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(352) Group II mGluR 2 knock out rats show increased sensitivity of TRPA1 function
S64
Abstracts
The Journal of Pain
(352) Group II mGluR 2 knock out rats show increased sensitivity of TRPA1 function
(354) Group III mGluR8 activation attenuates MO (TRPA1)induced nociceptive behaviors
S Carlton and S Zhou; University of Texas Medical Branch, Galveston, TX
R Govea, G Hargett, and S Carlton; University of Texas Medical Branch, Galveston, TX
Group II metabotropic glutamate receptors (mGluR2/3) produce neuronal depression when activated. We have demonstrated these receptors are expressed by primary sensory neurons, exert endogenous, activity-dependent inhibition of TRPV1 receptors, thus inhibit transmission of nociceptive input to the cord. We obtained mGluR2 knock out (KO) rats from Transposagen Biopharmaceuticals, Inc., breeding heterozygous pairs to obtain homozygous WT and KO. Genome-specific primers were used to amplify the Grm2 (C407X allele) genome sequence and products were directly sequenced to genotype animals derived from the heterozygous intercrosses. WTs and KOs were run through a battery of tests to determine if there were any changes in sensitivity to mechanical, heat, cold or chemical stimuli. We previously reported there was no change in mechanical (von Frey) or heat sensitivity in the KO compared to WT rats. However there was a change in cold sensitivity, tested with cold plate, cold water tail dip and acetone. Based on these data, we determined whether KO rats showed increased sensitivity to TRPA1 activation using icilin, mustard oil (MO) or cinnamaldehyde (CM) as agonists. Topical application of 1mM icilin or 10mM MO resulted in significantly increased flinching in KO compared to WT. This increased sensitivity was confirmed at the cellular level, with DRG cells from KOs demonstrating a significantly greater magnitude of response to 10 uM icilin, 100 uM MO or 100 uM CM compared to WT. Based on the knowledge that TRPA1 is required for histamine-independent itch, we determined whether chloroquine-induced itching behavior was also increased in mGluR2 KO rats. Injection of chloroquine did not produce robust itching behavior in the KO rat but in fact itching behavior was significantly decreased in the KO compared to WT. The data suggest that knockout of group II mGluR2 effects some functions of TRPA1 activation but not all.
(353) Changes in the topographic distribution of primary afferents associated with spinal cord injury-induced neuropathic pain M Detloff, A Naqvi, V Ninan, M McMullen, K Vannix, D Quiros Molina, and J Houle; Drexel University College of Medicine, Philadelphia, PA Spinal cord injury (SCI) impairs sensory fiber transmission causing chronic debilitating neuropathic pain, yet cellular and molecular mechanisms responsible for its development and sustainment remain elusive. The 4 classes of sensory afferents that terminate in distinct lamina of the dorsal horn of the spinal cord respond to specific neurotrophic factors. Peptidergic c-fibers display extensive arborization into deep dorsal horn that correlates with the development of neuropathic pain, yet reorganization or plasticity of the Ab, Ad or non-peptidergic c- fibers after SCI has not been examined. The purpose of this study was to determine changes in the topographic distribution of Ab, Ad, peptidergic and non-peptidergic c-fibers in the spinal cord after SCI. Adult, female, Sprague-Dawley rats (n=45) received a unilateral cervical spinal cord contusion (Infinite Horizons, 200 kdyne). Three days before sacrifice, cholera toxin-b was administered to the median and ulnar nerves to trace and identify Ab fibers. BBB, Forelimb Locomotor Scale (FLS), plantar heat and von Frey testing occurred preoperatively and weekly post SCI. At-level allodynia is observed in a subset of rats by 14 days that persists for the duration of the study. At 3, 7, 14, and 35 days post SCI, rats were sacrificed; cervical spinal cord and dorsal root ganglia were dissected, and Ab, Ad, peptidergic c, and non-peptidergic c primary afferents were visualized via immunocytochemistry. We are currently analyzing changes in the topographic distribution and density of afferents over time SCI in rats with and without tactile allodynia. These data will allow us to better understand the effect of SCI on the afferent plasticity and the development of neuropathic pain. Supported from grants from Craig H. Neilsen Foundation #220798 (MRD) and NIH NINDS NS055976 (JDH).
