Opioid receptor expressing neurons of the central amygdala gate behavioral effects of ketamine in mice.

Ketamine has anesthetic, analgesic, and antidepressant properties which may involve multiple neuromodulatory systems. In humans, the opioid receptor (OR) antagonist naltrexone blocks the antidepressant effect of ketamine. It is unclear whether naltrexone blocks a direct effect of ketamine at ORs, or whether normal functioning of the OR system is required to realize the full antidepressant effects of treatment. In mice, the effect of ketamine on locomotion, but not analgesia or the forced swim test, was sensitive to naltrexone and was therefore used as a behavioral readout to localize the effect of naltrexone in the brain. We performed whole-brain imaging of cFos expression in ketamine-treated mice, pretreated with naltrexone or vehicle, and identified the central amygdala (CeA) as the area with greatest difference in cFos intensity. CeA neurons expressing both µOR (MOR) and PKCµ were strongly activated by naltrexone but not ketamine, and selectively interrupting MOR function in the CeA either pharmacologically or genetically blocked the locomotor effects of ketamine. These data suggest that MORs expressed in CeA neurons gate behavioral effects of ketamine but are not direct targets of ketamine.

Brief lifestyle interventions for prediabetes in primary care: a service evaluation.

BACKGROUND: The increasing number of cases of prediabetes in the UK is concerning, particularly in Wales where there is no standard programme of support. The aim of the current service evaluation was to examine the effectiveness of brief lifestyle interventions on glucose tolerance in people at risk of developing type 2 diabetes. METHODS: In this pragmatic service evaluation clinical data on people deemed at risk of developing type 2 diabetes were evaluated from two GP clusters. Patients (n = 1207) received a single 15 to 30-min, face-to-face, consultation with a health care practitioner. Interventions were assessed by changes in HbA1c and distribution across the HbA1c ranges 12 months following intervention. Statistical significance of reversion to normoglycaemia and development of diabetes were assessed through comparison with expected rates without intervention. RESULTS: Between baseline and 12-month follow-up HbA1c fell from 43.85 ± 1.57 mmol/mol (6.16 ± 0.14%) to 41.63 ± 3.84 mmol/mol (5.96 ± 0.35%), a decrease of 2.22 mmol/mol (0.20%) (95% CI 2.01 (0.18%), 2.42 (0.22%); p < 0.0001). The proportion of people with normal glucose tolerance at 12 months (0.50 95%CI 0.47, 0.52) was significantly larger than the lower (0.06 (p < 0.0001) and the upper (0.19 (p < 0.0001)) estimates based on no intervention. CONCLUSION: Results indicate significant improvement in glucose tolerance across GP clusters. The brief intervention has the potential to offer a robust and effective option to support people at risk of developing type 2 diabetes. Further research in the form of a randomised trial is needed to confirm this and identify those likely to benefit most from this intervention.

A Standardized Strategy for Simultaneous Quantification of Urine Metabolites to Validate Development of a Biomarker Panel Allowing Comprehensive Assessment of Dietary Exposure.

SCOPE: Metabolites derived from individual foods found in human biofluids after consumption could provide objective measures of dietary intake. For comprehensive dietary assessment, quantification methods would need to manage the structurally diverse mixture of target metabolites present at wide concentration ranges. METHODS AND RESULTS: A strategy for selection of candidate dietary exposure biomarkers is developed. An analytical method for 62 food biomarkers is validated by extensive analysis of chromatographic and ionization behavior characteristics using triple quadrupole mass spectrometry. Urine samples from two food intervention studies are used: a controlled, inpatient study (n = 19) and a free-living study where individuals (n = 15) are provided with food as a series of menu plans. As proof-of-principle, it is demonstrated that the biomarker panel could discriminate between menu plans by detecting distinctive changes in the concentration in urine of targeted metabolites. Quantitative relationships between four biomarker concentrations in urine and dietary intake are shown. CONCLUSION: Design concepts for an analytical strategy are demonstrated, allowing simultaneous quantification of a comprehensive panel of chemically-diverse biomarkers of a wide range of commonly-consumed foods. It is proposed that integration of self-reported dietary recording tools with biomarker approaches will provide more robust assessment of dietary exposure.

Peripheral Nerve Stimulation for Pudendal Neuralgia: A Technical Note.

