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Nitin Agarwal,
Pal Pacher,
Irmgard Tegeder,
Fumimasa Amaya,
Cristina E Constantin,
Gary J Brenner,
Tiziana Rubino,
Christoph W Michalski,
Giovanni Marsicano,
Krisztina Monory, [......],
Claudiu Marian,
Sandor Batkai,
Daniela Parolaro,
Michael J Fischer,
Peter Reeh,
George Kunos,
Michaela Kress,
Beat Lutz,
Clifford J Woolf,
Rohini Kuner
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Nitin Agarwal,
Pal Pacher,
Irmgard Tegeder,
Fumimasa Amaya,
Cristina E Constantin,
Gary J Brenner,
Tiziana Rubino,
Christoph W Michalski,
Giovanni Marsicano,
Krisztina Monory, [......],
Claudiu Marian,
Sandor Batkai,
Daniela Parolaro,
Michael J Fischer,
Peter Reeh,
George Kunos,
Michaela Kress,
Beat Lutz,
Clifford J Woolf,
Rohini Kuner
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Nitin Agarwal,
Pal Pacher,
Irmgard Tegeder,
Fumimasa Amaya,
Cristina E Constantin,
Gary J Brenner,
Tiziana Rubino,
Christoph W Michalski,
Giovanni Marsicano,
Krisztina Monory, [......],
Claudiu Marian,
Sandor Batkai,
Daniela Parolaro,
Michael J Fischer,
Peter Reeh,
George Kunos,
Michaela Kress,
Beat Lutz,
Clifford J Woolf,
Rohini Kuner
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Nitin Agarwal,
Pal Pacher,
Irmgard Tegeder,
Fumimasa Amaya,
Cristina E Constantin,
Gary J Brenner,
Tiziana Rubino,
Christoph W Michalski,
Giovanni Marsicano,
Krisztina Monory, [......],
Claudiu Marian,
Sandor Batkai,
Daniela Parolaro,
Michael J Fischer,
Peter Reeh,
George Kunos,
Michaela Kress,
Beat Lutz,
Clifford J Woolf,
Rohini Kuner
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Ceng Luo,
Vijayan Gangadharan,
Kiran Kumar Bali,
Rou-Gang Xie, Nitin Agarwal,
Martina Kurejova,
Anke Tappe-Theodor,
Irmgard Tegeder,
Susanne Feil,
Gary Lewin,
Erika Polgar,
Andrew J Todd,
Jens Schlossmann,
Franz Hofmann,
Da-Lu Liu,
San-Jue Hu,
Robert Feil,
Thomas Kuner,
Rohini Kuner
[show abstract]
[hide abstract]
ABSTRACT: Synaptic long-term potentiation (LTP) at spinal neurons directly communicating pain-specific inputs from the periphery to the brain has been proposed to serve as a trigger for pain hypersensitivity in pathological states. Previous studies have functionally implicated the NMDA receptor-NO pathway and the downstream second messenger, cGMP, in these processes. Because cGMP can broadly influence diverse ion-channels, kinases, and phosphodiesterases, pre- as well as post-synaptically, the precise identity of cGMP targets mediating spinal LTP, their mechanisms of action, and their locus in the spinal circuitry are still unclear. Here, we found that Protein Kinase G1 (PKG-I) localized presynaptically in nociceptor terminals plays an essential role in the expression of spinal LTP. Using the Cre-lox P system, we generated nociceptor-specific knockout mice lacking PKG-I specifically in presynaptic terminals of nociceptors in the spinal cord, but not in post-synaptic neurons or elsewhere (SNS-PKG-I(-/-) mice). Patch clamp recordings showed that activity-induced LTP at identified synapses between nociceptors and spinal neurons projecting to the periaqueductal grey (PAG) was completely abolished in SNS-PKG-I(-/-) mice, although basal synaptic transmission was not affected. Analyses of synaptic failure rates and paired-pulse ratios indicated a role for presynaptic PKG-I in regulating the probability of neurotransmitter release. Inositol 1,4,5-triphosphate receptor 1 and myosin light chain kinase were recruited as key phosphorylation targets of presynaptic PKG-I in nociceptive neurons. Finally, behavioural analyses in vivo showed marked defects in SNS-PKG-I(-/-) mice in several models of activity-induced nociceptive hypersensitivity, and pharmacological studies identified a clear contribution of PKG-I expressed in spinal terminals of nociceptors. Our results thus indicate that presynaptic mechanisms involving an increase in release probability from nociceptors are operational in the expression of synaptic LTP on spinal-PAG projection neurons and that PKG-I localized in presynaptic nociceptor terminals plays an essential role in this process to regulate pain sensitivity.
