An excess of cosmic ray electrons at energies of 300-800 GeV.

Galactic cosmic rays consist of protons, electrons and ions, most of which are believed to be accelerated to relativistic speeds in supernova remnants. All components of the cosmic rays show an intensity that decreases as a power law with increasing energy (for example as E(-2.7)). Electrons in particular lose energy rapidly through synchrotron and inverse Compton processes, resulting in a relatively short lifetime (about 10(5) years) and a rapidly falling intensity, which raises the possibility of seeing the contribution from individual nearby sources (less than one kiloparsec away). Here we report an excess of galactic cosmic-ray electrons at energies of approximately 300-800 GeV, which indicates a nearby source of energetic electrons. Such a source could be an unseen astrophysical object (such as a pulsar or micro-quasar) that accelerates electrons to those energies, or the electrons could arise from the annihilation of dark matter particles (such as a Kaluza-Klein particle with a mass of about 620 GeV).

The multispanning membrane protein Ste24p catalyzes CAAX proteolysis and NH2-terminal processing of the yeast a-factor precursor.

Saccharomyces cerevisiae Ste24p is a multispanning membrane protein implicated in the CAAX proteolysis step that occurs during biogenesis of the prenylated a-factor mating pheromone. Whether Ste24p acts directly as a CAAX protease or indirectly to activate a downstream protease has not yet been established. In this study, we demonstrate that purified, detergent-solubilized Ste24p directly mediates CAAX proteolysis in a zinc-dependent manner. We also show that Ste24p mediates a separate proteolytic step, the first NH(2)-terminal cleavage in a-factor maturation. These results establish that Ste24p functions both as a bona fide COOH-terminal CAAX protease and as an a-factor NH(2)-terminal protease. Importantly, this study is the first to directly demonstrate that a eukaryotic multispanning membrane protein can possess intrinsic proteolytic activity.

The yeast a-factor transporter Ste6p, a member of the ABC superfamily, couples ATP hydrolysis to pheromone export.

ATP-binding cassette (ABC) proteins transport a diverse collection of substrates. It is presumed that these proteins couple ATP hydrolysis to substrate transport, yet ATPase activity has been demonstrated for only a few. To provide direct evidence for such activity in Ste6p, the yeast ABC protein required for the export of a-factor mating pheromone, we established conditions for purification of Ste6p in biochemical quantities from both yeast and Sf9 insect cells. The basal ATPase activity of purified and reconstituted Ste6p (V(max) = 18 nmol/mg/min; K(m) for MgATP = 0.2 mm) compares favorably with several other ABC proteins and was inhibited by orthovanadate in a profile diagnostic of ABC transporters (apparent K(I) = 12 microm). Modest stimulation (approximately 40%) was observed upon the addition of a-factor either synthetic or in native form. We also used an 8-azido-[alpha-(32)P]ATP binding and vanadate-trapping assay to examine the behavior of wild-type Ste6p and two different double mutants (G392V/G1087V and G509D/G1193D) shown previously to be mating-deficient in vivo. Both mutants displayed a diminished ability to hydrolyze ATP, with the latter uncoupled from pheromone transport. We conclude that Ste6p catalyzes ATP hydrolysis coupled to a-factor transport, which in turn promotes mating.

Biochemical studies of Zmpste24-deficient mice.

Genetic studies in Saccharomyces cerevisiae identified two genes, STE24 and RCE1, involved in cleaving the three carboxyl-terminal amino acids from isoprenylated proteins that terminate with a CAAX sequence motif. Ste24p cleaves the carboxyl-terminal "-AAX" from the yeast mating pheromone a-factor, whereas Rce1p cleaves the -AAX from both a-factor and Ras2p. Ste24p also cleaves the amino terminus of a-factor. The mouse genome contains orthologues for both yeast RCE1 and STE24. We previously demonstrated, with a gene-knockout experiment, that mouse Rce1 is essential for development and that Rce1 is entirely responsible for the carboxyl-terminal proteolytic processing of the mouse Ras proteins. In this study, we cloned mouse Zmpste24, the orthologue for yeast STE24 and showed that it could promote a-factor production when expressed in yeast. Then, to assess the importance of Zmpste24 in development, we generated Zmpste24-deficient mice. Unlike the Rce1 knockout mice, Zmpste24-deficient mice survived development and were fertile. Since no natural substrates for mammalian Zmpste24 have been identified, yeast a-factor was used as a surrogate substrate to investigate the biochemical activities in membranes from the cells and tissues of Zmpste24-deficient mice. We demonstrate that Zmpste24-deficient mouse membranes, like Ste24p-deficient yeast membranes, have diminished CAAX proteolytic activity and lack the ability to cleave the amino terminus of the a-factor precursor. Thus, both enzymatic activities of yeast Ste24p are conserved in mouse Zmpste24, but these enzymatic activities are not essential for mouse development or for fertility.