BACKGROUND: Pudendal neuropathy is a chronic, disabling form of perineal pain that involves the pudendal nerve, a mixed somatic and autonomic nerve that originates from sacral nerve roots. Peripheral nerve stimulation of the pudendal nerve can be useful to decrease symptom burden in patients who have failed initial conservative treatment modalities. METHODS: In this manuscript, we describe an approach to the placement of a peripheral nerve stimulator for the treatment of pudendal neuralgia. We present a case of complex pelvic neuropathy and review the factors that lead to successful placement. Technical aspects of stimulator placement and ultrasound landmarks are reviewed. RESULTS: A lateral to medial approach with ultrasound guidance at the level of the ischial spine is likely to facilitate proper lead placement along the course of the pudendal nerve. Aftercare and adherence to postimplant activity restrictions-particularly avoiding use of the extremes of hip flexion and extension for four weeks-lead to the absence of lead migration. CONCLUSIONS: Pudendal nerve stimulation is an emerging technique for neuromodulation of refractory pudendal neuralgia. Ultrasound-guided pudendal nerve stimulation is a viable technique for neuromodulation of pudendal neuralgia. Optimization of patient selection, ultrasound guidance, and proper adherence to postimplant activity restrictions may be helpful for long-term therapeutic success.

P2X4 Receptors on Muscle Macrophages Are Required for Development of Hyperalgesia in an Animal Model of Activity-Induced Muscle Pain.

Activity-induced pain is common in those with chronic musculoskeletal pain and limits participation in daily activities and exercise. Our laboratory developed a model of activity-induced pain and shows that depletion of muscle macrophages prevents development of hyperalgesia. Adenosine triphosphate (ATP) is released from fatiguing muscle and activates purinergic receptors (P2X), and P2X4 receptors are expressed on macrophages. We hypothesized that exercise releases ATP to activate P2X4 receptors on muscle macrophages, which subsequently release interleukin-1ß (IL-1ß) to produce hyperalgesia. In an animal model of activity-induced pain, using male and female C57BL6/J mice, we show increased expression of P2X4 on muscle macrophages, and blockade of P2X4 receptors in muscle prevented development of hyperalgesia. Using a lentivirus expressing an artificial micro-RNA to P2X4 under the control of a CD68 promoter, we decreased expression of P2X4 mRNA in cultured macrophages, decreased expression of P2X4 protein in muscle macrophages in vivo, and prevented development of activity-induced hyperalgesia. We further show that macrophages primed with LPS differentially released IL-1ß when treated with ATP in neutral or acidic pH. Lastly, blockade of IL-1ß in muscle prevented development of hyperalgesia in this model. Thus, our data suggest that P2X4 receptors could be a valid pharmacological target to control activity-induced muscle pain experienced by patients with chronic musculoskeletal pain.

No Influence of Low-, Medium-, or High-Dose Tyrosine on Exercise in a Warm Environment.

PURPOSE: Tyrosine administration may counter exercise fatigue in a warm environment, but the typical dose is inconclusive, with little known about higher doses. We explored how three tyrosine doses influenced the circulating ratio of tyrosine/amino acids competing for brain uptake and hypothesized that a medium and high dose would enhance exercise performance in a warm environment. METHODS: Eight recreationally trained, non-heat-acclimated male individuals (mean ± SD age, 23 ± 4 yr; stature, 181 ± 7 cm; body mass, 76.1 ± 5.9 kg; peak oxygen uptake, 4.1 ± 0.5 L·min) performed a peak oxygen uptake test, two familiarization trials, then four experimental trials in a randomized order separated by 7 d. Before exercise, subjects drank 2 × 300 mL sugar-free drinks delivering 0 (PLA), 150 (LOW), 300 (MED), or 400 (HIGH) mg·kg body mass tyrosine in a double-blind fashion. Subjects performed a 60-min constant intensity cycling then a simulated time trial in 30°C and 60% relative humidity. RESULTS: Time trial performance (P = 0.579) was not influenced by tyrosine ingestion. The plasma ratio of tyrosine/∑(free-tryptophan, leucine, isoleucine, valine, phenylalanine, methionine), a key determinant of brain tyrosine influx, increased relative to PLA (P < 0.001). The increase was similar (P > 0.05) in MED (7.7-fold) and HIGH (8.2-fold), and greater than that in LOW (5.3-fold; P < 0.05). No differences existed between trials in core and skin temperature, heart rate, RPE, or thermal sensation (P > 0.05). CONCLUSION: Exercise performance in a warm environment was not influenced by tyrosine availability in recreationally trained male individuals. The results provide novel data informing future studies, on the tyrosine dose maximizing the circulating ratio of tyrosine/amino acids competing for brain uptake.

