Papers by Elizabeth Burcher
Peptides, 1992
Contractile responses to neurokinin A (NKA), neuropeptide gamma(NP gamma), and the NK2 receptor-s... more Contractile responses to neurokinin A (NKA), neuropeptide gamma(NP gamma), and the NK2 receptor-selective analogs [Lys5,MeLeu9,Nle10]NKA(4-10) and MDL 28,564 were determined in the endothelium-denuded rabbit pulmonary artery. Responses to NKA, NP gamma, and [Lys5,MeLeu9,Nle10]NKA(4-10) were antagonized by the NK2 receptor antagonist MDL 29,913, with pA2 values of 6.67, 6.46, and 7.32, respectively. Autoradiographic studies failed to demonstrate any specific binding sites for [125I]-iodohistidyl NKA (INKA) over the pulmonary artery. These data suggest the presence in rabbit pulmonary artery of an unusual "nonclassical" NK2 receptor subtype, which appears to lack affinity for INKA.
Basic and Clinical Pharmacology and Toxicology, 2010
Bju Int, 2007
To examine the expression of muscarinic M2 and M3 receptors in human bladder detrusor and mucosa,... more To examine the expression of muscarinic M2 and M3 receptors in human bladder detrusor and mucosa, from controls and patients with idiopathic detrusor overactivity (IDO), as antimuscarinic agents are the primary pharmacological treatment for IDO. Biopsies from the bladder body were collected at cystoscopy from 20 women with urodynamically confirmed refractory IDO (age range 25-86 years); biopsies were also collected from 30 asymptomatic female controls (age range 32-87 years). Samples were collected into RNA extraction medium and dissected into mucosa (urothelium plus lamina propria) and detrusor. RNA was extracted and the expression of M2 and M3 receptor mRNA determined by quantitative competitive reverse transcription-polymerase chain reaction. Results were normalized to beta-actin expression in the same sample. Expression of M3 receptor mRNA, in mucosa of IDO patients (median 0.057 pg M3/100 ng total RNA; interquartile range 0.03-0.13, 12 samples), was four times (P = 0.039, Mann-Whitney) lower than from the control (median 0.22 pg M3/100 ng total RNA; 0.13-0.51, 11 samples). The expression of muscarinic M3 receptor mRNA was higher (14-35 times) in detrusor (control median 3.17; 26 samples) than in mucosa and did not change in IDO (median 2.03; 14 samples). M2 expression was not significantly different with region or with IDO. These data show that M3 muscarinic receptor mRNA expression was significantly less in mucosa from IDO patients than from age-matched controls. The role of mucosal M3 receptors is unknown at present and elucidation of this role might provide a greater understanding of the aetiology of IDO.
Molecular Pharmacology
In homogenates of guinea pig lung, binding of 125I-Bolton-Hunter-labeled substance P (BHSP), Bolt... more In homogenates of guinea pig lung, binding of 125I-Bolton-Hunter-labeled substance P (BHSP), Bolton-Hunter-labeled eledoisin (BHELE), and [125I]iodohistidyl neurokinin A (INKA) was investigated. Equilibrium dissociation constants (derived from "cold" saturation experiments) for BHSP, INKA, and BHELE were 0.96 +/- 0.15, 1.61 +/- 0.26, and 1.98 +/- 0.12 nM, respectively. Specific binding of all three radioligands was increased 2-3-fold by 10 microM phosphoramidon. The rank order of potency of unlabeled tachykinins in competing against BHSP was substance P (SP) greater than [Sar9,Met(O2)11]-SP greater than SP methyl ester greater than neuropeptide gamma greater than neurokinin A greater than or equal to neurokinin B = kassinin greater than or equal to eledoisin greater than or equal to scyliorhinin II much greater than neuropeptide K, indicating binding to sites with the general characteristics of NK1 receptors. Similar rank potency orders were observed for INKA and BHELE, showing binding to NK1 sites, rather than to NK2 or NK3 sites, which are labeled with high affinity by these radioligands in other tissues. For all radioligands, competition curves for SP and the NK1-selective agonist [Sar9,Met(O2)11]-SP could be resolved into two components, representing high and low affinity binding sites. These were present in the approximate ratios 2:3 (for BHSP), 1:1 (for INKA), and 8:1 (for BHELE). Other agonist competition curves also yielded high and low affinity components. The data suggest that BHSP and INKA bind partly and BHELE predominantly to high affinity NK1 receptors. The nature of the low affinity site(s) could be another tachykinin receptor or a low affinity state of the NK1 receptor. Binding to a "classical" NK2 receptor is unlikely, because selective NK2 receptor antagonists and analogs were very weak competitors. Our data suggest that, in addition to the NK1 receptor, another type of tachykinin receptor may exist in this tissue. The inability to detect NK2 binding sites is strikingly at variance with functional studies.
