William R. Lariviere

The melanocortin-1 receptor gene mediates female-specific mechanisms of analgesia in mice and humans

Sex specificity of neural mechanisms modulating nociceptive information has been demonstrated in rodents, and these qualitative sex differences appear to be relevant to analgesia from κ-opioid receptor agonists, a drug class reported to be clinically effective only in women. Via quantitative trait locus mapping followed by a candidate gene strategy using both mutant mice and pharmacological tools, we now demonstrate that the melanocortin-1 receptor (Mc1r) gene mediates κ-opioid analgesia in female mice only. This finding suggested that individuals with variants of the human MC1R gene, associated in our species with red hair and fair skin, might also display altered κ-opioid analgesia. We found that women with two variant MC1R alleles displayed significantly greater analgesia from the κ-opioid, pentazocine, than all other groups. This study demonstrates an unexpected role for the MC1R gene, verifies that pain modulation in the two sexes involves neurochemically distinct substrates, and represents an example of a direct translation of a pharmacogenetic finding from mouse to human.

Heritability of nociception. III. Genetic relationships among commonly used assays of nociception and hypersensitivity

&NA; We and others have previously demonstrated that nociception in the mouse is heritable. A genetic correlation analysis of 12 common measures of nociception among a common set of inbred strains revealed three major clusters (or ‘types’) of nociception in this species. In the present study, we re‐evaluated the major types of nociception and their interrelatedness using ten additional assays of nociception and hypersensitivity, including: three thermal assays (tail withdrawal from 47.5°C water or −15°C ethanol; tail flick from radiant heat), two chemical assays of spontaneous nociception (bee venom test; capsaicin test) and their subsequent thermal hypersensitivity states (including contralateral hypersensitivity in the bee venom test), a mechanical nociceptive assay (tail‐clip test), and a mechanical hypersensitivity assay (intrathecal dynorphin). Confirming our earlier findings, the results demonstrate distinct thermal and chemical nociceptive types. It is now clear that mechanical hypersensitivity and thermal hypersensitivity are genetically dissociable phenomena. Furthermore, we now see at least two distinct types of thermal hypersensitivity: afferent‐dependent, featuring a preceding significant period of spontaneous nociceptive behavior associated with afferent neural activity, and non‐afferent‐dependent. In conclusion, our latest analysis suggests that there are at least five fundamental types of nociception and hypersensitivity: (1) baseline thermal nociception; (2) spontaneous responses to noxious chemical stimuli; (3) thermal hypersensitivity; (4) mechanical hypersensitivity; and (5) afferent input‐dependent hypersensitivity.

Genetic basis of pain variability: recent advances

An estimated 15–50% of the population experiences pain at any given time, at great personal and societal cost. Pain is the most common reason patients seek medical attention, and there is a high degree of individual variability in reporting the incidence and severity of symptoms. Research suggests that pain sensitivity and risk for chronic pain are complex heritable traits of polygenic origin. Animal studies and candidate gene testing in humans have provided some progress in understanding the heritability of pain, but the application of the genome-wide association methodology offers a new tool for further elucidating the genetic contributions to normal pain responding and pain in clinical populations. Although the determination of the genetics of pain is still in its infancy, it is clear that a number of genes play a critical role in determining pain sensitivity or susceptibility to chronic pain. This review presents an update of the most recent findings that associate genetic variation with variability in pain and an overview of the candidate genes with the highest translational potential.

High-throughput behavioral phenotyping in the expanded panel of BXD recombinant inbred strains

Genetic reference populations, particularly the BXD recombinant inbred (BXD RI) strains derived from C57BL/6J and DBA/2J mice, are a valuable resource for the discovery of the bio‐molecular substrates and genetic drivers responsible for trait variation and covariation. This approach can be profitably applied in the analysis of susceptibility and mechanisms of drug and alcohol use disorders for which many predisposing behaviors may predict the occurrence and manifestation of increased preference for these substances. Many of these traits are modeled by common mouse behavioral assays, facilitating the detection of patterns and sources of genetic coregulation of predisposing phenotypes and substance consumption. Members of the Tennessee Mouse Genome Consortium (TMGC) have obtained phenotype data from over 250 measures related to multiple behavioral assays across several batteries: response to, and withdrawal from cocaine, 3,4‐methylenedioxymethamphetamine; “ecstasy” (MDMA), morphine and alcohol; novelty seeking; behavioral despair and related neurological phenomena; pain sensitivity; stress sensitivity; anxiety; hyperactivity and sleep/wake cycles. All traits have been measured in both sexes in approximately 70 strains of the recently expanded panel of BXD RI strains. Sex differences and heritability estimates were obtained for each trait, and a comparison of early (N = 32) and recent (N = 37) BXD RI lines was performed. Primary data are publicly available for heritability, sex difference and genetic analyses using the MouseTrack database, and are also available in GeneNetwork.org for quantitative trait locus (QTL) detection and genetic analysis of gene expression. Together with the results of related studies, these data form a public resource for integrative systems genetic analysis of neurobehavioral traits.