Alvimopan* (ADL 8-2698) is a novel peripheral opioid antagonist.

Alvimopan (ADL 8-2698; Adolor Corporation, Exton, PA, USA) is a novel, peripherally restricted opioid antagonist. After oral administration, it has activity specific to the gastrointestinal (GI) tract. ADL 8-2698 has low systemic absorption and a high affinity for mu-opioid receptors. In healthy subjects, ADL 8-2698 antagonized loperamide-induced changes in GI transit and prevented morphine-induced delays in oral-cecal transit time without antagonizing centrally mediated opioid effects, such as analgesia or pupillary constriction. In the treatment of opioid naive patients who underwent surgery and received opioids for acute pain, oral ADL 8-2698 (6.0 mg) improved the management of postoperative ileus (POI) by shortening the time to achieve normal bowel function and, ultimately, hospital stay. Postoperative nausea and vomiting and the overall incidence of all GI side effects were reduced in patients treated with ADL 8-2698 for POI. Analgesia was not compromised, because there were no changes in median opioid consumption or Visual Analog Scale (VAS) pain scores in patients treated with ADL 8-2698 versus patients treated with placebo. No drug-related side effects were observed in acute pain postsurgical patients in the initial POI study. In patients treated with opioids for chronic pain or opioid addiction, lower doses of oral ADL 8-2698 (0.5 to 3.0 mg) reversed opioid bowel dysfunction (OBD) and normalized GI activity. These effects were evident without compromising opioid analgesia or inducing central nervous system symptoms of withdrawal. Some chronic opioid patients receiving apparently supramaximal doses of ADL 8-2698 (> or = 3.0 mg) reported localized GI side effects, possibly indicative of a localized GI withdrawal response. The most common side effects of ADL 8-2698 in chronic pain patients with OBD were abdominal pain, flatulence, and diarrhea. These effects were not observed in most OBD patients receiving lower doses of ADL 8-2698. Overall, ADL 8-2698 was well tolerated in clinical trials. Further studies to evaluate the efficacy and safety of ADL 8-2698 in clinical practice are in progress.

Reconstitution of the Ste24p-dependent N-terminal proteolytic step in yeast a-factor biogenesis.

The yeast mating pheromone a-factor precursor contains an N-terminal extension and a C-terminal CAAX motif within which multiple posttranslational processing events occur. A recently discovered component in a-factor processing is Ste24p/Afc1p, a multispanning endoplasmic reticulum membrane protein that contains an HEXXH metalloprotease motif. Our in vivo genetic characterization of this protein has demonstrated roles for Ste24p in both the N-terminal and C-terminal proteolytic processing of the a-factor precursor. Here, we present evidence that the N-terminal proteolysis of the a-factor precursor P1 can be accurately reconstituted in vitro using yeast membranes. We show that this activity is dependent on Ste24p and is abolished by mutation of the Ste24p HEXXH metalloprotease motif or by mutation of the a-factor P1 substrate at a residue adjacent to the N-terminal P1 cleavage site. We also demonstrate that N-terminal proteolysis of the P1 a-factor precursor requires Zn(2+) as a co-factor and can be inhibited by the addition of the metalloprotease inhibitor 1,10-orthophenanthroline. Our results are consistent with Ste24p itself being the P1-->P2 a-factor protease or a limiting activator of this activity. Interestingly, we also show that the human Ste24 homolog expressed in yeast can efficiently promote the N-terminal processing of a-factor in vivo and in vitro, thus establishing a-factor as a surrogate substrate in the absence of known human substrates. The results reported here, together with the previously reported in vitro reconstitution of Ste24p-dependent CAAX processing, provide a system for examining the potential bifunctional roles of yeast Ste24p and its homologs.

