Endothelial ILK induces cardioprotection by preventing coronary microvascular dysfunction and endothelial-to-mesenchymal transition.

Endothelial dysfunction is an early event in coronary microvascular disease. Integrin-linked kinase (ILK) prevents endothelial nitric oxide synthase (eNOS) uncoupling and, thus, endothelial dysfunction. However, the specific role of endothelial ILK in cardiac function remains to be fully elucidated. We hypothesised that endothelial ILK plays a crucial role in maintaining coronary microvascular function and contractile performance in the heart. We generated an endothelial cell-specific ILK conditional knock-out mouse (ecILK cKO) and investigated cardiovascular function. Coronary endothelial ILK deletion significantly impaired cardiac function: ejection fraction, fractional shortening and cardiac output decreased, whilst left ventricle diastolic internal diameter decreased and E/A and E/E' ratios increased, indicating not only systolic but also diastolic dysfunction. The functional data correlated with extensive extracellular matrix remodelling and perivascular fibrosis, indicative of adverse cardiac remodelling. Mice with endothelial ILK deletion suffered early ischaemic-like events with ST elevation and transient increases in cardiac troponins, which correlated with fibrotic remodelling. In addition, ecILK cKO mice exhibited many features of coronary microvascular disease: reduced cardiac perfusion, impaired coronary flow reserve and arterial remodelling with patent epicardial coronary arteries. Moreover, endothelial ILK deletion induced a moderate increase in blood pressure, but the antihypertensive drug Losartan did not affect microvascular remodelling whilst only partially ameliorated fibrotic remodelling. The plasma miRNA profile reveals endothelial-to-mesenchymal transition (endMT) as an upregulated pathway in endothelial ILK conditional KO mice. Our results show that endothelial cells in the microvasculature in endothelial ILK conditional KO mice underwent endMT. Moreover, endothelial cells isolated from these mice and ILK-silenced human microvascular endothelial cells underwent endMT, indicating that decreased endothelial ILK contributes directly to this endothelial phenotype shift. Our results identify ILK as a crucial regulator of microvascular endothelial homeostasis. Endothelial ILK prevents microvascular dysfunction and cardiac remodelling, contributing to the maintenance of the endothelial cell phenotype.

Previous outpatient antibiotic use in patients admitted to hospital for COPD exacerbations: room for improvement.

PURPOSE: Several studies have analyzed factors associated to hospitalization in chronic obstructive pulmonary disease (COPD) patients. However, data are lacking on the quality of treatment received by patients prior to hospital admission. The present study analyzed how often patients requiring hospitalization for a COPD exacerbation had received previous treatment for the exacerbation, particularly antibiotics. METHODS: This was a multicenter, cross-sectional, observational study conducted in 30 Spanish hospitals among COPD patients aged >40 years who were hospitalized for an acute exacerbation. Patients were grouped according to whether or not they had received treatment prior to admission and, subsequently, according to whether or not they had received antibiotics. Patient eligibility for antibiotic therapy was assessed using both national and European guidelines. RESULTS: The study population consisted of 298 patients, of which 277 (93 %) were men, with a mean [standard deviation (SD)] age of 69.1 (9.5) years. One hundred and thirty-three patients (45 %) had received treatment prior to admission; among these, 76/133 (57 %) had received antibiotic therapy. However, 81-91 % of these patients fulfilled criteria for this therapy. Antibiotic use was significantly associated with yellow or green-yellow sputum prior to the exacerbation, a higher number of exacerbations in the previous year, more visits to emergency departments, and bronchiectasis. On the other hand, 10-20 % of patients who did receive antibiotics were not eligible for this therapy according to guidelines. CONCLUSIONS: This study demonstrates a low rate of previous outpatient treatment and antibiotic use among patients with a COPD exacerbation requiring hospital admission.

The prognostic impact of circulating miRNAs in patients with advanced esophagogastric cancer during palliative chemotherapy.

