Biomedical subjects

I P Chessell

Publications and source records attributed to I P Chessell.

At least 19 recordsLinked to original sources

PubMed2006

Attenuation of experimental arthritis in TRPV1R knockout mice.

The Transient Receptor Potential Vanilloid 1 (TRPV1R) is a ligand-gated, non-selective cation channel expressed predominantly by sensory neurons. TRPV1Rs respond to a variety of noxious stimuli including capsaicin, intense heat and acid. These factors, combined with behavioral studies, show that TRPV1Rs are involved in nociception. The aim of our study was to determine whether TRPV1Rs play a role in the development and maintenance of inflammation and mechanical hyperalgesia by studying the development of unilateral joint inflammation in TRPV1R-/- mice. Knee joints of TRPV1R-/- or wild-type (WT) mice were injected with FCA (200 microg) under temporary anesthesia, and the resulting inflammation and hyperalgesia measured for 35 days. Histological analysis was performed on joints at the end of the study. TRPV1R-/- mice developed mild joint swelling which was significantly less than that obtained in WT mice (P < 0.05, Mann-Whitney). The ratio of the weight distribution between the hind limbs in TRPV1R-/- mice was also significantly less than in WT mice (P < 0.05, Mann-Whitney). Neither swelling nor hypersensitivity was completely absent in the knockout mice, indicating either that other mechanisms are involved or that a compensatory mechanism operates in TRPV1R-/- mice. These results suggest that TRPV1 receptors are important for the development of joint inflammation and the associated mechanical hypersensitivity observed in this model.

Animals↗

PubMed2004

Isolectin B4 binding neurons are not present in the rat knee joint.

Small-diameter sensory neurons are key contributors in joint pain and have been implicated in the pathogenesis of rheumatoid arthritis (RA). Small-diameter sensory neurons can be separated into at least two distinct populations, which include isolectin B4 (IB4)-binding and tyrosine receptor kinase (trk) A-expressing. While trkA-expressing neurons have been identified in the rat knee joint there are no data, we are aware of, to suggest that IB4-binding neurons are also present. We aimed to determine whether or not there exists a population of IB4-binding neurons in the rat knee joint. Retrograde nerve tracing with fluoro-gold (FG) was used to identify the complete population of knee joint afferents in the lumbar dorsal root ganglia (DRG) L3 and L4 of female Wistar rats. IB4 conjugated to fluorescein isothiocyanate (FITC) was used to identify the cell bodies of IB4-binding neurons in the DRG. Of 1096 FG-labeled cell bodies in the DRG of knee joint injected animals (n=4), none were double labeled with FITC. Injection of FG into skin over the medial aspect of the rat knee (n=3) showed 48% of these cutaneous afferents in L3 and L4 DRG were double-labeled with FG and FITC. A complete absence of IB4-binding neurons in the rat knee joint makes it unlikely that this predominantly cutaneous, IB4-binding population of afferent neurons could have any significant influence in chronic inflammatory joint disease. This suggests that trkA-expressing neurons are the sole population of small-diameter sensory neurons in the knee joint and implies a significant role for these afferents in the progression of RA.

Animals↗

PubMed2001

Inhibition of C-fibre mediated sensory transmission in the rat following intraplantar formalin.

Using extracellular recordings from deep dorsal horn neurones in the anaesthetized rat, the effects of intraplantar (i.pl.) administration of formalin were examined. Phasic patterns of dorsal horn firing were observed which temporally concur with previously described behavioural responses following i.pl. formalin. However, unexpectedly, formalin abolished C-fibre mediated transmission, and significantly reduced responses to noxious mechanical stimulation, while electrically evoked A-fibre responses were unaffected. This suggests that whilst the first phase formalin response is mediated by direct afferent stimulation, the second phase behavioural response is dependant on a central hyperexcitability of the recipient second-order dorsal horn neurones, which may be maintained by peripheral input only from A-fibres. This study therefore provides further evidence for a centrally-mediated secondary component of the formalin model, and for the first time, describes loss of high-threshold C-fibre input to the spinal cord during this phase.

Action Potentials↗

PubMed2001

Anandamide activates peripheral nociceptors in normal and arthritic rat knee joints.

