Negatively charged amino acids near and in transient receptor potential (TRP) domain of TRPM4 channel are one determinant of its Ca2+ sensitivity.

Transient receptor potential (TRP) channel melastatin subfamily member 4 (TRPM4) is a broadly expressed nonselective monovalent cation channel. TRPM4 is activated by membrane depolarization and intracellular Ca(2+), which is essential for the activation. The Ca(2+) sensitivity is known to be regulated by calmodulin and membrane phosphoinositides, such as phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2). Although these regulators must play important roles in controlling TRPM4 activity, mutation analyses of the calmodulin-binding sites have suggested that Ca(2+) binds to TRPM4 directly. However, the intrinsic binding sites in TRPM4 remain to be elucidated. Here, by using patch clamp and molecular biological techniques, we show that there are at least two functionally different divalent cation-binding sites, and the negatively charged amino acids near and in the TRP domain in the C-terminal tail of TRPM4 (Asp-1049 and Glu-1062 of rat TRPM4) are required for maintaining the normal Ca(2+) sensitivity of one of the binding sites. Applications of Co(2+), Mn(2+), or Ni(2+) to the cytosolic side potentiated TRPM4 currents, increased the Ca(2+) sensitivity, but were unable to evoke TRPM4 currents without Ca(2+). Mutations of the acidic amino acids near and in the TRP domain, which are conserved in TRPM2, TRPM5, and TRPM8, deteriorated the Ca(2+) sensitivity in the presence of Co(2+) or PI(4,5)P2 but hardly affected the sensitivity to Co(2+) and PI(4,5)P2. These results suggest a novel role of the TRP domain in TRPM4 as a site responsible for maintaining the normal Ca(2+) sensitivity. These findings provide more insights into the molecular mechanisms of the regulation of TRPM4 by Ca(2+).

Different mechanisms of extracellular adenosine accumulation by reduction of the external Ca(2+) concentration and inhibition of adenosine metabolism in spinal astrocytes.

Extracellular adenosine is a neuromodulator in the central nervous system. Astrocytes mainly participate in adenosine production, and extracellular adenosine accumulates under physiological and pathophysiological conditions. Inhibition of intracellular adenosine metabolism and reduction of the external Ca(2+) concentration ([Ca(2+)]e) participate in adenosine accumulation, but the precise mechanisms remain unclear. This study investigated the mechanisms underlying extracellular adenosine accumulation in cultured rat spinal astrocytes. The combination of adenosine kinase and deaminase (ADK/ADA) inhibition and a reduced [Ca(2+)]e increased the extracellular adenosine level. ADK/ADA inhibitors increased the level of extracellular adenosine but not of adenine nucleotides, which was suppressed by inhibition of equilibrative nucleoside transporter (ENT) 2. Unlike ADK/ADA inhibition, a reduced [Ca(2+)]e increased the extracellular level not only of adenosine but also of ATP. This adenosine increase was enhanced by ENT2 inhibition, and suppressed by sodium polyoxotungstate (ecto-nucleoside triphosphate diphosphohydrolase inhibitor). Gap junction inhibitors suppressed the increases in adenosine and adenine nucleotide levels by reduction of [Ca(2+)]e. These results indicate that extracellular adenosine accumulation by ADK/ADA inhibition is due to the adenosine release via ENT2, while that by reduction of [Ca(2+)]e is due to breakdown of ATP released via gap junction hemichannels, after which ENT2 incorporates adenosine into the cells.

Contribution of cysteine aminotransferase and mercaptopyruvate sulfurtransferase to hydrogen sulfide production in peripheral neurons.

