Bradford D. Fischer

Lack of evidence for opioid tolerance or dependence in rhesus monkeys following high-dose anabolic-androgenic steroid administration.

Prolonged use of high-dose anabolic-androgenic steroids (AAS) may induce a dependence syndrome, and emerging evidence suggests that AAS effects on endogenous opioid systems may contribute to AAS abuse. The present study tested the hypothesis that high dose AAS treatment enhances endogenous opioid activity in rhesus monkeys as revealed by 1) tolerance to the antinociceptive effects of the mu opioid agonist morphine and 2) physical dependence as indicated by evidence of opioid withdrawal following administration of the opioid antagonist naloxone. Three rhesus monkeys were treated for 14 days with 3.2 mg/kg/day testosterone propionate, and the effects of morphine (0.32-10 mg/kg) and naloxone (0.01-0.32 mg/kg) were examined both before and during treatment. Morphine antinociception was evaluated using a warm-water tail-withdrawal procedure, and naloxone-precipitated withdrawal was evaluated using checked behavioral signs and measures of ventilatory rate. Chronic testosterone administration for 14 days produced a 100-fold increase in mean plasma testosterone levels. However, testosterone treatment did not significantly alter the antinociceptive effects of morphine, and naloxone did not precipitate signs of opioid withdrawal either before or during testosterone treatment. These data do not support the hypothesis that high-dose AAS treatment enhances endogenous opioid activity in rhesus monkeys in a way that produces opioid tolerance or dependence.

Attenuation of morphine antinociceptive tolerance by a CB1 receptor agonist and an NMDA receptor antagonist: Interactive effects

CB(1) cannabinoid (CB(1)) receptor agonists and N-Methyl-d-Aspartate (NMDA) receptor antagonists attenuate the development of morphine antinociceptive tolerance. The present study used dose-addition analysis to evaluate CB(1)/NMDA receptor interactions on this endpoint. Chronic morphine administration (5 days, 100 mg/kg, twice daily) resulted in a 2.8-fold rightward shift in the morphine dose-effect curve. Co-administration of either the CB(1) receptor agonist CP-55940 (5-(1,1-Dimethylheptyl)-2-[5-hydroxy-2-(3-hydroxypropyl)cyclohexyl]phenol; 0.32-1.0 mg/kg) or the NMDA receptor antagonist (-)-6-phosphonomethyl-deca-hydroisoquinoline-3-carboxylic acid (LY235959; 1.0-3.2 mg/kg) with morphine dose-dependently attenuated morphine tolerance. The relative potency of each drug alone was quantified using a defined level of effect (one-quarter log shift in the morphine dose-effect curve), resulting in equieffective doses of 0.42 mg/kg and 1.1 mg/kg for CP-55940 and LY235959, respectively. Subsequent experiments assessed CP-55940/LY235959 interactions using a fixed-proportion design. Co-administration of CP-55940/LY235959 mixtures (1:1, 1:3.2, or 1:10 CP-55940/LY235959) with morphine dose-dependently attenuated morphine tolerance. Isobolographic and dose-addition analysis were used to statistically compare the experimentally determined potency for each mixture (z(mix)) with predicted additive potency (z(add)). Mixtures of 1:1 and 1:3.2 CP-55940/LY235959 produced additive effects (z(add) = z(mix)), while the mixture of 1:10 CP-55940/LY235959 produced a supra-additive effect (z(add) > z(mix)). These results suggest that CP-55940 and LY235959 produce additive or supra-additive attenuation of morphine antinociceptive tolerance after repeated morphine administration, depending on their relative concentrations.

