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Paper Session 318: Genetics and Pain A. Molecular and Cellular Biology A06 - Regulation of Gene Expression (318/610) RNA binding and transport proteins staufen and fragile x mental retardation protein are expressed by sensory neurons and localize to peripheral axons of nociceptors
Abstracts
A30 - Other (318/614) Variations of catecholamine metabolism genes induce inter-individual variation in pain perception in humans
T Price, K Hargreaves, C Flores, F Cervero; McGill University, Montreal, QC, Canada Neuronal proteins have been traditionally viewed as being derived solely from the soma, wherein the preponderance of ribosomal/translational machinery resides. However, accumulating evidence indicates that non-somal sites, such as the dendrite and axon, are capable of a more autonomous role in terms of new protein synthesis. While this mechanism allows for more rapid, on-demand regulation of neuronal structure and function, it remains unclear precisely how the requisite RNA is trafficked into the axon for this purpose. Comprising the longest axons in the neuroaxis, primary afferent neurons of the dorsal root (DRG) and trigeminal (TG) ganglia would be greatly advantaged by such extra-somal protein synthesis, given the relatively greater time involved for new protein trafficking to occur to distal sites. Therefore, we hypothesized that staufen (stau) and fragile x mental retardation protein (FMRP), two important RNA transport granule proteins, might subserve a similar role in the PNS. In support of this hypothesis, we report here that mRNA transcripts encoding stau1 and 2 were expressed by rat DRG and TG neurons and that stau2 mRNA was expressed by human TG neurons. In cultured rat TG neurons, stau1 immunoreactivity was observed in neurite extensions, including transient receptor potential vanilloid type 1 (TRPV1)- and tau-immunoreactive neurites. Stau2 protein was detected in the sciatic nerve and dorsal roots, while stau2 mRNA was restricted to neuronal somata in the DRG. FMRP protein was also found in these tissues, although FMRP mRNA was not restricted to the DRG. Stau2 and FMRP immunoreactivity colocalized to TRPV1-immunoreactive sensory axons of the sciatic nerve, indicating that these proteins are found in the peripheral axons of nociceptors. Based on these findings, we propose that stau proteins and FMRP are attractive candidates to subserve RNA transport in sensory neurons, linking somal transcription to axonal translation.
P Mittal, H Kim, R Dionne; NINR, Bethesda, MD Based on the roles of catecholamines in the central nervous system, we have examined the effects of variations of loci in the monoamine oxidase A (MAOA) and B (MAOB) on acute clinical pain responses in humans. Healthy subjects (60 females and 52 males) whose self-reported ethnicity is European American participated in this study. Subjects requiring removal of third molars that included at least one bony impacted mandibular third molar, underwent standardized surgery. Prior to surgery, patients received intravenous midazolam and appropriate nerve blocks with local anesthetic. For single nucleotide polymorphisms (SNPs) genotyping, Assays-on-Demand or Assays-by-Design SNP Genotyping Products were used. Following PCR, fluorescence of each well was measured using the ABI Prism 7900 Sequence Detection System. From the genomic sequences including their flanking regions, 8 SNPs from MAOA and 3 SNPs from MAOB were screened. Clinically induced pain was patient reported using a paper and pencil visual analogue scale (VAS). After the extraction of the impacted third molars, pain was recorded every 20 minutes by VAS until subjects requested analgesic medication as the local anesthesia was eliminated and post-operative pain onset occurred. The maximum post-operative pain rating, onset time of post-operative pain, and the analgesic onset time after medication were used as measures of clinical pain. In the responses to clinically induced acute pain, SNP1 (rs3788862), variant homozygotes (A/A, 69.6 ⫾ 14.8mm) and heterozygotes (G/A, 67.8 ⫾ 18.8mm) showed higher pain ratings compared to non-variant homozygotes (G/G, 53.5 ⫾ 12.8 mm) (p ⫽ 0.005) in females. Seven remaining SNPs from MAOA also showed similar association with maximum post-operative pain ratings in females (p ⬍ 0.05) while no association was found in males. These results along with the previously reported effect of COMT genetic variation on pain sensitivity support the role of catecholamines in variations of pain perception in humans.