Hydrogen sulfide (H2 S) is a gaseous neuromodulator produced from L-cysteine. H2 S is generated by three distinct enzymatic pathways mediated by cystathionine γ-lyase (CSE), cystathionine ß-synthase (CBS), and mercaptopyruvate sulfurtransferase (MPST) coupled with cysteine aminotransferase (CAT). This study investigated the relative contributions of these three pathways to H2 S production in PC12 cells (rat pheochromocytoma-derived cells) and the rat dorsal root ganglion. CBS, CAT, and MPST, but not CSE, were expressed in the cells and tissues, and appreciable amounts of H2 S were produced from L-cysteine in the presence of α-ketoglutarate, together with dithiothreitol. The production of H2 S was inhibited by a CAT inhibitor (aminooxyacetic acid), competitive CAT substrates (L-aspartate and oxaloacetate), and RNA interference (RNAi) against MPST. Immunocytochemistry revealed a mitochondrial localization of MPST in PC12 cells and dorsal root ganglion neurons, and the amount of H2 S produced by CAT/MPST at pH 8.0, a physiological mitochondrial matrix pH, was comparable to that produced by CSE and CBS in the liver and the brain, respectively. Furthermore, H2 S production was markedly increased by alkalization. These results indicate that CAT and MPST are primarily responsible for H2 S production in peripheral neurons, and that the regulation of mitochondrial metabolism may influence neuronal H2 S generation. In the peripheral nervous system, hydrogen sulfide (H2 S) has been implicated in neurogenic pain or hyperalgesia. This study provides evidence that H2 S is synthesized in peripheral neurons through two mitochondrial enzymes, cysteine aminotransferase (CAT) and mercaptopyruvate sulfurtransferase (MPST). We propose that mitochondrial metabolism plays key roles in the physiology and pathophysiology of the peripheral nervous system via regulation of neuronal H2 S production.

Elucidation of the mechanism of reaction between S2O8(2-), Selenite and Mn2+ in aqueous solution and limestone-gypsum FGD liquor.

The mechanism of reaction between peroxodisulfate ion (S2O8(2-)), selenite (Se(IV)O3(2-)) and Mn(2+) as an inhibitor of selenite oxidation was studied using aqueous solutions composed of commercial reagents, as well as limestone-gypsum flue gas desulfurization (FGD) liquors sampled from coal fired power plants. The oxidation of selenite to selenate (Se(VI)O4(2-)) is promoted by the sulfate ion radical (SO4(-)) which results from decomposition of S2O8(2-). In the presence of Mn(2+), selenite oxidation was prevented due to the difference in rates of reaction with SO4(-). The ratio of the oxidation rate constants of selenite and Mn(2+) with SO4(-) was determined over a temperature range of 40-60 °C, and was found to be little influenced by the various coexisting components in FGD liquors.

Chronic repetitive transcranial magnetic stimulation failed to change dopamine synthesis rate: preliminary L-[ß-11C]DOPA positron emission tomography study in patients with depression.

We have examined the effects of repetitive transcranial magnetic stimulation (rTMS) on central dopaminergic function in patients with depression using positron emission tomography with L-[ß-11C]DOPA, a ligand to assess the rate of endogenous dopamine synthesis. Eight patients were treated with 10-daily sessions of rTMS over the left dorsolateral prefrontal cortex. Positron emission tomography scanning was performed in each patient twice, before the first session and 1 day after the last session. Although four out of eight patients responded to rTMS, there were no changes in the striatal dopamine synthesis rate (k) following rTMS. These results suggest that chronic rTMS had a limited effect on the dopaminergic system.

Involvement of transient receptor potential vanilloid subtype 1 in analgesic action of methylsalicylate.

Methylsalicylate (MS) is a naturally occurring compound that is used as a major active ingredient of balms and liniments supplied as topical analgesics. Despite the common use of MS as a pain reliever, the underlying molecular mechanism is not fully understood. Here we characterize the action of MS on transient receptor potential V1 (TRPV1). In human embryonic kidney 293 cells expressing human TRPV1 (hTRPV1), MS evoked increases of [Ca(2+)](i), which declined regardless of its continuous presence, indicative of marked desensitization. TRPV1 antagonists dose-dependently suppressed the MS-induced [Ca(2+)](i) increase. MS simultaneously elicited an inward current and increase of [Ca(2+)](i) in the voltage-clamped cells, suggesting that MS promoted Ca(2+) influx through the activation of TRPV1 channels. MS reversibly inhibited hTRPV1 activation by polymodal stimuli such as capsaicin, protons, heat, anandamide, and 2-aminoethoxydiphenyl borate. Because both the stimulatory and inhibitory actions of MS were exhibited in capsaicin- and allicin-insensitive mutant channels, MS-induced hTRPV1 activation was mediated by distinct channel regions from capsaicin and allicin. In cultured rat sensory neurons, MS elicited a [Ca(2+)](i) increase in cells responding to capsaicin. MS significantly suppressed nocifensive behavior induced by intraplantar capsaicin in rats. The present data indicate that MS has both stimulatory and inhibitory actions on TRPV1 channels and suggest that the latter action may partly underlie the analgesic effects of MS independent of inhibition of cyclooxygenases in vivo.

