William Maixner

Human Catechol-O-Methyltransferase Haplotypes Modulate Protein Expression by Altering mRNA Secondary Structure

Catechol-O-methyltransferase (COMT) is a key regulator of pain perception, cognitive function, and affective mood. Three common haplotypes of the human COMT gene, divergent in two synonymous and one nonsynonymous position, code for differences in COMT enzymatic activity and are associated with pain sensitivity. Haplotypes divergent in synonymous changes exhibited the largest difference in COMT enzymatic activity, due to a reduced amount of translated protein. The major COMT haplotypes varied with respect to messenger RNA local stem-loop structures, such that the most stable structure was associated with the lowest protein levels and enzymatic activity. Site-directed mutagenesis that eliminated the stable structure restored the amount of translated protein. These data highlight the functional significance of synonymous variations and suggest the importance of haplotypes over single-nucleotide polymorphisms for analysis of genetic variations.

GTP cyclohydrolase and tetrahydrobiopterin regulate pain sensitivity and persistence.

We report that GTP cyclohydrolase (GCH1), the rate-limiting enzyme for tetrahydrobiopterin (BH4) synthesis, is a key modulator of peripheral neuropathic and inflammatory pain. BH4 is an essential cofactor for catecholamine, serotonin and nitric oxide production. After axonal injury, concentrations of BH4 rose in primary sensory neurons, owing to upregulation of GCH1. After peripheral inflammation, BH4 also increased in dorsal root ganglia (DRGs), owing to enhanced GCH1 enzyme activity. Inhibiting this de novo BH4 synthesis in rats attenuated neuropathic and inflammatory pain and prevented nerve injury–evoked excess nitric oxide production in the DRG, whereas administering BH4 intrathecally exacerbated pain. In humans, a haplotype of the GCH1 gene (population frequency 15.4%) was significantly associated with less pain following diskectomy for persistent radicular low back pain. Healthy individuals homozygous for this haplotype exhibited reduced experimental pain sensitivity, and forskolin-stimulated immortalized leukocytes from haplotype carriers upregulated GCH1 less than did controls. BH4 is therefore an intrinsic regulator of pain sensitivity and chronicity, and the GTP cyclohydrolase haplotype is a marker for these traits.

Gender differences in the responses to noxious stimuli

Many clinical pain conditions, including migraine, fibromyalgia, and temporomandibular disorders, occur more frequently among females than males. Greater pain sensitivity among females has been considered as one possible explanation for these differences. Despite considerable clinical and experimental research on the topic, no consensus has emerged on the existence or nature of gender differences in response to noxious stimuli. In this Focus article the authors take the position that females exhibit greater sensitivity to noxious stimuli than males. In support of this position, they review the experimental literature on gender and pain responses. Then, they present and discuss a schematic model of several systems involved in the transmission and modulation of nociceptive information, which may contribute to gender-associated differences in pain sensitivity. Finally, the authors highlight several issues to be addressed by future research in this area.

Sensitivity of patients with painful temporomandibular disorders to experimentally evoked pain

&NA; Temporomandibular disorders (TMD) represent a group of chronic painful conditions involving the muscles of mastication and the temporomandibular joint. We determined whether patients with painful TMD are more sensitive to noxious stimuli than age‐matched control subjects. Fifty‐two TMD patients (16 with muscle pain and 36 with combined muscle and joint pain) and 23 age‐matched and gender‐matched volunteers participated. Forearm thermal pain threshold and tolerance values were determined. A submaximal effort tourniquet procedure was used to evoke ischemic muscle pain. Relative to control subjects, TMD patients had significantly lower thermal pain threshold, ischemic pain threshold, and ischemic pain tolerance values; and thermal pain tolerance values also tended to be lower. Pain sensitivity did not differ between the two groups of TMD patients. Furthermore, the submaximal effort tourniquet procedure, which is capable of altering acute orofacial pain (Sigurdsson and Maixner, 1994) did not produce a consistent reduction in orofacial pain associated with TMD. We concluded that TMD patients are more sensitive to noxious stimuli than pain‐free controls. These findings provide additional evidence that TMD is a psychophysiological disorder of the central nervous system which modulates emotional, physiological and neuroendocrine responses to emotional and physical stressors.

