A biomarker panel predicts progression of Barrett's esophagus to esophageal adenocarcinoma.

Progression from Barrett's esophagus (BE) to esophageal adenocarcinoma (EAC) is uncommon but the consequences are serious. Predictors of progression are essential to optimize resource utilization. This study assessed the utility of a promising panel of biomarkers applicable to routine paraffin embedded biopsies (FFPE) to predict progression of BE to EAC in a large population-based, nested case-control study.We utilized the Amsterdam-based ReBus nested case-control cohort. BE patients who progressed to high-grade dysplasia (HGD)/EAC (n = 130) and BE patients who never progressed (n = 130) were matched on age, sex, length of the BE segment, and duration of endoscopic surveillance. All progressors had minimum 2 years of endoscopic surveillance without HGD/EAC to exclude prevalent neoplasia. We assessed abnormal DNA content, p53, Cyclin A, and Aspergillus oryzae lectin (AOL) in FFPE sections. We performed conditional logistic regression analysis to estimate odds ratio (OR) of progression based on biomarker status.Expert LGD (OR, 8.3; 95% CI, 1.7-41.0), AOL (3 vs. 0 epithelial compartments abnormal; OR, 3.6; 95% CI, 1.2-10.6) and p53 (OR, 2.3; 95% CI, 1.2-4.6) were independently associated with neoplastic progression. Cyclin A did not predict progression and DNA ploidy analysis by image cytometry was unsuccessful in the majority of cases, both were excluded from the multivariate analysis. The multivariable biomarker model had an area under the receiver operating characteristic curve of 0.73.Expert LGD, AOL, and p53 independently predict neoplastic progression in BE patients and are applicable to routine practice. These biomarkers can aid in selecting patients for endoscopic ablation or more intensive surveillance.

Assessing the understanding of biobank participants.

Biobanks have been developed as a tool to better understand the genetic basis of disease by linking DNA samples to corresponding medical information. The broad scope of such projects presents a challenge to informed consent and participant understanding. To address this, 200 telephone interviews were conducted with participants in the NUgene Project, Northwestern University's biobank. Interviews included a modified version of the "quality of informed consent measure" (QuIC) and semi-structured questions which were analyzed thematically for 109 of the interviews. The QuIC, originally applied to cancer clinical trials, objectively assessed some of the components of informed consent for a biobank, and interview questions provided rich data to assist in interpreting participant understanding. The best understood domains included: the nature of the study, benefit to future patients, and the voluntary nature of participation. Lower knowledge scores included: potential risks and discomforts, experimental nature of the research, procedures in the event of study-related injury, and confidentiality issues. Qualitatively, confidentiality protections of the study were described as good by most (>50%). Although some cited concerns with employer (12%) or insurance discrimination (25%), most considered the risks to privacy low (25%) or none (approximately 60%). Only 10% of participants explicitly stated they had no expectation for personal benefit, and when asked whether they expected to be contacted with study results, respondents were split between having no expectation (39%), being hopeful for results (37%) and expecting to be contacted with results (12%). These findings are informative to those establishing and implementing biobanks, and to the IRBs reviewing such studies.

A visual analogue scale and a Likert scale are simple and responsive tools for assessing dysphagia in eosinophilic oesophagitis.

BACKGROUND: While symptom scores have been developed to evaluate dysphagia in eosinophilic oesophagitis (EoE), their complexity may limit clinical use. AIM: To evaluate a visual analogue scale (VAS) and a 10-point Likert scale (LS) for assessment of dysphagia severity before and after EoE treatment. METHODS: We conducted a prospective cohort study enrolling consecutive adults undergoing out-patient endoscopy. Incident cases of EoE were diagnosed per consensus guidelines. At diagnosis and after 8 weeks of treatment, symptoms were measured using the VAS, LS and the Mayo Dysphagia Questionnaire (MDQ). The percentage change in scores before and after treatment were compared overall, in treatment responders (<15 eos/hpf) and non-responders, and in patients without baseline dilation. RESULTS: In 51 EoE cases, the median VAS decreased from 3.6 at baseline to 1.4 post-treatment (71% decrease), the LS decreased from 6 to 2 (67%) and the MDQ decreased from 20 to 10 (49%). The VAS correlated with both the LS (R = 0.77; P < 0.0001) and MDQ (R = 0.46, P = 0.001). After stratification by histological response, the LS decreased 70% in responders vs. 13% in non-responders (P = 0.02). In patients who did not receive baseline dilation, both the VAS and LS decreased significantly more in the histological responders. CONCLUSIONS: Both the VAS and LS were responsive to successful treatment as measured by histologic improvement. Because the VAS and LS are simple to administer and are responsive to treatment, they can provide an efficient and objective method for assessing dysphagia severity in EoE in clinical practice.