The prognosis of patients with advanced oesophageal cancer (EC) and gastric cancer (GC) is poor. Circulating microRNAs (ci-miRNAs) may have prognostic and predictive value to improve patient selection for palliative treatment. The purpose of this study is to assess the prognostic and predictive value of specific ci-miRNAs in plasma of patients with EC and GC treated with first-line palliative gemcitabine and cisplatin. Droplet digital PCR (ddPCR) was used to quantify miR-200c-3p, miR-375, miR-21-5p, miR-148a-3p, miR-146a-5p, miR-141-3p and miR-218-5p in plasma from 68 patients. ci-miRNA expression was analyzed in relation to overall survival (OS), progression-free survival (PFS), and response to chemotherapy. ci-miRNA levels were detectable in 36 baseline (71%) samples and in 14 (47%) follow-up samples. Increased circulating miR-200c-3p in GC showed a trend (p = 0.06) towards a shorter OS. High circulating miR-375 was associated with a longer OS (p = 0.02) in patients with esophageal adenocarcinoma (EAC). No significant difference was observed in ci-miRNA expression between paired pre- and on-treatment samples. ci-miRNA expression was not associated with response to chemotherapy. ci-miRNAs can be measured in plasma samples of patients treated with first-line palliative chemotherapy using ddPCR despite prolonged storage in heparin. Elevated circulating miR-375 might be a prognostic marker for patients with EAC.

Bisphenol A induces coronary endothelial cell necroptosis by activating RIP3/CamKII dependent pathway.

Epidemiological studies link long term exposure to xenoestrogen Bisphenol-A to adverse cardiovascular effects. Our previous results show that BPA induces hypertension by a mechanism involving CamKII activation and increased redox stress caused by eNOS uncoupling. Recently, CamKII sustained activation has been recognized as a central mediator of programmed cell death in cardiovascular diseases, including necroptosis. However, the role of necroptosis in cardiac response to BPA had not yet been explored. Mice exposed to BPA for 16 weeks showed altered heart function, electrical conduction, and increased blood pressure. Besides, a stress test showed ST-segment depression, indicative of cardiac ischemia. The hearts exhibited cardiac hypertrophy and reduced vascularization, interstitial edema, and large hemorrhagic foci accompanied by fibrinogen deposits. BPA initiated a cardiac inflammatory response, up-regulation of M1 macrophage polarization, and increased oxidative stress, coinciding with the increased expression of CamKII and the necroptotic effector RIP3. In addition, cell death was especially evident in coronary endothelial cells within hemorrhagic areas, and Evans blue extravasation indicated a vascular leak in response to Bisphenol-A. Consistent with the in vivo findings, BPA increased the necroptosis/apoptosis ratio, the expression of RIP3, and CamKII activation in endothelial cells. Necrostatin-1, an inhibitor of necroptosis, alleviated BPA induced cardiac dysfunction and prevented the inflammatory and hemorrhagic response in mice. Mechanistically, silencing of RIP3 reversed BPA-induced necroptosis and CamKII activation in endothelial cells, while inhibition of CamKII activation by KN-93 had no effect on RIP3 expression but decreased necroptotic cell death suggesting that BPA induced necroptosis is mediated by a RIP 3/CamKII dependent pathway. Our results reveal a novel pathogenic role of BPA on the coronary circulation. BPA induces endothelial cell necroptosis, promotes the weakening of coronary vascular wall, which caused internal ventricular hemorrhages, delaying the reparative process and ultimately leading to cardiac dysfunction.

Efficacy of moxifloxacin in the treatment of bronchial colonisation in COPD.

This study was designed to investigate the efficacy of moxifloxacin for the eradication of bacterial colonisation of the airways in patients with moderate-to-severe chronic obstructive pulmonary disease (COPD). Out of 119 stable patients with COPD screened, 40 (mean age 69 yrs, mean forced expiratory volume in 1 s 50% predicted) were colonised with potentially pathogenic microorganisms (PPMs) and were included in a randomised, double-blind, placebo-controlled trial with moxifloxacin 400 mg daily for 5 days. Eradication rates were 75% with moxifloxacin and 30% with placebo at 2 weeks (p = 0.01). Bacterial persistence at 8 weeks was still higher (not significantly) in the placebo arm (five (25%) out of 20 versus one (5%) out of 20; p = 0.18). The frequencies of acquisition of a new PPM were high and similar in both treatment groups; consequently, the prevalence of colonisation at 8 weeks was also similar between treatment arms. No difference was found in the number of patients with exacerbations during the 5-month follow-up. Only the acquisition of a new PPM during follow-up showed a statistically significant relationship with occurrence of an exacerbation. Moxifloxacin was effective in eradicating PPMs in patients with positive sputum cultures. However, most patients were recolonised after 8 weeks of follow-up. Acquisition of a new strain of bacteria was associated with an increased risk of developing an exacerbation.

