Geoffrey D. Clarke

Reduced response to the formalin test and lowered spinal NMDA glutamate receptor binding in adenosine A2A receptor knockout mice

Abstract Adenosine is a neuromodulator with complex effects on pain pathways. Mice lacking the adenosine A2A receptor are hypoalgesic, and have altered analgesic responses to receptor‐selective opioid agonists. These and other findings suggest a role for the adenosine A2A receptor in sensitizing afferent fibres projecting to the spinal cord. To test this hypothesis formalin (20 μl, 5%) was injected into the paw and nociceptive responses were measured in wildtype and adenosine A2A receptor knockout mice. There was a significant reduction in nociception associated with sensory nerve activation in the knockout mice as measured by time spent biting/licking the formalin‐injected paw and number of flinches seen during the first phase, but only the number of flinches was reduced during the second inflammatory phase. In addition, the selective adenosine A2A antagonist SCH58261 (3 and 10 mg/kg) also antagonised both phases of the formalin test. We also labelled NMDA glutamate and NK1 receptors in spinal cord sections as an indirect measure of nociceptive transmission from peripheral sites to the spinal cord. [3H]‐Substance P binding to NK1 receptors was unaltered but there was a substantial reduction in binding of [3H]‐MK801 to NMDA glutamate receptors in all regions of the spinal cord from knockout mice. The decrease in NMDA glutamate receptor binding may reflect reduced peripheral sensory input to the spinal cord during development and could relate to the hypoalgesia in this genotype. These results support a key role for the adenosine A2A receptor in peripheral nociceptive pathways.

Comparative effects of selective κ-opioid receptor agonists on dopamine levels in the dorsal caudate of freely moving rats

Microdialysis was utilized to evaluate the effects of selective kappa-opioid receptor agonists on dopamine levels in the dorsal caudate of conscious rats. Subcutaneous administration of equivalent antinociceptive doses of spiradoline--(+/-)-(5 alpha, 7 alpha, 8 beta)-3, 4-dichloro-N-methyl-N-[7-(1-pyrrolidinyl)-1-oxaspiro[4,5]dec-8-yl] benzeneacetamide--(U62066; 12 mg/kg), BRL 52656--(2S)-1-[(4- trifluoromethylphenyl)acetyl]-2-[(1-pyrrolidinyl)methyl]piperidine--(2 mg/kg) and enadoline--(-)-(5 beta, 7 beta, 8 alpha)-N-methyl-N-[7-(1- pyrrolidinyl)-1-oxaspiro[4,5]dec-8-yl]benzo[b]furan-4-acetamide-- (CI-977; 0.1 mg/kg) produced similar, statistically significant decreases in dorsal caudate dopamine levels; BRL 53001--(2S)-2- (dimethylaminomethyl)-1-[(5,6,7,8-tetrahydro-5-oxo-2-naphthyl)ace tyl] piperidine--(12 mg/kg) was, however, without effect. At a higher dose (36 mg/kgP, BRL 53001 also caused a significant reduction in dopamine levels. BRL 52974--4-(1-pyrrolidinylmethyl) 5-[(3,4-dichlorophenyl)acetyl]-4,5,6,7-tetrahydroimidazo[4,5-c] pyridine, a selective kappa-opioid receptor agonist with limited ability to cross the blood brain barrier or produce antinociceptive effects, had no effect on dopamine levels at 10 mg/kg s.c. Overall, these findings suggest that selective kappa-opioid receptor agonists decrease dopamine levels in the dorsal caudate of rats via a central locus of action. Furthermore, compared to other kappa-opioid receptor agonists, BRL 53001 appears to have a reduced propensity to decrease dopamine levels at equianalgesic doses.

Comparison of a novel fast-dissolving acetaminophen tablet formulation (FD-APAP) and standard acetaminophen tablets using gamma scintigraphy and pharmacokinetic studies

Context: Acetaminophen (paracetamol, APAP) is widely used to relieve mild-to-moderate pain and reduce fever. Absorption of the drug can be impacted by dosage form; this may have implications for pain relief in some individuals, potentially accounting for suboptimal efficacy in analgesia. Objective: To assess the disintegration and dissolution of a new fast-dissolving acetaminophen tablet formulation (FD-APAP) and the impact on pharmacokinetic and pharmacodynamic parameters. Materials and methods: Two randomized, single-center, open-label, single-dose, two-way crossover studies in healthy subjects to compare FD-APAP (2 × 500 mg tablets) with standard acetaminophen (2 × 500 mg tablets). Gamma scintigraphy was used to assess tablet disintegration (Study 1, N = 24), and plasma profiles were evaluated in the fasted state (Study 2, N = 40). Results: In Study 1, the mean time to complete disintegration (12.9 vs. 69.6 min, P < 0.0001) and onset of disintegration were both significantly faster with FD-APAP than with standard acetaminophen (P < 0.0001). For Study 2, median Tmax was significantly faster for FD-APAP (0.50 vs. 0.67 h, P < 0.01) and AUC0–30 min was significantly greater (4.51 vs. 2.74, P < 0.05). AUC0–t and AUC0–inf were comparable between the two study treatments. Discussion: Despite the absence of comparative clinical data, the FD-APAP formulation may be expected to overcome some of the issues associated with the slow and variable absorption of standard acetaminophen tablet formulations, improving therapeutic outcome and avoiding the need to switch to alternative therapeutic options. Conclusion: Compared with standard acetaminophen, the FD-APAP formulation results in significantly faster onset of disintegration and more rapid absorption.