The effects of the endogenous cannabinoid anandamide were studied on peripheral, polymodal nociceptors recorded from normal and chronically inflamed (Freund's adjuvant) knee joint afferents in rats anaesthetized with pentobarbitone. Anandamide (860 nmol) caused a rapid, short lasting excitation of a sub-population of capsaicin-sensitive nociceptive afferents in normal knee joints (7.2+/-2.3 impulses s(-1); n=15 units from five animals). In arthritic joints there were 9.7+/-3.0 impulses s(-1) (n=11 from six animals), which was not significantly different from normal joints. The excitation was dose dependent (8.6 - 2900 nmol) and mediated by activation of the vanilloid receptor (VR(1)) as it was abolished by the VR1 antagonist capsazepine (1 mg kg(-1)). Our results show that anandamide, at high doses, can activate nociceptive afferents innervating the rat knee joints, in contrast with its widely described analgesic actions.

Animals↗

PubMed2000

Functional characterization of the P2X(4) receptor orthologues.

1. The aim of this study was to functionally characterize the recombinant mouse P2X(4) receptor and to compare its pharmacological properties with those of the human and rat orthologues. 2. Whole cell recordings were made from rafts of HEK-293 cells stably expressing recombinant mouse, rat or human P2X(4) receptors, using Cs-aspartate containing electrodes (3 - 8 MOmega) in a HEPES-buffered extracellular medium. 3. The agonist potency of ATP at the three species orthologues was similar, with mean EC(50) values of 2.3 microM, 1.4 microM and 5.5 microM, respectively. 4. Adenosine-5'-tetraphosphate (AP4) acted as a partial agonist with respect to ATP at the mouse and human P2X(4) receptors (EC(50)=2.6 and 3.0 microM), but was significantly less potent at the rat orthologue (EC(50)=20.0 microM). alpha,beta-methylene adenosine-5'-triphosphate (alpha,beta-meATP) also acted as a partial agonist, producing 29% of the maximum response at the mouse P2X(4) and 24% at the human P2X(4) receptor. 5. In contrast to the other species orthologues, alpha,beta-meATP failed to elicit a significant agonist response at rat P2X(4) receptors, and was found to act as an antagonist, with an IC(50) of 4.6 microM, against 10 microM ATP. 6. Mouse P2X(4) receptors were found to be sensitive to the antagonist, pyridoxalphosphate-6-azophenyl-2',4'-disulphonic acid (PPADS) (IC(50)=10.5 microM), as were human P2X(4) receptors (IC(50)=9.6 microM). The rat receptor however, showed a low sensitivity to PPADS (IC(50)>100 microM). 7. All three orthologues were relatively suramin-insensitive (IC(50)>100 microM) and insensitive to 1-[N, O-Bis(5-isoquinoline sulphonyl)benzyl]-2-(4-phenylpiperazine)ethyl]-5-isoquinoline sulphonamide (KN-62; IC(50)>3 microM). 8. Our results suggest that the pharmacological properties of the mouse receptor are most similar to the human P2X(4) receptor, and differ markedly from the rat receptor.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed2000

Apparent species differences in the kinetic properties of P2X(7) receptors.

1. Apparent species differences in the responses of recombinant P2X(7) receptors to repeated application of 2'- and 3'-O-(4-benzoylbenzoyl)-ATP (BzATP) have been investigated. 2. Repeated application of 100 microM BzATP resulted in a progressive increase in current magnitude (current growth) at mouse and human, but not rat P2X(7) receptors. 3. Current growth was thought to reflect progressive dilation of the P2X(7) ion-channel to a pore permeable to large molecules (MW<900), suggesting that channel dilation was not occurring at the rat P2X(7) receptor. However, 100 microM BzATP produced a rapid influx of YO-PRO-1 (MW375) in cells expressing rat or human P2X(7) receptors. 4. There were, however, species differences in agonist potency such that 100 microM BzATP was a supra-maximal concentration at rat, but not human or mouse, P2X(7) receptors. Importantly, when sub-maximal concentrations of BzATP or ATP were examined, current growth occurred at rat P2X(7) receptors. 5. The rate of current growth and YO-PRO-1 accumulation increased with agonist concentration and appeared more rapid at rat and human, than at mouse P2X(7) receptors. 6. The potency of BzATP and ATP was 1.5 - 10 fold lower in naïve cells than in cells repeatedly exposed to ATP. 7. This study demonstrates that current growth occurs at mouse, rat and human P2X(7) receptors but only when using sub-maximal concentrations of agonist. Previously, current growth was thought to reflect the progressive increase in pore diameter of the P2X(7) receptor ion channel, however, the results of this study suggest a progressive increase in agonist potency may also contribute.