PLoS Biology 03/2012; 10(3):e1001283. · 11.45 Impact Factor
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Anke Tappe-Theodor,
Cristina E Constantin,
Irmgard Tegeder,
Stefan G Lechner,
Michiel Langeslag,
Peter Lepcynzsky,
Richard I Wirotanseng,
Martina Kurejova, Nitin Agarwal,
Gergely Nagy,
Andrew Todd,
Nina Wettschureck,
Stefan Offermanns,
Michaela Kress,
Gary R Lewin,
Rohini Kuner
[show abstract]
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ABSTRACT: Peripheral injury or inflammation leads to a release of mediators capable of binding to a variety of ion channels and receptors. Among these are the 7-transmembrane receptors (G protein-coupled receptors) coupling to G(s), G(i/o), G₁₂/₁₃, or G(q/11) G proteins. Each of the G protein-coupled receptor pathways is involved in nociceptive modulation and pain processing, but the relative contribution of individual signaling pathways in vivo has not yet been worked out. The G(q)/G₁₁ signaling branch is of particular interest because it leads to the activation of phospholipase C-β, protein kinase C, the release of calcium from intracellular stores, and it modulates extracellular regulated kinases. To investigate the contribution of the entire G(q/11)-signaling pathway in nociceptors towards regulation of pain, we generated double-deficient mice lacking G(q/11) selectively in nociceptors using a conditional gene-targeting approach. We observed that nociceptor-specific loss of G(q) and G₁₁ results in reduced pain hypersensitivity following paw inflammation or spared nerve injury. Surprisingly, our behavioral and electrophysiological experiments also indicated defects in basal mechanical sensitivity in G(q/11) mutant mice, suggesting a novel function for G(q/11) in tonic modulation of acute nociception. Patch-clamp recordings revealed changes in voltage-dependent tetrodotoxin-resistant and tetrodotoxin-sensitive sodium channels in nociceptors upon a loss of G(q/11), whereas potassium currents remained unchanged. Our results indicate that the functional role of the G(q)/G₁₁ branch of G-protein signaling in nociceptors in vivo not only spans sensitization mechanisms in pathological pain states, but is also operational in tonic modulation of basal nociception and acute pain.
Pain 11/2011; 153(1):184-96. · 5.78 Impact Factor
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Vijayan Gangadharan,
Rui Wang,
Bettina Ulzhöfer,
Ceng Luo,
Rita Bardoni,
Kiran Kumar Bali, Nitin Agarwal,
Irmgard Tegeder,
Ullrich Hildebrandt,
Gergely G Nagy,
Andrew J Todd,
Alessia Ghirri,
Annette Häussler,
Rolf Sprengel,
Peter H Seeburg,
Amy B MacDermott,
Gary R Lewin,
Rohini Kuner
[show abstract]
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ABSTRACT: α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid-type (AMPA-type) glutamate receptors (AMPARs) play an important role in plasticity at central synapses. Although there is anatomical evidence for AMPAR expression in the peripheral nervous system, the functional role of such receptors in vivo is not clear. To address this issue, we generated mice specifically lacking either of the key AMPAR subunits, GluA1 or GluA2, in peripheral, pain-sensing neurons (nociceptors), while preserving expression of these subunits in the central nervous system. Nociceptor-specific deletion of GluA1 led to disruption of calcium permeability and reduced capsaicin-evoked activation of nociceptors. Deletion of GluA1, but not GluA2, led to reduced mechanical hypersensitivity and sensitization in models of chronic inflammatory pain and arthritis. Further analysis revealed that GluA1-containing AMPARs regulated the responses of nociceptors to painful stimuli in inflamed tissues and controlled the excitatory drive from the periphery into the spinal cord. Consequently, peripherally applied AMPAR antagonists alleviated inflammatory pain by specifically blocking calcium-permeable AMPARs, without affecting physiological pain or eliciting central side effects. These findings indicate an important pathophysiological role for calcium-permeable AMPARs in nociceptors and may have therapeutic implications for the treatment chronic inflammatory pain states.