Journal of Pharmacology and Experimental Therapeutics
The binding of [3H]11-OH-delta 9-tetrahydrocannabinol-1, 1-dimethyl-heptyl (THC-DMH), a recently ... more The binding of [3H]11-OH-delta 9-tetrahydrocannabinol-1, 1-dimethyl-heptyl (THC-DMH), a recently synthesized cannabinoid analog, was characterized in an in vitro brain slice binding assay and compared to that obtained with [3H]CP-55,940, the radiolabeled ligand used originally to characterize cannabinoid binding sites. The binding of both [3H]CP-55,940 and [3H]11-OH-delta 9-THC-DMH exhibited high affinity (Kd of 19 +/- 3 and 29 +/- 9 nM, respectively), and was saturable, reversible and specific. Values of maximal concentration of receptors determined for [3H]11-OH-delta 9-THC-DMH and [3H]CP-55,940 were 4.0 +/- 0.3 and 3.0 +/- 0.5 pmol/mg of protein, respectively. The distribution of [3H]11-OH-delta 9-THC-DMH and [3H]CP-55,940 binding in 30-microns rat brain sections was then compared by autoradiographic analysis. The binding of both ligands was densest in the basal ganglia (substantia nigra pars reticulata, globus pallidus, entopeduncular nucleus and regions of the caudate putamen) and cerebellum (molecular layer). Low levels of binding were observed in discrete brain regions including the brain stem (medulla and pons), thalamic nuclei, hypothalamus, corpus callosum and the deep nuclear layer of the cerebellum. Intermediate levels of binding were seen in layers I and VI of the cortex, and the dentate gyrus and CA pyramidal cell regions of the hippocampus. The ability of selected cannabinoid analogs to compete with [3H]11-OH-delta 9-THC-DMH binding was determined. The Ki's were correlated to the in vivo potencies for producing catalepsy, antinociception, hypothermia and decreasing spontaneous locomotor activity in mice (correlation coefficients > 0.86).(ABSTRACT TRUNCATED AT 250 WORDS)
Journal of Pharmacology and Experimental Therapeutics
ABSTRACT
Clinical and Experimental Pharmacology and Physiology
1. Neurokinin (NK)A is the endogenous ligand for the tachykinin NK2 receptor. In the present stud... more 1. Neurokinin (NK)A is the endogenous ligand for the tachykinin NK2 receptor. In the present study, tachykinins and selective receptor agonists were tested as contractile agonists in human colon circular muscle and [125I]-NKA was used to localize binding sites in human colon. 2. In strips of circular muscle, removal of mucosa and submucosa significantly (P < 0.05) increased the potency and the maximum response achieved by NKA. 3. The rank order of potency of tachykinin and selective receptor agonists in contracting circular muscle strips was NKA > or = [Lys5,MeLeu9,Nle10]NKA(4-10) > or = neuropeptide (NP)gamma > or = [betaAla8]NKA(4-10) > NKB > substance P (SP) > senktide approximate to [Pro9]SP. 4. Specific binding sites for [125I]-NKA were densely localized over circular muscle and muscularis mucosae. Weak specific binding was seen on longitudinal muscle and taenia coli, whereas no binding sites were seen on mucosa, ganglia or blood vessels. 5. In circular muscle, the selective NK2 receptor agonist [LysS,MeLeu9,Nle10]NKA(4-10) produced weak increases (maximum 37%) in inositol monophosphate formation with a pD2 of 6.8+/-0.51 (n = 3). Carbachol (100 micromol/L) was also a weak stimulant (maximum 45%). These agonists were over 10-fold more efficacious in stimulation of inositol monophosphate in rat urinary bladder. 6. In conclusion, [125I]-NKA binding sites localized on human colon circular muscle were characterized as NK2 receptors. Functionally, the tachykinin NK2 receptor is mediating circular smooth muscle contraction. Although the human NK2 receptor is coupled to the phosphatidylinositol pathway, other second messenger mechanisms may also operate in this tissue.