Contribution of GABAA receptors containing α3 subunits to the therapeutic-related and side effects of benzodiazepine-type drugs in monkeys

RationaleExperimental evidence suggests that the differential behavioral effects of benzodiazepines depend on their relative actions at γ-aminobutyric acid type A (GABAA) receptors that contain either an α1, α2, α3, or α5 subunit.ObjectivesThe present study was aimed at understanding the role of α3 subunit-containing GABAA (α3GABAA) receptors by examining the behavioral pharmacology of TP003 (4,2′-difluoro-5′-[8-fluoro-7-(1-hydroxy-1-methylethyl)imidazo[1,2-a]pyridine-3-yl]biphenyl-2-carbonitrile), which shows functional selectivity for α3GABAA receptors.MethodsFirst, a conflict procedure was used to assess the anxiolytic-like effects of TP003 and a representative clinically available benzodiazepine. TP003 was also administered before daily periods of sucrose pellet availability to evaluate potential hyperphagic effects. In separate experiments, observable behavioral effects were used to assess the motor and sedative effects of TP003.ResultsAdministration of TP003 produced robust anti-conflict effects without the rate-decreasing effects that were observed with the representative benzodiazepine. Unlike the reported effects of benzodiazepines, TP003 did not enhance palatable food consumption. However, increases in observable sleep-associated posture were induced by TP003, as were decreases in some species-typical behaviors (vocalization, locomotion, and environment-directed behaviors). When evaluated for its ability to induce a procumbent posture, TP003 failed to produce an effect.ConclusionsBased on conflict and observation tests in monkeys, our results suggest that TP003 may have anxiolytic properties but lack ataxic, hyperphagic, and pronounced sedative effects characteristic of classical benzodiazepines. TP003 did induce myorelaxant-like effects and had relatively mild sedative effects. Collectively, these results suggest that α3GABAA receptors play an important role in the anxiolytic-like and motor effects of benzodiazepine-type drugs.

Anticonflict and Reinforcing Effects of Triazolam + Pregnanolone Combinations in Rhesus Monkeys

Combinations of positive modulators of benzodiazepine and neuroactive steroid sites on GABAA receptors have been shown to act in an additive or supra-additive manner depending on the endpoint under study, but they have not been assessed on experimentally induced conflict or drug self-administration. The present study examined the interactive effects of the benzodiazepine triazolam and the neuroactive steroid pregnanolone in a rhesus monkey conflict procedure (a model of anxiolysis) and on a progressive-ratio schedule of drug self-administration (a model of abuse potential). Both triazolam and pregnanolone decreased rates of nonsuppressed responding, whereas only triazolam consistently increased rates of suppressed responding (i.e., had an anticonflict effect). Fixed-ratio mixtures of triazolam and pregnanolone also decreased rates of nonsuppressed responding and did so in an additive manner. In contrast, mixtures of triazolam and pregnanolone produced either additive or supra-additive rate-increasing effects on suppressed responding, depending on the proportion of drugs in the mixture. Both triazolam and pregnanolone were self-administered significantly, and triazolam and pregnanolone mixtures had either proportion-dependent additive or infra-additive reinforcing effects. These results suggest that combinations of triazolam and pregnanolone may have enhanced anxiolytic effects with reduced behavioral disruption and abuse potential compared with either drug alone.

Reinforcing effects of compounds lacking intrinsic efficacy at α1 subunit-containing GABAA receptor subtypes in midazolam- but not cocaine-experienced rhesus monkeys.

Benzodiazepines are prescribed widely but their utility is limited by unwanted side effects, including abuse potential. The mechanisms underlying the abuse-related effects of benzodiazepines are not well understood, although α1 subunit-containing GABAA receptors have been proposed to have a critical role. Here, we examine the reinforcing effects of several compounds that vary with respect to intrinsic efficacy at α2, α3, and α5 subunit-containing GABAA receptors but lack efficacy at α1 subunit-containing GABAA receptors (‘α1-sparing compounds’): MRK-623 (functional selectivity for α2/α3 subunit-containing receptors), TPA023B (functional selectivity for α2/α3/α5 subunit-containing receptors), and TP003 (functional selectivity for α3 subunit-containing receptors). The reinforcing effects of the α1-sparing compounds were compared with those of the non-selective benzodiazepine receptor partial agonist MRK-696, and non-selective benzodiazepine receptor full agonists, midazolam and lorazepam, in rhesus monkeys trained to self-administer midazolam or cocaine, under a progressive-ratio schedule of intravenous (i.v.) drug injection. The α1-sparing compounds were self-administered significantly above vehicle levels in monkeys maintained under a midazolam baseline, but not under a cocaine baseline over the dose ranges tested. Importantly, TP003 had significant reinforcing effects, albeit at lower levels of self-administration than non-selective benzodiazepine receptor agonists. Together, these results suggest that α1 subunit-containing GABAA receptors may have a role in the reinforcing effects of benzodiazepine-type compounds in monkeys with a history of stimulant self-administration, whereas α3 subunit-containing GABAA receptors may be important mediators of the reinforcing effects of benzodiazepine-type compounds in animals with a history of sedative-anxiolytic/benzodiazepine self-administration.

