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H Giles

Publications and source records attributed to H Giles.

At least 19 recordsLinked to original sources
PubMed2001

Discovery of 4-[3-(trans-3-dimethylaminocyclobutyl)-1H-indol-5-ylmethyl]-(4S)-oxazolidin-2-one (4991W93), a 5HT(1B/1D) receptor partial agonist and a potent inhibitor of electrically induced plasma extravasation.

Utilizing a pharmacophoric model of binding of 3-(2-aminoethyl)indoles to 5HT(1B/1D) receptors, we identified the 3-aminocyclobutyl group as a potential ethylamine isostere. A novel multidimensional chemometric approach was used to predict the intrinsic activity (degree of agonism) at the receptor. A qualitative model for pharmacokinetic properties was then used to guide the synthesis toward molecules likely to have oral bioavailability in humans. A novel synthetic route to 3-(3-dimethylaminocyclobutyl)indoles was developed. Analogues showed generally lower intrinsic activity at 5HT(1B/1D) receptors than their ethylamine counterparts. 4-[3-(trans-3-Dimethylaminocyclobutyl)-1H-indol-5-ylmethyl]-(4S)-oxazolidin-2-one (4991W93, 1) was identified as a partial agonist against 5HT(1B/1D) receptors, with low intrinsic activity. This molecule also has significant activity against 5HT(1F) receptors but is selective over other 5HT receptors. In addition this compound was found to be an exceptionally potent inhibitor of electrically induced plasma extravasation. Compound 1 may have utility in the treatment and prophylaxis of migraine.

Administration, Oral↗
PubMed2001

Older adults' trait ratings of three age-groups around the Pacific rim.

In this paper, we assess the traits that older adults associate with younger, middle-aged, and older adults in five Pacific Rim nations from Western and Eastern cultural traditions (Australia, People's Republic of China (PRC), Hong Kong, Philippines, Thailand). We find cross-cultural trends which replicate patterns found in the US context. In most cultures, attractiveness, strength, activity, liberalism, health, and flexibility are seen to decline with increasing age. Kindness assessments are positively associated with age across cultures. Mixed patterns are found with assessments of wisdom and generosity, with respondents from the PRC and Hong Kong being notably more negative about increasing age than other respondents. Implications for the aging process across cultures are discussed, and suggestions made for future research.

Journal Article↗
PubMed2001

Paediatric walk-out patients: characteristics and outcomes.

OBJECTIVE: To investigate the characteristics and outcomes of patients who walked out from a tertiary children's hospital emergency department (ED) without seeing a medical officer. METHODS: A prospective study of patients who walked-out from the ED of a children's hospital, without seeing a medical officer. Information collected at triage included demographics, presenting problems, time of arrival, time of departure and reason for leaving. Charts were reviewed and those at high risk of serious illness or adverse outcome were contacted by telephone within 24 h. Further information collected during follow up included outcome, such as adverse events and admission to hospital. Data were analysed by comparing the walk-out and non-walk-out groups with regard to demographic variables, presenting problems and outcomes. RESULTS: Over a 29 week period, 1037 (5.5%) patients walked out from the ED of the hospital without seeing a medical officer. Comparisons between the walk-out and non-walk-out patients indicated no differences in terms of demographics. However, significant differences were found between the triage categories, presenting problems and arrival time. Of these, 829 (79.9%) were followed up by telephone. This revealed the predominant presenting problem was non-urgent and infectious in nature and no adverse events occurred. The admission rate for walk-out patients (1.5%) was significantly lower in comparison with the non-walk-out group (6.9%; odds ratio 0.2; 95% confidence interval 0.1-0.3). Walk-out patients who were eventually hospitalized had a shorter mean length of stay than non-walk-out patients (20.4 vs 34.8 h, respectively; t = 17.78, P < 0.0001). CONCLUSIONS: Medical resources are limited and, therefore, some extended waiting in the ED is necessary. Paediatric patients who walk-out of the ED without seeing a medical officer have simple illnesses that resolve without medical intervention or adverse events.

Australia↗
PubMed2001

The benefits of using clinical pathways for managing acute paediatric illness in an emergency department.