Our lab has demonstrated that peripheral group III mGluRs are anti-hyperalgesic. Recently published data indicates that peripheral group III mGluRs negatively modulate TRPV1 activity. Preliminary data suggest that the group III mGluRs exert inhibitory control over another member of the TRP family, TRPA1. This TRP channel is expressed by nociceptors and is responsive to thermal (noxious cold temperatures) and mechanical input. Currently, no studies have examined the affect of group III mGluR8 agonists on TRPA1 activation. In the present study, we use a group III mGluR8 selective agonist, DCPG, to investigate the role of mGluR8 in modulating TRPA1 activity. For the calcium imaging study, the dorsal root ganglion (DRG) cells were exposed to the following conditions: 30 mM MO alone, 10 mM DCPG alone, or 30 mM MO + 10 mM DCPG. The first experiments demonstrated there was no change in calcium mobilization in the MO + DCPG group compared to MO within the first 3 min. Therefore, subsequent experiments will be carried out to a longer time point (4 to 5 min post-MO application). We expect DCPG to prevent later increases in calcium mobilization. For the behavioral study, nociceptive behaviors (flinching and paw withdrawal threshold [PWT]) were observed following: PBS + 10 mM MO; PBS + 1% DMSO; 30 mM DCPG + 10 mM MO; and 300 mM UBP1112 + 30 mM DCPG + 10mM MO. Results indicate that DCPG significantly attenuates MO-induced flinching and reverses MO-induced decrease in PWT, UBP blocks this DCPG-induced effect. These experiments provide lines of evidence for the functional coupling of group III mGluRs and TRPA1 ion channels. Additionally, these studies demonstrate that group III agonists may be effective in treatment of cold and mechanical allodynia which can develop as a result of inflammation, nerve injury, chemotherapy or other disease states.
E33 Sleep and Pain (355) Fatigue is associated with enhanced retrospective report of pain, but not enhanced spinal nociception, in a study of experimental pain and the menstrual cycle M Payne, E Bartley, S Palit, K Kerr, B Kuhn, J DelVentura, Y Guereca, J Rhudy, and E Terry; The University of Tulsa, Tulsa, OK Correlational and experimental evidence suggests that sleep disturbance can enhance pain. The present study examined the relationship between daytime fatigue and experimental pain sensitivity in 58 healthy, pain-free, normallycycling women. Pain sensitivity was assessed from electrocutaneous pain threshold/tolerance, nociceptive flexion reflex (NFR) threshold (a physiological measure of spinal nociception), and retrospective reports of sensory and affective pain in response to electrocutaneous stimulations (assessed from McGill Pain Questionnaire-Short Form). Testing sessions occurred during the midfollicular (days 5-8), ovulatory (1-2 days after luteinizing hormone surge), and late-luteal (days 23-28) phases of the menstrual cycle (testing order counterbalanced). Daytime fatigue was assessed from the Fatigue Severity Scale at the beginning of each testing session. Multilevel ANOVAs revealed an association between fatigue and retrospective sensory pain ratings (B=0.106, P<.001), but relationships with other pain outcomes were non-significant (Ps>.05). Moreover, the Menstrual Phase x Fatigue interaction was non-significant in all models (Ps>.05), suggesting that phase did not moderate fatigue-pain relationships. These data indicate that increased levels of fatigue may lead to increased sensory pain, but the effect may be more pronounced in retrospective pain reports given that there were no relationships with electric threshold or tolerance. Moreover, the lack of relationship with NFR suggests that any fatigue-related amplification of pain is likely to occur at supraspinal levels, perhaps via memorial mechanisms.