Chronic exposure to tumor necrosis factor in vivo induces hyperalgesia, upregulates sodium channel gene expression and alters the cellular electrophysiology of dorsal root ganglion neurons

The goal of these studies was to investigate the links between chronic exposure to the pro-inflammatory cytokine tumor necrosis factor (TNF), hyperalgesia and the excitability of dorsal root ganglion (DRG) sensory neurons. We employed transgenic mice that constitutively express TNF (TNFtg mice), a well-established model of chronic systemic inflammation. At 6 months of age, TNFtg mice demonstrated increased sensitivity to both mechanical and thermal heat stimulation relative to aged-matched wild-type controls. These increases in stimulus-evoked behaviors are consistent with nociceptor sensitization to normal physiological stimulation. The mechanisms underlying nociceptor sensitization were investigated using single-cell analysis to quantitatively compare gene expression in small-diameter (<30μm) DRG neurons. This analysis revealed the upregulation of mRNA encoding for tetrodotoxin-resistant (TTX-R) sodium (Na+) channels (Nav1.8, Nav1.9), Na+ channel β subunits (β1-β3), TNF receptor 1 (TNFR1) and p38α mitogen-activated protein kinase in neurons of TNFtg mice. Whole-cell electrophysiology demonstrated a corresponding increase in TTX-R Na+ current density, hyperpolarizing shifts in activation and steady-state inactivation, and slower recovery from inactivation in the TNFtg neurons. Increased overlap of activation and inactivation in the TNFtg neurons produces inward Na+ currents at voltages near the resting membrane potential of sensory neurons (i.e. window currents). The combination of increased Na+ current amplitude, hyperpolarized shifts in Na+ channel activation and increased window current predicts a reduction in the action potential threshold and increased firing of small-diameter DRG neurons. Together, these data suggest that increases in the expression of Nav1.8 channels, regulatory β1 subunits and TNFR1 contribute to increased nociceptor excitability and hyperalgesia in the TNFtg mice.

Pharmacological and antihyperalgesic properties of the novel α2/3 preferring GABAA receptor ligand MP-III-024

γ-Aminobutyric acid type A (GABAA) receptors are located in spinal nociceptive circuits where they modulate the transmission of pain sensory signals from the periphery to higher centers. Benzodiazepine-type drugs bind to GABAA receptors containing α1, α2, α3, and α5 subunits (α1GABAA, α2GABAA, α3GABAA and α5GABAA receptors, respectively) through which they inhibit the transmission of these signals. In the present study we describe the novel benzodiazepine site positive allosteric modulator modulator methyl 8-ethynyl-6-(pyridin-2-yl)-4H-benzo[f]imidazo[1,5-a][1,4]diazepine-3-carboxylate (MP-III-024). MP-III-024 displayed preference for α2GABAA and α3GABAA receptors relative to α1GABAA and α5GABAA receptors as well as an improved metabolic profile relative to subtype-selective positive modulators that are available currently. Administration of MP-III-024 resulted in a dose- and time-dependent reversal of mechanical hyperalgesia. On locomotor activity and schedule-controlled responding, MP-III-024 was ineffective across the doses tested. These data provide further evidence that α2GABAA and α3GABAA receptors play an important role in the antihyperalgesic effects and may not be involved in some of the undesired effects of benzodiazepine-like drugs. Further, these findings suggest that MP-III-024 is a suitable research tool for investigating the role of α2GABAA and α3GABAA receptors in the behavioral properties of benzodiazepine-like drugs in mice.

