Novel P2X7 receptor antagonists ease the pain.

In recent months, a series of chemically diverse antagonists has been identified for the ATP-gated P2X(7) receptor. In particular, two classes of highly-selective competitive P2X(7) antagonists have been developed by Michael Jarvis and his colleagues at Abbott Laboratories. These di-substituted tetrazole and cyanoguanidine derivatives are outstanding for a number of reasons (not least their stability, selectivity, potency and, of course, reversibility); most exciting is their near equal potency at human and rodent P2X(7) isoforms. Armed with drugs such as A740003 and newer A438079, Jarvis and colleagues have explored the role of P2X(7) receptors in the onset and persistence of chronic pain in animal models. Their findings - and applicability to the human condition - are reviewed in this current issue of British Journal of Pharmacology. This accompanying Commentary describes the progress made by Jarvis and others in developing novel P2X(7) antagonists for pain relief.

Image quality and dose comparison among screen-film, computed, and CT scanned projection radiography: applications to CT urography.

PURPOSE: To evaluate image quality and dose for abdominal imaging techniques that could be used as part of a computed tomographic (CT) urographic examination: screen-film (S-F) radiography or computed radiography (CR), performed with moving and stationary grids, and CT scanned projection radiography (CT SPR). MATERIALS AND METHODS: An image quality phantom underwent imaging with moving and stationary grids with both a clinical S-F combination and CR plate. CT SPR was performed with six CT scanners at various milliampere second and kilovolt peak settings. Entrance skin exposure (ESE); spatial, contrast, and temporal resolutions; geometric accuracy; and artifacts were assessed. RESULTS: S-F or CR images, with either grid, provided image quality equivalent to that with the clinical standard, S-F with a moving grid. ESE values for both S-F and CR were 435 mR (112.2 microC/kg [1 mR = 0.258 microC/kg]) with a moving grid and 226 mR (58.3 microC/kg) with a stationary grid. All CT SPR images provided inferior spatial resolution compared with S-F or CR images. High-contrast objects generated substantial artifacts on CT SPR images. Compared with S-F, CR and CT SPR provided improved resolution of small low-contrast objects. The contrast between iodine and soft-tissue-mimicking structures on CT SPR images acquired at 80 kVp was twice that at 120 kVp. CT SPR images with acceptable noise levels required a midline ESE value of approximately 300 mR (77.4 microC/kg) at 80 kVp. CONCLUSION: S-F and CR provided better spatial resolution than did CT SPR. However, CT SPR provided improved low-contrast resolution compared with S-F, at exposures comparable to those used for S-F or CR.

An auxiliary CT tabletop for radiography at the time of CT.

An auxiliary CT tabletop was designed and manufactured such that radiographic images might be acquired, with use of a ceiling-mounted X-ray tube, without removing the patient from the CT table. The tabletop required no modifications to the original CT table housing and did not produce artifacts in the CT images. Radiographs obtained with the overhead X-ray tube and auxiliary tabletop demonstrated image quality equivalent to traditional radiographs.

Embedded MR fluoroscopy: high temporal resolution real-time imaging during high spatial resolution 3D MRA acquisition.

A method termed "embedded fluoroscopy" for simultaneously acquiring a real-time sequence of 2D images during acquisition of a 3D image is presented. The 2D images are formed by periodically sampling the central phase encodes of the slab-select direction during the 3D acquisition. The tradeoffs in spatial and temporal resolution are quantified by two parameters: the "redundancy" (R), the fraction of the 3D acquisition sampled more than once; and the "effective temporal resolution" (T), the time between temporal updates of the central views. The method is applied to contrast-enhanced MR angiography (CE-MRA). The contrast bolus dynamics are portrayed in real time in the 2D image sequence while a high-resolution 3D image is being acquired. The capability of the 2D acquisition to measure contrast enhancement with only a 5% degradation of the spatial resolution of the 3D CE-MR angiogram is shown theoretically. The method is tested clinically in 15 CE-MRA patient studies of the carotid and renal arteries.

Selective agonism of group I P2X receptors by dinucleotides dependent on a single adenine moiety.