Adenosine Triphosphate↗

PubMed2000

Antagonist effects on human P2X(7) receptor-mediated cellular accumulation of YO-PRO-1.

We have examined the interaction of P2 antagonists with the human P2X(7) receptor by studying their effect on 2' and 3'-O-benzoyl-benzoyl-ATP (DbATP) stimulated cellular accumulation of the fluorescent, DNA binding dye, YO-PRO-1 (MW=375Da). In suspensions of HEK293 cells expressing human recombinant P2X(7) receptors, DbATP produced time and concentration-dependent increases in YO-PRO-1 fluorescence. This response presumably reflects YO-PRO-1 entry through P2X(7) receptor channels and binding to nucleic acids. When studies were performed in a NaCl-free, sucrose-containing buffer, full concentration-effect curves to DbATP could be constructed. The P2 antagonists, pyridoxalphosphate-6-azophenyl-2', 4'-disulphonic acid (PPADS) and periodate oxidized ATP (oATP), reduced the potency of DbATP and decreased its maximum response. 1-[N,O-bis(1, 5-isoquinolinesulphonyl)-N-methyl-L-tyrosyl]-4-phenylpiperazine (KN62) and its analogue, KN04, reduced the potency of DbATP. Schild slopes for KN62 and KN04 were shallow and exhibited a plateau at concentrations of compound greater than 1 microM, indicating that these compounds were not competitive antagonists. Calmidazolium and a monoclonal antibody to human P2X(7) receptors attenuated DbATP-stimulated YO-PRO-1 accumulation but they were not competitive antagonists and only produced 2 - 3 fold decreases in the potency of DbATP. The effects of PPADS and KN62 were partially reversible whereas those of oATP were not. PPADS protected cells against the irreversible antagonist effects of oATP suggesting a common site of action. In contrast KN62 was not effective suggesting that it may bind at a different site to oATP and PPADS. This study has demonstrated that P2X(7) receptor function can be quantified by measuring DbATP stimulated YO-PRO-1 accumulation and has provided additional information about the interaction of P2 receptor antagonists with the human P2X(7) receptor.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed2000

Expression of P2X(7) purinoceptors on human lymphocytes and monocytes: evidence for nonfunctional P2X(7) receptors.

Lymphocytes from normal subjects and patients with B-chronic lymphocytic leukemia (B-CLL) show functional responses to extracellular ATP characteristic of the P2X(7) receptor (previously termed P2Z). These responses include opening of a cation-selective channel/pore that allows entry of the fluorescent dye ethidium and activation of a membrane metalloprotease that sheds the adhesion molecule L-selectin. The surface expression of P2X(7) receptors was measured in normal leucocytes, platelets, and B-CLL lymphocytes and correlated with their functional responses. Monocytes showed four- to fivefold greater expression of P2X(7) than B, T, and NK lymphocytes, whereas P2X(7) expression on neutrophils and platelets was weak. All cell types demonstrated abundant intracellular expression of this receptor. All 12 subjects with B-CLL expressed lymphocyte P2X(7) at about the same level as B lymphocytes from normal subjects. P2X(7) function, measured by ATP-induced uptake of ethidium, correlated closely with surface expression of this receptor in normal and B-CLL lymphocytes and monocytes (n = 47, r = 0.70; P < 0.0001). However, in three patients the ATP-induced uptake of ethidium into the malignant B lymphocytes was low or absent. The lack of P2X(7) function in these B lymphocytes was confirmed by the failure of ATP to induce Ba(2+) uptake into their lymphocytes. This lack of function of the P2X(7) receptor resulted in a failure of ATP-induced shedding of L-selectin, an adhesion molecule that directs the recirculation of lymphocytes from blood into the lymph node.

Adenosine Triphosphate↗

PubMed1999

Excitatory effect of P2X receptor activation on mesenteric afferent nerves in the anaesthetised rat.