Regulatory Peptides, 1988
Substance p (SP) i/xm21oreactive fibres are found outside the smooth muscle and beneath tb~ epith... more Substance p (SP) i/xm21oreactive fibres are found outside the smooth muscle and beneath tb~ epithelium of the guinea-pig vas deferens (i, 2). After capsaicin pre~nent, all SP-immmoreactive neurons disappear (2). On the isolated guinea-pig vas deferens, SP potentiates the "twitch response" caused by transmural nerve stin~tlation and also causes an increase in tone (3). The first but not the second action is blocked by phenoxybenzamine, and may be mediated by presynapti~2~_-i receptors. Here we investigated the localization of binding sites for [ I]-Bolton-Hunter SP (H~SP) in normal, capsaicintreated and vehicle-treated animals, using in vitro labelling and autoradiography.
British Journal of Pharmacology
The bladder urothelium is now known to have active properties. Our aim was to investigate the con... more The bladder urothelium is now known to have active properties. Our aim was to investigate the contractile properties of the urinary mucosa in response to the tachykinin neurokinin A (NKA) and carbachol. Discrete concentration-response curves for carbachol and NKA were obtained in matched strips of porcine detrusor, mucosa and intact bladder, suspended in organ baths. The effects of inhibitors and tachykinin receptor antagonists were studied on NKA-mediated contractions in mucosal strips. Intact sections of bladder and experimental strips were processed for histology and immunohistochemistry. All types of strips contracted to both carbachol and NKA. Mucosal responses to NKA (pD2 7.2) were higher than those in intact strips and were inhibited by the NK2 receptor antagonist SR48968 (pKB 9.85) but not the NK1 receptor antagonist SR140333, tetrodotoxin or indomethacin. Immunostaining for smooth muscle actin and vimentin occurred under the urothelium and on blood vessels. Desmin immunosta...
Pulmonary pharmacology, 1994
Tachykinins are potent bronchoconstrictors. Here we have used novel highly selective agonists and... more Tachykinins are potent bronchoconstrictors. Here we have used novel highly selective agonists and antagonists to characterize tachykinin receptors mediating bronchoconstriction and hypotension in anaesthetized, paralysed guinea-pigs. Total lung resistance (RL) and dynamic lung compliance (Cdyn) were measured using an on-line, breath-by-breath method to analyse lung mechanics. The effects of iv injections of neuropeptide gamma (NP gamma), [Sar9,Met(O2)11]-SP (Sar-SP) (NK-1 agonist), [Lys5,MeLeu9,Nle10]-NKA(4-10) (Lys-NKA) (NK-2 agonist) and senktide (NK-3 agonist) were compared, before and after iv injection of the NK-1 antagonist CP 96345 or the NK-2 antagonists MDL 29913 and SR 48968. NP gamma, Sar-SP and Lys-NKA, but not senktide, increased RL and decreased Cdyn, accompanied by a fall in blood pressure (BP). The potency for increase of RL was Lys-NKA > NP gamma congruent to Sar-SP, for reduction of Cdyn was Lys-NKA = Sar-SP > NP gamma, and for fall in BP was Sar-SP > NP g...
The Biochemical journal, 1992
A tachykinin peptide was isolated from an extract of the intestine of the European green frog, Ra... more A tachykinin peptide was isolated from an extract of the intestine of the European green frog, Rana ridibunda, and its primary structure was established as: His-Lys-Leu-Asp-Ser-Phe-Ile-Gly-Leu-Met.CONH2. This sequence was confirmed by chemical synthesis and shows two amino acid substitutions (leucine for threonine at position 3 and isoleucine for valine at position 7) compared with neurokinin A. Binding parameters for synthetic [Leu3,Ile7]neurokinin A and mammalian tachykinins were compared using receptor-selective radioligands and crude membranes from tissues enriched in the NK1, NK2 and NK3 receptors. [Leu3,Ile7]Neurokinin A was approx. 3-fold less potent than substance P in inhibiting the binding of 125I-labelled [Sar9,Met(O2)11]substance P (labelled with Bolton-Hunter reagent) to rat submandibular gland (NK1 receptor), 8-fold less potent than neurokinin A in inhibiting the binding of [2-[125I]iodohistidine1]neurokinin A to rat stomach fundus (NK2 receptor) and 6-fold less potent...