The aim of this study was to provide an evaluation of the overall effectiveness of using a number of clinical pathways in treating common acute paediatric conditions in an emergency department. This was a before and after study conducted on the effectiveness of three clinical pathways (gastroenteritis, asthma, and croup) in the emergency department of the Children's Hospital at Westmead, conducted over two separate yearly periods January to December 1996 and January to December 1999 representing before and after the introduction of clinical pathways in the emergency department. The main outcomes of the effectiveness of the pathways, namely admission to an in-patient bed, length of hospital stay and re-presentation after discharge from the ED were compared. Other outcomes of interest such as parental satisfaction and patient waiting times were also presented. Any deviation from a key clinical pathway process was reported. A total of 2854 children were managed by a clinical pathway compared to 2680 children managed before clinical pathways were introduced. The admission rate was reduced by threefold (9.1% compared to 23.6%) with a twofold reduction in length of hospital stay (32.7 h compared to 17.5 h). In 3.6% of children using a clinical pathway an unscheduled medical visit or re-presentation to the emergency department occurred after discharge, compared to 4.9% before the use of clinical pathways. No adverse events were reported in these children. In 76 cases deviation from a clinical pathway process was reported. High parental satisfaction was reported for clinical pathways throughout the study. Clinical pathways in this emergency department allowed rapid stabilisation of children, reducing admission rate, with a shortened length of hospital stay and few patients re-presenting after discharge and were well accepted by parents.

Asthma↗
PubMed2001

Spontaneous contractions of myometrium from humans, non-human primate and rodents are sensitive to selective oxytocin receptor antagonism in vitro.

OBJECTIVES: To determine whether: 1. oxytocin receptor antagonists influence spontaneous contractions of myometrium from humans, non-human primates and rodents (in vitro), and 2. vasopressin V1a receptor antagonism is important for inhibition of spontaneous contractions in human myometrium. DESIGN: In vitro pharmacology of spontaneous contractions of myometrium from humans and animals. SETTING: The research laboratories of a university department of obstetrics and gynaecology and a pharmaceutical industry research centre. INTERVENTIONS: Samples of human myometrium were obtained at caesarean section. Tissue strips were suspended in organ baths for isometric force recording. Cumulative concentration effect curves to a selective oxytocin receptor antagonist (L-371,257) and a mixed oxytocin/vasopressin V1a receptor antagonist (atosiban) were obtained. The effect of L-371,257 was also determined in myometrium from non-pregnant rats and marmosets. MAIN OUTCOME MEASURES: The inhibition of spontaneous myometrial contractions in vitro. RESULTS: L-371,257 and atosiban significantly inhibited spontaneous activity of human myometrium in a concentration-related manner (P < 0.05), although the effect was more pronounced with L-371,257. Spontaneous contractions of myometrium from non-pregnant rats and marmosets were also inhibited by L-371,257 (atosiban was not tested). CONCLUSIONS: Spontaneous contractions of myometrium from humans, marmosets and rats are, at least in part, dependent on oxytocin receptor activity, in vitro. L-371,257 and atosiban may be inverse agonists. Selective non-peptide oxytocin receptor antagonists may be effective tocolytics.

Animals↗
PubMed2000

Use of a dual firefly and Renilla luciferase reporter gene assay to simultaneously determine drug selectivity at human corticotrophin releasing hormone 1 and 2 receptors.

The aim of this study was to investigate and validate the use of a dual glow-signal luciferase reporter gene assay to simultaneously evaluate drug activity at two different seven-transmembrane receptor subtypes. Stable cell lines (CHO) transfected with either human corticotrophin releasing hormone 1 (hCRH1) receptors and a firefly luciferase reporter gene or hCRH2 and a Renilla luciferase reporter gene were created to provide different luciferase readouts for CRH1 and CRH2 receptors, respectively. Cells were combined for stimulation and measurement of luciferase luminescence in a 96-well plate format assay. The nonselective CRH agonists rat/human CRH and sauvagine caused concentration-dependent increases in luminescence via activation of CRH1 (firefly luciferase; pEC50 = 8.40 +/- 0.06 and 8.39 +/- 0.08, respectively, n = 8) and CRH2 (Renilla luciferase; pEC50 = 8.89 +/- 0.14 and 8.92 +/- 0.13, respectively, n = 8) receptors. The nonselective CRH antagonist astressin blocked these agonist-induced increases in luciferase at both CRH1 and CRH2 receptors. The selective CRH1 antagonist CP154,526 blocked r/hCRH- and sauvagine-induced increases in luciferase at CRH1 receptors only. These data report the expected pharmacology for CRH1 and CRH2 receptors. This assay enabled two receptor subtypes to be studied simultaneously in the same 96-well plate and generated robust data with low variability. It has the potential advantage of significant time and cost savings, with application to both basic research and compound screening.

