Ordered nanopore arrays with large interpore distances via one-step anodization.

Preparation of pre-patterned alumina substrates using bottom-up techniques compatible with nanotechnology applications is still a challenge. We present a novel methodology to achieve superior order in 'anodic' alumina with large interpore distances by a convenient one-step anodization process. The use of transparent insulators renders such anodic layers applicable as templates for nanostructured photovoltaic or photoelectrochemical devices.

Direct visualization of the interfacial position of colloidal particles and their assemblies.

A method for direct visualization of the position of nanoscale colloidal particles at air-water interfaces is presented. After assembling hard (polystyrene, poly(methyl methacrylate), silica) or soft core-shell gold-hydrogel composite (Au@PNiPAAm) colloids at the air-water interface, butylcyanoacrylate is introduced to the interface via the gas phase. Upon contact with water, an anionic polymerization reaction of the monomer is initiated and a film of poly(butylcyanoacrylate) (PBCA) is generated, entrapping the colloids at their equilibrium position at the interface. We apply this method to investigate the formation of complex, binary assembly structures directly at the interface, to visualize soft, nanoscale hydrogel colloids in the swollen state, and to visualize and quantify the equilibrium position of individual micro- and nanoscale colloids at the air-water interface depending of the amount of charge present on the particle surface. We find that the degree of deprotonation of the carboxyl group shifts the air-water contact angle, which is further confirmed by colloidal probe atomic force microscopy. Remarkably, the contact angles determined for individual colloidal particles feature a significant distribution that greatly exceeds errors attributable to the size distribution of the colloids. This finding underlines the importance of accessing soft matter on an individual particle level.

Switching light with light--advanced functional colloidal monolayers.

Colloidal monolayers comprising of highly ordered two dimensional crystals are of high interest to generate surface patterns for a variety of different applications. Mostly, unfunctionalized polymer or silica colloids are assembled into monolayers. However, the incorporation of functional molecules into such colloids offers a convenient possibility of implementing additional properties to the two-dimensional crystal. Here, we present the formation of novel functional colloidal monolayers with photoswitchable fluorescence. The miniemulsion polymerization technique was used to incorporate an appropriate dye system of a perylene-based fluorophore and a bis-arylethene as a photochrome in polymeric colloids in defined ratios. Upon irradiation with UV or visible light the photochrome reversibly isomerizes from the ring-closed form, which is able to absorb light of the emission wavelength of the fluorescent dye and the ring-open form, which is not. The fluorescence emission of the dye can thus be reversibly switched on and off with light even when embedded in colloids. The colloids were self-assembled at the air-water interface to produce hexagonally ordered functional monolayers and more complex binary crystals. We investigate in detail the influence of the polymeric matrix on the switching properties of the fluorophore/photochrome system and find that the rate constants for the photoswitching, which all lie in the same range, are less influenced by the polymeric environment than expected. We demonstrate the reversible switching of the fluorescence emission in self-assembled colloidal monolayers. The arrangement of broadly distributed functional colloids into ordered monolayers with high addressability was obtained by the formation of binary colloidal monolayers.

Topical capsaicin for pain management: therapeutic potential and mechanisms of action of the new high-concentration capsaicin 8% patch.

Topical capsaicin formulations are used for pain management. Safety and modest efficacy of low-concentration capsaicin formulations, which require repeated daily self-administration, are supported by meta-analyses of numerous studies. A high-concentration capsaicin 8% patch (Qutenza™) was recently approved in the EU and USA. A single 60-min application in patients with neuropathic pain produced effective pain relief for up to 12 weeks. Advantages of the high-concentration capsaicin patch include longer duration of effect, patient compliance, and low risk for systemic effects or drug-drug interactions. The mechanism of action of topical capsaicin has been ascribed to depletion of substance P. However, experimental and clinical studies show that depletion of substance P from nociceptors is only a correlate of capsaicin treatment and has little, if any, causative role in pain relief. Rather, topical capsaicin acts in the skin to attenuate cutaneous hypersensitivity and reduce pain by a process best described as 'defunctionalization' of nociceptor fibres. Defunctionalization is due to a number of effects that include temporary loss of membrane potential, inability to transport neurotrophic factors leading to altered phenotype, and reversible retraction of epidermal and dermal nerve fibre terminals. Peripheral neuropathic hypersensitivity is mediated by diverse mechanisms, including altered expression of the capsaicin receptor TRPV1 or other key ion channels in affected or intact adjacent peripheral nociceptive nerve fibres, aberrant re-innervation, and collateral sprouting, all of which are defunctionalized by topical capsaicin. Evidence suggests that the utility of topical capsaicin may extend beyond painful peripheral neuropathies.