Journal of the Autonomic Nervous System, 1996
We have studied the expression, distribution and function of receptors for the sensory neurotrans... more We have studied the expression, distribution and function of receptors for the sensory neurotransmitters, substance P (SP) and calcitonin gene-related peptide (CGRP) in the dilator muscle and arterioles of the rat iris. Immunohistochemical studies showed that the sensory fibres containing these peptides are distributed throughout the connective tissue stroma of the iris and in association with the larger arterioles, but do not come into close association with the dilator muscle cells. Using reverse-transcription polymerase chain reaction, we have shown that both NK1 and NK3 receptor message is expressed by iris tissue, comprising both dilator muscle and stromal tissue. Binding sites for the NK1 agonist, [Sar9, Met(O2)11]-substance P (SarSP), and for CGRP are confined to the stromal layer and to the larger arterioles within that layer and do not appear to be associated with the dilator muscle itself. Application of either SarSP or CGRP produced both a vasodilatation and an inhibition of sympathetic nerve-induced vasoconstriction of the larger arterioles. Neither SarSP nor CGRP altered the resting tone of the dilator nor were they capable of modulating the contractions due to sympathetic nervous activity. These results suggest that the sensory fibres perform an efferent role in the larger irideal arterioles while their presence in the irideal stroma appears not to modulate the activity of the dilator muscle.
Digestive Diseases and Sciences, 2015
Prostaglandin E2 (PGE2) is the dominant prostaglandin in the colon and is associated with colonic... more Prostaglandin E2 (PGE2) is the dominant prostaglandin in the colon and is associated with colonic inflammation. PGE2 levels are regulated not only by cyclooxygenases (COX-1 and COX-2) but also by 15-hydroxyprostaglandin dehydrogenase (15-PGDH), the major PGE2-degrading enzyme. Information about the involvement of 15-PGDH in colonic inflammation is sparse. We thus aimed to determine the gene expression and immunoreactivity (IR) of COX-1, COX-2, and 15-PGDH in colonic mucosa from patients with diverse inflammatory disorders: ulcerative colitis (UC), Crohn's disease (CD), and acute diverticular disease (DD). RNA from human colonic mucosa was extracted and assessed for gene expression by real-time PCR. Intact colon sections were processed for immunohistochemistry with immunostaining of the mucosal areas quantified using ImageJ. In colonic mucosa of both UC and CD, COX-2 mRNA and COX-2-IR were significantly increased, whereas 15-PGDH mRNA and 15-PGDH-IR were significantly reduced. In macroscopically undamaged acute DD mucosa, the opposite findings were seen: for both gene expression and immunoreactivity, there was a significant downregulation of COX-2 and upregulation of 15-PGDH. COX-1 mRNA and COX-1-IR remained unchanged in all diseases. Our study for the first time demonstrated differential expression of the PGE2-related enzymes COX-2 and 15-PGDH in colonic mucosa from UC, CD, and acute DD. The reduction of 15-PGDH in IBD provides an additional mechanism for PGE2 increase in IBD. With respect to DD, alterations of PGE2-related enzymes suggest that a low PGE2 level may precede the onset of inflammation, thus providing new insight into the pathogenesis of DD.
European Urology Supplements, 2006
Regulatory Peptides, 2000
Neurotensin (NT) is an important regulatory peptide in the mammalian gastrointestinal tract. The ... more Neurotensin (NT) is an important regulatory peptide in the mammalian gastrointestinal tract. The influence of NT on intestinal contractility has been investigated in rodents, but few studies exist in humans and little information is available about its role in gastrointestinal diseases. In this study, normal colon was obtained from patients undergoing resection for colon carcinoma (30-79 years, n=32). Radioligand binding with [125l]NT in normal sigmoid colon showed a high affinity binding site (Kd 0.96 +/-0.4 nM, Bmax 3.5 +/-0.4 fmol/mg). Binding was inhibited by NT(8-13) > NT = SR14294A > neuromedin N 7 SR48692, indicating binding to the NT1 receptor. Autoradiographic binding sites for [125l]NT were seen over the circular muscle and myenteric ganglia. The potency of NT to elicit a contractile response in circular muscle strips from sigmoid and ascending colon was similar (pD2 7.1+/-0.3 nM, 6.8 +I-0.4 nM respectively), but the efficacy of NT was greater in sigmoid colon, compared to ascending. Responses to NT were significantly potentiated in the presence of indomethacin in ascending colon, and significantly reduced in the presence of the tachykinin NK2 receptor antagonist MEN10627 in sigmoid colon. Together these results suggest that NT responses are mediated by specific NT receptors located on the smooth muscle and myenteric ganglia, which induce direct contraction as well as release of mediators such as tachykinins and relaxant prostanoids. The mechanism of action and efficacy of NT differs in the proximal compared to the distal colon. Contractile responses to circular muscle from abnormal colon (diverticular disease, 33-74 years, n=7; idiopathic chronic constipation, 30-73 years, n=7) were compared with normal circular muscle from age-and region-matched specimens. Preliminary results suggest that responses to NT were significantly reduced in diverticular disease and idiopathic constipation, suggesting a role for NT in both diseases.