Animals↗
PubMed2000

Pharmacological characterization of human recombinant melatonin mt(1) and MT(2) receptors.

We have pharmacologically characterized recombinant human mt(1) and MT(2) receptors, stably expressed in Chinese hamster ovary cells (CHO-mt(1) and CHO-MT(2)), by measurement of [(3)H]-melatonin binding and forskolin-stimulated cyclic AMP (cAMP) production. [3H]-melatonin bound to mt(1) and MT(2) receptors with pK(D) values of 9.89 and 9.56 and B(max) values of 1.20 and 0.82 pmol mg(-1) protein, respectively. Whilst most melatonin receptor agonists had similar affinities for mt(1) and MT(2) receptors, a number of putative antagonists had substantially higher affinities for MT(2) receptors, including luzindole (11 fold), GR128107 (23 fold) and 4-P-PDOT (61 fold). In both CHO-mt(1) and CHO-MT(2) cells, melatonin inhibited forskolin-stimulated accumulation of cyclic AMP in a concentration-dependent manner (pIC(50) 9.53 and 9.74, respectively) causing 83 and 64% inhibition of cyclic AMP production at 100 nM, respectively. The potencies of a range of melatonin receptor agonists were determined. At MT(2) receptors, melatonin, 2-iodomelatonin and 6-chloromelatonin were essentially equipotent, whilst at the mt(1) receptor these agonists gave the rank order of potency of 2-iodomelatonin>melatonin>6-chloromelatonin. In both CHO-mt(1) and CHO-MT(2) cells, melatonin-induced inhibition of forskolin-stimulated cyclic AMP production was antagonized in a concentration-dependent manner by the melatonin receptor antagonist luzindole, with pA(2) values of 5.75 and 7.64, respectively. Melatonin-mediated responses were abolished by pre-treatment of cells with pertussis toxin, consistent with activation of G(i)/G(o) G-proteins. This is the first report of the use of [(3)H]-melatonin for the characterization of recombinant mt(1) and MT(2) receptors. Our results demonstrate that these receptor subtypes have distinct pharmacological profiles.

Animals↗
PubMed2000

The effects of a seamless model of management on the quality of care for emergency department patients.

The aim of this study was to examine the effectiveness of the seamless model of management on the quality of care for emergency department (ED) patients. This was a pre- and postintervention study of comparing post intervention data with the baseline on several variables of interest. The intervention was the seamless model of ED management, designed and implemented as a response to the challenge of increasing work load at the ED. Information on patients' waiting times, critical care performance, patients' satisfaction and staff morale was collected at baseline and postintervention for comparison. The results indicated significant improvements on all outcome measures were found postintervention. The average waiting time reduced by 40%, from 92.1 to 55.3 min, as well as the time to craniotomy from 120 to 45 min for more severe patients. Complaints from patients were reduced by 80% and staff morale had improved with a 48% reduction in sick days and a 80% reduction in staff resignations. Ambulatory patients presenting to the department saw a 20% improvement in their waiting time. We report a successful restructuring process that used routinely collected clinical and administrative data to highlight problems. Using these data and through a systematic planning process, appropriate strategies for restructuring were developed by emergency staff in partnership with the hospital executive. Significant improvements in waiting time and patient care were clearly demonstrated.

Attitude of Health Personnel↗
PubMed1999

Signalling by CXC-chemokine receptors 1 and 2 expressed in CHO cells: a comparison of calcium mobilization, inhibition of adenylyl cyclase and stimulation of GTPgammaS binding induced by IL-8 and GROalpha.