Review of the role of the central serotonergic neuronal system in blood pressure regulation.

Alterations in the dynamics of brain serotonin biosynthesis can lead to changes in cardiovascular function. It appears that the activation of cerebral serotonin receptors produces a pressor effect in normotensive rats but produces a depressor effect in normotensive cats or dogs. On the other hand, reductions in the levels of serotonin can prevent the onset of hypertension in some experimental hypertensive models and lower the blood pressure of organisms with established hypertension. The ability of brain serotonin to modulate arterial blood pressure may be mediated by the influences of the serotonergic neuronal systems on efferent sympathetic activity. Finally, the reduction in sympathetic outflow produced by increasing brain serotonin levels in dogs protects the heart against ventricular fibrillation and may, therefore, constitute a reasonable adjunct in the management of high-risk, cardiac-arrest patients.

Estrogen desensitizes 5-HT(1A) receptors and reduces levels of G(z), G(i1) and G(i3) proteins in the hypothalamus.

The present study investigated whether estrogen would desensitize hypothalamic serotonin(1A) (5-HT(1A)) receptors by examining the neuroendocrine response to 8-OH-DPAT, a 5-HT(1A) agonist. Rats were ovariectomized, allowed to recover for 5 days, then given 2 daily injections of estradiol benzoate or vehicle (10 microg/day, s.c.). Twenty-four hours after the second injection, rats were challenged with a sub-maximal dose of 8-OH-DPAT (50 microg/kg, sc) or saline 15 min prior to sacrifice. 8-OH-DPAT produced a significant increase in plasma oxytocin, ACTH and corticosterone levels in ovariectomized rats. While estrogen treatment for 2 days did not alter basal hormone levels, it did significantly reduce the magnitude of oxytocin, ACTH and corticosterone responses to 8-OH-DPAT. The reduction in hormone responses was accompanied by a significant reduction in hypothalamic levels of G(z), G(i1) and G(i3) proteins (by 50%, 30% and 50%, respectively). These findings suggest that a reduction in these G proteins may contribute to the mechanisms underlying estrogen-induced desensitization of 5-HT(1A) receptors. The desensitization of 5-HT(1A) receptors has been suggested to underlie the therapeutic effects of antidepressant 5-HT uptake inhibitors (SSRIs). Thus, the present results suggest that estrogen or estrogen-like substances in combination with SSRIs may prove effective in developing novel therapeutic strategies for neuropsychiatric disorders in women.

5-Hydroxytryptamine release in vivo from a cytoplasmic pool: studies on the 5-HT behavioural syndrome in reserpinized rats.

Treatment of rats with reserpine in order to disrupt vesicular amine storage reduces 5-hydroxytryptamine (5-HT) levels throughout brain by 90-95%. Despite the drastic reduction in brain 5-HT content by reserpine, the 5-HT releasing drug p-chloramphetamine (PCA) produces a behavioural syndrome in reserpine-treated rats which is not different from that observed in normal animals given PCA. Prior treatment of reserpinized rats with p-chlorophenylalanine (PCPA), the irreversible tryptophan hydroxylase inhibitor which inhibits the synthesis of new 5-HT, prevents the PCA-induced behavioural syndrome. The 5-HT receptor antagonist methergoline, blocks the PCA effect in reserpine-treated rats. Treatment of reserpinized rats with pargyline, a non-selective inhibitor of monoamine oxidase, in order to increase cerebral 5-HT levels, shifts the PCA dose-response curve for inducing the 5-HT behavioural syndrome to the left. The specific 5-HT uptake blocker, fluoxetine, protects normal and reserpine-treated rats from the 5-HT depleting effects of PCA but does not always prevent the PCA-induced 5-HT behavioural syndrome. These results indicate that PCA releases 5-HT into the synapse from a small cytoplasmic pool which is resistant to reserpine and suggest that this newly synthesized compartment of 5-HT represents the 'functional' transmitter pool.