Acid Sensing Ion Channel 1a (ASIC1a) Mediates Activity-induced Pain by Modulation of Heteromeric ASIC Channel Kinetics.

Chronic muscle pain is acutely worsened by exercise. Acid sensing ion channels (ASIC) are heteromeric channels expressed in muscle sensory neurons that detect decreases in pH. We have previously shown ASIC3 is important in activity-induced hyperalgesia. However, ASICs form heteromers with ASIC1a being a key component in sensory neurons. Therefore, we studied the role of ASIC1a in mice using behavioral pharmacology and genetic deletion in a model of activity-induced hyperalgesia. We found ASIC1a-/- mice developed mechanical hyperalgesia similar to wild-type mice, but antagonism of ASIC1a, with psalmotoxin, prevented development of mechanical hyperalgesia in wild-type mice, but not in ASIC1a-/- mice. To explain this discrepancy, we then performed electrophysiology studies of ASICs and examined the effects of psalmotoxin on ASIC heteromers. We expressed ASIC1a, 2 and 3 heteromers or ASIC1 and 3 heteromers in CHO cells, and examined the effects of psalmotoxin on pH sensitivity. Psalmotoxin significantly altered the properties of ASIC hetomeric channels. Specifically, in ASIC1a/2/3 heteromers, psalmotoxin slowed the kinetics of desensitization, slowed the recovery from desensitization, and inhibited pH-dependent steady-state desensitization, but had no effect on pH-evoked current amplitudes. We found a different pattern in ASIC1a/3 heteromers. There was a significant leftward shift in the pH dose response of steady-state desensitization and decrease in pH-evoked current amplitudes. These results suggest that blockade of ASIC1a modulates the kinetics of heteromeric ASICs to prevent development of activity-induced hyperalgesia. These data suggest ASIC1a is a key subunit in heteromeric ASICs and may be a pharmacological target for treatment of musculoskeletal pain.

Reduced Arch Length as a Factor for 4-Implant Immediate Function in the Maxilla: A Technical Note and Report of 39 Patients Followed for 5 Years.

PURPOSE: To determine whether maxillary arch length deficiency treated with a V-4 implant placement method is adequate for immediate functional loading during a 5-year follow-up. MATERIALS AND METHODS: Thirty-nine patients were treated with maxillary immediate function from January 3, 2011 to February 28, 2011 and followed for a period of 5 years. Arch length after implant placement was measured retrospectively around the arch from the anterior sinus wall to the contralateral anterior sinus wall in a mid-alveolar arc on the occlusal view of the preoperative computed tomogram. Eight patients with an osseous arch length shorter than 45 mm were treated with a V-4 pattern. Thirty-one patients with an arch length longer than 45 mm were treated with an M-4 placement pattern. The aim was to determine whether immediate function with a shorter arch length could be obtained on the day of surgery using a V-4 placement and whether implant stability would persist during the 5-year follow-up. Any surgical events, including lost implants, were recorded in the charts. Late follow-up was performed by panoramic films. RESULTS: During the 2-month treatment period, 39 patients (8 with V-4 placement and and 31 with M-4 placement) received maxillary treatment. The 8 patients in the V-4 group had an arch length average of 36.0 mm available for osseointegration. The 31 patients in the M-4 group had an arch length average of 56.6 mm. There were 7 implant losses (and replacements) during the 5-year follow-up, 1 in the V-4 group and 6 in the M-4 group. CONCLUSION: When the arch length bone available for osseointegration is shorter than 45 mm, a V-4 placement strategy might enable successful 4-implant fixed denture support for immediate function.

Regular physical activity prevents chronic pain by altering resident muscle macrophage phenotype and increasing interleukin-10 in mice.