Abstracts
The Journal of Pain
(352) Group II mGluR 2 knock out rats show increased sensitivity of TRPA1 function
(354) Group III mGluR8 activation attenuates MO (TRPA1)induced nociceptive behaviors
S Carlton and S Zhou; University of Texas Medical Branch, Galveston, TX
R Govea, G Hargett, and S Carlton; University of Texas Medical Branch, Galveston, TX
Group II metabotropic glutamate receptors (mGluR2/3) produce neuronal depression when activated. We have demonstrated these receptors are expressed by primary sensory neurons, exert endogenous, activity-dependent inhibition of TRPV1 receptors, thus inhibit transmission of nociceptive input to the cord. We obtained mGluR2 knock out (KO) rats from Transposagen Biopharmaceuticals, Inc., breeding heterozygous pairs to obtain homozygous WT and KO. Genome-specific primers were used to amplify the Grm2 (C407X allele) genome sequence and products were directly sequenced to genotype animals derived from the heterozygous intercrosses. WTs and KOs were run through a battery of tests to determine if there were any changes in sensitivity to mechanical, heat, cold or chemical stimuli. We previously reported there was no change in mechanical (von Frey) or heat sensitivity in the KO compared to WT rats. However there was a change in cold sensitivity, tested with cold plate, cold water tail dip and acetone. Based on these data, we determined whether KO rats showed increased sensitivity to TRPA1 activation using icilin, mustard oil (MO) or cinnamaldehyde (CM) as agonists. Topical application of 1mM icilin or 10mM MO resulted in significantly increased flinching in KO compared to WT. This increased sensitivity was confirmed at the cellular level, with DRG cells from KOs demonstrating a significantly greater magnitude of response to 10 uM icilin, 100 uM MO or 100 uM CM compared to WT. Based on the knowledge that TRPA1 is required for histamine-independent itch, we determined whether chloroquine-induced itching behavior was also increased in mGluR2 KO rats. Injection of chloroquine did not produce robust itching behavior in the KO rat but in fact itching behavior was significantly decreased in the KO compared to WT. The data suggest that knockout of group II mGluR2 effects some functions of TRPA1 activation but not all.
(353) Changes in the topographic distribution of primary afferents associated with spinal cord injury-induced neuropathic pain M Detloff, A Naqvi, V Ninan, M McMullen, K Vannix, D Quiros Molina, and J Houle; Drexel University College of Medicine, Philadelphia, PA Spinal cord injury (SCI) impairs sensory fiber transmission causing chronic debilitating neuropathic pain, yet cellular and molecular mechanisms responsible for its development and sustainment remain elusive. The 4 classes of sensory afferents that terminate in distinct lamina of the dorsal horn of the spinal cord respond to specific neurotrophic factors. Peptidergic c-fibers display extensive arborization into deep dorsal horn that correlates with the development of neuropathic pain, yet reorganization or plasticity of the Ab, Ad or non-peptidergic c- fibers after SCI has not been examined. The purpose of this study was to determine changes in the topographic distribution of Ab, Ad, peptidergic and non-peptidergic c-fibers in the spinal cord after SCI. Adult, female, Sprague-Dawley rats (n=45) received a unilateral cervical spinal cord contusion (Infinite Horizons, 200 kdyne). Three days before sacrifice, cholera toxin-b was administered to the median and ulnar nerves to trace and identify Ab fibers. BBB, Forelimb Locomotor Scale (FLS), plantar heat and von Frey testing occurred preoperatively and weekly post SCI. At-level allodynia is observed in a subset of rats by 14 days that persists for the duration of the study. At 3, 7, 14, and 35 days post SCI, rats were sacrificed; cervical spinal cord and dorsal root ganglia were dissected, and Ab, Ad, peptidergic c, and non-peptidergic c primary afferents were visualized via immunocytochemistry. We are currently analyzing changes in the topographic distribution and density of afferents over time SCI in rats with and without tactile allodynia. These data will allow us to better understand the effect of SCI on the afferent plasticity and the development of neuropathic pain. Supported from grants from Craig H. Neilsen Foundation #220798 (MRD) and NIH NINDS NS055976 (JDH).