The Journal of clinical investigation 03/2011; 121(4):1608-23. · 15.39 Impact Factor
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ABSTRACT: gamma-aminobutyric acid (GABA) is an important inhibitory neurotransmitter which mainly mediates its effects on neurons via ionotropic (GABA(A)) and metabotropic (GABA(B)) receptors. GABA(B) receptors are widely expressed in the central and the peripheral nervous system. Although there is evidence for a key function of GABA(B) receptors in the modulation of pain, the relative contribution of peripherally- versus centrally-expressed GABA(B) receptors is unclear.
In order to elucidate the functional relevance of GABA(B) receptors expressed in peripheral nociceptive neurons in pain modulation we generated and analyzed conditional mouse mutants lacking functional GABA(B1) subunit specifically in nociceptors, preserving expression in the spinal cord and brain (SNS-GABA(B1)-/- mice). Lack of the GABA(B1) subunit precludes the assembly of functional GABA(B) receptor. We analyzed SNS-GABA(B1)-/- mice and their control littermates in several models of acute and neuropathic pain. Electrophysiological studies on peripheral afferents revealed higher firing frequencies in SNS-GABA(B1)-/- mice compared to corresponding control littermates. However no differences were seen in basal nociceptive sensitivity between these groups. The development of neuropathic and chronic inflammatory pain was similar across the two genotypes. The duration of nocifensive responses evoked by intraplantar formalin injection was prolonged in the SNS-GABAB(1)-/- animals as compared to their control littermates. Pharmacological experiments revealed that systemic baclofen-induced inhibition of formalin-induced nociceptive behaviors was not dependent upon GABA(B1) expression in nociceptors.
This study addressed contribution of GABA(B) receptors expressed on primary afferent nociceptive fibers to the modulation of pain. We observed that neither the development of acute and chronic pain nor the analgesic effects of a systematically-delivered GABA(B) agonist was significantly changed upon a specific deletion of GABA(B) receptors from peripheral nociceptive neurons in vivo. This lets us conclude that GABA(B) receptors in the peripheral nervous system play a less important role than those in the central nervous system in the regulation of pain.
Molecular Pain 11/2009; 5:68. · 3.53 Impact Factor
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ABSTRACT: The peptide endothelin-1 (ET1), which was originally identified as a vasoconstrictor, has emerged as a critical regulator of a number of painful conditions, including inflammatory pain and tumor-associated pain. There is considerable pharmacological evidence supporting a role for endothelin A receptors (ET(A)) in mediating ET1-induced pro-algesic functions. ET(A) receptors are expressed in small-diameter nociceptive neurons, but also found in a variety of other cell types in peripheral tissues, including immune cells, keratinocytes, endothelial cells, which have the potential to modulate nociception. To elucidate the functional contribution of ET(A) receptors expressed in sensory neurons towards the functions of the ET1 axis in pathological pain states, we undertook a conditional gene deletion approach to selectively deplete expression of ET(A) in sensory nerves, preserving expression in non-neural peripheral tissues; the expression of ET(B) remained unchanged. Behavioural and pharmacological experiments showed that only late nociceptive hypersensitivity caused by ET1 is abrogated upon a loss of ET(A) receptors on nociceptors and further suggest that ET1-induced early nociceptive hypersensitivity involves activation of ET(A) as well as ET(B) receptors in non-neural peripheral cells. Furthermore, in the context of alleviation of cancer pain and chronic inflammatory pain by ET(A) receptor antagonists, we observed in corresponding mouse models that the contribution of ET(A) receptors expressed in nociceptors is most significant. These results help understand the role of ET(A) receptors in complex biological processes and peripheral cell-cell interactions involved in inflammatory and tumor-associated pain.
