DE 19933,926 A 1, 2001 EP OO76530 9, 1982 EP O 640 601 B1 12, 1996 v. 20, p. 99-112 (2008).* Mearin et al. "Levo Sulpiride and cisapride . . 2004156779 (2004).* Cisapride RN 81098-60-4 (1984).* Barnes, N.M., B. Costall, R.J. Naylor, F.D. Tattersall (Apr. 5, 1998) “Identification of 5-HT3 recognition sites in the ferret area postrema” J. Pharm. Pharmacol. 40:586-588. Decktor, Dennis, Robert G. Pendleton, Anne T. Elnitsky, Ann M. Jenkins, Anthony P. McDowell (1988) “Effect of metoclopramide, bethanechol and the cholecystokinin receptor antagonist, L-364,718, on gastric emptying in the rat European Journal of Pharmacology 147:313-316. Stacher, Georg, Gabriele Gaupmann, Gerda Mittelbach, Christa Scheider, Hermann Steinringer, Brigitte Langer (Nov. 1987) “Effects of Oral Cisapride on Interdigestive jejunal Motor Activity, Psychomotor Function, and Side-Effect Profile in Healthy Man” Digestive Diseases and Sciences 32(11): 1223-1230. Van Daele, Georges H.P. Marcel F.L. De Bruyn, Francois M. Som men et al. (1986) “Synthesis of Cisapride, a Gastrointestinal Stimu lant Derived From Cis-4-Amino-3-Methoxypiperidine” Drug Devel opment Research 8:225-232. EMBASE No.

Transgenic Drosophila melanogaster carrying the human gene for alpha synuclein is an animal model for the study of Parkinson's Disease. Climbing activity in these flies is reduced as a result of the effect of this protein on the locomotor activity of the transgenic fly. L-DOPA and gamma amino butyric acid (GABA) reverse the loss of this activity when placed in the food fed to these flies. While muscimol, a GABA(A) receptor agonist has no effect in this system, baclofen and the allosteric agonists CG 7930 and GS 39783 which affect the GABA(B) receptor reverse this activity. This latter effect is eliminated when these compounds are fed in conjunction with the GABA(B) receptor antagonist 2-hydroxysaclofen. In addition, fendiline which is a Ca(++) receptor blocker also reverses the loss of climbing ability. Because there is a calcium channel close to the GABA(B) receptor on the cell surface, these data are indicative of a relationship between the roles of the GABA(B) receptor, the calcium channel and the effect of alpha-synuclein on the motor activity of the transgenic fly.

Adult transgenic fruit flies (Drosophila melanogaster) carrying the human gene coding for the protein alpha-synuclein were tested for geotactic, locomotor, and phototactic behaviors as well as alpha-synuclein expression. Specific assays in adult conscious flies can be used to determine geotactic, phototactic, and locomotor behaviors. The presence of endogenous alpha-synuclein in the fly brain was assessed via immunological blot assays. The expression of monomeric human alpha-synuclein (19 kDa) in the brain of flies containing the homozygous human UAS wild-type alpha-synuclein transgene was shown by specific immunological blotting procedures. Behavioral testing indicated that motor rather than sensory functions were principally affected by the presence of alpha-synuclein, mimicking its effects in Parkinson's disease. These data illustrate that the human gene coding for alpha-synuclein can be expressed in the fly and produce behavioral effects in accord with behaviors seen in patients with Parkinson's disease.

Tyrosine hydroxylase (TH), the enzyme which catalyzes the conversion of tyrosine to L-DOPA and is rate limiting in catecholamine biosynthesis, is biochemically expressed in late stage wild-type Drosophila oocytes as well as in early embryogenesis. Null mutant alleles of TH (pale) are embryonic lethals with death occurring in the late embryonic or early larval periods of development. Staging of embryos demonstrated that inhibition of the enzymatic activity of TH by alpha-methyl-p-tyrosine (alphaMT) retards the progression of embryos primarily during the organogenesis stages of embryonic development, with lesser effects on earlier and later stages. On the other hand, time of gene action studies with a conditional temperature sensitive pale mutant (ple(ts1)) at its restrictive temperature (29 degrees C) indicate an onset of tyrosine hydroxylase gene action beginning in the oocyte stage of development. Thus, maternal as well as embryonic effects on the secretion and/or functionality of this enzyme may play roles in the early developmental program of the organism.