Animal models of rheumatoid pain: experimental systems and insights

Severe chronic pain is one of the hallmarks and most debilitating manifestations of inflammatory arthritis. It represents a significant problem in the clinical management of patients with common chronic inflammatory joint conditions such as rheumatoid arthritis, psoriatic arthritis and spondyloarthropathies. The functional links between peripheral inflammatory signals and the establishment of the neuroadaptive mechanisms acting in nociceptors and in the central nervous system in the establishment of chronic and neuropathic pain are still poorly understood, representing an area of intense study and translational priority. Several well-established inducible and spontaneous animal models are available to study the onset, progression and chronicization of inflammatory joint disease, and have been instrumental in elucidating its immunopathogenesis. However, quantitative assessment of pain in animal models is technically and conceptually challenging, and it is only in recent years that inflammatory arthritis models have begun to be utilized systematically in experimental pain studies using behavioral and neurophysiological approaches to characterize acute and chronic pain stages. This article aims primarily to provide clinical and experimental rheumatologists with an overview of current animal models of arthritis pain, and to summarize emerging findings, challenges and unanswered questions in the field.

Irritable Bowel Syndrome: Clinical Manifestations, Dietary Influences, and Management

Irritable bowel syndrome (IBS) is a functional gastrointestinal disorder that is characterized by symptoms of chronic abdominal pain and altered bowel habits in the absence of an overtly identifiable cause. It is the most commonly diagnosed functional gastrointestinal disorder, accounting for about one third of gastroenterology visits. It generally presents as a complex of symptoms, including psychological dysfunction. Hypersensitivity to certain foods, especially foods that contain high amounts of fructose, plays a role in the pathophysiology of IBS. Elevated consumption of high-fructose corn syrup (HFCS) has been discussed in this aspect. The treatment options for IBS are challenging and varied. In addition to dietary restrictions for HFCS-induced IBS, such as low-FODMAP (Fermentable Oligosaccharides, Disaccharide, Monosaccharides, and Polyols) diets, existing drug therapies are administered based on the predominant symptoms and IBS-subtype. Patients with IBS are likely to suffer from issues, such as anxiety, depression, and post-traumatic-stress disorder. Biopsychosocial factors particularly socioeconomic status, sex, and race should, thus, be considered for diagnostic evaluation of patients with IBS.

Antagonism of triazolam self-administration in rhesus monkeys responding under a progressive-ratio schedule: In vivo apparent pA2 analysis

BACKGROUND Conventional benzodiazepines bind non-selectively to GABAA receptors containing α1, α2, α3, and α5 subunits (α1GABAA, α2GABAA, α3GABAA, and α5GABAA receptors, respectively), and the role of these different GABAA receptor subtypes in the reinforcing effects of benzodiazepines has not been characterized fully. We used a pharmacological antagonist approach with available subtype-selective ligands to evaluate the role of GABAA receptor subtypes in the reinforcing effects of the non-selective conventional benzodiazepine, triazolam. METHODS Rhesus monkeys (n=4) were trained under a progressive-ratio schedule of intravenous midazolam delivery and dose-response functions were determined for triazolam, in the absence and presence of flumazenil (non-selective antagonist), βCCT and 3-PBC (α1GABAA-preferring antagonists), and XLi-093 (α5GABAA-selective antagonist). RESULTS Flumazenil, βCCT and 3-PBC shifted the dose-response functions for triazolam to the right in a surmountable fashion, whereas XLi-093 was ineffective. Schild analyses revealed rank orders of potencies of flumazenil=βCCT>3-PBC. Comparison of potencies between self-administration and previous binding studies with human cloned GABAA receptor subtypes suggested that the potencies for βCCT and 3-PBC were most consistent with binding at α2GABAA and α3GABAA receptors, but not α1GABAA or α5GABAA receptor subtypes. CONCLUSIONS Our findings were not entirely consistent with blockade of α1GABAA receptors and are consistent with the possibility of α2GABAA and/or α3GABAA subtype involvement in antagonism of the reinforcing effects of triazolam. The α5GABAA receptor subtype likely does not play a substantial role in self-administration under these conditions.