A07 - Regulating Neuronal Excitability
B. Systems (Physiology, Anatomy, Animal Models)
(318/612) The effects of genetic variation in serotonin and dopamine transporters on acute clinical pain in humans
B16 - Psychophysics/Hyperalgesia
K Ayres, H Kim, R Dionne; NINR, Bethesda, MD Catecholamine transporters play a significant role in the nervous system by reuptake of dopamine and serotonin. Genetic variations in transporter genes may affect catecholamine levels in the nervous system and contribute to inter-individual variations in pain sensitivity. We investigated the association between single nucleotide polymorphisms (SNP) in serotonin and dopamine transporter genes and acute clinical pain. Healthy European American subjects (60 females and 52 males) that participated in this study received third molar extraction by the same surgeon and were asked to report the level of pain every 20 minutes using a standard 10 cm visual analogue scale (VAS). Clinical pain was recorded postoperatively as the local anesthetic dissipated and subjects requested analgesic medications. The maximum post-operative pain rating, onset time of post-operative pain, and the analgesic onset time after medication were used as measures of clinical pain. By using PCR and the ABI Prism 7900 Sequence Detection system, 5 frequent SNPs from serotonin transporter gene and 4 frequent SNPs from dopamine transporter gene were genotyped from white blood cells. Variant homozygotes (T/T) of a SNP (rs2066713) in serotonin transporter gene showed longer time to onset of post-operative pain (165 ⫾ 50 minutes) compared to heterozygotes (T/C, 133 ⫾ 23 minutes) and non-variant homozygotes (C/C, 121 ⫾ 39 minutes) (p ⫽ 0.016) in males while no significant association was found in females. SNPs from dopamine transporter gene did not show any significant association with pain sensitivity after oral surgery. This suggests that genetic polymorphisms of catecholamine transporter genes may affect on acute clinical pain in humans.
(318/677) Association of A118G single nucleotide polymorphism of the u opioid receptor gene (OPRM) with experimental pain in a multi-ethnic sample B Hastie, L Kaplan, D Herrera, C Campbell, K Virtusio, J Mogil, M Wallace, R Fillingim; University of Florida College of Dentistry, Gainesville, FL Emerging literature has demonstrated vast inter-individual variation as well as substantial ethnic differences in pain perception. Recent limited evidence suggests that differences in pain may be partially mediated by single nucleotide polymorphisms (SNPs) of identified “pain genes.” Namely, individuals with the rare allele of A118G SNP on the OPRM have exhibited significantly higher pressure pain thresholds compared to those with two common alleles. Frequencies of the A118G allele range from 20-30% in the general population, though allele frequencies have been shown to vary by ethnicity. We sought to replicate the previous association of the A118G SNP to experimental pain responses and to extend the investigation by examining ethnic differences in allelic frequency and associations with pain responses. 132 healthy young adults (52% female; mean age ⫽ 24.5) representing three ethnic groups (40 African Americans, AA; 41 Hispanics, H; and 51 non-Hispanic whites, W) underwent several experimental pain modalities (thermal, pressure, ischemic, cold pressor pain) to test for differences in pain responses across ethnic group. Genotyping of the A118G SNP was also conducted. Results revealed only one AA with the rare allele but comparable allele frequencies across H and W (29.3% vs 27.5%, respectively). Further analysis of Hispanic and non-Hispanic whites revealed significant race by genotype interactions on thermal and ischemic pain measures (Ps⬍.05), and a similar but non-significant pattern emerged for pressure pain. Examination of the means indicated that the rare allele was associated with lower pain sensitivity among non-Hispanic whites, while Hispanics with the rare allele showed higher pain sensitivity. This extends our previous results by demonstrating ethnicity-dependent association of OPRM genotype with pain sensitivity. The much lower frequency of the rare allele in African Americans is consistent with prior results. These findings have potentially important implications for understanding the contributions of ethnicity and genetics to pain responses.