The effect of interleukin-8 (IL-8) and growth-related oncogene alpha (GROalpha) on [35S]-guanosine 5'-O-(3-thiotriphosphate) ([35S]GTPgammaS) binding, forskolin-stimulated cyclic AMP accumulation and cytosolic calcium concentration were determined in recombinant CHO cells expressing HA-tagged CXC-chemokine receptors 1 and 2 (CXCR1 and CXCR2). Radioligand binding assays confirmed that the binding profiles of the recombinant receptors were similar to those of the native proteins. IL-8 displaced [125I]-IL-8 binding to CXCR1 and CXCR2 with pKi values of 8.89+/-0.05 and 9.27+/-0.03, respectively. GROalpha, a selective CXCR2 ligand, had a pKi value of 9.66+/-0.39 at CXCR2 but a pKi>8 at CXCR1. Calcium mobilization experiments were also consistent with previous reports on native receptors. Activation of both receptors resulted in stimulation of [35S]GTPgammaS binding and inhibition of adenylyl cyclase. A comparison of the functional data at CXCRI showed that a similar potency order (IL-8> >GROalpha) was obtained in all three assays. However, at CXCR2 whilst the potency orders for calcium mobilization and inhibition of adenylyl cyclase were similar (IL-8 > or = GROalpha), the order was reversed for stimulation of [35S]GTPgammaS binding (GROalpha > IL-8). All of the functional responses at both receptors were inhibited by pertussis toxin (PTX), suggesting coupling to a Gi/Go protein. However, the calcium mobilization induced by IL-8 at CXCR1 was not fully inhibited by PTX, suggesting an interaction with a G-protein of the Gq family. Our results with pertussis toxin also suggested that, in the [35S]GTPgammaS binding assay, CXCR1 displays some constitutive activity. Thus, we have characterized the binding and several functional responses at HA-tagged CXCRs 1 and 2 and have shown that their pharmacology agrees well with that of the native receptors. We also have preliminary evidence that CXCR1 displays constitutive activity in our cell line and that CXCR2 may traffic between different PTX sensitive G-proteins.

Adenylate Cyclase Toxin↗
PubMed1998

Pharmacological characterisation of melatonin mt1 receptor-mediated stimulation of [35S]-GTPgammaS binding.

The activation of G-proteins by melatonin mt1 receptors was studied by measuring [35S]-guanosine-5'-(3-thiotriphosphate) ([35S]-GTPgammaS) binding to membranes prepared from Chinese hamster ovary (CHO) cells stably expressing human mt1 receptors. Melatonin stimulated [35S]-GTPgammaS binding in a concentration-dependent manner (pEC50, 8.77+/-0.02). The optimal (212+/-4%) increase over basal levels of binding (basal = 100%) was observed following incubation of membranes (12.5 microg protein/well) for 120 min at 30 degrees with [35S]-GTPgammaS (0.1 nM), in the presence of GDP (10 microM), NaCl (100 mM), and MgCl2 (10 mM). Melatonin analogues stimulated [35S]-GTPgammaS binding with a rank order (2-iodomelatonin > melatonin = S20098 > GR196429 > 6-chloromelatonin = 6-hydroxymelatonin >> N-acetylserotonin > or = GR135531 = mt1 luzindole = 5-HT = 0), which was identical to their affinities for the high affinity state of the receptor (correlation coefficient 0.94). All agonists evoked similar maximum increases in [35S]-GTPgammaS binding. EC50 values were 14- to 63-fold lower than binding affinities. The melatonin receptor antagonist luzindole (0.1-10 microM) evoked a parallel rightward shift in the melatonin concentration-response curve, with a pKB of 7.19+/-0.13, which is similar to its affinity in radioligand binding studies for human mt1 receptors. Stimulation of [35S]-GTPgammaS binding was abolished by pretreatment of cells with pertussis toxin (18 hr, 100 ng/mL) prior to preparation of membranes. Melatonin was without effect in CHO cells which lacked the mt1 receptor. Thus, melatonin and melatonin analogues stimulate [35S]-GTPgammaS binding with a profile which is consistent with binding to mt1 receptors causing activation of Gi/Go G-proteins.

Animals↗
PubMed1997

Receptor specificity and trigemino-vascular inhibitory actions of a novel 5-HT1B/1D receptor partial agonist, 311C90 (zolmitriptan).