We have investigated the activity of naturally occurring high-performance liquid chromatography-purified diadenosine polyphosphates (Ap(n)A, n = 5-6), adenosine polyphospho guanosines (Ap(n)G, n = 5-6), and diguanosine polyphosphates (Gp(n)G, n = 5-6) under voltage-clamp conditions at recombinant rat P2X1-4 purinoceptor subtypes expressed in Xenopus laevis oocytes. At rP2X1 and rP2X3 receptors, Ap(n)As and Ap(n)Gs evoked concentration-dependent inward currents. Gp(n)Gs were not active at these receptors. At rP2X2 and rP2X4 receptors, dinucleotides did not show significant activity. For the rP2X1 receptor, Ap(n)As and Ap(n)Gs were partial agonists; for the P2X3 receptor, only Ap5G was full agonist, whereas the other tested substances were partial agonists. The rank order of potency at rP2X1 was ATP > or = Ap6A > or = Ap5A > or = Ap6G > or = Ap5G, and rank order of efficacy was ATP > or = Ap5A > or = Ap6A > Ap5G > Ap6G, whereas at rP2X3 the rank order of potency was ATP > Ap5G > or = Ap5A > or = Ap6A > or = Ap6G and the rank order of efficacy was ATP approximately Ap5G > or = Ap5A approximately Ap6A > or = Ap6G. For rP2X1 and rP2X3 it is evident that receptor agonism depended on the presence of at least one adenine moiety in the dinucleotide, while the presence of a guanine moiety had a significant impact and decreased agonist efficacy. The data suggest that naturally occurring Ap(n)As and Ap(n)Gs may play an important physiological role in different human tissues and systems by activating group I P2X receptors.

Reflections on the purinergic hypothesis: the Burnstock Festschrift in the millennial year.

Few have made such an impact as Geoffrey Burnstock in their scientific field. As the originator of the purinergic hypothesis, Burnstock has been central to the development of our understanding of the P2 receptor family and of the role of extracellular ATP in cell-to-cell signalling. In this millennial year, Burnstock has been awarded the Queen's medal from The Royal Society and Lifetime Achievement Award from the American Gastroenterology Association. Thus, it was my privilege to join Alan North in organising and producing the Burnstock Festschrift (Purines and the Autonomic Nervous System; from controversy to the clinic, in [J. Auton. Nerv. Syst. Vol. 81 (2000)]) to honour not only Geoffrey Burnstock's successes in this millennial year, but a lifetime of achievements spanning some 40 years in the field of purine signalling.

Structure-activity relationships of pyridoxal phosphate derivatives as potent and selective antagonists of P2X1 receptors.

Novel analogues of the P2 receptor antagonist pyridoxal-5'-phosphate 6-azophenyl-2',5'-disulfonate (2) were synthesized and studied as antagonists in functional assays at recombinant rat P2X1, P2X2, and P2X3 receptors expressed in Xenopus oocytes (ion flux stimulation) and at turkey erythrocyte P2Y1 receptors (phospholipase C activation). Selected compounds were also evaluated as antagonists of ion flux and the opening of a large pore at the recombinant human P2X7 receptor. Modifications were made in the 4-aldehyde and 5'-phosphate groups of the pyridoxal moiety: i.e. a CH2OH group at the 4-position in pyridoxine was either condensed as a cyclic phosphate or phosphorylated separately to form a bisphosphate, which reduced potency at P2 receptors. 5-Methylphosphonate substitution, anticipated to increase stability to hydrolysis, preserved P2 receptor potency. At the 6-position, halo, carboxylate, sulfonate, and phosphonate variations made on the phenylazo ring modulated potency at P2 receptors. The p-carboxyphenylazo analogue, 4, of phosphate 2 displayed an IC50 value of 9 nM at recombinant P2X1 receptors and was 1300-, 16-, and > 10,000-fold selective for P2X1 versus P2X2, P2X3, and P2Y1 subtypes, respectively. The corresponding 5-methylphosphonate was equipotent at P2X1 receptors. The 5-methylphosphonate analogue containing a 6-[3,5-bis(methylphosphonate)]phenylazo moiety, 9, had IC50 values of 11 and 25 nM at recombinant P2X1 and P2X3 receptors, respectively. The analogue containing a phenylazo 4-phosphonate group, 11, was also very potent at both P2X1 and P2X3 receptors. However, the corresponding 2,5-disulfonate analogue, 10, was 28-fold selective for P2X1 versus P2X3 receptors. None of the analogues were more potent at P2X7 and P2Y1 receptors than 2, which acted in the micromolar range at these two subtypes.

International union of pharmacology. XXIV. Current status of the nomenclature and properties of P2X receptors and their subunits.