Ste6p mutants defective in exit from the endoplasmic reticulum (ER) reveal aspects of an ER quality control pathway in Saccharomyces cerevisiae.

We are studying the intracellular trafficking of the multispanning membrane protein Ste6p, the a-factor transporter in Saccharomyces cerevisiae and a member of the ATP-binding cassette superfamily of proteins. In the present study, we have used Ste6p as model for studying the process of endoplasmic reticulum (ER) quality control, about which relatively little is known in yeast. We have identified three mutant forms of Ste6p that are aberrantly ER retained, as determined by immunofluorescence and subcellular fractionation. By pulse-chase metabolic labeling, we demonstrate that these mutants define two distinct classes. The single member of Class I, Ste6-166p, is highly unstable. We show that its degradation involves the ubiquitin-proteasome system, as indicated by its in vivo stabilization in certain ubiquitin-proteasome mutants or when cells are treated with the proteasome inhibitor drug MG132. The two Class II mutant proteins, Ste6-13p and Ste6-90p, are hyperstable relative to wild-type Ste6p and accumulate in the ER membrane. This represents the first report of a single protein in yeast for which distinct mutant forms can be channeled to different outcomes by the ER quality control system. We propose that these two classes of ER-retained Ste6p mutants may define distinct checkpoint steps in a linear pathway of ER quality control in yeast. In addition, a screen for high-copy suppressors of the mating defect of one of the ER-retained ste6 mutants has identified a proteasome subunit, Hrd2p/p97, previously implicated in the regulated degradation of wild-type hydroxymethylglutaryl-CoA reductase in the ER membrane.

Endoplasmic reticulum membrane localization of Rce1p and Ste24p, yeast proteases involved in carboxyl-terminal CAAX protein processing and amino-terminal a-factor cleavage.

Proteins terminating in the CAAX motif, for example Ras and the yeast a-factor mating pheromone, are prenylated, trimmed of their last three amino acids, and carboxyl-methylated. The enzymes that mediate these activities, collectively referred to as CAAX processing components, have been identified genetically in Saccharomyces cerevisiae. Whereas the Ram1p/Ram2p prenyltransferase is a cytosolic soluble enzyme, sequence analysis predicts that the other CAAX processing components, the Rce1p and Ste24p proteases and the Ste14p methyltransferase, contain multiple membrane spans. To determine the intracellular site(s) at which CAAX processing occurs, we have examined the localization of the CAAX proteases Rce1p and Ste24p by subcellular fractionation and indirect immunofluorescence. We find that both of these proteases are associated with the endoplasmic reticulum (ER) membrane. In addition to having a role in CAAX processing, the Ste24p protease catalyzes the first of two cleavage steps that remove the amino-terminal extension from the a-factor precursor, suggesting that the first amino-terminal processing step of a-factor maturation also occurs at the ER membrane. The ER localization of Ste24p is consistent with the presence of a carboxyl-terminal dilysine ER retrieval motif, although we find that mutation of this motif does not result in mislocalization of Ste24p. Because the ER is not the ultimate destination for a-factor or most CAAX proteins, our results imply that a mechanism must exist for the intracellular routing of CAAX proteins from the ER membrane to other cellular sites.

The Saccharomyces cerevisiae prenylcysteine carboxyl methyltransferase Ste14p is in the endoplasmic reticulum membrane.