Tyrosine hydroxylase (TH), the enzyme which catalyzes the conversion of tyrosine to l-DOPA and is the rate limiting step in catecholamine biosynthesis, is genetically expressed during development in Drosophila. Null mutant alleles of the single copy gene which codes for this enzyme are developmentally lethal as is a conditional TH mutant at its restrictive temperature. In adult flies, inhibition of TH by alpha-methyl-p-tyrosine (αMT) decreases locomotor activity in a dose-dependent manner. This behavioral effect is accompanied by reductions in brain levels of dopamine, the primary CNS catecholamine in Drosophila, and can be prevented by the coadministration of l-DOPA. Similar effects are found with reserpine and at the restrictive temperature in flies with a temperature conditional mutation for TH. In agreement with published studies in mammals, inhibition of TH by αMT during Drosophila development results in enhanced expression of this enzyme in the progeny of surviving adults. This biochemical outcome is accompanied behaviorally by increased sensitivity to the locomotor effects of both αMT and reserpine, drugs which act via depletion of brain catecholamines. Since TH is the rate limiting enzyme responsible for the conversion of tyrosine to l-DOPA and l-DOPA is converted to dopamine by aromatic amino acid decarboxylase (AAAD), the results indicate that depletion of catecholamine levels in the fly embryo results in increased dopamine biosynthesis in the next generation accompanied by alterations in behavior.

Behavioral observations of Drosophila are as old as some of the early genetic studies of this organism. Because the genetics of Drosophila is studied extensively, the appearance of these specific behavioral patterns in adults and larvae has almost immediately led to investigations of their genetic control. Application of Drosophila to the study of movement disorders research must begin with a detailed understanding of fly behavioral genetics. There are obvious complications in all organisms that result in problems for studying the control of behavioral and motor patterns. Drosophila , however, still possess important advantages for these studies. The central nervous system is amenable to study. While it is complex, it is still possible to sort out changes within the brain and the peripheral nervous system. More importantly, however, is the wealth of knowledge associated with the genetic control of behavioral responses in Drosophila . The genetic manipulations resulting in mutagenesis and transgenesis have become standard, and it is probable that deliberate interference with gene activity and intentional interference with physiological processes in Drosophila can lead to a fuller understanding of behavioral and motor functions in higher organisms.

The brain of the adult fruit fly, Drosophila melanogaster, contains tyrosine hydroxylase, the rate-limiting enzyme required for catecholamine biosynthesis, as well as dopa decarboxylase. Catecholamines, principally dopamine, are also present. We have previously shown that pharmacological inhibition of tyrosine hydroxylase with α-methyl-p-tyrosine results in a dose-related inhibition of locomotor activity in adult organisms. Similar results were found with reserpine, a well-known inhibitor of catecholamine uptake into storage granules. The drug-induced inhibition could be prevented in each case by the concomitant administration of l-dopa. The single-copy gene coding for tyrosine hydroxylase in Drosophila is pale (ple). Both null and temperature-sensitive loss of function mutant alleles of ple are recessive embryonic lethals. Heterozygous null mutant flies have normal locomotor activity demonstrating that only a single dose of the wild type form of ple is required to support normal function. Both hemizygous and homozygous temperature-sensitive ple mutants (plets1) also show normal locomotor activity at the permissive temperature for this mutant allele (18°C), which progressively declines as the temperature is increased to its restrictive level (29°C). These abnormal locomotor effects are reversible by l-dopa. Thus the effects on locomotor activity resulting from the pharmacological inhibition of catecholamine synthesis or storage are the same as those resulting from lack of tyrosine hydroxylase expression. These findings indicate that brain catecholamine loss decreases locomotor activity in the fly, as it does in mammals, and demonstrate the ability of functional genomic studies to mimic that of pharmacological inhibition of enzyme function or other similar processes.