Inhibition of tryptophan hydroxylase by benserazide and other catechols.

Tryptophan hydroxylase (L-tryptophan, tetrahydropteridine:oxygen oxidoreductase [5-hydroxylating]; EC 1.14.16.4; TPH), the initial and rate-limiting enzyme in the biosynthesis of the neurotransmitter serotonin, was inhibited directly by benserazide, an inhibitor of aromatic-L-amino-acid decarboxylase (3,4-dihydroxy-L-phenylalanine carboxy-lyase; EC 4.1.1.28; AAAD). Benserazide was a competitive inhibitor for the pterin cofactor tetrahydrobiopterin and an uncompetitive inhibitor for the substrate tryptophan. NSD 1015, another decarboxylase inhibitor, did not directly inhibit TPH. Other compounds with catechol moieties in their structures such as 3,4-dihydroxyphenylalanine (DOPA), dopamine, apomorphine, and SKF 38393 were also found to be potent inhibitors of TPH. These results indicate that drugs or neurotransmitters with catechol structures directly inhibit the activity of TPH and add to a growing body of evidence indicating that endogenous dopamine can exert untoward effects on serotonin neurons, including inhibition of TPH. Furthermore, the use of decarboxylase inhibitors to cause the accumulation of 5-hydroxytryptophan as an in vivo measure of TPH activity could be problematic, particularly when drugs with catechol structures or dopamine-releasing compounds are also administered.

5-HT3- and 5-HT2C-antagonist properties of cyamemazine: significance for its clinical anxiolytic activity.

RATIONALE: Cyamemazine is a neuroleptic compound which possesses anxiolytic properties in humans. On the other hand, 5-HT3- and 5-HT2C-receptors have been implicated in anxiety disorders and a previous binding study has shown that cyamemazine possesses high affinity for both serotonin receptor types. OBJECTIVE: The present study was undertaken to establish whether cyamemazine antagonizes 5-HT3- and/or 5-HT2C-mediated responses, and whether it compares with reference compounds. METHODS: Cyamemazine was tested for its ability to antagonize: (i) 5-HT3-dependent contraction of the isolated guinea-pig ileum and bradycardic responses in the rat and (ii) 5-HT2C-dependent phospholipase C (PLC) stimulation in rat brain membranes. RESULTS: In isolated guinea-pig ileum, cyamemazine potently and competitively antagonized 5-HT-dependent contractions (pA2 = 7.52 +/- 0.08; n = 5). In this test, cyamemazine was 5-7 times more potent (pIC50 = 6.75 +/- 0.13) than tropisetron (pIC50 = 6.02 +/- 0.04). In rats, cyamemazine i.v. antagonized 5-HT-dependent bradycardic responses with ID50% = 3.2 +/- 1.5 mg/kg (n = 4). Finally, in rat brain membranes cyamemazine antagonized 5-HT2C-dependent PLC stimulation with Ki = 424 nM (mianserin exhibits a Ki = 113 nM). CONCLUSIONS: Cyamemazine behaves as an antagonist at both 5-HT3- and 5-HT2C-receptors, which compares well with reference compounds. These 5-HT3- and 5-HT2C-antagonistic actions of cyamemazine can be involved, at least in part, in its beneficial therapeutic actions in anxiety disorders.

Cocaine and serotonin neurochemistry.

The neurochemical interactions between cocaine and the serotonin (5-HT) neuronal system are the subject of this Commentary. Attempts to understand the mechanisms through which the euphoric and rewarding properties of cocaine are mediated have focused on the brain dopamine system, with particular emphasis on the dopamine uptake site. However, increasing amounts of research have led to the realization that cocaine also exerts a powerful influence over the serotonin neuronal system. These data are briefly discussed from the standpoint of how cocaine alters the functional or synaptic activity of serotonin. Recent evidence also indicates that the reinforcing properties of cocaine can be reduced by treatments which interact directly or indirectly with the serotonin neuronal system.