1. We examined the effects of P2X purinoceptor agonists and P2 purinoceptor antagonists on mesenteric afferent nerves supplying the jejunum in the pentobarbitone sodium-anaesthetised rat. 2. ATP (0. 01-10 mg kg-1, i.a.) and alpha,beta-methylene-ATP (1-30 microg kg-1, i.a.) each induced dose-dependent increases in afferent nerve discharge and intrajejunal pressure. The effect on afferent nerves comprised an early (< 2 s after administration) intense burst of activity followed by a later increase (> 2 s after administration), less pronounced in comparison, which coincided with elevated intrajejunal pressure. 3. Pyridoxalphosphate-6-azophenyl-2', 4'-disulphonic acid (20 mg kg-1, i.v.) and suramin (80 mg kg-1, i.v. ) each antagonised both the early and later increases in afferent nerve discharge elicited by alpha,beta-methylene-ATP (30 microg kg-1, i.a.). 4. Co-administration of omega-conotoxin MVIIA and omega-conotoxin SVIB (each at 25 microg kg-1, i.v.), or treatment with the selective 5-HT3 receptor antagonist alosetron (30 microg kg-1, i.v.), did not affect the rapid burst of afferent nerve activity elicited by alpha,beta-methylene-ATP (30 microg kg-1, i.a.). However, toxin treatment did attenuate the elevations in intrajejunal pressure and the corresponding later phases of evoked afferent discharge, while alosetron inhibited basal afferent nerve activity. 5. In summary, ATP and alpha,beta-methylene-ATP each evoke excitation of mesenteric afferent nerves in the anaesthetised rat. We propose that the early increase in mesenteric afferent nerve activity represents a direct effect on the nerve ending, mediated by P2X receptors, whereas the later increase reflects activation of mechanosensitive fibres secondary to elevated intrajejunal pressure.

Action Potentials↗

PubMed1999

Ionic effects on human recombinant P2X7 receptor function.

The actions of monovalent and divalent ions on the P2X7 receptor have been assessed by measuring their effect on responses to the P2 receptor agonist, 2'- and 3'-O-(4-benzoyl-benzoyl)-ATP (DbATP), in HEK293 cells expressing the human recombinant P2X7 receptor. In these cells, DbATP increased the cellular accumulation of the DNA binding, fluorescent dye, YO-PRO-1. The potency of DbATP to elicit this effect was decreased by both calcium and magnesium ions. In addition, when the pH was increased above 8 or reduced below 6.5, the potency of DbATP was less than obtained at pH 7.5. Monovalent ions also affected the P2X7 receptor such that the potency of DbATP was 19-fold higher in NaCl-free buffer containing 280 mM sucrose (pEC50=6.48) than in 140 mM NaCl containing buffer (pEC50=5.19). Monovalent cations differentially affected the potency of DbATP. Thus, when the chloride concentration was maintained at 140 mM, pEC50 values for DbATP were 6.14, 5.87 and 5.19 when the counter cation was 140 mM choline, potassium or sodium, respectively. Monovalent anions also differentially affected the potency of DbATP and in the presence of 140 mM sodium ions, pEC50 values for DbATP were 6.14, 6.07, 5.19 and 4.53, respectively, when the counter anion was 140 mM aspartate, glutamate, chloride or iodide. The inhibitory effect of monovalent anions on P2X7 receptor function was also observed in electrophysiological studies. Thus in sodium glutamate containing buffer the potency of DbATP (pEC50=5.55) was approximately 22-fold higher than in NaCl containing buffer (pEC50=4.20). This study has demonstrated that P2X7 receptor function can be markedly affected by a wide range of ions and that physiological concentrations of sodium and chloride ions, as well as divalent cations, contribute to the low potency of ATP as an agonist at this receptor.

Adenosine Triphosphate↗

PubMed1999

Pharmacological characterization of ATP- and LPS-induced IL-1beta release in human monocytes.