The human gene that codes for the protein alpha-synuclein has been transferred into the Drosophila melanogaster genome. The transgenic flies recapitulate some of the essential features of Parkinson's disease. These include the degeneration of certain dopaminergic neurons in the brain accompanied by the appearance of age-dependent abnormalities in locomotor activity. In the present study, we tested the locomotor response of these transgenic flies to prototypes of the major classes of drugs currently used to treat this disorder. A time course study was first conducted to determine when impaired locomotor activity appeared relative to normal "wild-type" flies. A climbing or negative geotaxis assay measuring the ability of the organisms to climb up the walls of a plastic vial was used. Based on the results obtained, normal and transgenic flies were treated with each of the drugs in their food for 13 days and then assayed. The activity of transgenic flies treated with L-DOPA was restored to normal. Similarly, the dopamine agonists pergolide, bromocriptine, and 2,3,4,5-tetrahydro-7,8-dihydroxy- 1-phenyl-1H-3-benzazepine (SK&F 38393) were substantially effective. Atropine, the prototypical muscarinic cholinergic receptor antagonist, was also effective but to a lesser extent than the other antiparkinson compounds. p-Chlorophenylalanine, an inhibitor of serotonin synthesis, was without beneficial effect as was alpha-methyl-p-tyrosine, an inhibitor of tyrosine hydroxylase, the rate-limiting step in catecholamine biosynthesis. This behavioral study further demonstrates the utility of this model in studying Parkinson's disease and reinforces the concept that inhibition of the action of alpha-synuclein may be useful in its treatment as may dopamine D(1) receptor agonists.

Inhibition of tyrosine hydroxylase, the rate-limiting enzyme in catecholamine biosynthesis, by α-methyl-p-tyrosine (αMT) at media concentrations of 0.3–3.0 mM, markedly inhibited (>90%) fly reproduction and development as evidenced by progeny count. Under these conditions adult spontaneous locomotor activity (SMA) was also dose-dependently reduced. However, no significant effects on behavior were observed at 0.3 mM. The behavioral effects of αMT were prevented by coadministration of L-DOPA. Similar effects on behavior and development were produced by reserpine at doses of 0.1–1.0 mM. The higher doses affected behavior, but no behavioral effects occurred at 0.1 mM. Rescue from the effects of reserpine was also achieved with L-DOPA. Treatment at 1 mM with the mammalian catecholamine receptor blockers propranolol (β1β2), raclopride (D2), SK&F 83566 (D1), prazosin (α1), and rauwolscine (α2) did not affect behavior. The two α adrenoceptor inhibitors, however, markedly decreased fly development. These results suggest that the receptor mechanisms mediating the effects of catecholamines on behavior and development are different. The locomotor behavior of adult progeny of parents treated with the lowest doses of αMT and reserpine described above was markedly suppressed by these drug treatments. This result indicates an increased sensitivity of the progeny to compounds which share the ability to deplete tissue catecholamines. Thus, alterations in catecholamine function during development may have behavioral consequences in surviving organisms. Drug Dev. Res. 50:142–146, 2000. © 2000 Wiley-Liss, Inc.

Drosophila melanogaster, maintained on culture media containing alpha-methyl-p-tyrosine (alpha-MT) at millimolar levels for 7 days, fail to produce viable progeny. Lesser concentrations delay development. This effect of alpha-MT, an inhibitor of tyrosine hydroxylase (TH), is partially reversible by co-administration of L-dihydroxyphenylalanine, the product of TH. Potent inhibitors of other steps in the pathway for catecholamine biosynthesis are inactive except for a partial effect with dopamine B-hydroxylase inhibition. The effect of alpha-MT is due to a combination of ovulation suppression coupled with decreases in embryonic and larval viability. Effects similar to those of alpha-MT are found with millimolar levels of reserpine, prazosin and to a lesser extent, rauwolscine. No significant effects are found with propranolol, chlorpromazine, sulpiride and SK&F 83566. Mutant alleles of the gene coding for TH are known to be lethal at the embryonic stage when homozygous in both Drosophila and mice. Taken together, these results indicate that in addition to their established roles in the nervous system catecholamines function in animal development via an action mediated through alpha-adrenoceptors.

AbstractThe (R)‐enantiomer of the NSAID ketoprofen was administered orally at 20 mg/kg to a series of 8 animal species. In all species, a highly significant degree of inversion occurred after 1 h which varied from 27% (gerbil) to 73% (dog) and persisted or increased in plasma samples obtained 3 h after drug administration. Although the (R)‐enantiomer was inactive as an inhibitor of cyclooxygenase, the analgesic effects of that isomer was almost the same as the (S)‐isomer in animal analgesic assays, following oral administration of the drugs to mice and rats. Taken together, the present results suggest that (R)‐ketoprofen administered alone functioned primarily as a prodrug for (S)‐ketoprofen under the experimental conditions of this study. © 1995 Wiley‐Liss, Inc.