1. 311C90 (zolmitriptan zomig: (S)-4[[3-[2-(dimethylamino)ethyl]-1H-indol-5-yl]methyl]-2-oxazolidinone) is a novel 5-HT1B/1D receptor agonist with proven efficacy in the acute treatment of migraine. Here, we describe the receptor specificity of the drug and its actions on trigeminal-evoked plasma protein extravasation into the dura mater of the anaesthetized guinea-pig. 2. At the "5-HT1B-like' receptor mediating vascular contraction (rabbit saphenous vein), the compound was a potent (p[A50] = 6.79 +/- 0.06) partial agonist achieving 77 +/- 4% of the maximum effect to 5-hydroxytryptamine (5-HT). In the same experiments, sumatriptan (p[A50] = 6.48 +/- 0.04) was half as potent as 311C90 and produced 97 +/- 2% of the 5-HT maximum effect. Studies in which receptor inactivation methods were used to estimate the affinity (pKA) and efficacy relative to 5-HT (tau rel) for each agonist confirmed that 311C90 exhibits higher affinity than sumatriptan (pKA = 6.63 +/- 0.04 and 6.16 +/- 0.03, respectively) and that both drugs are partial agonists relative to 5-HT (tau rel = 0.61 +/- 0.03 and 0.63 +/- 0.10, respectively, compared to 5-HT = 1.0). 3. Consistent with its effects in rabbit saphenous vein, 311C90 also produced concentration-dependent contractions of primate basilar artery and human epicardial coronary artery rings. In basilar artery, agonist potency (p[A50] = 6.92 +/- 0.07) was similar to that demonstrated in rabbit saphenous vein, again being 2-3 fold higher than for sumatriptan (p[A50] = 6.46 +/- 0.03). Both agonists produced about 50% of the maximum response obtained with 5-HT in the same preparations. In rings of human coronary artery, the absolute potency of 311C90 and sumatriptan was higher than in primate basilar artery (p[A50] = 7.3 +/- 0.1 and 6.7 +/- 0.1, respectively), but maximum effects relative to 5-HT were lower (37 +/- 8% and 35 +/- 7%, respectively). In both types of vessel, the inability of 5-HT1B/1D agonists to achieve the same maximum as the endogenous agonist 5-HT is explained by the additional presence of 5-HT2A receptors. 4. 311C90 displayed high affinity at human recombinant 5-HT1D (formerly 5-HT1D alpha) and 5-HT1B (formerly 5-HT1D beta) receptors in transfected CHO-K1 cell membranes (pIC50 values = 9.16 +/- 0.12 and 8.32 +/- 0.09, respectively). In intact cells, the drug produced concentration-dependent inhibition of forskolin-stimulated adenylyl cyclase (p[A50] = 9.9 and 9.5, respectively) achieving the same maximum effect as 5-HT. Excepting human recombinant 5-HT1A and 5-ht1F receptors at which the drug behaved as an agonist with modest affinity (pIC50 = 6.45 +/- 0.11 and 7.22 +/- 0.12, respectively), 311C90 exhibited low, or no detectable affinity (pKi or pKB < or = 5.5) at numerous other monoamine receptors, including other 5-HT receptor subtypes. 5. When administered to anaesthetized guinea-pigs ten minutes before unilateral electrical stimulation of the trigeminal ganglion (1.2 mA, 5 Hz, 5 ms, 5 min), 311C90 (3-30 micrograms kg-1, i.v.) caused a dose-dependent inhibition of [125I]-albumin extravasation within the ipsilateral dura mater. At the same doses, the drug also produced dose-dependent falls in cranial vascular conductance (32.3 +/- 7.5% at 30 micrograms kg-1), as measured in the ear by laser doppler flowmetry. 6. These results show that 311C90, a novel member of the 5-HT1B/1D agonist drug class, exhibits a high degree of pharmacological specificity. Its potent partial agonist action at "5-HT1B-like' receptors in intracranial arteries, coupled with potent agonism at 5-HT1D and 5-HT1B receptors and an ability to inhibit neurogenic plasma protein extravasation in the dura, are consistent with its utility as an effective acute treatment for migraine.

Animals↗
PubMed1997

Toxicologic effects of an oligodeoxynucleotide phosphorothioate and its analogs following intravenous administration in rats.