Digestive diseases and sciences, 2002
Many studies have indicated changes in neuropeptides in inflammatory bowel disease (IBD), but wit... more Many studies have indicated changes in neuropeptides in inflammatory bowel disease (IBD), but with contradictory results. Nerve growth factor also has a potential role in the maintenance of enteric nerves and may be associated with IBD. A quantitative immunohistochemical method was used to measure area density of immunoreactive nerves in the colonic mucosa of surgical specimens. No significant differences in immunoreactivity for substance P, vasoactive intestinal polypeptide, growth associated protein 43, and the neurotrophin receptor p75 were seen in the control, Crohn's, and ulcerative colitis groups. Compared to age-matched normal colon (N = 18), there was an increase in neutrophil number in Crohn's (P < 0.05) and ulcerative colitis (P < 0.01) (both N = 9). There were positive correlations (P < 0.05) between neutrophil number and growth associated protein, between p75 and substance P immunoreactive nerves in ulcerative colitis, and between p75 and vasoactive inte...
Peptides, 1993
Neurokinin A (NKA), substance P (SP), and neurokinin B (NKB) enhanced the contractile force of ut... more Neurokinin A (NKA), substance P (SP), and neurokinin B (NKB) enhanced the contractile force of uterine preparations from estrogen-treated rats. Neurokinin A was more and NKB less potent than SP. The actions of SP were enhanced by phosphoramidon (1 microM) but were unaffected by captopril (10 microM) or bestatin (10 microM). The actions of the peptides were enhanced in the combined presence of phosphoramidon, captopril, and bestatin; the potency order remained NKA &gt; SP &gt; NKB. Atropine inhibited responses to NKB but not to NKA, and slightly reduced those to SP. Specific binding of [125I]-iodohistidyl-neurokinin A (INKA) to uterine membranes was displaced by the tachykinins with a potency order of NKA &gt; SP &gt; NKB. These findings indicate that in the rat uterus 1) tachykinins act at an NK-2 receptor, and that another tachykinin receptor on cholinergic nerves may also be present; and 2) endopeptidase-24.11 participates in the inactivation of the tachykinins.
Synapse, 1988
Whole mounts of guinea pig ileum submucosa were incubated with radiolabeled tachykinins, and bind... more Whole mounts of guinea pig ileum submucosa were incubated with radiolabeled tachykinins, and binding sites were visualized using autoradiography. Very dense specific binding for [1251]-Bolton-Hunter substance P (BHSP) was observed over ganglia of the submucous plexus, with weaker binding over internodal strands. Dense specific binding was also seen over occasional strands of circular muscle, with weak binding over clumps of mucosa. Although very weak binding was seen over some large blood vessels, no binding was associated with smaller blood vessels. Localization of binding was absent in whole-mounts coincubated with 1 p M substance P, used to define nonspecific binding. Localization of BHSP-specific binding was also abolished in whole-mounts coincubated with 1 nM substance P, but not with 1 nM neurokinin B, suggesting that binding was probably to a n NK-1 tachykinin receptor. In whole-mounts incubated in [ 1251]-iodohistidyl neurokinin A (INKA) or [1251]-Bolton-Hunter neurokinin B (BHNKB), no specific binding over ganglia was observed. These binding sites €or BHSP are probably identical with the neuronal substance P receptors mediating mucosal ion transport.
Uploads
Papers by Elizabeth Burcher