Paper Session 318: Genetics and Pain A. Molecular and Cellular Biology A06 - Regulation of Gene Expression (318/610) RNA binding and transport proteins staufen and fragile x mental retardation protein are expressed by sensory neurons and localize to peripheral axons of nociceptors
Abstracts
A30 - Other (318/614) Variations of catecholamine metabolism genes induce inter-individual variation in pain perception in humans
T Price, K Hargreaves, C Flores, F Cervero; McGill University, Montreal, QC, Canada Neuronal proteins have been traditionally viewed as being derived solely from the soma, wherein the preponderance of ribosomal/translational machinery resides. However, accumulating evidence indicates that non-somal sites, such as the dendrite and axon, are capable of a more autonomous role in terms of new protein synthesis. While this mechanism allows for more rapid, on-demand regulation of neuronal structure and function, it remains unclear precisely how the requisite RNA is trafficked into the axon for this purpose. Comprising the longest axons in the neuroaxis, primary afferent neurons of the dorsal root (DRG) and trigeminal (TG) ganglia would be greatly advantaged by such extra-somal protein synthesis, given the relatively greater time involved for new protein trafficking to occur to distal sites. Therefore, we hypothesized that staufen (stau) and fragile x mental retardation protein (FMRP), two important RNA transport granule proteins, might subserve a similar role in the PNS. In support of this hypothesis, we report here that mRNA transcripts encoding stau1 and 2 were expressed by rat DRG and TG neurons and that stau2 mRNA was expressed by human TG neurons. In cultured rat TG neurons, stau1 immunoreactivity was observed in neurite extensions, including transient receptor potential vanilloid type 1 (TRPV1)- and tau-immunoreactive neurites. Stau2 protein was detected in the sciatic nerve and dorsal roots, while stau2 mRNA was restricted to neuronal somata in the DRG. FMRP protein was also found in these tissues, although FMRP mRNA was not restricted to the DRG. Stau2 and FMRP immunoreactivity colocalized to TRPV1-immunoreactive sensory axons of the sciatic nerve, indicating that these proteins are found in the peripheral axons of nociceptors. Based on these findings, we propose that stau proteins and FMRP are attractive candidates to subserve RNA transport in sensory neurons, linking somal transcription to axonal translation.
P Mittal, H Kim, R Dionne; NINR, Bethesda, MD Based on the roles of catecholamines in the central nervous system, we have examined the effects of variations of loci in the monoamine oxidase A (MAOA) and B (MAOB) on acute clinical pain responses in humans. Healthy subjects (60 females and 52 males) whose self-reported ethnicity is European American participated in this study. Subjects requiring removal of third molars that included at least one bony impacted mandibular third molar, underwent standardized surgery. Prior to surgery, patients received intravenous midazolam and appropriate nerve blocks with local anesthetic. For single nucleotide polymorphisms (SNPs) genotyping, Assays-on-Demand or Assays-by-Design SNP Genotyping Products were used. Following PCR, fluorescence of each well was measured using the ABI Prism 7900 Sequence Detection System. From the genomic sequences including their flanking regions, 8 SNPs from MAOA and 3 SNPs from MAOB were screened. Clinically induced pain was patient reported using a paper and pencil visual analogue scale (VAS). After the extraction of the impacted third molars, pain was recorded every 20 minutes by VAS until subjects requested analgesic medication as the local anesthesia was eliminated and post-operative pain onset occurred. The maximum post-operative pain rating, onset time of post-operative pain, and the analgesic onset time after medication were used as measures of clinical pain. In the responses to clinically induced acute pain, SNP1 (rs3788862), variant homozygotes (A/A, 69.6 ⫾ 14.8mm) and heterozygotes (G/A, 67.8 ⫾ 18.8mm) showed higher pain ratings compared to non-variant homozygotes (G/G, 53.5 ⫾ 12.8 mm) (p ⫽ 0.005) in females. Seven remaining SNPs from MAOA also showed similar association with maximum post-operative pain ratings in females (p ⬍ 0.05) while no association was found in males. These results along with the previously reported effect of COMT genetic variation on pain sensitivity support the role of catecholamines in variations of pain perception in humans.