Serotonin neurochemistry revisited: A new look at some old axioms.

A number of principles have long guided research into serotonin neurochemistry and some of them are presently reconsidered with a slightly different outlook. An increasing amount of experimental data does not always support these older axioms and as more empirical observations are made, some of these may require modification. For example, increases in 5-HT produced by injections of l-tryptophan do not necessarily indicate that tryptophan hydroxylase is less than saturated with its substrate in vivo; injections of l-tryptophan increase 5-HT and its turnover but do not appear to increase its release; 5-HT release can apparently occur from the cytoplasm and need not involve exocytosis; presynaptic autoreceptors do not appear to modulate 5-HT release; and, 5-HIAA most accurately reflects monoamine oxidase activity, not release of 5-HT or activity of 5-HT neurons.

Influence of tetrahydrobiopterin on serotonin synthesis, metabolism and release in synaptosomes.

The effects of (6R)-5,6,7,8-tetrahydro-l-biopterin (BH(4)) on the uptake of tryptophan, its conversion to serotonin (5-HT) and 5-hydroxyindoleacetic acid and on the basal release of 5-HT was studied in rat brain synaptosomes. When BH(4), the essential cofactor for tryptophan hydroxylase, was incubated with synaptosomes in concentrations varying from 10 to 200 ?M, there was no effect on 5-HT formation, metabolism or release. Concentrations of 1-2 mM BH(4) had strong inhibitory effects on 5-HT synthesis. The incubation of synaptosomes with tryptophan increased the synthesis of 5-HT, but BH(4) did not further increase this effect. BH(4) was taken up into synaptosomes in a concentration-dependent manner under all incubation conditions and was stable in the chemically reduced (tetrahydro-) form. These results indicate that increases in the synaptosomal concentration of BH(4) do not increase the synthesis and release of 5-HT. It is concluded from the present results that tryptophan hydroxylase is saturated with BH(4) in synaptosomes.

Antihypertensive effects of L-tryptophan are not mediated by brain serotonin.

L-Tryptophan produces a significant antihypertensive effect in spontaneously hypertensive rats while D-tryptophan does not change blood pressure. Both isomers of tryptophan significantly increase brain serotonin to the same extent at a time when the antihypertensive effect of L-tryptophan is maximal. Thus, the antihypertensive effects of L-tryptophan do not appear to be mediated by brain serotonin.

Pressor effects of dorsal raphe stimulation and intrahypothalamic application of serotonin in the spontaneously hypertensive rat.

Electrical stimulation of the dorsal raphe nucleus or direct microinjection of serotonin into the preoptic region of the anterior hypothalamus produces a transient rise in arterial blood pressure in both spontaneously hypertensive rats (SRH) and Wistar--Kyoto (WKY) controls. SRHs are more responsive to raphe stimulation but are somewhat less responsive to serotonin injections when compared to WKYs. The serotonin antagonist metergoline blocks the pressor response to serotonin in both strains. These results suggest that the central serotonergic neuronal system play a similar, but not identical, role in blood pressure modulation in hypertensive and normotensive rats.

A disconnection analysis of hippocampal function.

A disconnection analysis determined the extent to which the fornix, hippocampus and entorhinal cortex are components of the same functional system in tasks that require working memory. Preoperatively, rats were trained to perform accurately on a radial arm maze. Then various combinations of unilateral and bilateral lesions were placed in the fornix and entorhinal cortex, either with or without a transection of the hippocampal commissures. When the lesions left intact at least one pathway through the hippocampus interconnecting the fornix and entorhinal cortex, rats performed normally. Either an uncrossed pathway (following a unilateral lesion of the fornix, transection of the hippocampal commissures, and an ipsilateral lesion of the entorhinal cortex) or a crossed pathway (following a unilateral lesion of the fornix and a contralateral lesion of the entorhinal cortex, leaving the hippocampal commissures intact) was sufficient. When the lesions produced a complete bilateral disconnection of the fornix and entorhinal cortex, rats performed poorly. The results indicate that the hippocampal system provides a functional connection between the subcortical structures associated with the fornix and the neocortical structures associated with the entorhinal cortex, and that without this connection normal processing of working memory can not occur.