The (R)-enantiomer of the NSAID ketoprofen was administered orally at 20 mg/kg to a series of 8 animal species. In all species, a highly significant degree of inversion occurred after 1 h which varied from 27% (gerbil) to 73% (dog) and persisted or increased in plasma samples obtained 3 h after drug administration. Although the (R)-enantiomer was inactive as an inhibitor of cyclooxygenase, the analgesic effects of that isomer was almost the same as the (S)-isomer in animal analgesic assays, following oral administration of the drugs to mice and rats, Taken together, the present results suggest that (R)-ketoprofen administered alone functioned primarily as a prodrug for (S)-ketoprofen under the experimental conditions of this study. (C) 1995 Wiley-Liss, Inc.

The addition of angiotensin II (AII) and angiotensin III (AIII) to isolated tissue baths produced the same maximal contractile response of rabbit aortic strips. AIII was about 10 times less potent, the slope of its concentration-response curve was less steep and its rate of onset slower than that of AII. The responses of both AII and AIII were inhibited with equal potency by the surmountable AII antagonist Phe4, Tyr8-AII and its unsurmountable analog Sar1, Leu8-AII but the kinetic patterns of inhibition by both were less well defined with the agonist AIII than with AII. The addition of AIII to tissues which had exhibited a maximal response to AII did not increase the level of contraction, in contrast to the case when norepinephrine was added to tissues contracted by AII. Both AII and AIII displaced [125I]AII binding from rabbit adrenal membranes; AIII was 6 times less potent than AII but displayed competitive kinetics as an inhibitor of [125I]AII binding. In further studies two binding sites for [125I]AII were identified in adrenal membranes, having KD values of 2.0 +/- 0.2 and 19.6 +/- 2.3 nM, respectively. Each site was inhibited by both AII and AIII and the ratio of the apparent Ki values for the two hormones was not significantly different. The Hill coefficient for the high affinity site was, however, lower for AIII than AII. We interpret our data to suggest that AII and AIII act on the same receptors. AIII apparently binds less efficiently than does AII in both rabbit adrenal membranes and rabbit aortic strips.

To exploit the well documented effect of 2,3-diphosphoglyceric acid (2,3-DPG) in enhancing oxygen delivery by human erythrocytes, we have investigated whether the DPG synthase/phosphatase enzyme system can be targeted to increase DPG levels in the cell. The hydrolytic activity (phosphatase) of the DPG metabolizing enzyme complex exhibits a marked dependence on a physiological effector, 2-phosphoglycolate. Little phosphatase activity is detected in the absence of this activator irrespective of the concentrations of the substrate. The phosphoglycolate-dependent phosphatase activity is competitively inhibited by a glycolytic intermediate, 3-phosphoglyceric acid (3-PGA). The 3-PGA inhibition persists when the 2,3-DPG concentration is raised to saturation level. In contrast, 3-PGA enhances the DPG synthase activity in a dose-dependent manner. In intact red cells, one-half of the cellular DPG content is depleted after 6 hr at 37 degrees C in glucose-free medium. The rate of 2,3-DPG degradation is accelerated when the cellular level of phosphoglycolate is increased by incubation with exogenous glycolate. Together, these results indicate that 2,3-DPG content in erythrocytes can be directly regulated through modulation of phosphatase/synthase activities. In support of this notion, a pyruvate kinase inhibitor, L-alanine, increases by 2-fold the cellular 3-PGA level. This is accompanied by a significant increase (30%) in 2,3-DPG content in human red blood cells. It is postulated that the DPG-promoting action of 3-PGA is mediated through simultaneous phosphatase inhibition and synthase activation. Furthermore, as a result of increased DPG accumulation, the oxygen-hemoglobin dissociation curve in L-alanine-treated cells is rightward shifted by 2.5 torr.