Immunostaining in whole-mount lipid-cleared peripheral nerves and dorsal root ganglia after neuropathy in mice.

Immunohistochemical characterization of primary afferent fibers (intact or after nerve damage) is traditionally performed in thin sections from dorsal root ganglia (DRGs) or in teased fibers, as light scattering in whole-mounts compromises visualization. These procedures are time-consuming, require specific equipment and advanced experimental skills. Lipid-clearing techniques are increasing in popularity, but they have never been used for the peripheral nervous system. We established a modified, inexpensive clearing method based on lipid-removal protocols to make transparent peripheral nerve tissue (inCLARITY). We compared retrograde-labeling and free-floating immunostaining with cryo-sections. Confocal microscopy on whole-mount transparent DRGs showed neurons marked with retrograde tracers applied to experimental neuromas (Retrobeads, Fluoro-ruby, Fluoro-emerald, DiI, and Fluoro-gold). After immunostaining with calcitonin gene-related peptide (peptidergic) or isolectin IB4 (non-peptidergic), nociceptors were visualized. Immunostaining in transparent whole-mount nerves allows simultaneous evaluation of the axotomized branches containing the neuroma and neighboring intact branches as they can be mounted preserving their anatomical disposition and fiber integrity. The goal of our study was to optimize CLARITY for its application in peripheral nerve tissues. The protocol is compatible with the use of retrograde tracers and improves immunostaining outcomes when compared to classical cryo-sectioning, as lack of lipids maximizes antibody penetration within the tissue.

Hyperpolarization-activated channels shape temporal patterns of ectopic spontaneous discharge in C-nociceptors after peripheral nerve injury.

BACKGROUND: Neuropathic pain is thought to be mediated by aberrant impulses from sensitized primary afferents, and the temporal summation of the discharges might also influence nociceptive processing. Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels (Ih current) generate rhythmic activity in neurons within the central nervous system and contribute to nociceptors excitability in neuropathic pain. METHODS: We searched for single fibres with ectopic spontaneous discharges from an in vitro preparation in mice containing a neuroma formed in a peripheral branch of the saphenous nerve together with the undamaged branches. RESULTS: Both damaged (axotomized) and undamaged fibres (putative intact) developed ectopic spontaneous activity with different temporal spike trains: Clock-like, Irregular or Bursts. The Ih current blocker, ZD7288, significantly suppressed ectopic spontaneous discharges in nociceptive fibres (3/5 Aδ- and 24/31 C-units and 1 nonclassified) by 64%. Additionally, ZD7288 changed the spike patterns of 5/7 Clock-like and 3/4 Burst units to Irregular. Exogenous cAMP produced a significant ~65% increase in the ectopic firing in 5 Irregular fibres, which was restored by ZD7288. In six additional fibres (three Clock-like and three Irregular), exogenous cAMP had no further effect, but co-application with ZD7288 decreased their discharge by half. These units showed significant higher levels of discharges than the cAMP-sensitive ones. CONCLUSIONS: Our data suggest that HCN channels modulate ectopic spontaneous firing in C-nociceptors and shape their temporal patterns of discharge which will, ultimately, modify the nociceptive message received and processed by second-order neurons. SIGNIFICANCE: We show an involvement of HCN channels in the modulation of ectopic spontaneous discharges from C-nociceptors. This finding exposes a mechanism of nociceptive transmission enhancement and highlights the clinical relevance of peripheral HCN blockade for spontaneous pain relief during neuropathy.

The teratogenicity of allopurinol: A comprehensive review of animal and human studies.