Importance of spinal noradrenergic pathways in cardiovascular reflexes and central actions of clonidine and alpha-methyldopa in the rabbit.

We have examined in conscious rabbits the chronic effects of 6-hydroxydopamine (6-OHDA)-induced local lesions of the spinal noradrenaline (NA) pathways on (i) resting mean arterial pressure (MAP) and heart rate (HR), (ii) the nasopharyngeal pressor response, (iii) the sympathetic component of the baroreceptor-heart rate reflex (iv) the acute responses to intracisternal (i.c.) clonidine and alpha-methyldopa (alpha-MD), and (v) the acute NA release response produced by i.e. 6-OHDA. One month after injection of 6-OHDA (40 nmol in 4 microliters) into the first cervical spinal cord segment (C1), the NA content was reduced to 29% in C2, 45% in T4 and 61% in L3 with little non-specific damage. Basal MAP was 14% higher (P less than 0.05) than in sham-operated rabbits suggesting increased vasoconstrictor tone. Basal cardiac sympathetic tone was enhanced, but a corresponding increase in cardiac vagal tone resulted in little net effect on resting HR in the spinal NA-depleted group. Spinal NA lesions attenuated the nasopharyngeal pressor reflex by 27% in baroreceptor-intact rabbits and by 38% in sino-aortically denervated (SAD) animals. The lesion did not affect HR range, gain and BP50 of the sympathetic baroreflex. In SAD rabbits, the acute MAP responses to i.c. 6-OHDA (early hypotension, late hypertension) were not affected by spinal NA depletion, but the early fall in HR (cardiac sympathetic inhibition) was abolished. The hypotension produced by i.c. clonidine or alpha-MD was not affected by the lesion, probably because many of the NA terminals in the lower thoracic and upper lumbar cord were still intact. Our results suggest that intraspinal NA fibers have a tonic inhibitory action on spinal preganglionic vasoconstrictor and cardiac motoneurons. The spinal NA neurons affecting vasomotor tone (but not cardiac sympathetic tone) are in turn inhibited by higher vasomotor centers receiving projections from the arterial and trigeminal afferents and thereby participate in vasoconstrictor reflexes.

Effects of combined serotonin depletion and lesions of the nucleus basalis magnocellularis on acquisition of a complex spatial discrimination task in the rat.

The purpose of the present experiment was to determine the effects of lesions of cholinergic neurons originating from the nucleus basalis magnocellularis (NBM), alone or in combination with central serotonin depletion, on learning and memory in rats trained in the Stone 14-unit T-maze--a complex, positively-reinforced spatial discrimination task. Lesion of cholinergic neurons within the NBM was accomplished by bilateral infusion of ibotenic acid. Serotonin depletion was accomplished by the systemic administration of p-chloroamphetamine (PCA). The results show that PCA-induced serotonin depletion enhanced learning. This effect was completely prevented by NBM lesions, despite the fact that NBM lesions alone did not affect the performance of rats in this task. The results of this study support the view that the cholinergic and serotonergic systems may functionally interact in learning and memory processes. The significance of this interaction in the etiology and treatment of dementia should be further investigated.

Effect of corticosterone on serotonin and catecholamine receptors and uptake sites in rat frontal cortex.

The effects of corticosterone (1 mg/kg per day for 7 days) on serotonin 5-HT1A, 5-HT2A, 5-HT uptake sites, and alpha 2-adrenergic receptor sites were measured. Corticosterone treatment significantly decreased the number of 5-HT1A receptor sites (Bmax = 108 +/- 8.20 fmol/mg protein and 152.31 +/- 13.36 fmol/mg protein in corticosterone- and vehicle-treated rats, respectively). No significant differences were found in other measures. It is possible that corticosteroids exert some of their behavioral effects via regulation of 5-HT1A sites in frontal cortex.

Preprotachykinin and preproenkephalin mRNA expression within striatal subregions in response to altered serotonin transmission.