Pain 10/2009; 148(2):206-14. · 5.78 Impact Factor
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Matthias Schweizerhof,
Sebastian Stösser,
Martina Kurejova,
Christian Njoo,
Vijayan Gangadharan, Nitin Agarwal,
Martin Schmelz,
Kiran Kumar Bali,
Christoph W Michalski,
Stefan Brugger,
Anthony Dickenson,
Donald A Simone,
Rohini Kuner
[show abstract]
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ABSTRACT: Pain is one of the most severe and debilitating symptoms associated with several forms of cancer. Various types of carcinomas and sarcomas metastasize to skeletal bones and cause spontaneous bone pain and hyperalgesia, which is accompanied by bone degradation and remodeling of peripheral nerves. Despite recent advances, the molecular mechanisms underlying the development and maintenance of cancer-evoked pain are not well understood. Several types of non-hematopoietic tumors secrete hematopoietic colony-stimulating factors that act on myeloid cells and tumor cells. Here we report that receptors and signaling mediators of granulocyte- and granulocyte-macrophage colony-stimulating factors (G-CSF and GM-CSF) are also functionally expressed on sensory nerves. GM-CSF sensitized nerves to mechanical stimuli in vitro and in vivo, potentiated CGRP release and caused sprouting of sensory nerve endings in the skin. Interruption of G-CSF and GM-CSF signaling in vivo led to reduced tumor growth and nerve remodeling, and abrogated bone cancer pain. The key significance of GM-CSF signaling in sensory neurons was revealed by an attenuation of tumor-evoked pain following a sensory nerve-specific knockdown of GM-CSF receptors. These results show that G-CSF and GM-CSF are important in tumor-nerve interactions and suggest that their receptors on primary afferent nerve fibers constitute potential therapeutic targets in cancer pain.
Nature medicine 07/2009; 15(7):802-7. · 27.14 Impact Factor
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Matthias Schweizerhof,
Sebastian St|[ouml]|sser,
Martina Kurejova,
Christian Njoo,
Vijayan Gangadharan, Nitin Agarwal,
Martin Schmelz,
Kiran Kumar Bali,
Christoph W Michalski,
Stefan Brugger,
Anthony Dickenson,
Donald A Simone,
Rohini Kuner
Nature Medicine 06/2009; 15(7):802-807. · 22.46 Impact Factor
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ABSTRACT: The key role of Cyclin-dependent kinase 5 (Cdk5) in neuronal function has been well established but understanding of its importance in sensory pathways is in its infancy. Recently we described the important role of Cdk5 in pain signaling. Our studies indicated that conditional deletion of Cdk5 in small sensory neurons causes hypoalgesia. In current study, we identified development of atypical non-healing skin lesions in these mutant mice during the general colony maintenance. Detailed examination of these lesions clearly distinguishes them from ulcerative dermatitis. Here we hypothesize that these skin lesions are due to general sensation loss in these mice as evident from deep skin scratches that turn into unhealed wounds.