ATP acts as a humoral mediator to control cell function extracellularly. The receptors that mediate the actions of ATP belong to two classes, the metabotropic P2Y receptors and the transmitter-gated, ion channel P2X receptors. This review describes the structure, distribution, function, and ligand recognition characteristics of P2X receptors, which comprise seven distinct subunits that can function as both homo- and hetero- polymers. The pharmacology of P2X receptors is complicated by marked differences between species orthologues. The current nomenclature is based largely on recombinant receptor studies and detailed knowledge of endogenous P2X receptors in native tissues is limited because of lack of good selective agonists and antagonists for each receptor type.

High-spatial-resolution contrast-enhanced MR angiography of the renal arteries: a prospective comparison with digital subtraction angiography.

PURPOSE: To evaluate a high-spatial-resolution three-dimensional (3D) contrast material-enhanced magnetic resonance (MR) angiographic technique for detecting proximal and distal renal arterial stenosis. MATERIALS AND METHODS: Twenty-five patients underwent high-spatial-resolution small-field-of-view (FOV) 3D contrast-enhanced MR angiography of the renal arteries, which was followed several minutes later by more standard, large-FOV 3D contrast-enhanced MR angiography that included the distal aorta and iliac arteries. For both acquisitions, MR fluoroscopic triggering and an elliptic centric view order were used. Two readers evaluated the MR angiograms for grade and hemodynamic significance of renal arterial stenosis, diagnostic quality, and presence of artifacts. MR imaging results for each patient were compared with those of digital subtraction angiograms. RESULTS: The high-spatial-resolution small-FOV technique provided high sensitivity (97%) and specificity (92%) for the detection of renal arterial stenosis, including all four distal stenoses encountered. The portrayal of the segmental renal arteries was adequate for diagnosis in 19 (76%) of 25 patients. In 12% of the patients, impaired depiction of the segmental arteries was linked to motion. CONCLUSION: The combined high-spatial-resolution small-FOV and large-FOV MR angiographic examination provides improved spatial resolution in the region of the renal arteries while maintaining coverage of the abdominal aorta and iliac arteries.

Coexpression of P2X(3) and P2X(2) receptor subunits in varying amounts generates heterogeneous populations of P2X receptors that evoke a spectrum of agonist responses comparable to that seen in sensory neurons.

Using voltage-clamp procedures on Xenopus oocytes, agonist-evoked ionic currents by P2X receptors resulting from the coexpression of P2X(2) and P2X(3) subunits were compared against the agonist responses of homomeric P2X(2) and P2X(3) receptors. With the quantity of P2X(3) mRNA kept constant and quantity of P2X(2) mRNA progressively increased, expressed P2X receptors changed from a P2X(3)-like receptor to a P2X(2)-like receptor. In all cases, however, agonist-evoked responses comprised biphasic (fast and slow) currents-the former showing the properties of P2X(3) receptors and latter consistent with the presence of P2X(2) and P2X(2/3) receptors. Using desensitization procedures, the P2X(3)-like fast current was selectively removed to allow the slow current to be studied in isolation. P2X(2/3) receptors were then characterized by slowly inactivating inward currents that were reproducible within 30 s of washout and whose pharmacological profile [selective agonists, Ap(5)A > alpha,beta-methylene ATP >> beta,gamma-methylene ATP > UTP; antagonists, TNP-ATP >> suramin > or = Reactive blue-2 (RB-2)] contrasted with the profile of P2X(2) receptors (Ap(5)A, alpha,beta-methylene ATP, beta,gamma-methylene ATP, and UTP inactive; antagonists, RB-2 > TNP-ATP > suramin). Thus, our experiments reveal that coexpression of two P2X subunits, which of themselves can generate functional homomeric receptors, results in a complex population of heterogeneous P2X receptors-in this case P2X(2), P2X(3), and P2X(2/3) receptors. Depending on the relative levels of P2X subunit coexpression, the operational profile of the resultant P2X receptors can change from one phenotype to another. This spectrum may explain the variability of agonist responses in small sensory neurons that also express P2X(2) and P2X(3) subunits in different amounts.

Building community capacity in public health: the role of action-oriented partnerships.

Public health practice increasingly is concerned with the capacity and performance of communities to identify, implement, strengthen, and sustain collective efforts to improve health. The authors developed ways to assist local Turning Point partnerships to improve their community public health system as a secondary outcome of their work on the expressed needs of the community. Using focus groups, meeting minutes, attendance records, and meeting observation, the authors fed information back to the partnerships on systems change. A public health systems improvement plan supportive of local partnerships' work on specific health issues was funded and the collaborative research agenda was further refined.