[Vaginal prolapse and rectal obstruction. Treated with a vaginal vault mesh colpo suspension and laparoscopic resection rectopexy]. / Vaginalprolaps und Defäkationsobstruktion. Kombination einer Netzkolposuspension mit der laparoskopischen Resektionsrektopexie.

BACKGROUND: Stool outlet obstruction with incomplete or complete rectal prolapse combined with vaginal vault prolapse is a severe form of pelvic floor insufficiency. Combining laparoscopic resection rectopexy with a vaginal vault mesh colpo suspension is a possible way of correcting this defect. METHOD: The safety of the combination was evaluated in 18 patients. RESULTS: The procedure was performed successfully with no complications in 16 of the 18 patients. One patient suffered intraoperative rectal injury and therefore received no polypropylene mesh, and one showed intraoperative bleeding requiring transfusion. No secondary surgery was required. Hospital stay lasted an average of 11.4 days (range 8-20) and the urinary catheters could be removed after an average of 4.3 days (range 2-10). No urinary disturbances were noted at the time of hospital release. Short-term mild fever appeared in 28% of cases (5/18). There were two urinary tract infections. No disturbance in healing and no anastomotic insufficiency were observed. The duration of postoperative antibiotic therapy averaged 3 days (range 0-8). CONCLUSION: The combination of laparoscopic resection rectopexy with a vaginal vault mesh colpo suspension might be safe. The close contact between the mesh and anastomosis might induce no increase in insufficiency. Long-term outcome must still be evaluated.

Investigation of the prostacyclin (IP) receptor antagonist RO1138452 on isolated blood vessel and platelet preparations.

BACKGROUND AND PURPOSE: The current study examined the utility of the recently described prostacyclin (prostanoid IP) receptor antagonist RO1138452 (2-(4-(4-isopropoxybenzyl)-phenylamino) imidazoline) as a tool for classifying prostanoid receptors. EXPERIMENTAL APPROACH: pA(2) values were determined on isolated smooth muscle and platelet preparations. KEY RESULTS: RO1138452 antagonized relaxation of human pulmonary artery, guinea-pig aorta and rabbit mesenteric artery induced by the selective IP agonist cicaprost. Schild plots had slopes close to unity, generating pA(2) values of 8.20, 8.39 and 8.12 respectively. Non-surmountable antagonism was sometimes found with the higher concentrations of RO1138452, attributable to the EP(3) contractile action of cicaprost. RO1138452 did not block relaxation of guinea-pig trachea induced by the EP(2)-selective agonist butaprost. In contrast, there was a modest inhibition of butaprost-induced relaxation of human pulmonary artery by RO1138452, implying activation of both EP(2) and IP receptors by butaprost. RO1138452 did not affect relaxation induced by PGE(2) (EP(4) agonist) and substance P (NK(1)/endothelium-dependent agonist) in rabbit mesenteric artery. In human and rat platelet-rich plasmas, RO1138452 antagonized cicaprost-induced inhibition of platelet aggregation in a surmountable manner; pA(2) values may have been affected by binding of RO1138452 to plasma protein. RO1138452 did not affect the inhibitory actions of PGD(2) (DP(1) agonist) and NECA (adenosine A(2A) agonist) in human platelets. CONCLUSIONS AND IMPLICATIONS: The data indicate that RO1138452 is a potent and selective IP receptor antagonist. RO1138452 represents an important addition to our armoury of prostanoid receptor antagonists and a potential clinical agent in situations where prostacyclin has a pathophysiological function.

[Open versus laparoscopic operation for perforated appendicitis--a comparative study]. / Konventionelle oder laparoskopische Operation der perforierten Appendizitis - Eine vergleichende Studie.