A07 - Regulating Neuronal Excitability
B. Systems (Physiology, Anatomy, Animal Models)
(318/612) The effects of genetic variation in serotonin and dopamine transporters on acute clinical pain in humans
B16 - Psychophysics/Hyperalgesia
K Ayres, H Kim, R Dionne; NINR, Bethesda, MD Catecholamine transporters play a significant role in the nervous system by reuptake of dopamine and serotonin. Genetic variations in transporter genes may affect catecholamine levels in the nervous system and contribute to inter-individual variations in pain sensitivity. We investigated the association between single nucleotide polymorphisms (SNP) in serotonin and dopamine transporter genes and acute clinical pain. Healthy European American subjects (60 females and 52 males) that participated in this study received third molar extraction by the same surgeon and were asked to report the level of pain every 20 minutes using a standard 10 cm visual analogue scale (VAS). Clinical pain was recorded postoperatively as the local anesthetic dissipated and subjects requested analgesic medications. The maximum post-operative pain rating, onset time of post-operative pain, and the analgesic onset time after medication were used as measures of clinical pain. By using PCR and the ABI Prism 7900 Sequence Detection system, 5 frequent SNPs from serotonin transporter gene and 4 frequent SNPs from dopamine transporter gene were genotyped from white blood cells. Variant homozygotes (T/T) of a SNP (rs2066713) in serotonin transporter gene showed longer time to onset of post-operative pain (165 ⫾ 50 minutes) compared to heterozygotes (T/C, 133 ⫾ 23 minutes) and non-variant homozygotes (C/C, 121 ⫾ 39 minutes) (p ⫽ 0.016) in males while no significant association was found in females. SNPs from dopamine transporter gene did not show any significant association with pain sensitivity after oral surgery. This suggests that genetic polymorphisms of catecholamine transporter genes may affect on acute clinical pain in humans.
(318/677) Association of A118G single nucleotide polymorphism of the u opioid receptor gene (OPRM) with experimental pain in a multi-ethnic sample B Hastie, L Kaplan, D Herrera, C Campbell, K Virtusio, J Mogil, M Wallace, R Fillingim; University of Florida College of Dentistry, Gainesville, FL Emerging literature has demonstrated vast inter-individual variation as well as substantial ethnic differences in pain perception. Recent limited evidence suggests that differences in pain may be partially mediated by single nucleotide polymorphisms (SNPs) of identified “pain genes.” Namely, individuals with the rare allele of A118G SNP on the OPRM have exhibited significantly higher pressure pain thresholds compared to those with two common alleles. Frequencies of the A118G allele range from 20-30% in the general population, though allele frequencies have been shown to vary by ethnicity. We sought to replicate the previous association of the A118G SNP to experimental pain responses and to extend the investigation by examining ethnic differences in allelic frequency and associations with pain responses. 132 healthy young adults (52% female; mean age ⫽ 24.5) representing three ethnic groups (40 African Americans, AA; 41 Hispanics, H; and 51 non-Hispanic whites, W) underwent several experimental pain modalities (thermal, pressure, ischemic, cold pressor pain) to test for differences in pain responses across ethnic group. Genotyping of the A118G SNP was also conducted. Results revealed only one AA with the rare allele but comparable allele frequencies across H and W (29.3% vs 27.5%, respectively). Further analysis of Hispanic and non-Hispanic whites revealed significant race by genotype interactions on thermal and ischemic pain measures (Ps⬍.05), and a similar but non-significant pattern emerged for pressure pain. Examination of the means indicated that the rare allele was associated with lower pain sensitivity among non-Hispanic whites, while Hispanics with the rare allele showed higher pain sensitivity. This extends our previous results by demonstrating ethnicity-dependent association of OPRM genotype with pain sensitivity. The much lower frequency of the rare allele in African Americans is consistent with prior results. These findings have potentially important implications for understanding the contributions of ethnicity and genetics to pain responses.