Allopurinol is widely used in the management of multiple disorders including gout, kidney stones and inflammatory bowel disease. Despite of long-term experience, its safety in pregnancy has been debated due to reports on possible teratogenicity. We aimed to review the literature on the safety of allopurinol in pregnancy and offspring. In animals, allopurinol induced species-specific reproductive toxicity. In humans, a total of 53 allopurinol exposed infants were reported in the literature. Major congenital malformations were reported in two cases with a comparable pattern of multiple abnormalities. Five other infants had minor birth defects. In conclusion, the association between allopurinol and teratogenicity appears to be weak and limited to two reports with uncertain causality. However, the available data are insufficient to make a certain judgement, and as allopurinol treatment evolves, report and prospective follow-up of all exposed infants (i.e. deviant and normal cases) should be encouraged.

GABAergic structures in the ventral part of the oral pontine reticular nucleus: An ultrastructural immunogold analysis.

GABA mediates inhibitory effects in neurons of the ventral part of the oral pontine reticular nucleus (vRPO). Evidence increasingly suggests that GABA plays an important role in the modulation of rapid eye movement (REM) sleep generation in the cat vRPO. Here, we investigate the anatomical substrate of this modulation using GABA immunocytochemistry. Immunoperoxidase labeling revealed a few small GABA-immunoreactive cell bodies scattered throughout the vRPO. The numerical densities of all vRPO synapses and the GABA-immunoreactive synapses were estimated, at the electron microscopical level, by using a combination of the physical disector and the post-embedding immunogold techniques. We estimated that 30% of all vRPO synaptic terminals were immunoreactive to GABA. Our findings support the hypothesis that vRPO neuron activity is significantly controlled by inhibitory GABAergic terminals that directly target somata and the different parts of the dendritic tree, including distal regions. GABAergic input could inhibit vRPO REM sleep-inducing neurons during other states of the sleep-wakefulness cycle such as wakefulness or non-REM sleep.

[Attitudes toward the diagnosis of chronic obstructive pulmonary disease in primary care]. / Problemas con el diagnóstico de la EPOC en atención primaria.

OBJECTIVE: Although the prevalence of chronic obstructive pulmonary disease (COPD) has increased among women, it is still considered a disease that mainly affects men. This study aimed to identify the diagnostic attitudes of primary care physicians toward patients with COPD according to gender and spirometric results. METHODS: A representative sample of 839 primary care physicians participated in the study. Each physician dealt with 1 of 8 hypothetical cases based on a patient diagnosed with COPD. In half the cases, the physician was told the patient was a man. The other half of the physicians were told the same patient was a woman. After presentation of the medical history and results of physical examination, the physicians were asked to state a probable diagnosis and indicate the diagnostic tests that were necessary. They were then told the results of spirometry, which indicated obstruction ranging from moderate to severe. Negative results of bronchodilator tests and oral corticosteroid tests were then communicated. RESULTS: COPD was more likely to be the preliminary diagnosis for male patients than for females (odds ratio [OR], 1.55; 95% confidence interval [CI], 1.15-2.1). This gender bias disappeared once the physicians were shown the abnormal results of spirometry. Patients with severe obstruction were more likely to be diagnosed with COPD than those with moderate obstruction (OR, 1.5; 95% CI, 1.08-2.09). CONCLUSIONS: There is gender bias in the diagnosis of COPD. Patients with moderate obstruction are often believed not to have COPD. These biases may compromise the early diagnosis of the disease in a group of patients with ever increasing risk.

Spontaneous activity in C-fibres after partial damage to the saphenous nerve in mice: Effects of retigabine.