INTRODUCTION: Laparoscopic therapy of complicated appendicitis is still discussed controversially. This retrospective study compared the clinical results of open and laparoscopic appendectomy in a single centre. PATIENTS AND METHODS: Within a period of three years (1999 to 2001) 493 patients with suspected acute appendicitis were operated prospectively at a German district hospital (250 open appendectomies, 243 laparoscopic appendectomies). Twenty percent of the patients in every group had a complicated appendicitis (48 open, 44 laparoscopic appendectomies) and were analysed retrospectively considering demographic data, operative time, body mass index, preoperative inflammatory parameters (white blood cell count, C-reactive protein and body temperature) and postoperative complication rate. RESULTS: Both groups were comparable with regard to demographic data. One patient of the laparoscopic group needed an open operation (conversion rate 2.3 %). The body mass index of the laparoscopic group was significantly higher (26.3 vs. 24.1 kg/m(2)). Preoperative white blood cell count, C-reactive protein, body temperature as well as postoperative antibiotic therapy and analgesics requirement were comparable in both groups. There was no significant difference between the length of operative time (open 48 min, laparoscopic 53.5 min). The postoperative hospital stay was significantly shorter in the laparoscopic group (8 vs. 9 days, p = 0.032). Complication rate was significantly lower in the laparoscopic group (11.5 vs. 35 %, p = 0.014). CONCLUSION: Laparoscopic appendectomy is a safe procedure for the treatment of complicated appendicitis with a significantly decreased complication rate and shorter postoperative stay.

[Minimally invasive video-assisted thyroid resection (MIVAT)--a well accepted operative procedure]. / Die minimalinvasive video-assistierte Schilddrüsenresektion (MIVAT)--ein Verfahren mit hoher Patientenakzeptanz.

INTRODUCTION: We report on our results of MIVAT operations. METHOD: Nineteen Patients including 15 females and 4 males were operated with MIVAT, corresponding to 11 % of all patients undergoing an operation for benign goitre in our hospital. A single node of the thyroid gland within 30 mm and enlargement of a thyroid lobe up to 25 ml were the selection criteria. The median age was 34 years (range 16-61). We performed 2 thyroidectomies, 6 Hartley-Dunhill resections, 9 hemihyroidectomies and 2 enucleations. Eighteen patients (95 %) were followed-up postoperatively. The median follow-up period was 8 months (range 1 -21). RESULTS: The median operation time was 83 min (range 60-124). The median preoperative thyroid gland volume was 25 ml (range 10-54) measured by ultrasound. The resected thyroid tissue ranged from 12 to 51 g (median 29 g). A statistical significant difference (P = 0.04 paired t-test) between the preoperative and postoperative estimated serum-calcium levels (2.31 vs. 2.25 mM) was observed. But both parameters were inside the physiological range. Paresis of the recurrent laryngeal nerve was not observed. 89 % of the patients rated the postoperative cosmetic result as very good and all patients would prefer this operation procedure once again. CONCLUSION: The MIVAT procedure is a safe operation also in small groups with excellent cosmetic results postoperatively and a high acceptance by the patients.

Meckel's diverticulum: a disease associated with a colored clinical picture.

Meckel's diverticulum is reported in only 1 or 2% of the population. In most cases, it is free of clinical symptoms. The diagnostic modalities are effective in only 60-70% of all cases. The diagnostic laparoscopy is a safe and effective method for patients suffering from unclear abdominal pain with the option of a definitive surgical therapy. We describe the case of a 10-year-old girl with recurrent abdominal pain caused by a chronic subileus due to a Meckel's diverticulum in combination with a fibrous cord from the base of diverticulum to the mesenterial root. Both were resected in a laparoscopic technique.

A stable prostacyclin analog enhances ectopic activity in rat sensory neurons following neuropathic injury.