The aim of the present study is to evaluate the in vivo toxicologic effects of a phosphorothioate oligodeoxynucleotide (PS oligo) and three of its analogs [PS oligo containing four methylphosphonate linkages at the 3' and 5'-ends (MBO 1), PS oligo containing four 2'-O-methylribonucleosides at both the 3'- and 5'-ends (MBO 2), and PS oligo containing an 8 bp loop region at the 3'-end (self-stabilized oligo)]. Oligodeoxynucleotides were administrated intravenously to male and female rats at doses of 3, 10, and 30 mg/kg/day for 14 days. Rats were killed on day 15, blood samples were collected for hematology and clinical chemistry determinations, and tissues, including lymph nodes, spleens, livers, and kidneys, were subjected to pathologic examinations. The toxicity profiles of the four oligodeoxynucleotides were very similar, but differed in magnitude. In terms of the severity of the abnormalities caused by the oligodeoxynucleotides, the order was MBO 2 > PS oligo > self-stabilized oligo > MBO 1. Alterations in hematology parameters included thrombocytopenia, anemia, and neutropenia. Abnormalities in clinical chemistry parameters observed with PS oligo or MBO 2 were dose-dependent elevation of liver transaminases and reduction of the levels of alkaline phosphatase, albumin, and total protein. In addition, MBO 2 caused elevation of the total bilirubin level in male rats at the 30 mg/kg dose. No major alterations in hematology or clinical chemistry were observed in rats receiving MBO 1 or self-stabilized oligo. Dose-dependent enlargements of spleen, liver, and kidney were observed, especially in rats receiving PS oligo and MBO 2. Pathologic studies showed a generalized hyperplasia of the reticuloendothelial (RE) system in the tissues examined. Alterations in the spleen were mainly RE cell hyperplasia and hematopoietic cell proliferation. In addition to RE cell hyperplasia, lymph nodes showed necrosis, hepatocytes showed cytologic alterations and necrosis, and kidneys showed renal tubule regeneration. The severity of pathologic changes observed was oligodeoxynucleotide dependent, in the order of MBO 2 > PS oligo > self-stabilized oligo > MBO 1.

Animals↗
PubMed1996

Biochemical and molecular characterization of acetylcholinesterase from the hagfish Myxine glutinosa.

To obtain information about the evolution of the cholinesterases, we investigated the cholinesterase activity of an agnathan vertebrate, the hagfish Myxine glutinosa. On the basis of evidence from enzymology, pharmacology, and molecular biology, we conclude that the cholinesterase activity is due to acetylcholinesterase (AChE). The enzyme hydrolyzes acetylthiocholine preferentially and exhibits substrate inhibition. The hydrolysis of both acetylthiocholine and butyrylthiocholine are inhibited in parallel by cholinesterase inhibitors, with the AChE-specific drug BW284c51 being the most potent; however, this drug and propidium, a peripheral anionic site ligand, are much weaker inhibitors of the hagfish enzyme than of Torpedo AChE. We used sequential extraction, collagenase digestion, and velocity sedimentation on sucrose gradients to determine that the AChE from the skeletal muscle of the hagfish is present in both globular and asymmetric forms. We also used the polymerase chain reaction with degenerate oligonucleotide probes and genomic DNA to obtain a 1 kb gene fragment for hagfish AChE. The enzyme has an acyl binding site typical of other vertebrate AChE, but lacks two aromatic residues implicated in the function of the peripheral anionic subsite. We discuss the relevance of our findings to the evolution of the cholinesterases in the vertebrates.

Acetylcholinesterase↗
PubMed1996

Synthesis of Hexahydrocyclopentimidazol-2-(1H)-one derivatives displaying selective DP-receptor agonist properties.

The rationale for investigating conformationally restricted analogues of BW245C as DP-receptor ligands and the syntheses of three such racemic bicyclic imidazolidinone analogues are described. Compounds 7 (BW587C), 8 (BW480C85), and 9 (BW572C85) were found to be potent inhibitors of human platelet aggregation and selective DP-receptor agonists in washed platelet and jugular vein isolated tissue assays.

Animals↗
PubMed1996

Characterization of a 5-HT1B receptor on CHO cells: functional responses in the absence of radioligand binding.