BACKGROUND: Spontaneous pain is the most devastating positive symptom in neuropathic pain patients. Recent data show a direct relationship between spontaneous discharges in C-fibres and spontaneous pain in neuropathic patients. Unfortunately, to date there is a lack of experimental animal models for drug testing. METHODS: We recorded afferent fibres from a new experimental model in vitro. The preparation contains a neuroma formed in a peripheral branch of the saphenous nerve together with the undamaged branches, which maintain intact terminals in a skin flap. RESULTS: Fibres with stable rates of ectopic spontaneous discharges were found among axotomized (5 A- and 18 C-fibres, mean discharge 0.48 ± 0.08 Hz) and 'putative intact' fibres (12 C-fibres, mean discharge 0.28 ± 0.08 Hz). A proportion (~9%) of axotomized fibres had mechanical receptive fields in the skin far beyond the site of injury. Collision experiments demonstrated that action potentials evoked from neuroma and skin travelled by the same fibre, indicating functional cross-talk between neuromatose and putative intact fibres. Retigabine, the specific Kv7 channel opener, depressed spontaneous discharges by 70% in 15/18 units tested. In contrast, responses to mechanical stimulation of the skin were unaltered by retigabine. CONCLUSIONS: Partial damage to a peripheral nerve may increase the incidence of spontaneous activity in C-fibres. Retigabine reduced spontaneous activity but not stimulus-evoked activity, suggesting an important role for ion channels in the control of spontaneous pain and demonstrating the utility of the model for the testing of compounds in clinically relevant variables. WHAT DOES THIS STUDY ADD?: Our in vitro experimental model of peripheral neuropathy allows for pharmacological characterization of spontaneously active fibres. Using this model, we show that retigabine inhibits aberrant spontaneous discharges without altering physiological responses in primary afferents.

Effects of centrally acting analgesics on spinal segmental reflexes and wind-up.

BACKGROUND: The spinal cord is a prime site of action for analgesia. Here we characterize the effects of established analgesics on segmental spinal reflexes. The aim of the study was to look for the pattern of action or signature of analgesic effects on these reflexes. METHODS: We used a spinal cord in vitro preparation of neonate mice to record ventral root responses to dorsal root stimulation. Pregabalin, clonidine, morphine and duloxetine and an experimental sigma-1 receptor antagonist (S1RA) were applied to the preparation in a cumulative concentration protocol. Drug effects on the wind-up produced by repetitive stimulation of C-fibres and on responses to single A- and C-fibre intensity stimuli were analysed. RESULTS: All compounds produced a concentration-dependent inhibition of total spikes elicited by repetitive stimulation. Concentrations producing ∼50% reduction in this parameter were (in µM) clonidine (0.01), morphine (0.1), pregabalin (1), duloxetine (10) and S1RA (30). At these concentrations clonidine, pregabalin and S1RA had significant effects on the wind-up index and little depressant effects on responses to single stimuli. Morphine and duloxetine did not depress wind-up index and showed large effects on responses to single stimuli. None of the compounds had strong effects on the amplitude of the non-nociceptive monosynaptic reflex. CONCLUSIONS: morphine and duloxetine had general depressant effects on spinal reflexes, whereas the effects of clonidine, pregabalin and S1RA appeared to be restricted to signals originated by strong repetitive activation of C-fibres. Results are discussed in the context of reported behavioural effects of the compounds studied.

Ultrastructural synaptic organization of axon terminals in the ventral part of the cat oral pontine reticular nucleus.

In an attempt to contribute to the current knowledge of the brainstem reticular formation synaptic organization, the ultrastructure and distribution of synaptic terminal profiles on neurons in the ventral part of the oral pontine reticular nucleus (vRPO), the rapid eye movement (REM) sleep-induction site, were studied quantitatively. Terminals with asymmetric contacts and rounded vesicles were classified according to vesicle density as type I or II (high or low density, respectively). The area, apposed perimeter length, and mitochondrial area of type I terminals, on average, were significantly smaller than those of type II terminals. Type III and IV terminals had symmetric contacts and oval and/or flattened vesicles; type III terminals formed synapses between them and on initial axons. Type V and VI terminals showed characteristics intermediate to those of asymmetric and symmetric synapses. Interestingly, some terminal features were related to both terminal area and postsynaptic dendritic diameter. The percentages of different synapses sampled on somata were as follows: asymmetric synapses (usually formed by type II terminals; mean +/- S.D.), 26.4% +/- 3%; symmetric synapses, 46.7% +/- 5.2%; and intermediate synapses, 26.9% +/- 6.1%. The percentages of different synapses sampled on dendrites were asymmetric synapses, 62.1% +/- 9%; symmetric synapses, 25.6% +/- 8.1%; and intermediate synapses, 12.3% +/- 1.7%. Comparison between large- and small-diameter dendrites revealed that the percentages of symmetric synapses and type II terminals decreased, whereas the percentages of type I terminals increased as postsynaptic dendritic diameters became smaller. Synaptic density was approximately four times lower on somata than on dendrites. The vRPO synaptic organization reflects some patterns that are similar to those found in other regions of the central nervous system as well as specific synaptic patterns that are probably related to its functions: the generation and maintenance of REM sleep and the control of eye movement or limb muscle tone.