Genetic analysis in the Collaborative Cross breeding population

Genetic reference populations in model organisms are critical resources for systems genetic analysis of disease related phenotypes. The breeding history of these inbred panels may influence detectable allelic and phenotypic diversity. The existing panel of common inbred strains reflects historical selection biases, and existing recombinant inbred panels have low allelic diversity. All such populations may be subject to consequences of inbreeding depression. The Collaborative Cross (CC) is a mouse reference population with high allelic diversity that is being constructed using a randomized breeding design that systematically outcrosses eight founder strains, followed by inbreeding to obtain new recombinant inbred strains. Five of the eight founders are common laboratory strains, and three are wild-derived. Since its inception, the partially inbred CC has been characterized for physiological, morphological, and behavioral traits. The construction of this population provided a unique opportunity to observe phenotypic variation as new allelic combinations arose through intercrossing and inbreeding to create new stable genetic combinations. Processes including inbreeding depression and its impact on allelic and phenotypic diversity were assessed. Phenotypic variation in the CC breeding population exceeds that of existing mouse genetic reference populations due to both high founder genetic diversity and novel epistatic combinations. However, some focal evidence of allele purging was detected including a suggestive QTL for litter size in a location of changing allele frequency. Despite these inescapable pressures, high diversity and precision for genetic mapping remain. These results demonstrate the potential of the CC population once completed and highlight implications for development of related populations.

The nociceptive and anti-nociceptive effects of bee venom injection and therapy: A double-edged sword

Bee venom injection as a therapy, like many other complementary and alternative medicine approaches, has been used for thousands of years to attempt to alleviate a range of diseases including arthritis. More recently, additional theraupeutic goals have been added to the list of diseases making this a critical time to evaluate the evidence for the beneficial and adverse effects of bee venom injection. Although reports of pain reduction (analgesic and antinociceptive) and anti-inflammatory effects of bee venom injection are accumulating in the literature, it is common knowledge that bee venom stings are painful and produce inflammation. In addition, a significant number of studies have been performed in the past decade highlighting that injection of bee venom and components of bee venom produce significant signs of pain or nociception, inflammation and many effects at multiple levels of immediate, acute and prolonged pain processes. This report reviews the extensive new data regarding the deleterious effects of bee venom injection in people and animals, our current understanding of the responsible underlying mechanisms and critical venom components, and provides a critical evaluation of reports of the beneficial effects of bee venom injection in people and animals and the proposed underlying mechanisms. Although further studies are required to make firm conclusions, therapeutic bee venom injection may be beneficial for some patients, but may also be harmful. This report highlights key patterns of results, critical shortcomings, and essential areas requiring further study.

Pain sensitivity and vasopressin analgesia are mediated by a gene-sex-environment interaction

Quantitative trait locus mapping of chemical/inflammatory pain in the mouse identified the Avpr1a gene, which encodes the vasopressin-1A receptor (V1AR), as being responsible for strain-dependent pain sensitivity to formalin and capsaicin. A genetic association study in humans revealed the influence of a single nucleotide polymorphism (rs10877969) in AVPR1A on capsaicin pain levels, but only in male subjects reporting stress at the time of testing. The analgesic efficacy of the vasopressin analog desmopressin revealed a similar interaction between the drug and acute stress, as desmopressin inhibition of capsaicin pain was only observed in nonstressed subjects. Additional experiments in mice confirmed the male-specific interaction of V1AR and stress, leading to the conclusion that vasopressin activates endogenous analgesia mechanisms unless they have already been activated by stress. These findings represent, to the best of our knowledge, the first explicit demonstration of analgesic efficacy depending on the emotional state of the recipient, and illustrate the heuristic power of a bench-to-bedside-to-bench translational strategy.