Eukaryotic proteins containing a C-terminal CAAX motif undergo a series of posttranslational CAAX-processing events that include isoprenylation, C-terminal proteolytic cleavage, and carboxyl methylation. We demonstrated previously that the STE14 gene product of Saccharomyces cerevisiae mediates the carboxyl methylation step of CAAX processing in yeast. In this study, we have investigated the subcellular localization of Ste14p, a predicted membrane-spanning protein, using a polyclonal antibody generated against the C terminus of Ste14p and an in vitro methyltransferase assay. We demonstrate by immunofluorescence and subcellular fractionation that Ste14p and its associated activity are localized to the endoplasmic reticulum (ER) membrane of yeast. In addition, other studies from our laboratory have shown that the CAAX proteases are also ER membrane proteins. Together these results indicate that the intracellular site of CAAX protein processing is the ER membrane, presumably on its cytosolic face. Interestingly, the insertion of a hemagglutinin epitope tag at the N terminus, at the C terminus, or at an internal site disrupts the ER localization of Ste14p and results in its mislocalization, apparently to the Golgi. We have also expressed the Ste14p homologue from Schizosaccharomyces pombe, mam4p, in S. cerevisiae and have shown that mam4p complements a Deltaste14 mutant. This finding, plus additional recent examples of cross-species complementation, indicates that the CAAX methyltransferase family consists of functional homologues.

Synthesis and opioid activity of [D-Pro10]dynorphin A-(1-11) analogues with N-terminal alkyl substitution.

Several N-terminal di- and monoalkylated derivatives of [D-Pro10]dynorphin A-(1-11) were synthesized in order to explore the structure-activity relationships for antagonist vs agonist activity at kappa-opioid receptors. N,N-Dialkylated and N-monoalkylated (alkyl = allyl, benzyl, and cyclopropylmethyl (CPM) tyrosine derivatives were prepared from tyrosine tert-butyl ester and the corresponding alkyl halides. [D-Pro10]Dyn A-(2-11) was prepared by solid phase synthesis using Fmoc-protected amino acids, and the tyrosine derivatives were coupled to the peptide with BOP ((benzotriazol-1-yloxy)tris(dimethylamino)phosphonium hexafluorophosphate). Both the degree of substitution and the identity of the alkyl group affected kappa-receptor affinity, selectivity, and efficacy. All of the N-monoalkylated derivatives exhibited much higher affinity (Ki < 0.05 nM) for kappa receptors in the guinea pig cerebellum and greatly enhanced kappa-receptor selectivity (Ki ratio (kappa/mu) > 200) compared to the N,N-dialkyl [D-Pro10]Dyn A-(1-11) analogues, although one disubstituted analogue, N,N-diCPM[D-Pro10]Dyn A-(1-11), retained high affinity (Ki = 0.19 nM) for kappa receptors. Thus the introduction of the second alkyl group at the N-terminus lowered kappa-receptor affinity and selectivity. The N-allyl and N-CPM analogues were moderately potent agonists in the guinea pig ileum (GPI) assay, while the N-benzyl derivative was a weak agonist in this assay. In vivo in the phenylquinone abdominal stretching assay the N-CPM analogue exhibited potent antinociceptive activity (ED50 = 1.1 micrograms/mouse), while N-allyl[D-Pro10]Dyn A-(1-11) exhibited weak antinociceptive activity (ED50 = 27 micrograms/mouse). For the N,N-dialkyl derivatives the identity of the N-terminal alkyl group affected the efficacy observed in the smooth muscle assays. The N,N-diCPM analogue exhibited negligible agonist activity, and N,N-diallyl[D-Pro10]Dyn A-(1-11) showed weak antagonist activity against Dyn A-(1-13)NH2 in the GPI. In contrast, the N,N-dibenzyl compound showed appreciable opioid agonist activity in this assay. In vivo the N,N-diallyl analogue exhibited weak antinociceptive activity (ED50 = 26 micrograms/mouse in the phenylquinone abdominal stretching assay). The N-monoalkylated peptides are among the most kappa-selective opioid peptides reported to date, showing comparable or greater selectivity and higher affinity than the kappa-selective non-peptide agonists U-50,488 and U-69,593. The N,N-diCPM and N,N-diallyl peptides are lead compounds in the development of peptide-based kappa-receptor antagonists.

Ionic milieu controls the compartment-specific activation of pro-opiomelanocortin processing in AtT-20 cells.