A morphologic analysis of neurons and neuropil in the dorsal lateral geniculate nucleus of aged rats.

Light and electron microscopy were used to investigate the morphology of neuropil and neuronal cell bodies of the dorsal lateral geniculate nucleus (LGNd) of aged rats. Light microscopic examination reveals that, despite the optic tract showing signs of degeneration, the LGNd is scarcely affected. Thus, a slight but significant reduction in the diameters of both soma and nuclei is observed in aged neurons of the LGNd. Ultrastructural analysis demonstrates a few degenerating profiles of the neuropil. Neurons resembling relay cells exhibit typical features of aged neurons. Cells showing a very infolded nucleus, most of the ER cisternae connected with the nuclear envelope, abundant free polyribosomes and subsurface cisterns associated with mitochondria are similar to interneurons of adult rats. Therefore, aging and partial loss of visual input appear to induce small changes in the morphology of most of LGNd neurons.

An electron microscopic study of astroglia and oligodendroglia in the lateral geniculate nucleus of aged rats.

The ultrastructural features of glial cells in the lateral geniculate nucleus of aged rats have been studied. Abundant filaments as well as heterogeneous dense bodies are observed in the majority of astrocytes. They frequently surround both axons and nerve terminals showing signs of degeneration. In addition, some degenerating myelinated axons are seen in phase suggestive of engulfment by astrocyte processes. Oligodendrocytes display broad processes containing an organelle-rich cytoplasm and a continuity between their plasma membrane and the outer myelin lamellae which partially ensheath the adjacent axons. Multivesicular bodies and pleomorphic dense inclusions, composed of amorphous material as well as laminated structures, are also present in oligodendrocytes. The significance of these morphological features is discussed in relation to process of normal ageing.

The effect of age on the monoamines of the hypothalamus.

Measurement of dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC) homovanillic acid (HVA), 3-methoxytyramine (3-MT), noradrenaline (NA), 3-methoxy-4-hydroxyphenyl glycol (MHPG) and serotonin (5-HT) and its main metabolite, 5-hydroxyindol-3-acetic acid (5-HIAA) was assessed in hypothalamus and median eminence of aged rats. Age-related changes were not observed in the concentration of NA and its metabolites in median eminence. In contrast, there was a significant NA decrease in aged hypothalamus compared with 12 months (no differences were found compared with 3 months). No significant differences were found in DA concentration and its metabolites in hypothalamus but DA decreased significantly in aged median eminence compared with 12 months. The ratio 5-HIAA/5-HT, indicative of 5-HT turnover, appeared to increase in the hypothalamus and median eminence of the aged rat. Morphological dissimilarities between hypothalamus of young and aged rats were demonstrated using serotonin-immunocytochemistry. A degeneration of the serotoninergic system, denoted by the appearance of enlarged or swollen varicosities, was observed in the hypothalamus of the aged rat. These aberrant serotoninergic fibers may reflect the local degeneration of serotoninergic hypothalamic afferents during ageing. Such differential age-dependent alterations of the serotoninergic system might be responsible for at least some of the functional deficits in aged animals.

Substance P, calcitonin gene related peptide and PGE2 co-released from the mouse colon: a new model to study nociceptive and inflammatory responses in viscera, in vitro.