Antinociceptive effects of non-steroidal anti-inflammatory drugs in a rat model of unilateral hindpaw inflammation.

The antinociceptive activity of intramuscular 6-methoxy-2-naphthylacetic acid (6-MNA), the active metabolite of nabumetone, was examined in a rat model of unilateral hindpaw inflammation/hyperalgesia and compared with that of three other non-steroidal anti-inflammatory drugs (NSAIDs)--diclofenac, naproxen and piroxicam. Over a dose range of 10-100 mg/kg i.m., 6-MNA produced a dose-dependent increase in the withdrawal threshold to a noxious mechanical stimulus applied to the inflamed paw; however, a higher dose (300 mg/kg) produced no further increase in antinociceptive activity. Peak effects occurred 30 min after intramuscular injection. Diclofenac, naproxen and piroxicam produced antinociceptive effects that were qualitatively similar to those of 6-MNA. There was an indication of quantitative differences between the four NSAIDs in terms of potency and efficacy although this was not statistically significant. The rapid onset of effect and the lack of correlation between the relative antinociceptive effects of the four NSAIDs and the anti-inflammatory activities (assayed as inhibition of carrageenan-induced rat paw oedema) suggest that their pain relieving properties may not be entirely the result of their anti-inflammatory effects. These experimental data support the therapeutic value of 6-MNA as an analgesic in conditions of inflammatory pain.

Deletion of the adenosine A(2A) receptor in mice enhances spinal cord neurochemical responses to an inflammatory nociceptive stimulus.

Knockout mice lacking the adenosine A(2A) receptor are less sensitive to nociceptive stimuli, and this may be due to the presence of pronociceptive A(2A) receptors on sensory nerves. In support of this hypothesis, we have recently shown that in A(2A) receptor knockout mice there are marked reductions in the changes of two markers of spinal cord neuronal activity, [(3)H]MK801 binding to NMDA receptors and uptake of [(14)C]-2-deoxyglucose, in response to formalin injection. We now report that following a more prolonged inflammatory stimulus, consisting of intraplantar injections of PGE(2) and paw pressure, there was in contrast an increase in [(3)H]MK801 binding and [(14)C]-2-deoxyglucose uptake in the spinal cords of the A(2A) receptor knockout mice which was much greater than in the wild-type mice. This increase suggests that when there is a pronounced inflammatory component to the stimulus, loss of inhibitory A(2A) receptors on inflammatory cells outweighs the loss of pronociceptive A(2A) receptors on peripheral nerves so that overall there is an increase in nociceptive signalling. This implies that although A(2A) antagonists have antinociceptive effects they may have only limited use as analgesics in chronic inflammatory pain.

Genetic deletion of the adenosine A(2A) receptor in mice reduces the changes in spinal cord NMDA receptor binding and glucose uptake caused by a nociceptive stimulus

Mice lacking the adenosine A(2A) receptor are less sensitive to nociceptive stimuli, and A(2A) receptor antagonists have antinociceptive effects. We have previously shown a marked reduction in the behavioural responses to formalin injection in A(2A) receptor knockout mice. This may be due to the presence of pronociceptive A(2A) receptors on sensory nerves, and if so spinal cords from A(2A) receptor knockout mice may have altered neurochemical responses to a nociceptive stimulus. We tested this hypothesis by studying two parameters known to change with spinal cord activity, NMDA glutamate receptor binding and [(14)C]-2-deoxyglucose uptake, following intraplantar formalin injection in wild-type and A(2A) receptor knockout mice. In naïve untreated A(2A) knockout mice [(14)C]-2-deoxyglucose uptake in all regions of the spinal cord was significantly lower compared to the wild-type, similar to the reduced NMDA receptor binding that we have previously observed. Following formalin treatment, there was an decrease in [(3)H]-MK801 binding to NMDA receptors and an increase in [(14)C]-2-deoxyglucose uptake in the spinal cords of wild-type mice, and these changes were significantly reduced in the A(2A) knockout mice. In addition to altered behavioural responses, there are therefore corresponding reductions in spinal cord neurochemical changes induced by formalin in mice lacking adenosine A(2A) receptors. These observations support the hypothesis that activation of A(2A) receptors enhances nociceptive input into the spinal cord and suggests a possible role for A(2A) antagonists as analgesics.