The effects of lowered serotonin (5-hydroxytryptamine; 5-HT) neurotransmission on preprotachykinin (PPT) and preproenkephalin (PPE) mRNA levels were examined in subregions of the striatum. Adult male rats were treated systemically with para-chlorophenylalanine (pCPA; 350 mg/kg single i.p. injection) which reduced forebrain 5-HT amounts to approximately 20% of saline-injected controls at 24 and 48 h. As measured by Northern analysis, PPT and PPE mRNA levels were elevated 50% and 160% respectively in the anterior ventromedial striatum (region included nucleus accumbens). PPT mRNA levels were raised 90% in posterior striatum (at the level of the globus pallidus) by 48 h post-pCPA injection. To determine if increased PPT and PPE mRNA levels represented a transient response to brief 5-HT inhibition, additional experiments were performed to provide continual suppression of 5-HT within the striatum. First, rats received daily intraperitoneal injections of saline or the 5-HT1A receptor agonist, 8-OH-DPAT (1 mg/kg), for 7 days to reduce 5-HT release from raphestriatal terminals. In a parallel experiment, the serotonin neurotoxin, 5,7-dihydroxytryptamine (5,7-DHT, 5 micrograms), was stereotaxically injected into the striatum as a means to permanently remove 5-HT terminals. Although levels of each mRNA species were differentially sensitive to 5,7-DHT or 8-OH-DPAT, PPT and PPE mRNAs were lowered between 30-55% within the anterior dorsolateral and ventromedial striatum. Although these results support previous studies suggesting an overall positive regulatory role of serotonin on striatal tachykinin biosynthesis, PPT and PPE gene regulation in certain striatal subregions may by differentially sensitive to lowered 5-HT neurotransmission. This suggestion is supported by observations that acute systemic stimulation of 5-HT2A/C receptors with DOI (7 mg/kg single i.p. injection) raised PPT and PPE mRNA levels within anterior dorsolateral (30-60%) and posterior (100-200%) striata, but not within the anterior ventromedial striatum.

alpha-Methyldopa metabolism in central serotonergic nerve terminals: effects on serotonin levels, synthesis and release.

The direct effects of in vivo methyldopa administration on serotonin (5-HT) neurochemistry was investigated. Specifically the ability of methyldopa to alter nerve terminal-associated 5-HT synthesis, storage and release and the possibility that 5-HT nerve terminals accumulate methyldopamine (the product of decarboxylation of methyldopa) was investigated. Synaptosomes isolated from rats given 200 mg/kg of methyldopa (calculated as the free amino acid) 2 h prior to killing exhibited a 25% reduction in intrasynaptosomal 5-HT and a 15% reduction in 5-HT synthesis when compared to synaptosomes from saline-treated animals. In addition a 15% reduction in synaptosomal tryptophan levels was observed. Despite these changes there was no apparent decrease in basal or depolarization-induced 5-HT release from synaptosomes of methyldopa-treated rats. The presence of methyldopamine within 5-HT-containing synaptosomes was confirmed by demonstrating that p-chloroamphetamine, a selective 5-HT releasing agent, could release both methyldopamine and 5-HT from synaptosomes and that this release could be selectively antagonised by fluoxetine, a selective 5-HT uptake inhibitor. The significance of these data with respect to the involvement of 5-HT neurons in the hypotensive action of methyldopa is discussed.

Blood pressure responses to local application of serotonergic agents in the nucleus tractus solitarii.

The effects of serotonin applied directly to the region of the medulla oblongata which contains the nucleus tractus solitarii, nucleus intercalatus, nucleus originis dorsalis vagi and the nucleus originis nervi hypoglossi were investigated in anesthetized rats. A dose dependent increase in blood pressure with variable changes in heart rate was observed after unilateral application of serotonin into the nucleus tractus solitarius. The serotonergic antagonists, 2-bromolysergic acid diethylamide (locally applied) and metergoline (systemically administered) significantly attenuated the serotonin-induced pressor response. Fluoxetine, a serotonin uptake inhibitor, significantly enhanced the magnitude of the pressor response, but did not prolong it. The present data suggest that enhancement of serotonergic activity in the region of the nucleus tractus solitarius produces a neurogenic pressor response.