Our lab has demonstrated that peripheral group III mGluRs are anti-hyperalgesic. Recently published data indicates that peripheral group III mGluRs negatively modulate TRPV1 activity. Preliminary data suggest that the group III mGluRs exert inhibitory control over another member of the TRP family, TRPA1. This TRP channel is expressed by nociceptors and is responsive to thermal (noxious cold temperatures) and mechanical input. Currently, no studies have examined the affect of group III mGluR8 agonists on TRPA1 activation. In the present study, we use a group III mGluR8 selective agonist, DCPG, to investigate the role of mGluR8 in modulating TRPA1 activity. For the calcium imaging study, the dorsal root ganglion (DRG) cells were exposed to the following conditions: 30 mM MO alone, 10 mM DCPG alone, or 30 mM MO + 10 mM DCPG. The first experiments demonstrated there was no change in calcium mobilization in the MO + DCPG group compared to MO within the first 3 min. Therefore, subsequent experiments will be carried out to a longer time point (4 to 5 min post-MO application). We expect DCPG to prevent later increases in calcium mobilization. For the behavioral study, nociceptive behaviors (flinching and paw withdrawal threshold [PWT]) were observed following: PBS + 10 mM MO; PBS + 1% DMSO; 30 mM DCPG + 10 mM MO; and 300 mM UBP1112 + 30 mM DCPG + 10mM MO. Results indicate that DCPG significantly attenuates MO-induced flinching and reverses MO-induced decrease in PWT, UBP blocks this DCPG-induced effect. These experiments provide lines of evidence for the functional coupling of group III mGluRs and TRPA1 ion channels. Additionally, these studies demonstrate that group III agonists may be effective in treatment of cold and mechanical allodynia which can develop as a result of inflammation, nerve injury, chemotherapy or other disease states.
E33 Sleep and Pain (355) Fatigue is associated with enhanced retrospective report of pain, but not enhanced spinal nociception, in a study of experimental pain and the menstrual cycle M Payne, E Bartley, S Palit, K Kerr, B Kuhn, J DelVentura, Y Guereca, J Rhudy, and E Terry; The University of Tulsa, Tulsa, OK Correlational and experimental evidence suggests that sleep disturbance can enhance pain. The present study examined the relationship between daytime fatigue and experimental pain sensitivity in 58 healthy, pain-free, normallycycling women. Pain sensitivity was assessed from electrocutaneous pain threshold/tolerance, nociceptive flexion reflex (NFR) threshold (a physiological measure of spinal nociception), and retrospective reports of sensory and affective pain in response to electrocutaneous stimulations (assessed from McGill Pain Questionnaire-Short Form). Testing sessions occurred during the midfollicular (days 5-8), ovulatory (1-2 days after luteinizing hormone surge), and late-luteal (days 23-28) phases of the menstrual cycle (testing order counterbalanced). Daytime fatigue was assessed from the Fatigue Severity Scale at the beginning of each testing session. Multilevel ANOVAs revealed an association between fatigue and retrospective sensory pain ratings (B=0.106, P<.001), but relationships with other pain outcomes were non-significant (Ps>.05). Moreover, the Menstrual Phase x Fatigue interaction was non-significant in all models (Ps>.05), suggesting that phase did not moderate fatigue-pain relationships. These data indicate that increased levels of fatigue may lead to increased sensory pain, but the effect may be more pronounced in retrospective pain reports given that there were no relationships with electric threshold or tolerance. Moreover, the lack of relationship with NFR suggests that any fatigue-related amplification of pain is likely to occur at supraspinal levels, perhaps via memorial mechanisms.