Prostanoids sensitize sensory afferents during inflammation. However, their role in neuropathic pain is still unclear. We analyzed the actions of prostanoids, non-selective (indomethacin) or selective (celecoxib and NS-398) cyclooxygenase-2 (COX or COX-2) inhibitors, on the ectopic activity of dorsal root ganglia (DRG) and dorsal horn (DH) neurons in a model of neuropathic injury. Extracellular recordings of DRG and DH neurons and cardiovascular measurements were performed on anesthetized, paralyzed and artificially ventilated adult male Sprague-Dawley rats whose sciatic nerve had been transected. PGD(2), PGE(2), PGF(2alpha), carbaprostacyclin (cPGI(2); a stable prostacyclin analog), and carbocyclic thromboxane (cTXA(2)) were administered at cumulative doses (0.0001-5 mg/kg, i.p.) at 5 or 10 min intervals. Only cPGI(2) significantly increased the DRG and DH activity in a dose-dependent manner, with ED(50) values of 0.05 (0.01-0.96) and 0.69 (0.11-1.04) mg/kg, respectively. The other prostanoids did not significantly increase activity, although they reduced heart rate for up to 5 min following administration. Time course experiments with single doses of cPGI(2) (1 mg/kg, i.v.) increased DH discharge rate 3-17 min after injection. Indomethacin (3 mg/kg, s.c.), but not celecoxib or NS-398 (both at 6 mg/kg, s.c.), reduced both DRG and DH activity. Our results indicate that cPGI(2) excites DRG and DH neurons of neuropathic rats, and may suggest a role for IP prostanoid receptors in pain episodes associated with nerve injury. The inhibitory effect of indomethacin, but not celecoxib or NS-398, on ectopic activity may suggest that a tonic generation of PGI(2) by COX-1 could contribute to neuropathic pain.

Characterization of prostanoid receptor-evoked responses in rat sensory neurones.

1. Prostanoid receptor-mediated sensitization, or excitation, of sensory nerve fibres contributes to the generation of hyperalgesia. To characterize the prostanoid receptors present on sensory neurones, biochemical assays were performed on primary cultures of adult rat dorsal root ganglia (DRG) and the F-11 (embryonic rat DRG x neuroblastoma hybrid) cell line. 2. In DRG cultures, the IP receptor agonists, cicaprost and carbaprostacyclin (cPGI2) stimulated cyclic AMP accumulation. Prostaglandin E2 (PGE2) also increased cyclic AMP levels, but to a lesser extent, while carbocyclic thromboxane A2 (cTxA2), PGD2 and PGF2alpha had negligible effects. The rank order of agonist potency was cicaprost>PGE2=BMY45778=cPGI2=PGI2. In the F-11 cells, the rank order of agonist potency for the stimulation of cyclic AMP accumulation was: cicaprost>iloprost=cPGI2=PGI2=BMY45778>PGE2=cTXA2++ +. In DRG cultures, cicaprost induced significantly more accumulation of inositol phosphates than PGE2. 3. To examine the effects of prostanoids on C-fibre activity, extracellular recordings of d.c. potentials from the rat isolated vagus nerve were made with the 'grease-gap' technique. PGI2 (0.1 nM-10 microM) produced the largest depolarizations of the nerve. The rank order of agonist potency was: PGI2=cPGI2=PGE1>cTXA2>PGE2=PGD2=TXB2>PGF2alpha. 4. Prior depolarization of nerves with either forskolin (10 microM) or phorbol dibutyrate (1 microM) alone significantly reduced the response to PGI2 (10 microM), while simultaneous application of both forskolin and phorbol dibutyrate attenuated PGI2 responses almost completely. 5. Putative EP1 and/or TP receptor-selective antagonists had no effect on the responses to PGI2, cPGI2 or PGE2 in the three preparations studied. 6. Collectively, these data are consistent with a positive coupling of IP receptors to both adenylyl cyclase and phospholipase C in sensory neurones. These findings suggest that IP receptors play a major role in the sensitization of rat sensory neurones.

The role of IP prostanoid receptors in inflammatory pain.

Prostanoid receptor-mediated sensitization of sensory nerve fibres is a key contributor to the generation of hyperalgesia. It is generally thought that prostaglandin (PG) E2 is the principal pro-inflammatory prostanoid. Consequently, prostanoid EP receptors on sensory neurones have been identified as potential therapeutic targets. However, IP prostanoid receptors are also present on sensory neurones, and recent data from transgenic mice lacking the IP receptor demonstrate its importance in the induction of oedema and pain behaviour. PGI2, the primary endogenous agonist for the IP receptor, is rapidly produced following tissue injury or inflammation; thus, it may be of equal, or greater, importance than PGE2 during episodes of inflammatory pain. In this review, Keith Bley, John Hunter, Richard Eglen and Jacqueline Smith compare the roles of EP and IP receptors in nociception and suggest that the IP receptor constitutes a novel target for anti-nociceptive agents.