Role of adenosine in contrast media-induced acute renal failure in diabetes mellitus.

Increased release of renal adenosine and stimulation of renal adenosine receptors have been proposed to be major mechanisms in the development of contrast media-induced acute renal failure (CM-ARF). Patients with diabetes mellitus or preexisting renal disease who have reduced renal function have a markedly increased risk to develop CM-ARF. This increased risk to develop CM-ARF in patients with diabetes mellitus is linked to a higher sensitivity of the renal vasculature to adenosine, since experimental studies have shown increased adenosine-induced vasoconstriction in the kidneys of diabetic animals. Furthermore, recent evidence suggests that administration of adenosine receptor antagonists reduces the risk of development of CM-ARF in both diabetic and nondiabetic patients. The purpose of this review is to discuss the role of adenosine in the development of CM-ARF, particularly in the kidneys of diabetic patients, and to evaluate the therapeutic potential of adenosine receptor antagonists in the prevention of CM-ARF. Selective adenosine A1 receptor antagonists may provide a therapeutic tool to prevent CM-ARF in patients with diabetes mellitus and reduced renal function.

Identification of a locus for autosomal dominant polycystic liver disease, on chromosome 19p13.2-13.1.

Polycystic liver disease (PCLD) is characterized by the growth of fluid-filled cysts of biliary epithelial origin in the liver. Although the disease is often asymptomatic, it can, when severe, lead to complications requiring surgical therapy. PCLD is most often associated with autosomal dominant polycystic kidney disease (ADPKD); however, families with an isolated polycystic liver phenotype without kidney involvement have been described. The clinical presentation and histological features of polycystic liver disease in the presence or absence of ADPKD are indistinguishable, raising the possibility that the pathogenetic mechanisms in the diseases are interrelated. We ascertained two large families with polycystic liver disease without kidney cysts and performed a genomewide scan for genetic linkage. A causative gene, PCLD, was mapped to chromosome 19p13.2-13.1, with a maximum LOD score of 10.3. Haplotype analysis refined the PCLD interval to 12.5 cM flanked by D19S586/D19S583 and D19S593/D19S579. The discovery of genetic linkage will facilitate diagnosis and study of this underdiagnosed disease entity. Identification of PCLD will be instrumental to an understanding of the pathogenesis of cyst formation in the liver in isolated PCLD and in ADPKD.

Puboperineales: muscular boundaries of the male urogenital hiatus in 3D from magnetic resonance imaging.

PURPOSE: The aims of this report are 1) to extend our previous two-dimensional magnetic resonance imaging study to create a three-dimensional image of the pelvic floor, including the puboperinealis, the most anteromedial component of the levator ani; 2) to clarify the historical controversy about this particular component of the levator ani; and 3) to present clinical implications of this muscle with respect to urinary continence and radical prostatectomy. MATERIALS AND METHODS: We reused the axial magnetic resonance imaging series from 1 of 15 men in a previous series. Analyze AVWTM allowed creation of three-dimensional images. Further, a movie clip of all three-dimensional images was developed and placed at the manuscript-dedicated Web site: http://www.mayo. edu/ppmovie/pp.html. RESULTS: Our three-dimensional images show how the puboperinealis portion of the levator ani flanks the urethra as it courses from the pubis to its insertion in the perineal body. CONCLUSIONS: The puboperinealis corresponds to muscles previously designated as the levator prostatae, Wilson's muscle, pubourethralis, and levator urethrae, among others. The images suggest that the puboperinealis is the muscle most responsible for the quick stop phenomenon of urination in the male. Our study supports the suggestion that weakening of the puboperinealis by transection, traction injury, or denervation may affect urinary continence after radical prostatectomy.

Diinosine pentaphosphate: an antagonist which discriminates between recombinant P2X(3) and P2X(2/3) receptors and between two P2X receptors in rat sensory neurones.