Newly synthesized prohormones and their processing enzymes transit through the same compartments before being packaged into regulated secretory granules. Despite this coordinated intracellular transport, prohormone processing does not occur until late in the secretory pathway. In the mouse pituitary AtT-20 cell line, conversion of pro-opiomelanocortin (POMC) to mature adrenocorticotropic hormone involves the prohormone convertase PC1. The mechanism by which this proteolytic processing is restricted to late secretory compartments is unknown; PC1 activity could be regulated by compartment-specific activators/inhibitors, or through changes in the ionic milieu that influence its activity. By arresting transport in a semi-intact cell system, we have addressed whether metabolically labeled POMC trapped in early secretory compartments can be induced to undergo conversion if the ionic milieu in these compartments is experimentally manipulated. Prolonged incubation of labeled POMC trapped in the endoplasmic reticulum or Golgi/trans-Golgi network did not result in processing, thereby supporting the theory that processing is normally a post-Golgi/trans-Golgi network event. However, acidification of these compartments allowed effective processing of POMC to the intermediate and mature forms. The observed processing increased sharply at a pH below 6.0 and required millimolar calcium, regardless of the compartment in which labeled POMC resided. These conditions also resulted in the coordinate conversion of PC1 from the 84/87 kDa into the 74-kDa and 66-kDa forms. We propose that POMC processing is predominantly restricted to acidifying secretory granules, and that a change in pH within these granules is both necessary and sufficient to activate POMC processing.

Synthesis and targeting of insulin-like growth factor-I to the hormone storage granules in an endocrine cell line.

Export of growth factors is generally believed to be restricted to the constitutive secretory pathway, whereas peptide hormones are typically secreted in a regulated manner. Here we show that insulin-like growth factor (IGF)-I, a growth factor released constitutively from the liver, is synthesized and secreted from the mouse pituitary AtT-20 cell line via the regulated pathway. IGF-I production is 1500-fold less than the peptide hormone ACTH. Secretagogue induces IGF-I secretion in a manner similar to ACTH. Like ACTH, IGF-I is sorted into the regulated pathway >35-fold more efficiently than a constitutively secreted protein. Dense core granules isolated from cells transfected with a human IGF-I cDNA contain both ACTH and human IGF-I. AtT-20 cells also synthesize IGF-binding proteins, and at least one of these is secreted by the regulated pathway. Human IGF-I does not exhibit milieu-induced, concentration-dependent aggregation, in contrast to secretogranin II which sorts by a proposed aggregation mechanism. These data suggest that 1) growth factors are not solely released from tissues via the constitutive pathway, 2) IGF-I may contain information for correct granular targeting, and 3) IGF-I may be sorted by a mechanism distinct from that proposed for the secretogranins.

Evaluation of delayed treatment of focal cerebral ischemia with three selective kappa-opioid agonists in cats.

BACKGROUND AND PURPOSE: The purpose of this study was to determine the therapeutic efficacy of three kappa-opioid agonists used for delayed treatment of experimental focal cerebral ischemia. METHODS: Forty halothane-anesthetized cats underwent permanent occlusion of the right intracranial internal carotid, middle cerebral, and anterior cerebral arteries via a transorbital, microsurgical approach. Six hours after occlusion, animals received a blinded bolus injection, and a subcutaneous osmotic pump was implanted to provide continuous release for 7 days. The injection and pump contained either saline or one of three kappa-agonists: dynorphin (1-13), U-50,488, or DuP E3800. Survival, neurological function, tissue damage, and brain weight were assessed. RESULTS: As a group, kappa-agonist-treated animals had higher survival (P < .02), less tissue damage (P < .02), and lower brain weight (P < .05) than saline controls. U-50,488 more effectively improved survival (P < .03) than dynorphin (P < .07) or E3800 (P < .07). Each of the three kappa compounds improved tissue damage (dynorphin, P < .02; U-50,488, P < .05; E3800, P < .05). Greater improvement in neurological function was seen after treatment with dynorphin (P < .05) than with U-50,488 (P < .6) or E3800 (P < .7). The only significant reduction in brain weight was seen after dynorphin treatment (P < .01). CONCLUSIONS: Compounds that act at the kappa subclass of opiate receptors are effective in increasing survival, improving neurological function, and decreasing tissue damage and edema in a cat model of focal cerebral ischemia. The current study provides support for the benefits of treatment of acute cerebrovascular ischemia with kappa-opioid agonists. The agents may prove to be of superior clinical utility because of efficacy even when administered 6 hours after the onset of stroke.