QX-314 inhibits ectopic nerve activity associated with neuropathic pain.

In animal models of neuropathic pain, transection or constrictive injury to peripheral nerves produces ectopic discharges originating at both injury sites and related dorsal root ganglia (DRG). In addition, hyperexcitability is observed in associated dorsal horn (DH) neurons of the spinal cord. As ectopic discharges are inhibited by agents that block voltage-sensitive Na+ channels, it has been postulated that accumulation of Na+ channels in the membrane at nerve injury sites may contribute to the development of ectopic nerve activity (ENA). The goal of the present study was to compare the sensitivity of ENA to lidocaine and QX-314, a positively charged lidocaine derivative, which is frequently assumed to be membrane impermeant. Experiments were performed on adult male Sprague-Dawley rats in which the common sciatic nerve had been transected 4-10 days earlier. Extracellular microelectrode recordings were made from DRG and DH neurons, and neuronal activity was measured in fine bundles of microfilaments teased from sciatic nerves in anesthetized and paralyzed rats. Comparative effects on heart rate (HR) and mean blood pressure (MBP) were also studied. To confirm that externally applied QX-314 is able to inhibit high frequency activity in sensory nerves, QX-314 was superfused over isolated rat vagus nerves during stimulation of compound action potentials in C-fibers (C-spikes). As expected, intravenously administered lidocaine inhibited ENA at all three sites. Lidocaine ED50 values (expressed as mg/kg, with 95% confidence limits) were: 10.2 (7.8-13.3), 1.4 (0.8-2.4) and 0.9 (0.4-2.0) for neuromas, DRG and DH neurons, respectively. QX-314 also induced dose-dependent inhibition of ENA at neuromas and DRG, but produced only a small inhibition of DH neuron ENA. QX-314 had the following ED50 values (mg/kg) for neuromas, DRG and DH neurons, respectively: 2.3 (2.0-2.8), 6.9 (4.7-26.5) and 85.7. QX-314-mediated inhibition of DRG ENA had a slow onset and was long-lasting, relative to lidocaine. Lidocaine or QX-314 also significantly reduced HR and MBP in the same dose range as that which reduced ENA in DRG or neuromas. In isolated rat vagus nerve recordings, QX-314 induced marked use-dependent inhibition of C-spike amplitude, with IC50 values (microM) of 9000 (4600-18,000) and 350 (290-420) for low- (0.03 Hz) and high-frequency (30 Hz) C-spikes, respectively. These data support the hypothesis that Na+ channel accumulation contributes to the generation of ectopic discharges in neuromas and DRG, and suggests that intravenous QX-314 can acutely block Na+ channels at these sites.

Tetrodotoxin inhibits neuropathic ectopic activity in neuromas, dorsal root ganglia and dorsal horn neurons.

Neuropathic pain or persistent dysesthesias may be initiated by mechanical, chemical, or ischemic damage to peripheral sensory nerves. In animal models of neuropathic pain, transection or constrictive injury to peripheral nerves produces ectopic discharges originating at both injury sites and related dorsal root ganglia (DRG), and, consequently, hyperexcitability in associated dorsal horn (DH) neurons of the spinal cord. Since ectopic discharges are inhibited by agents that block voltage-sensitive Na+ channels, it has been postulated that accumulation of Na+ channels in the membrane at nerve injury sites may contribute to, or be responsible for, the development of ectopic neuronal activity (ENA). The present study therefore, tested the sensitivity of ENA to intravenously administered tetrodotoxin (TTX), an extremely potent and selective Na+ channel blocker. Comparative effects of TTX on cardiac parameters such as heart rate (HR) and diastolic blood pressure (DBP) were also studied. Experiments were performed on adult male Sprague-Dawley rats in which the common sciatic nerve had been transected 4-10 days earlier. Neuromal activity was measured in fine bundles of microfilaments teased from sciatic nerves, and extracellular microelectrode recordings were made from DRG and DH neurons. Cardiovascular parameters were recorded simultaneously. Intravenously administered TTX induced dose-dependent inhibition of ENA, with that originating from neuromas being the most sensitive; ED50 values (expressed as microg/kg, with 95% confidence limits) for neuromal, DRG and DH neuron activity were: 0.8 (0.6-1.2), 4.3 (2.2-8.4) and 36.2 (16.1-81.3), respectively. Inhibition of ENA in neuromas and DRG did not recover within 10 min after 100 or 300 microg/kg TTX. By comparison, the ED50 value for the initial decrease of HR was 17.9 (15.0-21.5) microg/kg, and partial recovery occurred within approximately 3 min. These data support the hypothesis that Na+ channel accumulation contributes to the generation of ectopic discharges in neuromas and DRG, and suggest that TTX-sensitive Na+ channels located at the nerve injury site and DRG play an important role in the genesis of neuropathic pain.