Cell cycle (Georgetown, Tex.) 04/2008; 7(6):750-3. · 5.36 Impact Factor
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Nitin Agarwal,
Pal Pacher,
Irmgard Tegeder,
Fumimasa Amaya,
Cristina E Constantin,
Gary J Brenner,
Tiziana Rubino,
Christoph W Michalski,
Giovanni Marsicano,
Krisztina Monory, [......],
Claudiu Marian,
Sandor Batkai,
Daniela Parolaro,
Michael J Fischer,
Peter Reeh,
George Kunos,
Michaela Kress,
Beat Lutz,
Clifford J Woolf,
Rohini Kuner
[show abstract]
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ABSTRACT: Although endocannabinoids constitute one of the first lines of defense against pain, the anatomical locus and the precise receptor mechanisms underlying cannabinergic modulation of pain are uncertain. Clinical exploitation of the system is severely hindered by the cognitive deficits, memory impairment, motor disturbances and psychotropic effects resulting from the central actions of cannabinoids. We deleted the type 1 cannabinoid receptor (CB1) specifically in nociceptive neurons localized in the peripheral nervous system of mice, preserving its expression in the CNS, and analyzed these genetically modified mice in preclinical models of inflammatory and neuropathic pain. The nociceptor-specific loss of CB1 substantially reduced the analgesia produced by local and systemic, but not intrathecal, delivery of cannabinoids. We conclude that the contribution of CB1-type receptors expressed on the peripheral terminals of nociceptors to cannabinoid-induced analgesia is paramount, which should enable the development of peripherally acting CB1 analgesic agonists without any central side effects.
Nature Neuroscience 08/2007; 10(7):870-9. · 15.53 Impact Factor
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Christoph W Michalski,
Tamara Laukert,
Danguole Sauliunaite,
Pál Pacher,
Frank Bergmann, Nitin Agarwal,
Yun Su,
Thomas Giese,
Nathalia A Giese,
Sándor Bátkai,
Helmut Friess,
Rohini Kuner
[show abstract]
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ABSTRACT: The functional involvement of the endocannabinoid system in modulation of pancreatic inflammation, such as acute pancreatitis, has not been studied to date. Moreover, the therapeutic potential of cannabinoids in pancreatitis has not been addressed.
We quantified endocannabinoid levels and expression of cannabinoid receptors 1 and 2 (CB1 and CB2) in pancreas biopsies from patients and mice with acute pancreatitis. Functional studies were performed in mice using pharmacological interventions. Histological examination, serological, and molecular analyses (lipase, myeloperoxidase, cytokines, and chemokines) were performed to assess disease pathology and inflammation. Pain resulting from pancreatitis was studied as abdominal hypersensitivity to punctate von Frey stimuli. Behavioral analyses in the open-field, light-dark, and catalepsy tests were performed to judge cannabinoid-induced central side effects.
Patients with acute pancreatitis showed an up-regulation of cannabinoid receptors and elevated levels of endocannabinoids in the pancreas. HU210, a synthetic agonist at CB1 and CB2, abolished abdominal pain associated with pancreatitis and also reduced inflammation and decreased tissue pathology in mice without producing central, adverse effects. Antagonists at CB1- and CB2-receptors were effective in reversing HU210-induced antinociception, whereas a combination of CB1- and CB2-antagonists was required to block the anti-inflammatory effects of HU210 in pancreatitis.
In humans, acute pancreatitis is associated with up-regulation of ligands as well as receptors of the endocannabinoid system in the pancreas. Furthermore, our results suggest a therapeutic potential for cannabinoids in abolishing pain associated with acute pancreatitis and in partially reducing inflammation and disease pathology in the absence of adverse side effects.
Gastroenterology 06/2007; 132(5):1968-78. · 11.68 Impact Factor
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Anke Tappe-Theodor, Nitin Agarwal,
István Katona,
Tiziana Rubino,
Lene Martini,
Jakub Swiercz,
Ken Mackie,
Hannah Monyer,
Daniela Parolaro,
Jennifer Whistler,
Thomas Kuner,
Rohini Kuner
[show abstract]
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ABSTRACT: Clinical usage of cannabinoids in chronic pain states is limited by their central side effects and the pharmacodynamic tolerance that sets in after repeated dosage. Analgesic tolerance to cannabinoids in vivo could be caused by agonist-induced downregulation and intracellular trafficking of cannabinoid receptors, but little is known about the molecular mechanisms involved. We show here that the type 1 cannabinoid receptor (CB1) interacts physically with G-protein-associated sorting protein 1 (GASP1), a protein that sorts receptors in lysosomal compartments destined for degradation. CB1-GASP1 interaction was observed to be required for agonist-induced downregulation of CB1 in spinal neurons ex vivo as well as in vivo. Importantly, uncoupling CB1 from GASP1 in mice in vivo abrogated tolerance toward cannabinoid-induced analgesia. These results suggest that GASP1 is a key regulator of the fate of CB1 after agonist exposure in the nervous system and critically determines analgesic tolerance to cannabinoids.