Using suitability profiles to better inform consumers' choice of commonly used over-the-counter analgesics

Objective To quantify the impact of labelled contraindications, precautions and warnings for use on the population that may take commonly used over‐the‐counter (OTC) analgesics.

A new rapidly absorbed paediatric paracetamol suspension. A six-way crossover pharmacokinetic study comparingthe rate and extent of paracetamol absorption from a new paracetamol suspension with two marketed paediatric formulations

The objective of this study was to compare the rate and extent of paracetamol absorption from the new Paracetamol pediatric suspension (PPS) with two marketed paracetamol suspensions: Children’s panadol (CP) and Panodil baby & infant (PBI). The study also assessed the effect on paracetamol absorption of light-calorie, low-fat food consumed 2 h before dosing. Twenty eight male adult volunteers received a single oral dose of 1000 mg of paracetamol from each of three treatments, in both fasted and fed states according to a randomized, single-center, open-label, six-way crossover study design. PPS was bioequivalent to both CP and PBI for AUC0–10 h, AUC0–inf and Cmax in both fasted and fed state. However, PPS had greater rate of paracetamol absorption and a faster speed of onset. Tmax for PPS was significantly shorter than for PBI in both fasted (p = 0.0005) and fed state (p = 0.0001). Median Tmax for PPS was also 10 min shorter than CP in fasted state. Time to reach minimum effective concentration (MEC) for PPS was significantly shorter than CP and PBI. Early paracetamol exposure of PPS was significantly higher than that of the two existing paracetamol products. Food had a significant effect in the early exposure and onset of therapeutic level of paracetamol from PPS. AUC0–30 min was significantly higher and time to reach plasma paracetamol at MEC level was significantly shorter than in the fasted state.

A Prospective Survey to Compare the Suitability Profiles of Over-the-Counter Ibuprofen and Paracetamol Use in a French General Practitioner-Controlled Population

AbstractBackground and objectives: The PAIN (Paracetamol, Aspirin and Ibuprofen New tolerability) study published in 1999 assessed the tolerability of over-the-counter (OTC) analgesics in a French general practitioner (GP)-controlled population and found no apparent difference between the tolerability of ibuprofen and paracetamol (acetaminophen). However, patient selection in that study could cast doubt over the relevance of the results to a more generalised OTC population. The aim of our survey was to prospectively determine what proportion of a French GP-controlled population is able to take ibuprofen and paracetamol in order to allow appropriate interpretation of the PAIN study. Methods: This was a prospective survey to identify all contraindications/warnings on product labelling for ibuprofen and paracetamol in 5000 patients aged 18–75 years. 100 French GPs each collected anonymised data on 50 patients over a 1-week period using a standardised form. GPs were stratified by geographical location and size of practice. Demographic data for patients were summarised. Contraindications/warnings for ibuprofen and paracetamol were summarised overall and by individual criterion. Results: The mean age of patients was 45.8 (± 16.5) years; 42% were males and 58% females. Of these patients, 49.9% would have been ineligible for the PAIN study because of at least one contraindication/warning for ibuprofen, whereas only 6.8% would have been ineligible because of at least one contraindication/warning for paracetamol. More specifically, contraindications to use of ibuprofen was noted in 12.3% of patients and 37.6% of patients should have consulted a doctor before use of this drug. In contrast, contraindications to use of paracetamol were noted in only 0.7% of patients and only 6.1% should have consulted a doctor before use of this drug. Conclusions: Our survey demonstrates how careful patient selection can limit the generalisability of a study. The PAIN study excluded individuals ‘at-risk’ from ibuprofen and the tolerability results of this study should therefore be interpreted with extreme caution because the patient population may not be representative of unsupervised OTC analgesic users.

C20orf9-003 (ACI-1), a gene localized on chromosome 20q13.12 encoding for a 49 kD cytoplasmic protein with a putative nucleotide binding site.

Murine NGD5 is a gene identified from NG108-15 cells which is postulated to be involved in opioid receptor function. Here we report the cloning and characterization of a cDNA C20orf9-003 (ACI-1) encoding the human orthologue of the mouse NGD5. Analysis of the genomic structure revealed that C20orf9-003 (ACI-1) contains 13 exons and 12 introns, spanning 52.5 kb of genomic DNA and is a variant of C20orf9. Chromosomal localization of human C20orf9-003 (ACI-1) assigned this gene to chromosome 20q13.12. Genes at this locus have been associated with the progression and possibly the development of various cancers. In addition several linkage studies support the possibility that one or more genes affecting obesity are located in 20q13. No function can be clearly assigned to C20orf9-003 (ACI-1), however, the protein has a cytoplasmic subcellular location and the secondary structure contains a Rossman fold like feature which is found in many nucleotide binding proteins.