Characterization of the electrophysiological, biochemical and behavioral actions of epibatidine.

Epibatidine has been reported to be a potent, nonopioid analgesic. In this study we further characterized its receptor interactions and its analgesic properties. Radioligand binding assays demonstrated that epibatidine has high affinity for nicotinic receptors (Ki = 0.12 nM) but low affinity for opioid and other receptors (Ki > 3.0 microM). In vitro functional assays demonstrated that the compound is a potent agonist at both neuronal and neuromuscular nicotinic receptors. Epibatidine depolarized rat isolated vagus nerve with an EC50 of 33.1 nM and contracted guinea pig ileum with an EC50 of 6.1 nM. Epibatidine contracted frog rectus abdominis muscle with an EC50 of 18.2 nM. In vivo, epibatidine demonstrated short-lived analgesic actions. Epibatidine (10 and 30 micrograms/kg), at 5 but not 20 min after dosing, increased the threshold for vocalization evoked by foot shock. Epibatidine, at 5 and 20 but not 60 min after dosing, also increased the latency to a nociceptive response in a hot-plate assay. Both (+)- and (-)-enantiomers of epibatidine were active in these assays. The action of epibatidine in the hot-plate test was reversed by the nicotinic receptor antagonist mecamylamine but not by the opioid receptor antagonist naloxone. In contrast to morphine, epibatidine failed to increase locomotor activity. These findings demonstrate that epibatidine is a potent agonist at both neuronal and neuromuscular nicotinic receptors. These findings also demonstrate a short-lived, naloxone-insensitive, analgesic action for both the (+)- and (-)-enantiomers of epibatidine.(ABSTRACT TRUNCATED AT 250 WORDS)

Characterization of 5-hydroxytryptamine-induced depolarizations in rat isolated vagus nerve.

An additional component of the depolarization induced by 5-hydroxytryptamine (5-HT) in the rat isolated vagus nerve has recently been attributed to activation of 5-HT4 receptors. To confirm and extend this finding, extracellular recordings of D.C. potentials were made using the 'grease-gap' technique during continuous superfusion of the isolated nerve. Beginning at 1 nM, 5-HT induced small depolarizations that displayed a slow onset. At concentrations > or = 1 microM, large depolarizations with rapid onset were elicited. In the presence of the 5-HT3 receptor antagonists, granisetron or ondansetron, 5-HT responses were diminished and exhibited an increased latency to peak. These small, slow depolarization were not reduced by 5-HT1 or 5-HT2 receptor antagonists, but were potently inhibited by the 5-HT4 receptor antagonist GR 113808 (pA2 = 9.3), and mimicked by 5-methoxytryptamine (pEC50 = 5.3). 5-HT4-mediated responses were larger at 37 degrees C than at 31 degrees C, but also showed marked diminution with repeated 5-HT applications at concentrations greater than 1 microM. Conversely, 5-HT3 receptor responses were potentiated at lower temperatures (< or = 31 degrees C). Consistent with the reported positive coupling of 5-HT4 receptors to adenylyl cyclase, forskolin and 8-Br-cAMP produced slowly developing depolarizations which were qualitatively similar to 5-HT4 receptor activation. Pre-depolarization of nerves with 10 microM forskolin or 300 microM 8-Br-cAMP diminished the effect of 5-HT4 receptors. This study has confirmed the presence of 5-HT4 receptors on the vagus nerve of the rat and defined some conditions that optimize their pharmacological isolation.(ABSTRACT TRUNCATED AT 250 WORDS)

RS 23597-190: a potent and selective 5-HT4 receptor antagonist.