1. Chinese hamster ovary (CHO) cells have been reported to be devoid of 5-HT receptors and have frequently been used as hosts for the expression of cloned 5-HT receptors. Unexpectedly, 5-HT was found to induce profound inhibition of forskolin-stimulated cyclic AMP production in these cells and the aim of this study was to classify the 5-HT receptor involved. 2. In CHO(dhfr-) cells 5-HT was a potent agonist and caused 80-100% inhibition of forskolin stimulated cyclic AMP production. A study using several 5-HT1 receptor agonists revealed the following potencies (p[A50]): RU24969 (9.09 +/- 0.17) > 5-carboxamidotryptamine (8.86 +/- 0.20) > 5-HT (8.07 +/- 0.05) > CP-93,129 (7.74 +/- 0.10) > sumatriptan (5.93 +/- 0.04). All five agonists achieved a similar maximum effect. Irreversible receptor alkylation studies yielded a pKA estimate of 7.04 +/- 0.34 for 5-HT. 3. The 5-HT1A/1B antagonist, (+/-)-cyanopindolol (4-100 nM), caused parallel rightward shifts of the 5-HT concentration-effect curve with no change in asymptote. Schild analysis yielded a pKB estimate of 8.69 +/- 0.09 (Schild slope 1.13 +/- 0.10). (+/-)-Cyanopindolol actually behaved as a partial agonist with an intrinsic activity of 0.2-0.5 and a p[A50] of 8.55. 4. 5-HT (0.01-10 microM) also elicited a concentration-dependent increase in intracellular [Ca2+] in CHO(dhfr-) cells thus demonstrating that dual coupling is not a phenomenon restricted to systems in which there is overexpression of transfected receptors. 5. This agonist and antagonist profile is consistent with the presence of a 5-HT1B receptor. 8-OH-DPAT (1 microM) and renzapride (3 microM) were without effect on forskolin-stimulated cyclic AMP production and ketanserin (0.3 microM) did not antagonize the inhibition produced by 5-HT, thus excluding the involvement of 5-HT1A, 5-HT4, and 5-HT2 receptors. 6. The possibility that expression of a 5-HT1B receptor was associated with the dhfr- mutation was excluded since RU24969, 5-HT and CP-93,129 were also potent agonists in unmutated, CHO-K1 cells: p[A50] 9.03 +/- 0.03, 8.34 +/- 0.05, 7.69 +/- 0.07 respectively, and (+/-)-cyanopindolol (0.1 microM) shifted the 5-HT curve to the right and yielded a pA2 estimate of 8.70 +/- 0.06. 7. Little or no specific binding of [3H]-5-HT (0.1-200 nM) or of the high affinity ligand [125I]-iodocyanopindolol (0.01-3 nM) to CHO(dhfr-) cell membranes could be detected. 5-HT also failed to elicit any increase in the binding of [35S]-GTP gamma S to CHO membranes. 8. In conclusion, cultured CHO cells express 5-HT1B receptors which are negatively coupled to adenylyl cyclase and positively coupled to increases in intracellular calcium. The absence of radioligand binding was unexpected in view of the high potency of 5-HT and the partial agonist activity of the normally 'silent' competitive antagonist, (+/-)-cyanopindolol. This implies very efficient receptor-effector coupling of a low density of 5-HT1B receptors. Clearly, the absence of detectable radioligand binding cannot be assumed to mean the absence of receptors capable of eliciting a significant functional response.

Animals↗
PubMed1995

Cloning and characterisation of the rabbit 5-HT1D alpha and 5-HT1D beta receptors.

The genes encoding the rabbit 5HT1D alpha and 5HT1D beta receptors have been cloned. The deduced amino acid sequence of these receptors shows 91-92% amino acid sequence identity with their human homologues, and similar high sequence identity with homologues from other species. The receptors were transiently expressed in COS-7 cells and exhibit a pharmacological profile closely resembling their human homologues, including a higher affinity of ketanserin for the 5-HT1D alpha subtype. However, sumatriptan had a lower affinity for both the rabbit receptors compared to their human counterparts. This may be accounted for by differences between the primary amino acid sequences of these species homologues.

Amino Acid Sequence↗