Sensitivity of patients with painful temporomandibular disorders to experimentally evoked pain: evidence for altered temporal summation of pain

&NA; Temporomandibular disorders (TMD) represent a group of chronic painful conditions involving the muscles of mastication and the temporomandibular joint. Several studies have reported that TMD is associated with enhanced sensitivity to experimental pain. Twenty‐three TMD subjects and 24 pain‐free matched control subjects participated in a set of studies which were designed to evaluate whether the temporal integrative aspects of thermal pain perception are altered in TMD patients compared with control subjects. Specifically, we have examined in both TMD patients and in age‐ and gender‐matched control subjects: (1) the time‐course and magnitude of perceived pain evoked by the application of sustained 7‐s noxious thermal stimuli (45–48°C) to the face and forearm, (2) the central summation of C‐fiber‐mediated pain produced by applying brief trains of noxious heat pulses to the skin overlying the ventral aspect of the right palm and (3) the ability to discriminate small increments in noxious heat applied to facial and volar forearm skin. Data collected from these studies indicate that TMD patients show enhanced temporal integration of thermal pain compared with control subjects. TMD patients show greater thermal C‐fiber‐mediated temporal summation than pain‐free subjects and they report a greater magnitude of sustained noxious heat pulses applied to either the face or the forearm than control subjects. In contrast to these findings, TMD and pain‐free subjects are equally able to discriminate and detect small increments of heat applied to noxious adapting temperatures. These findings suggest that the augmented temporal integration of noxious stimuli may result from alterations in central nervous system processes which contribute to the enhanced pain sensitivity observed in TMD patients.

Catechol-O-methyltransferase gene polymorphisms are associated with multiple pain-evoking stimuli

&NA; Variations in the gene encoding catechol‐O‐methyltransferase (COMT) are linked to individual differences in pain sensitivity. A single nucleotide polymorphism (SNP) in codon 158 (val158met), which affects COMT protein stability, has been associated with the human experience of pain. We recently demonstrated that three common COMT haplotypes, which affect the efficiency of COMT translation, are strongly associated with a global measure of pain sensitivity derived from individuals’ responses to noxious thermal, ischemic, and pressure stimuli. Specific haplotypes were associated with low (LPS), average (APS), or high (HPS) pain sensitivity. Although these haplotypes included the val158met SNP, a significant association with val158met variants was not observed. In the present study, we examined the association between COMT genotype and specific pain‐evoking stimuli. Threshold and tolerance to thermal, ischemic, and mechanical stimuli, as well as temporal summation to heat pain, were determined. LPS/LPS homozygotes had the least, APS/APS homozygotes had average, and APS/HPS heterozygotes had the greatest pain responsiveness. Associations were strongest for measures of thermal pain. However, the rate of temporal summation of heat pain did not differ between haplotype combinations. In contrast, the val158met genotype was associated with the rate of temporal summation of heat pain, but not with the other pain measures. This suggests that the val158met SNP plays a primary role in variation in temporal summation of pain, but that other SNPs of the COMT haplotype exert a greater influence on resting nociceptive sensitivity. Here, we propose a mechanism whereby these two genetic polymorphisms differentially affect pain perception.

Idiopathic pain disorders--pathways of vulnerability.

Idiopathic pain disorders (IPDs) consist of such conditions as temporomandibular joint disorders (TMJD), fibromyalgia syndrome (FMS), irritable bowel syndrome (IBS), chronic headaches, interstitial cystitis, chronic pelvic pain, chronic tinnitus, whiplash-associated disorders, and vulvar vestibulitis (VVS). IPDs commonly aggregate as ‘‘comorbid’’ conditions that are characterized by a complaint of pain as well as a mosaic of abnormalities in motor function, autonomic balance, neuroendocrine function, and sleep. Although the mechanisms that underlie the majority of these conditions are poorly understood, IPDs have been associated with a state of pain amplification and psychological distress (McBeth et al., 2001; Bradley and McKendree-Smith, 2002; Verne and Price, 2002; Gracely et al., 2004).