1. The pharmacological properties of RS 23597-190 (3-(piperdine-1-yl)-propyl-4-amino-5-chloro-2-methoxy benzoate hydrochloride) have been studied in vitro and in vivo. 2. RS 23597-190 competitively antagonized 5-HT4 receptor-mediated relaxations of rat, carbachol precontracted oesophageal muscularis mucosae, (pA2 = 7.8 +/- 0.1; Schild slope = 1.2 +/- 0.2). Affinity estimates (-log KB) at 5-HT4 receptors using either renzapride or SC-53116 as agonists yielded a -log KB value of 8.0 +/- 0.01. In contrast, RS 23597-190 failed to antagonize contractile responses to 5-HT of guinea-pig ileal 5-HT3 receptors, even at concentrations up to 10 microM. 3. Increases in short-circuit current, induced by 5-HT, were studied in guinea-pig ileal mucosal sheets. Concentration-response curves to 5-HT were biphasic, with the high potency phase to 5-HT inhibited by RS 23597-190 and mimicked by 5-methoxytryptamine. The -log KB value for RS 23597-190 at the high potency phase was 7.3 confirming that 5-HT4 receptors mediated the high potency phase. 4. In rat isolated vagus nerve, 5-HT elicited a slow, maintained depolarization at low concentrations and a rapid, transient depolarization at higher concentrations. The high potency, slow depolarizing phase to 5-HT was abolished selectively in the presence of 1 microM RS 23597-190 and the low potency phase was abolished selectively in the presence of 1 microM ondansetron. These data confirm that 5-HT4 and 5-HT3 receptors mediated slow and fast depolarization responses, respectively. 5. At 5-HT3 binding sites in membranes from NG 108-15 cells, labelled by [3H]-quipazine, RS 23597-190 exhibited an apparent affinity (- log Ki) of 5.7 +/- 0.1. At 5-HT3 receptors in membranes from rat cerebral cortex, labelled by [3H]-RS 42358-197, the apparent affinity (- log Ki) of RS 23597-190 was also 5.7 +/- 0.1. In both studies, Hill coefficients were not significantly different from unity. At 5-HT1A, 5-HT2,muscarinic M1, M2, M3, M4 and dopamine D1 and D2 receptors, RS 23597-190 exhibited low apparent affinities, with all - log Ki values less than 5.5.6. Intravenous infusion of RS 23597-190 in the conscious, restrained rat antagonized the von Bezold Jarisch reflex induced by 2-methyl 5-HT, with an ID50 of 300 microg kg-1 min-1, i.v. In the anaesthetized,bilaterally vagotomized micropig, RS 23597-190 (6 mg kg-1, i.v.) antagonized 5-HT-induced tachycardia with a half-life of 77 (63-99) min. Transient arrhythmic effects were noted after administration of the compound.7. In conclusion, RS 23597-190 acts as a high affinity, selective competitive antagonist at 5-HT4 receptors. Thus, the compound appears to be a useful tool for 5-HT4 receptor identification in vitro. In vivo, the compound is rapidly metabolized in pigs such that 5-HT4 blockade is not maintained. However,in the rat, when given by infusion, RS 23597-190 antagonizes 5-HT3 mediated responses, at doses consistent with a low affinity 5-HT3 receptor. These data suggest that, under appropriate experimental conditions, RS 23597-190 may also be used in vivo to characterize further 5-HT4 receptor function.

Inhibition of Ca2+ and K+ channels in sympathetic neurons by neuropeptides and other ganglionic transmitters.

Neuropeptides are known to modulate the excitability of frog sympathetic neurons by inhibiting the M-current and increasing the leak current, but their effects on Ca2+ channels are poorly understood. We compared effects of LHRH, substance P, epinephrine, and muscarine on Ca2+, K+, and leak currents in dissociated frog sympathetic neurons. At concentrations that inhibit M-current, LHRH and substance P strongly reduced N-type Ca2+ current and induced a leak conductance that may contribute to slow EPSPs. In contrast, muscarine produced little reduction of Ca2+ current, even in cells in which it strongly suppressed the M-current. We find that peptidergic inhibition of Ca2+ channels involves G proteins, but does not require protein kinases. In addition, it leads to reductions in Ca2(+)-activated K+ current and catecholamine release.