1. We have utilized the human monocytic cell line, THP-1, and freshly isolated adherent human monocytes with the compounds pyridoxalphosphate-6-azophenyl-2',4'-disuphonic acid (PPADS), oxidized ATP, and 1-(N, O-bis[5-isoquinolinesufonyll]-N-methyl-L-tyrosyl)-4-phenylpiper azi ne (KN-62) to pharmacologically characterize the P2 receptor involved in ATP-induced release of interleukin 1beta (IL-1beta). We have also investigated the involvement of P2 receptors in lipopolysaccharide (LPS)-induced IL-1beta release from both cell types. 2. ATP caused release of IL-1beta from LPS primed THP-1 cells in both a time- and concentration-dependent manner, with a minimal effective ATP concentration of 1 mM. Stimulation of cells with 5 mM ATP resulted in detectable concentrations of IL-1beta in cell supernatants within 30 min. 3. The ATP analogue benzoylbenzoyl ATP (DBATP), a P2X7 receptor agonist, was approximately 10 fold more potent than ATP at eliciting IL-1beta release. 4. KN-62 (1 micro M), PPADS (100 microM) or oxidized ATP (100 uM) significantly inhibited 5 mM ATP-induced IL-1beta release by 81, 90 and 66% respectively, but failed to significantly inhibit LPS-induced IL-1beta release in both THP-1 cells and in freshly isolated human monocytes. 5. In both THP-1 cells and freshly isolated human monocytes, addition of the ATP degrading enzyme apyrase (0.4 U ml(-1)) to cell supernatants prior to LPS activation failed to significantly inhibit the LPS-induced IL-1beta release. In addition there was no correlation between extracellular ATP concentrations and IL-1beta release in THP-1 cells when studied over a 6 h time period. 6. In conclusion our data confirm the involvement of P2X7 receptors in ATP-induced IL-1beta release in human monocytes. However no evidence was obtained which would support the involvement of either endogenous ATP release or P2X7 receptor activation as the mechanism by which LPS-induces IL-1beta release in either the THP-1 cell line or in freshly isolated human monocytes.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1998

Blockade of human P2X7 receptor function with a monoclonal antibody.

A monoclonal antibody (MoAb) specific for the human P2X7 receptor was generated in mice. As assessed by flow cytometry, the MoAb labeled human blood-derived macrophage cells natively expressing P2X7 receptors and cells transfected with human P2X7 but not other P2X receptor types. The MoAb was used to immunoprecipitate the human P2X7 receptor protein, and in immunohistochemical studies on human lymphoid tissue, P2X7 receptor labeling was observed within discrete areas of the marginal zone of human tonsil sections. The antibody also acted as a selective antagonist of human P2X7 receptors in several functional studies. Thus, whole cell currents, elicited by the brief application of 2',3'-(4-benzoyl)-benzoyl-ATP in cells expressing human P2X7, were reduced in amplitude by the presence of the MoAb. Furthermore, preincubation of human monocytic THP-1 cells with the MoAb antagonized the ability of P2X7 agonists to induce the release of interleukin-1beta.

Adenosine Triphosphate↗

PubMed1998

Cloning and functional characterisation of the mouse P2X7 receptor.

We have isolated a 1785-bp complementary DNA (cDNA) encoding the murine P2X7 receptor subunit from NTW8 mouse microglial cells. The encoded protein has 80%, and 85% homology to the human and rat P2X7 subunits, respectively. Functional properties of the heterologously expressed murine P2X7 homomeric receptor broadly resembled those of the P2X7 receptor in the native cell line. However, marked phenotypic differences were observed between the mouse receptor, and the other P2X7 receptor orthologues isolated with respect to agonist and antagonist potencies, and the kinetics of formation of the large aqueous pore.

Amino Acid Sequence↗

PubMed1998

Evidence that P2X purinoceptors mediate the excitatory effects of alphabetamethylene-ADP in rat locus coeruleus neurones.

