subunits. This suggests that other regulatory proteins are required for normal levels of Na(V)1.8 expression. Here we report the use of a yeast two-hybrid system and a rat dorsal root ganglion cDNA library to identify 28 different clones encoding proteins which interact with intracellular domains of Na(V)1.8. Many clones are expressed at high levels in small diameter DRG neurons as judged by in situ hybridization. Interacting proteins include cytoplasmic elements and linker proteins (e.g. beta-actin and moesin), enzymes (e.g. inositol polyphosphate 5-phosphatase and TAO2 thousand and one protein kinase), channels and membrane-associated proteins (voltage-dependent anion channel VDAC3V and tetraspanin), as well as motor proteins (dynein intermediate and light chain) and transcripts encoding previously undescribed proteins. Immunoprecipitation (pull-down) assays confirm that some of the proteins interact with, and may hence regulate, Na(V)1.8 in vivo.

Animals↗

PubMed2002

Annexin II light chain regulates sensory neuron-specific sodium channel expression.

The tetrodotoxin-resistant sodium channel Na(V)1.8/SNS is expressed exclusively in sensory neurons and appears to have an important role in pain pathways. Unlike other sodium channels, Na(V)1.8 is poorly expressed in cell lines even in the presence of accessory beta-subunits. Here we identify annexin II light chain (p11) as a regulatory factor that facilitates the expression of Na(V)1.8. p11 binds directly to the amino terminus of Na(V)1.8 and promotes the translocation of Na(V)1.8 to the plasma membrane, producing functional channels. The endogenous Na(V)1.8 current in sensory neurons is inhibited by antisense downregulation of p11 expression. Because direct association with p11 is required for functional expression of Na(V)1.8, disrupting this interaction may be a useful new approach to downregulating Na(V)1.8 and effecting analgesia.

Animals↗

PubMed2002

Sodium channels in primary sensory neurons: relationship to pain states.

Electrophysiological studies of dorsal root ganglion (DRG) neurons, and the results of PCR, Northern blot and in situ hybridization analyses have demonstrated the molecular diversity of Na+ channels that operate in sensory neurons. Several subtypes of alpha-subunit have been detected in DRG neurons and transcripts encoding all three beta-subunits are also present. Interestingly, one alpha subunit, Na(v)1.8, is selectively expressed in C-fibre and Adelta fibre associated sensory neurons that are predominantly involved in damage sensing. Another channel, Na(v).3, is selectively up regulated in a variety of models of neuropathic pain. In this review we focus on Na+ channels that are selectively expressed in DRG neurons as potential analgesic drug targets. In the absence of subtype specific inhibitors, the production of null mutant mice provides useful information on the specialized functions of particular Na+ channels. A refinement of this approach is to delete Na+ channel genes flanked by lox-P sites in the sensory ganglia of adult animals, using viruses to deliver the bacteriophage Cre recombinase enzyme.

Animals↗