1. We have compared the antagonist activity of trinitrophenyl-ATP (TNP-ATP) and diinosine pentaphosphate (Ip(5)I) on recombinant P2X receptors expressed in Xenopus oocytes with their actions at native P2X receptors in sensory neurones from dorsal root and nodose ganglia. 2. Slowly-desensitizing responses to alpha,beta-methylene ATP (alpha,beta-meATP) recorded from oocytes expressing P2X(2/3) receptors were inhibited by TNP-ATP at sub-micromolar concentrations. However, Ip(5)I at concentrations up to 30 microM was without effect. 3. Nodose ganglion neurones responded to alpha,beta-meATP with slowly-desensitizing inward currents. These were inhibited by TNP-ATP (IC(50), 20 nM), but not by Ip(5)I at concentrations up to 30 microM. 4. In DRG neurones that responded to ATP with a rapidly-desensitizing inward current, the response was inhibited by TNP-ATP with an IC(50) of 0.8 nM. These responses were also inhibited by Ip(5)I with an IC(50) of 0.1 microM. Both antagonists are known to inhibit homomeric P2X(3) receptors. 5. Some DRG neurones responded to alpha,beta-meATP with a biphasic inward current, consisting of transient and sustained components. While the transient current was abolished by 1 microM Ip(5)I, the sustained component remained unaffected. 6. In conclusion, Ip(5)I is a potent antagonist at homomeric P2X(3) receptors but not at heteromeric P2X(2/3) receptors, and therefore should be a useful tool for elucidating the subunit composition of native P2X receptors.

Coexpression of rat P2X2 and P2X6 subunits in Xenopus oocytes.

Transcripts for P2X(2) and P2X(6) subunits are present in rat CNS and frequently colocalize in the same brainstem nuclei. When rat P2X(2) (rP2X(2)) and rat P2X(6) (rP2X(6)) receptors were expressed individually in Xenopus oocytes and studied under voltage-clamp conditions, only homomeric rP2X(2) receptors were fully functional and gave rise to large inward currents (2-3 microA) to extracellular ATP. Coexpression of rP2X(2) and rP2X(6) subunits in Xenopus oocytes resulted in a heteromeric rP2X(2/6) receptor, which showed a significantly different phenotype from the wild-type rP2X(2) receptor. Differences included reduction in agonist potencies and, in some cases (e.g., Ap(4)A), significant loss of agonist activity. ATP-evoked inward currents were biphasic at the heteromeric rP2X(2/6) receptor, particularly when Zn(2+) ions were present or extracellular pH was lowered. The pH range was narrower for H(+) enhancement of ATP responses at the heteromeric rP2X(2/6) receptor. Also, H(+) ions inhibited ATP responses at low pH levels (

In search of selective P2 receptor ligands: interaction of dihydropyridine derivatives at recombinant rat P2X(2) receptors.

1,4-Dihydropyridines are regarded as privileged structures for drug design, i.e. they tend to bind to a wide variety of receptor sites. We have shown that upon appropriate manipulation of the substituent groups on a 1,4-dihydropyridine template, high affinity and selectivity for the A(3) subtype of adenosine receptors ('P1 receptors') may be attained. In the present study we have begun to extend this approach to P2 receptors which are activated by ATP and other nucleotides. Nicardipine, a representative dihydropyridine, used otherwise as an L-type calcium channel blocker, was shown to be an antagonist at recombinant rat P2X(2) (IC(50)=25 microM) and P2X(4) (IC(50) approximately 220 microM) receptors expressed in Xenopus oocytes. Thus, this class of compounds represents a suitable lead for enhancement of affinity through chemical synthesis. In an attempt to modify the 1,4-dihydropyridine structure with a predicted P2 receptor recognition moiety, we have replaced one of the ester groups with a negatively charged phosphonate group. Several 4-phenyl-5-phosphonato-1,4-dihydropyridine derivatives, MRS 2154 (2, 6-dimethyl), MRS 2155 (6-methyl-2-phenyl), and MRS 2156 (2-methyl-6-phenyl), were synthesized through three component condensation reactions. These derivatives were not pure antagonists of the effects of ATP at P2X(2) receptors, rather were either inactive (MRS 2156) or potentiated the effects of ATP in a concentration-dependent manner (MRS 2154 in the 0.3-10 microM range and MRS 2155 at >1 microM). Antagonism of the effects of ATP at P2X(2) receptor superimposed on the potentiation was also observed at >10 microM (MRS 2154) or 0.3-1 microM (MRS 2155). Thus, while a conventional dihydropyridine, nicardipine, was found to antagonize rat P2X(2) receptors ninefold more potently than P2X(4) receptors, the effects of novel, anionic 5-phosphonate analogues at the receptor were more complex.