RG 12915 [4-[N-(1-azabicyclo[2.2.2.]octan-3-(S)-yl)]2-chloro-cis 5a-(S)-9a-(S)-5a,6,7,8,9,9a-hexahydrobenzofurancarboxamide hydrochloride] is a potent and effective agent against cisplatin-induced emesis in the ferret after i.v. or p.o. administration. This agent (p.o.) is also highly protective against cisplatin-induced emesis in the dog, as well as cyclophosphamide/doxorubicin-induced emesis in the ferret. When administered either p.o. or i.v., RG 12915 has a lower ED50 value (0.004 mg/kg) than GR 38032F, BRL 43694 and metoclopramide for attenuating cisplatin-induced emetic episodes in the ferret. It also has a long duration of action against cisplatin-induced emesis in the ferret. In contrast to metoclopramide, RG 12915 lacks significant antidopaminergic activity both in vitro [( 3H]spiroperidol displacement), as well as in vivo (apomorphine-induced emesis). In radioligand binding assays, RG 12915 is a potent and selective displacer of binding of 5-hydroxytryptamine (5-HT)3 binding sites (IC50 value = 0.16 nM), whereas failing to displace binding of ligands for the alpha-1, alpha-2 and beta adrenergic, 5-HT1 or 5-HT2 or cholinergic-muscarinic sites with IC50 values less than 1 microM. At a p.o. dose (1 mg/kg) in which RG 12915 is highly protective against cisplatin-induced emesis in the dog, RG 12915 has no significant gastroprokinetic activity in the same species. In summary, RG 12915 is a potent and p.o. effective agent against cytotoxic drug-induced emesis in animal models. The antiemetic potency of RG 12915 against cisplatin is unrelated to antidopaminergic or gastroprokinetic activity, but may be related to its affinity for 5-HT3 binding sites.

The efficacy of various drugs used to treat ulcerative colitis, (sulfasalazine, 5-aminosalicylate, hydrocortisone) was investigated in a model of acetic acid-induced colitis in the rat. Subsequently, we tested the ability of antioxidant/5-lipoxygenase inhibitors (gossypol and nordihydroguiaretic acid [NDGA]) and a cyclooxygenase inhibitor (indomethacin) to attenuate the macroscopic colonic damage and/or neutrophil influx (myeloperoxidase activity [MPO]) associated with this model of colitis. Oral pretreatment with either sulfasalazine, gossypol, or NDGA significantly decreased colonic MPO activity induced by acetic acid. Intrarectal administration of such drugs resulted in an even larger reduction of the colonic inflammation, with gossypol being the most potent compound. Oral or intrarectal administration of corticosteroids (dexamethasone, hydrocortisone) also attenuated the parameters of acetic acid induced colitis. In contrast, pretreatment with indomethacin was ineffective, or when administered daily after colitis induction, indomethacin actually increased colonic neutrophil influx significantly. Our data suggest that both the route of drug administration and dosing regimen employed affect the antiinflammatory potency and/or efficacy of compounds on colitis induced by acetic acid in the rat. Drugs which were effective against this colitis may act by scavenging of oxygen derived free radicals.

Angiotensin II (AII) labeled with 125I binds to rabbit adrenal cortical membranes over a concentration range from 0.5 to 20 nM at an apparent single site with a KD of 5 nM. This binding was inhibited in a surmountable fashion with respect to AII by the peptide analogs sarcosine1 (Sar1),Leu8AII and Phe4, Tyr8 AII when added to the incubation media concomitant with AII addition. With a 30-min preincubation, however, the former inhibitor displayed nonsurmountable kinetics whereas the profile of the latter was unaffected. In rabbit aortic strips with the same preincubation time, the Sar1Leu8AII analog was a nonsurmountable antagonist of the contractile effect of AII whereas the inhibition produced by Phe4,Tyr8AII was surmountable by increasing agonist (AII) concentrations. The inhibitory effect of the former was maintained after repeated washing of the tissue whereas that of the latter was readily reversible. Addition of Phe4,Tyr8AII to the bath 5 min before preincubation protected the tissue from the prolonged AII inhibition by Sar1,Leu8AII. These findings indicate different kinetic modes of AII inhibition by these two antagonists. Phe4,Tyr8AII behaves as a reversible, competitive inhibitor of AII binding, whereas Sar1,Leu8AII combines with the AII receptor in a slowly dissociable manner and is therefore not readily displaced by AII.