Sex differences in temporal summation but not sensory-discriminative processing of thermal pain

&NA; Gender differences in experimental pain sensitivity have been widely investigated, and the results generally indicate that females exhibit greater sensitivity to noxious stimuli than males. However, results using thermal pain procedures have been inconsistent, with some studies reporting greater responses among females and other studies reporting no gender differences. The present study investigated gender differences in thermal pain perception using several different psychophysical procedures. Twenty‐seven females and 22 males underwent thermal testing, including: determination of thermal pain threshold and tolerance, a thermal discrimination procedure, real‐time magnitude estimates of heat pulses, and temporal summation of thermal pain. The results indicated lower thermal pain threshold and tolerance and greater temporal summation of thermal pain among females, but no gender differences in thermal discrimination or real‐time magnitude estimates of discrete heat pulses. These findings suggest that gender differences in thermal pain perception may be more robust for sustained, temporally dynamic thermal stimuli with a strong C‐fiber component.

Pain perception is altered by a nucleotide polymorphism in SCN9A

The gene SCN9A is responsible for three human pain disorders. Nonsense mutations cause a complete absence of pain, whereas activating mutations cause severe episodic pain in paroxysmal extreme pain disorder and primary erythermalgia. This led us to investigate whether single nucleotide polymorphisms (SNPs) in SCN9A were associated with differing pain perception in the general population. We first genotyped 27 SCN9A SNPs in 578 individuals with a radiographic diagnosis of osteoarthritis and a pain score assessment. A significant association was found between pain score and SNP rs6746030; the rarer A allele was associated with increased pain scores compared to the commoner G allele (P = 0.016). This SNP was then further genotyped in 195 pain-assessed people with sciatica, 100 amputees with phantom pain, 179 individuals after lumbar discectomy, and 205 individuals with pancreatitis. The combined P value for increased A allele pain was 0.0001 in the five cohorts tested (1277 people in total). The two alleles of the SNP rs6746030 alter the coding sequence of the sodium channel Nav1.7. Each was separately transfected into HEK293 cells and electrophysiologically assessed by patch-clamping. The two alleles showed a difference in the voltage-dependent slow inactivation (P = 0.042) where the A allele would be predicted to increase Nav1.7 activity. Finally, we genotyped 186 healthy females characterized by their responses to a diverse set of noxious stimuli. The A allele of rs6746030 was associated with an altered pain threshold and the effect mediated through C-fiber activation. We conclude that individuals experience differing amounts of pain, per nociceptive stimulus, on the basis of their SCN9A rs6746030 genotype.

Genetically determined P2X7 receptor pore formation regulates variability in chronic pain sensitivity

Chronic pain is highly variable between individuals, as is the response to analgesics. Although much of the variability in chronic pain and analgesic response is heritable, an understanding of the genetic determinants underlying this variability is rudimentary. Here we show that variation within the coding sequence of the gene encoding the P2X7 receptor (P2X7R) affects chronic pain sensitivity in both mice and humans. P2X7Rs, which are members of the family of ionotropic ATP-gated receptors, have two distinct modes of function: they can function through their intrinsic cationic channel or by forming nonselective pores that are permeable to molecules with a mass of up to 900 Da. Using genome-wide linkage analyses, we discovered an association between nerve-injury–induced pain behavior (mechanical allodynia) and the P451L mutation of the mouse P2rx7 gene, such that mice in which P2X7Rs have impaired pore formation as a result of this mutation showed less allodynia than mice with the pore-forming P2rx7 allele. Administration of a peptide corresponding to the P2X7R C-terminal domain, which blocked pore formation but not cation channel activity, selectively reduced nerve injury and inflammatory allodynia only in mice with the pore-forming P2rx7 allele. Moreover, in two independent human chronic pain cohorts, a cohort with pain after mastectomy and a cohort with osteoarthritis, we observed a genetic association between lower pain intensity and the hypofunctional His270 (rs7958311) allele of P2RX7. Our findings suggest that selectively targeting P2X7R pore formation may be a new strategy for individualizing the treatment of chronic pain.