Journal of Neuroscience 05/2007; 27(15):4165-77. · 7.11 Impact Factor
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ABSTRACT: Transient receptor potential vanilloid 1 (TRPV1), a ligand-gated cation channel highly expressed in small-diameter sensory neurons, is activated by heat, protons, and capsaicin. The phosphorylation of TRPV1 provides a versatile regulation of intracellular calcium levels and is critical for TRPV1 function in responding to a pain stimulus. We have previously reported that cyclin-dependent kinase 5 (Cdk5) activity regulates nociceptive signaling. In this article we report that the Cdk5-mediated phosphorylation of TRPV1 at threonine-407 can modulate agonist-induced calcium influx. Inhibition of Cdk5 activity in cultured dorsal root ganglia neurons resulted in a significant reduction of TRPV1-mediated calcium influx, and this effect could be reversed by restoring Cdk5 activity. Primary nociceptor-specific Cdk5 conditional-knockout mice showed reduced TRPV1 phosphorylation, resulting in significant hypoalgesia. Thus, the present study indicates that Cdk5-mediated TRPV1 phosphorylation is important in the regulation of pain signaling.
Proceedings of the National Academy of Sciences 02/2007; 104(2):660-5. · 9.68 Impact Factor
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ABSTRACT: Glutamatergic signaling and intracellular calcium mobilization in the spinal cord are crucial for the development of nociceptive plasticity, which is associated with chronic pathological pain. Long-form Homer proteins anchor glutamatergic receptors to sources of calcium influx and release at synapses, which is antagonized by the short, activity-dependent splice variant Homer1a. We show here that Homer1a operates in a negative feedback loop to regulate the excitability of the pain pathway in an activity-dependent manner. Homer1a is rapidly and selectively upregulated in spinal cord neurons after peripheral inflammation in an NMDA receptor-dependent manner. Homer1a strongly attenuates calcium mobilization as well as MAP kinase activation induced by glutamate receptors and reduces synaptic contacts on spinal cord neurons that process pain inputs. Preventing activity-induced upregulation of Homer1a using shRNAs in mice in vivo exacerbates inflammatory pain. Thus, activity-dependent uncoupling of glutamate receptors from intracellular signaling mediators is a novel, endogenous physiological mechanism for counteracting sensitization at the first, crucial synapse in the pain pathway. Furthermore, we observed that targeted gene transfer of Homer1a to specific spinal segments in vivo reduces inflammatory hyperalgesia. Thus, Homer1 function is crucially involved in pain plasticity and constitutes a promising therapeutic target for the treatment of chronic inflammatory pain.
Nature Medicine 07/2006; 12(6):677-81. · 22.46 Impact Factor
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ABSTRACT: The use of Cre-loxP technology for conditional mutagenesis in pain pathways had been restricted by the unavailability of mice expressing Cre recombinase selectively in functionally distinct components of the nociceptive system. Here we describe the generation of transgenic mouse lines which express Cre recombinase selectively in sensory ganglia using promoter elements of the Na(v)1.8 gene (Scn10a). Cre-mediated recombination was greatly evident in all nociceptive and thermoreceptive neurons of the dorsal root ganglia and trigeminal ganglia, but only in a small proportion of proprioceptive neurons. Cre-mediated recombination was not detectable in the brain, spinal cord, or any nonneural tissues and began perinatally after invasion of primary afferents into the developing spinal cord. Thus, these mice enable selective deletion of genes in subsets of sensory neurons and offer a wide scope for studying potential functions of genes in pain perception, independent of secondary effects arising from developmental defects or global gene ablation.
genesis 04/2004; 38(3):122-9. · 2.53 Impact Factor