Extracellular and whole-cell patch clamp recordings were used to study the excitatory responses elicited by purine nucleotides in pontine slices of the rat brain containing the locus coeruleus (LC). The P2 purinoceptor agonists, alphabeta-methyleneadenosine 5'-triphosphate (alphabetameATP) and adenosine 5'-O-(2-thiodiphosphate) (ADPalphabetaS), and a novel purinoceptor agonist, alphabeta-methyleneadenosine 5'-diphosphate (alphabetameADP), elicited concentration-dependent increases in the spontaneous firing rate over the concentration range (1-300 microM). On vagus nerve or dorsal root preparations alphabetameADP (100 microM) had no agonist activity. In the presence of both alphabetameATP (300 microM), ADPbetaS (300 microM) elicited a further and significant increase in the firing rate of the LC neurones, whilst neither alphabetameATP nor alphabetameADP (300 microM) elicited a further response. The P2 purinoceptor antagonists, suramin (100 microM) and pyridoxalphosphate-6-azophenyl-2',4'-disulphonic acid (PPADS; 30 microM), markedly attenuated responses to all three agonists. Whole-cell recording of membrane current showed that, at - 60 mV, alphabetameATP and alphabetameADP (both 100 microM) elicited inward currents of a similar magnitude, whilst the inward currents elicited by a lower concentration of ADPbetaS (30 microM) were larger and faded in the presence of this agonist. In the presence of tetrodotoxin and a combination of other neurotransmission blockers, both alphabetameATP and alphabetameADP still produced inward currents. Based on the known selectivity of the agonists used in this study, there appear to be two distinct P2 purinoceptor types present on neurones in the LC, which correspond to the P2X and P2Y types. The responses elicited by alphabetameADP appear to be mediated through a putative P2X purinoceptor, although further work is required to determine which P2X receptor subtype(s) are involved.

Adenosine Diphosphate↗

PubMed1998

Cibacron blue allosterically modulates the rat P2X4 receptor.

We have used whole-cell patch clamp electrophysiology to characterise the actions of the P2 antagonist, cibacron blue, on the rat recombinant P2X4 receptor, stably expressed in human embryonic kidney 293 (HEK293) cells. In single cells, adenosine triphosphate (ATP) evoked inward currents, but the response was subject to considerable run down which precluded obtaining quantitative data. However, when recordings were made from cells that were part of a group of 20-40 electrically coupled cells (cell rafts), run-down of current was not observed and reproducible responses could be obtained. When studied using cell rafts, cibacron blue was a weak antagonist of the rat P2X4 receptor (IC50 > 300 microM) when co-applied with ATP. However, when cell rafts were preincubated with low concentrations of cibacron blue (3-30 microM) for 5 min prior to ATP addition, cibacron blue increased responses to ATP by increasing its potency (up to 4-fold) without affecting the maximum current. Potentiation of ATP-evoked currents was also observed following washout of high, inhibitory concentrations of cibacron blue (300 microM). In contrast to these effects on P2X4 receptors, cibacron blue inhibited the ATP-induced response in both single cells and rafts of HEK293 cells expressing the P2X2 receptor (IC50 approximately 600-800 nM). The effects of cibacron blue on the P2X4 receptor were quantitatively similar to those of Zn2+ which also increased ATP-evoked currents by decreasing the EC50 of ATP (up to 3.5-fold). These data are consistent with the concept that cibacron blue, like zinc, allosterically regulates the function of the P2X4 receptor.

Adenosine Triphosphate↗

PubMed1998

Effects of antagonists at the human recombinant P2X7 receptor.

1. We have used whole-cell patch clamping methods to examine the properties of the recombinant human P2X7 (P2Z) receptor stably expressed in HEK-293 cells. 2. In an extracellular solution with lowered concentrations of divalent cations (zero Mg2+ and 0.5 mM Ca2+), both ATP and the nucleotide analogue, 2'- and 3'-O-(4-benzoylbenzoyl)-adenosine 5'-triphosphate (Bz-ATP) evoked concentration-dependent whole-cell inward currents with maxima of 4658+/-671 and 5385+/-990 pA, respectively, at a holding potential of -90 mV. Current-voltage relationships determined using 100 microM Bz-ATP reversed at -2.7+/-3.1 mV, and did not display significant rectification. 3. Repeated applications of 300 microM Bz-ATP produced inward currents with similar rise-times (approx. 450 ms, 5-95% current development) but with progressively slower 95-5% decay times, with the eighth application of this agonist yielding a decay time of 197% of the first application. 4. Concentration-effect curves to ATP and Bz-ATP produced estimated EC50 values of 780 and 52.4 microM, respectively. Consecutive concentration-effect curves to Bz-ATP produced curves with similar maxima and EC50 values. 5. The non-selective P2 antagonists, pyridoxal-phosphate-6-azophenyl-, 2',4'-disulphonic acid (PPADS) and suramin, both produced concentration-dependent increases in maximal inward currents to Bz-ATP, with IC50 concentrations of approximately 1 microM and 70 microM, respectively. The profile of antagonism produced by PPADS was not that of a competitive antagonist. 6. The isoquinolene derivatives 1-(N,O-bis[5-isoquinolinesulphonyl]-N-methyl-L-tyrosyl)-4-phenylpi perazine (KN-62) and calmidazolium both produced antagonism which was not competitive, with IC50 concentrations of approximately 15 and 100 nM, respectively. HMA (5-(N,N-hexamethylene)- amiloride) was also an effective antagonist at a concentration of 10 microM. The group IIb metal, copper, also displayed antagonist properties at the human P2X7 receptor, reducing the maximum response to Bz-ATP by about 50% at a concentration of 1 microM. 7. These data demonstrate that the human recombinant P2X7 receptor displays functional behaviour which is similar to the recombinant rat P2X7 receptor, but has a distinct pharmacological profile with respect to agonist and antagonist sensitivity.