Regular physical activity in healthy individuals prevents development of chronic musculoskeletal pain; however, the mechanisms underlying this exercise-induced analgesia are not well understood. Interleukin-10 (IL-10), an antiinflammatory cytokine that can reduce nociceptor sensitization, increases during regular physical activity. Since macrophages play a major role in cytokine production and are present in muscle tissue, we propose that physical activity alters macrophage phenotype to increase IL-10 and prevent chronic pain. Physical activity was induced by allowing C57BL/6J mice free access to running wheels for 8 weeks and compared to sedentary mice with no running wheels. Using immunohistochemical staining of the gastrocnemius muscle to label regulatory (M2, secretes antiinflammatory cytokines) and classical (M1, secretes proinflammatory cytokines) macrophages, the percentage of M2-macrophages increased significantly in physically active mice (68.5% ± 4.6% of total) compared with sedentary mice (45.8% ± 7.1% of total). Repeated acid injections into the muscle enhanced mechanical sensitivity of the muscle and paw in sedentary animals, which does not occur in physically active mice; no sex differences occur in either sedentary or physically active mice. Blockade of IL-10 systemically or locally prevented the analgesia in physically active mice, ie, mice developed hyperalgesia. Conversely, sedentary mice pretreated systemically or locally with IL-10 had reduced hyperalgesia after repeated acid injections. Thus, these results suggest that regular physical activity increases the percentage of regulatory macrophages in muscle and that IL-10 is an essential mediator in the analgesia produced by regular physical activity.

ASIC3 Is Required for Development of Fatigue-Induced Hyperalgesia.

An acute bout of exercise can exacerbate pain, hindering participation in regular exercise and daily activities. The mechanisms underlying pain in response to acute exercise are poorly understood. We hypothesized that proton accumulation during muscle fatigue activates acid-sensing ion channel 3 (ASIC3) on muscle nociceptors to produce hyperalgesia. We investigated the role of ASIC3 using genetic and pharmacological approaches in a model of fatigue-enhanced hyperalgesia. This model uses two injections of pH 5.0 saline into muscle in combination with an electrically induced fatigue of the same muscle just prior to the second injection of acid to induce mechanical hyperalgesia. We show a significant decrease in muscle force and decrease in muscle pH after 6 min of electrical stimulation. Genetic deletion of ASIC3 using knockout mice and pharmacological blockade of ASIC3 with APETx2 in muscle prevents the fatigue-enhanced hyperalgesia. However, ASIC3(-/-) mice and APETx2 have no effect on the fatigue response. Genetic deletion of ASIC3 in primary afferents innervating muscle using an HSV-1 expressing microRNA (miRNA) to ASIC3 surprisingly had no effect on the development of the hyperalgesia. Muscle fatigue increased the number of macrophages in muscle, and removal of macrophages from muscle with clodronate liposomes prevented the development of fatigue-enhanced hyperalgesia. Thus, these data suggest that fatigue reduces pH in muscle that subsequently activates ASIC3 on macrophages to enhance hyperalgesia to muscle insult.

Effect of Intramuscular Protons, Lactate, and ATP on Muscle Hyperalgesia in Rats.

Chronic muscle pain is a significant health problem leading to disability[1]. Muscle fatigue can exacerbate muscle pain. Metabolites, including ATP, lactate, and protons, are released during fatiguing exercise and produce pain in humans. These substances directly activate purinergic (P2X) and acid sensing ion channels (ASICs) on muscle nociceptors, and when combined, produce a greater increase in neuron firing than when given alone. Whether the enhanced effect of combining protons, lactate, and ATP is the sum of individual effects (additive) or more than the sum of individual effects (synergistic) is unknown. Using a rat model of muscle nociceptive behavior, we tested each of these compounds individually over a range of physiologic and supra-physiologic concentrations. Further, we combined all three compounds in a series of dilutions and tested their effect on muscle nociceptive behavior. We also tested a non-hydrolyzable form of ATP (α,ß-meATP) alone and in combination with lactate and acidic pH. Surprisingly, we found no dose-dependent effect on muscle nociceptive behavior for protons, lactate, or ATP when given alone. We similarly found no effect after application of each two-metabolite combination. Only pH 4 saline and α,ß-meATP produced hyperalgesia when given alone. When all 3 substances were combined, however, ATP (2.4µm), lactate (10mM), and acidic pH (pH 6.0) produced an enhanced effect greater than the sum of the effects of the individual components, i.e. synergism. α,ß me ATP (3nmol), on the other hand, showed no enhanced effects when combined with lactate (10mM) or acidic pH (pH 6.0), i.e. additive. These data suggest that combining fatigue metabolites in muscle produces a synergistic effect on muscle nociception.