Genotype-dependence of gabapentin and pregabalin sensitivity: the pharmacogenetic mediation of analgesia is specific to the type of pain being inhibited

&NA; The antiepileptic drug, gabapentin, and another structurally related compound, pregabalin, are increasingly employed in the pharmacotherapy of chronic pain states, although their primary mechanism of action remains a topic of active study. A genomic approach to the study of these drugs may elucidate their potentially novel mechanisms. We examined the heritability of sensitivity to analgesia from gabapentin and pregabalin as a precursor to linkage mapping efforts. Accordingly, 11 inbred mouse strains were tested for inhibition of nociception by gabapentin or pregabalin (50–300 mg/kg, i.p.) in two different preclinical assays of inflammatory pain, the formalin test (5% formalin; 20 &mgr;l) and zymosan thermal hyperalgesia on the paw‐withdrawal test (3 mg/ml zymosan; 20 &mgr;l). Significant strain‐dependence of drug action was noted in each case, indicating that sensitivity to these analgesics is heritable. Furthermore, the pattern of strain sensitivities to gabapentin and pregabalin were mostly similar, supporting the notion that they act via similar genetic and physiological mechanisms. However, there was virtually no correlation between strain sensitivities to pregabalin inhibition of formalin nociception and zymosan thermal hyperalgesia. In light of previous data from our laboratory and others regarding morphine analgesia, we now establish and empirically demonstrate the general principle that pharmacogenetic mechanisms underlying analgesic sensitivity are specific to the type of pain being inhibited. This has considerable implications for ongoing pharmacogenetic investigations and, more generally, for the choices of preclinical models of pain used in drug development.

Pain modality- and sex-specific effects of COMT genetic functional variants

&NA; The effects of COMT genetic polymorphism on pain and analgesia are modality – selective and gender – dependent, in both mouse and humanPlease approve Summary as edited. &NA; The enzyme catechol‐O‐methyltransferase (COMT) metabolizes catecholamine neurotransmitters involved in a number of physiological functions, including pain perception. Both human and mouse COMT genes possess functional polymorphisms contributing to interindividual variability in pain phenotypes such as sensitivity to noxious stimuli, severity of clinical pain, and response to pain treatment. In this study, we found that the effects of Comt functional variation in mice are modality specific. Spontaneous inflammatory nociception and thermal nociception behaviors were correlated the most with the presence of the B2 SINE transposon insertion residing in the 3′UTR mRNA region. Similarly, in humans, COMT functional haplotypes were associated with thermal pain perception and with capsaicin‐induced pain. Furthermore, COMT genetic variations contributed to pain behaviors in mice and pain ratings in humans in a sex‐specific manner. The ancestral Comt variant, without a B2 SINE insertion, was more strongly associated with sensitivity to capsaicin in female vs male mice. In humans, the haplotype coding for low COMT activity increased capsaicin‐induced pain perception in women, but not men. These findings reemphasize the fundamental contribution of COMT to pain processes, and provide a fine‐grained resolution of this contribution at the genetic level that can be used to guide future studies in the area of pain genetics.

Comt1 genotype and expression predicts anxiety and nociceptive sensitivity in inbred strains of mice

Catechol‐O‐methyltransferase (COMT) is a ubiquitously expressed enzyme that maintains basic biologic functions by inactivating catechol substrates. In humans, polymorphic variance at the COMT locus has been associated with modulation of pain sensitivity and risk for developing psychiatric disorders. A functional haplotype associated with increased pain sensitivity was shown to result in decreased COMT activity by altering mRNA secondary structure‐dependent protein translation. However, the exact mechanisms whereby COMT modulates pain sensitivity and behavior remain unclear and can be further studied in animal models. We have assessed Comt1 gene expression levels in multiple brain regions in inbred strains of mice and have discovered that Comt1 is differentially expressed among the strains, and this differential expression is cis‐regulated. A B2 short interspersed nuclear element (SINE) was inserted in the 3′‐untranslated region (3′‐UTR) of Comt1 in 14 strains generating a common haplotype that correlates with gene expression. Experiments using mammalian expression vectors of full‐length cDNA clones with and without the SINE element show that strains with the SINE haplotype (+SINE) have greater Comt1 enzymatic activity. +SINE mice also exhibit behavioral differences in anxiety assays and decreased pain sensitivity. These results suggest that a haplotype, defined by a 3′‐UTR B2 SINE element, regulates Comt1 expression and some mouse behaviors.