Adenosine Triphosphate↗

PubMed1998

P2X receptor-mediated excitation of nociceptive afferents in the normal and arthritic rat knee joint.

1. We tested the hypothesis that functional P2X receptors are present on peripheral terminals of primary afferent articular nociceptors in the rat knee joint. Neural activity was recorded extracellularly from the medial articular nerve innervating the knee joint in rats anaesthetized with pentobarbitone. 2. The selective P2X receptor agonist, alphabeta methylene ATP (alphabetameATP), and the endogenous ligand, ATP, caused a rapid short-lasting excitation of a sub-population of C and Adelta nociceptive afferent nerves innervating normal knee joints when injected intra-arterially or intra-articularly, and this effect was antagonized by the non-selective P2 receptor antagonist PPADS. 3. Induction of a chronic (14-21 days) unilateral inflammatory arthritis of the knee joint using locally injected Freund's adjuvant neither increased or decreased responsiveness of joint nociceptors to alphabetameATP or ATP. 4. Our results support the hypothesis that alphabetameATP-sensitive P2X receptors are expressed on peripheral nociceptive afferents in the rat knee joint suggesting that they may be involved in the initiation of nociception and pain.

Adenosine Triphosphate↗

PubMed1998

Adenosine A1 receptor-mediated excitation of nociceptive afferents innervating the normal and arthritic rat knee joint.

We tested the hypothesis that adenosine excites nociceptive primary afferents innervating the knee joint. Neuronal recordings were made from fine nerve filaments innervating the knee joint in rats anaesthetized with pentobarbitone. Drugs were injected close-arterially (i.a.) or into the articular space (i.art.). We studied normal and chronically inflamed arthritic joints, the latter 14-21 days after a single intra-articular injection of Freund's Complete Adjuvant, performed under halothane anaesthesia. Adenosine injected i.a. caused delayed (approximately 10 s) excitation of the majority of polymodal C-fibre afferents, and had similar effects when injected directly into the joint. Adenosine triphosphate (ATP) had biphasic effects on discharge, a fast (<1 s) excitation was followed by a delayed increase similar to that seen with adenosine. The adenosine A1 receptor agonists N6-cyclopentyladenosine (CPA) and N-[(1S,trans)-2-hydroxypentyl] adenosine (GR79236) also excited the C-fibre afferents. The A1 antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) antagonized the responses evoked by adenosine, CPA, and the delayed increase seen after ATP, indicating that excitation of the nociceptive afferents was mediated via adenosine A1 receptors. Adenosine and ATP evoked delayed excitatory effects of similar magnitude, regardless of whether or not the knee joint was chronically inflamed. The increased basal discharge observed in arthritic joints was unaffected by DPCPX, which implies that the increase in spontaneous activity associated with arthritis is unlikely to involve tonically released adenosine. The results support the hypothesis that adenosine excites primary afferent nociceptive nerve terminals in the rat knee joint, an effect mediated by adenosine A1 receptors. ATP, adenosine, and A1 receptors may play a role in generating the peripheral nociceptive (pain) signal.

Action Potentials↗