Synthesis and opioid activity of 7-oxygenated 2,3,4,4a,5,6,7,7a-octahydro-1H-benzofuro[3,2-e]isoquinolin-9-ols.

3-(Cyclopropylmethyl)-9-hydroxy-7-oxo-2,3,4,4a alpha,5,6,7,7a alpha- octahydro-1H-benzofuro[3,2-e]isoquinoline (4b) containing the ACNO ring system of morphine and a 7-keto function on ring C has been synthesized and found to possess potent PQW (ED50 = 0.15 mg/kg sc) and anti-Straub tail (ED50 = 0.02 mg/kg sc) activity. As compared to its 7-deoxy analog 1b, introduction of the 7-keto group did not significantly affect binding to any of the three opioid receptors (mu, kappa, and delta), but caused a 34-fold reduction in sigma-binding, suggesting reduced propensity to induce psychotomimetic effects. The C/D cis isomer of 4b (4c) was much less potent at the three opioid receptors, while displaying a slight increase in sigma affinity. Both 7-hydroxy derivatives 4e and 4f were active in anti-Straub tail assay (ED50 < or = 0.8 mg/kg sc), but only the alpha-isomer 4e demonstrated analgesic activity (PQW ED50 = 0.37 mg/kg sc) in the dose range tested. In guinea pig ileum preparations, 4e was characterized as a selective full agonist at the kappa opioid receptor (IC50 = 2.8 nM); while its beta-isomer 4f was a partial agonist (78% at 1 microM), with antagonist activity observed at both mu- and kappa-opioid receptors.

Assessing spiradoline-like discriminative effects of DuP 747: influence of route of administration.

DuP 747, trans-3,4-dichloro-N-methyl-N-[2-(pyrrolidin-1-yl)-1,2,3,4- tetrahydronaphthalen-1-yl]benzeacetamide methanesfonate, is a recently synthesized analgesic drug that binds with high affinity and selectivity to the kappa-opioid receptor. In order to determine if DuP 747 has kappa-like discriminative effects it was tested for stimulus generalization in rats trained to discriminate between SC injections of saline and 3.0 mg/kg of spiradoline, a potent kappa-opioid agonist. A range of drug doses was administered by each of several routes 30 min before a test session. Spiradoline occasioned orderly dose-dependent increases in spiradoline-appropriate lever selection after SC or IP administration, with ED50s of 0.65 and 1.75 mg/kg, respectively. In contrast, DuP 747 (1.0-30 mg/kg) occasioned little spiradoline-appropriate lever selection when administered SC, but was generalized from spiradoline partially when administered IP (ED50 = 5.9 mg/kg) or PO (ED50 = 59 mg/kg). The 5-hydroxy-desmethoxy metabolite of DuP 747, administered SC (0.3-10 mg/kg), occasioned selection of the saline-appropriate lever only. That DuP 747 had little spiradoline-like activity after SC administration suggests that a metabolite of DuP 747 was responsible for the spiradoline-appropriate responding that followed IP and PO administration of the drug, apparently a metabolite other than 5-hydroxy-desmethoxy-DuP 747.

DuP 734 [1-(cyclopropylmethyl)-4-(2'(4''-fluorophenyl)-2'-oxoethyl)- piperidine HBr], a sigma and 5-hydroxytryptamine2 receptor antagonist: receptor-binding, electrophysiological and neuropharmacological profiles.