Visceral inflammation is thought to play an important role in the sensitization of low and high threshold mechanosensory and polymodal afferents and to recruit silent nociceptors. Yet, little is known about the potential role of the mediators involved in nociceptor sensitization to mechanical stimulation as compared to heat sensitization in the skin. In the present study we developed a new isolated preparation of the mouse colon which allowed to apply controlled mechanical distensions. Excised segments of colon from CD mice were immersed in synthetic interstitial fluid (SIF) exposing the serosal surface during 5 min to different types of noxious stimuli; the increase in neuropeptide and PGE(2) release were analyzed (by EIA technique). Capsaicin, heat and pH 5.2 were able to induce significant increases in calcitonin gene related peptide (CGRP) release (14.6-, 5.1-, and 2.3-fold over baseline), however, only capsaicin induced a significant increase in substance P (SP) levels (1.8-fold over baseline). When pH 3.4 was used, a massive liberation of both CGRP and SP was obtained (14- and 15-fold from baseline) which was Ca(2+)-independent and not recovering, suggesting unphysiological release. Mechanical distensions in the noxious range (45, 60 and 90 mmHg) evoked a long-linear graded release of CGRP (1.3-, 1.6- and 2.6-fold over baseline) and of PGE(2) (1.9- 3.8-, 12.3-fold over baseline). Only the 90 mmHg distension evoked a significant increase of SP (1.9-fold over baseline). We conclude that the mouse colon preparation is a suitable model to study inflammatory and nociceptive mechanisms in viscera. Furthermore, a potentially important and yet unexplored role of PGE(2) in noxious visceral distension has been revealed.

Role of central and peripheral tachykinin NK1 receptors in capsaicin-induced pain and hyperalgesia in mice.

Substance P and its receptor (NK1) are thought to play an important role in pain and hyperalgesia. Here we have further examined this role by comparing the behavioural responses to intradermal capsaicin of mutant mice with a disruption of the NK1 receptor (NK1 KO) and wild-type (WT) mice. We have also evaluated the contribution of peripheral NK1 receptors to capsaicin-evoked behaviour by selective blockade of peripheral NK1 receptors in WT mice using a non-brain penetrant NK1 receptor antagonist. Injection of 6 microg capsaicin into the heel evoked paw licking with the same latency in WT and KO mice, but a significantly longer duration in WT mice. A higher dose (30 microg) evoked a similar duration of licking in both groups. There were no differences in mechanical sensitivity tested with von Frey hairs between WT and KO mice before capsaicin. Both capsaicin doses resulted in pronounced increases in responses to von Frey hairs (hyperalgesia) and novel responses to cotton wisps (allodynia) applied to the digits of the injected paw in WT mice, but no significant changes from baseline in KO mice. Selective blockade of peripheral NK1 receptors in WT mice resulted in a complete inhibition of capsaicin-evoked plasma extravasation, but the mechanical hyperalgesia induced by 30 microg capsaicin intraplantar was still significantly greater than that seen in KO mice. We conclude that the response to intradermal capsaicin is still present but abbreviated in mice lacking NK1 receptors, such that secondary hyperalgesia is not observed even after a high dose. Further, the lack of secondary hyperalgesia in NK1 KO mice is largely due to the loss of central rather than peripheral NK1 receptors. The phenotype of the NK1 KO mice is consistent with a loss of function of mechanically-insensitive nociceptors, and thus we propose that substance P may be expressed by this group of primary sensory neurones and required for their function.

Changes in neurotransmitters in superior colliculus after neonatal enucleation: biochemical and immunocytochemical studies.

The turnover rate of dopamine and serotonin and the level of glutamate in superior colliculus are increased in adult, neonatally enucleated rats compared with normal control adult animals. Moreover, immunocytochemical data showed that the stratum zonale and the stratum griseum superficiale of the superior colliculus, specifically of bienucleated rats, display a dense network of serotonin-immunoreactive fibres, suggesting an increase in serotoninergic innervation. At the electron microscope level, serotonin-immunoreactive fibres and large postsynaptic serotonin-immunoreactive profiles exhibiting microtubules could be observed in the stratum zonale and the stratum griseum superficiale of the bienucleated rat. These results suggest that neonatal enucleation produces reorganization of serotoninergic and glutamatergic inputs. It is possible that serotonin may exert a profound influence upon collicular function.