The dichotomized role for acid sensing ion channels in musculoskeletal pain and inflammation.

Chronic muscle pain affects between 11 and 24% of the world's population with the majority of people experiencing musculoskeletal pain at some time in their life. Acid sensing ion channels (ASICs) are important sensors of modest decreases in extracellular pH that occur within the physiological range. These decreases in extracellular pH occur in response to inflammation, fatiguing exercise, and ischemia. Further, injection of acidic saline into muscle produces enhanced nociceptive behaviors in animals and pain in human subjects. Of the different types of ASICs, ASIC3 and ASIC1 have been implicated in transmission of nociceptive information from the musculoskeletal system. The current review will provide an overview of the evidence for ASIC3 and ASIC1 in musculoskeletal pain in both inflammatory and non-inflammatory models. This article is part of the Special Issue entitled 'Acid-Sensing Ion Channels in the Nervous System'.

Anatomical and physiological factors contributing to chronic muscle pain.

Chronic muscle pain remains a significant source of suffering and disability despite the adoption of pharmacologic and physical therapies. Muscle pain is mediated by free nerve endings distributed through the muscle along arteries. These nerves project to the superficial dorsal horn and are transmitted primarily through the spinothalamic tract to several cortical and subcortical structures, some of which are more active during the processing of muscle pain than other painful conditions. Mechanical forces, ischemia, and inflammation are the primary stimuli for muscle pain, which is reflected in the array of peripheral receptors contributing to muscle pain-ASIC, P2X, and TRP channels. Sensitization of peripheral receptors and of central pain processing structures are both critical for the development and maintenance of chronic muscle pain. Further, variations in peripheral receptors and central structures contribute to the significantly greater prevalence of chronic muscle pain in females.

An overview of animal models of pain: disease models and outcome measures.

UNLABELLED: Pain is ultimately a perceptual phenomenon. It is built from information gathered by specialized pain receptors in tissue, modified by spinal and supraspinal mechanisms, and integrated into a discrete sensory experience with an emotional valence in the brain. Because of this, studying intact animals allows the multidimensional nature of pain to be examined. A number of animal models have been developed, reflecting observations that pain phenotypes are mediated by distinct mechanisms. Animal models of pain are designed to mimic distinct clinical diseases to better evaluate underlying mechanisms and potential treatments. Outcome measures are designed to measure multiple parts of the pain experience, including reflexive hyperalgesia measures, sensory and affective dimensions of pain, and impact of pain on function and quality of life. In this review, we discuss the common methods used for inducing each of the pain phenotypes related to clinical pain syndromes as well as the main behavioral tests for assessing pain in each model. PERSPECTIVE: Understanding animal models and outcome measures in animals will assist in translating data from basic science to the clinic.

Fatigue-enhanced hyperalgesia in response to muscle insult: induction and development occur in a sex-dependent manner.

Chronic muscle pain affects 20-50% of the population, is more common in women than men, and is associated with increased pain during physical activity and exercise. Muscle fatigue is common in people with chronic muscle pain, occurs in response to exercise, and is associated with release of fatigue metabolites. Fatigue metabolites can sensitize muscle nociceptors, which could enhance pain with exercise. Using a mouse model we tested whether fatigue of a single muscle, induced by electrical stimulation, resulted in enhanced muscle hyperalgesia and if the enhanced hyperalgesia was more pronounced in female mice. Muscle fatigue was induced in combination with a sub-threshold muscle insult (2 injections of pH 5.0 saline) in male and female mice. We show that male and female mice, fatigued immediately prior to muscle insult in the same muscle, develop similar muscle hyperalgesia 24 hours later. However, female mice also develop hyperalgesia when muscle fatigue and muscle insult occur in different muscles, and when muscle insult is administered 24 hours after fatigue in the same muscle. Further, hyperalgesia lasts significantly longer in females. Finally, muscle insult with or without muscle fatigue results in minimal inflammatory changes in the muscle itself, and sex differences are not related to estradiol (ovariectomy) or changes in brainstem activity (pNR1). Thus, the current model mimics muscle fatigue-induced enhancement of pain observed in chronic muscle pain conditions in the human population. Interactions between fatigue and muscle insult may underlie the development of chronic widespread pain with an associated female predominance observed in human subjects.