It has been suggested that sigma receptor antagonists may be useful as antipsychotic drugs and that 5-hydroxytryptamine (5-HT2) receptor antagonists produce improvements of the negative symptoms of schizophrenia. [1-(Cyclopropylmethyl)-4-(2'-(4''-fluorophenyl)-2'- oxoethyl)-piperidine HBr] (DuP 734) is a novel compound with high affinity for the sigma (Ki = 10 nM) and 5-HT2 (Ki = 15 nM) receptors, but low affinity for dopamine receptors (Ki > 1000 nM) as well as 33 other receptors, ion channels and second messenger systems in vitro. DuP 734 did not inhibit the synaptosomal uptake of dopamine, 5-HT or norepinephrine. Oral administration of DuP 734 potently blocked 5-hydroxy-L-trytophan (5-HTP)-induced head twitch in the rat (ED50 = 6.5 mumol/kg), indicating 5-HT2 antagonist activity. Extracellular single-unit recording studies demonstrated that DuP 734 antagonized the effect of the selective sigma ligand (+)-3-(3-hydroxyphenyl-N-(1-propyl) piperidine [(+)-3-PPP] on dopamine neuronal activity in the substantia nigra of the rat with an ED90 of 3.6 mumol/kg i.v. The sigma receptor agonists (+)-SKF 10,047 and phencyclidine both elicited rotational behavior in rats with unilateral lesion of the substantia nigra. The rotational behavior induced by either (+)-SKF 10,047 or phencyclidine was dose-dependently antagonized by DuP 734 with oral ED50 of 8.7 and 19.6 mumol/kg, respectively. The 5-HT2 receptor antagonist ICI 169,369, even at high doses (up to 33 mumol/kg, s.c.), did not antagonize the rotational behavior induced by (+)-SKF 10,047.(ABSTRACT TRUNCATED AT 250 WORDS)

Novel piperidine sigma receptor ligands as potential antipsychotic drugs.

sigma receptor ligands represent a new class of potential antipsychotic drugs. This paper presents the structure-activity relationships leading to novel disubstituted piperidine sigma ligands, which have little or no affinity for dopamine D2 receptors. Selectivity for sigma sites over dopamine D2 or serotonin 5-HT2 receptors appears to be governed by the chemical nature of the piperidine nitrogen substituent, its distance from the basic nitrogen, and its orientation relative to the other piperidine substituent. Several of these compounds have good oral potency in some animal models used to evaluate potential antipsychotic drugs. The N-cyclopropylmethyl ketones and ethers (e.g. 6i (DuP 734), 6q, 18a, and 18n) have the best in vivo potency. Compounds 6i (DuP 734) and 6q did not cause catalepsy in the rat, even at very high doses. On the basis of the pharmacology profiles of these sigma ligands, we propose these compounds may be effective antipsychotic drugs, which do not induce extrapyramidal side effects or tardive dyskinesia.

Anti-inflammatory and safety profile of DuP 697, a novel orally effective prostaglandin synthesis inhibitor.

DuP 697 (5-bromo-2[4-fluorophenyl]-3-[4-methylsulfonylphenyl]-thiophene) is a potent inhibitor of paw swelling in nonestablished and established adjuvant arthritis in rats (ED50 = 0.03 and 0.18 mg/kg/day, respectively). DuP 697 had no effect on phenylquinone writhing in rats (ED50 greater than 100 mg/kg), but was analgetic against inflammation-related pain in the Randall-Selitto assay (ED30 = 3.5 mg/kg) and was a very potent antipyretic agent (ED50 = 0.05 mg/kg). The drug was not ulcerogenic in rats at single doses up to 400 mg/kg. DuP 697 (5 mg/kg i.v.) did not alter renal blood flow or the renal vascular response to angiotensin II in furosemide-pretreated, volume-depleted rats. In contrast, indomethacin (5 mg/kg i.v.) decreased renal blood flow and potentiated the renal vascular response to angiotensin II in these animals. DuP 697 was a moderate inhibitor of bull seminal vesicle prostaglandin (PG) synthesis (IC50 = 2.4 X 10(-5) M) and a potent inhibitor of rat brain PG synthesis (IC50 = 4.5 X 10(-6) M) but was ineffective against rat kidney PG synthesis (IC50 7.5 X 10(-5) M). These differential effects of DuP 697 on PG synthesis by various tissues may account for its high potency as an anti-inflammatory and antipyretic agent and its minimal toxicity profile.