Lipopolysaccharides enhance the action of bradykinin in enteric neurons via secretion of interleukin-1beta from enteric glial cells.

Functional changes of the enteric nervous system have been observed under inflammatory states of inflammatory bowel disease increasing the endotoxin level. The aim of the present study was to determine the effect of lipopolysaccharides (LPS) on myenteric neuron-glia interaction in vitro. We examined the increase of the intracellular Ca(2+) concentration ([Ca(2+)](i)) and the release of interleukin-1beta (IL-1beta) or prostaglandin E(2) (PGE(2)) and COX-2 expression in myenteric plexus cells from the rat intestine induced by LPS. LPS potentiated BK-induced [Ca(2+)](i) increases in both myenteric neurons and enteric glial cells, which were suppressed by a B1R antagonist. Only in enteric glial cells, a B1R agonist increased [Ca(2+)](i). The effects of LPS were blocked by pretreatment with an interleukin-1 receptor antagonist or by reducing the density of enteric glial cells in culture. LPS prompted the release of IL-1beta from enteric glial cells. The augmenting effects of IL-1beta on the BK-induced neural [Ca(2+)](i) increase and PGE(2) release from enteric glial cells were abolished by a phospholipase A(2) (PLA(2)) inhibitor and a COX inhibitor, and partly suppressed by a COX-2 inhibitor. IL-1beta up-regulated the COX-2 expression in enteric glial cells. LPS promotes IL-1beta secretion from enteric glial cells, resulting in augmentation of the neural response to BK through PGE(2) release via glial PLA(2) and COX-2. The alteration of the regulatory effect of glial cells may be the cause of the changes in neural function in the enteric nervous system in inflammatory bowel disease.

Amelioration of anemia in the ICGN mouse, a renal anemia model, with a subcutaneous bolus injection of erythropoietin adsorbed to hydroxyapatite matrix.

The recombinant human erythropoietin (rhEPO) is used for the treatment of patients with renal anemia. However, rhEPO should be administered subcutaneously or intravenously three times a week. The repetitive injections of rhEPO result in burdens to patients. To resolve this problem, we investigated the sustaining release methods using an rhEPO-hydroxyapatite (HAp) made by spray-drying technique as the drug delivery system. Two types of rhEPO-HAp formulations were prepared; zinc (Zn) formulation and Zn and poly-L-lactic acid (PLA) formulation. These formulations were examined in genetically anemic model, ICGN (ICR-derived glomerulonephritis) mice. According to in vivo release test of rhEPO from HAp in ICGN mice, elevated plasma concentration of rhEPO could be maintained for more than 7 days. These mice showed the amelioration of anemia for more than 3 weeks post-administration without causing any side effect. In conclusion, Zn or Zn/PLA formulation of HAp was considered to be one of the useful carriers of rhEPO for long-term improvement of anemia.

Sustained efficacy of erythropoietin with a hydroxyapatite carrier administered in mice.

For chronic kidney disease patients with renal anemia, recombinant human erythropoietin (rHuEPO) is a very effective drug; however, the treatment regime is troublesome, requiring multiple administrations each week. In the present study, we examined the efficiency of hydroxyapatite (HAp) as a drug delivery carrier for the sustained release of erythropoietin (EPO) to reduce the frequency of administration. Spray-dried HAp microparticles, formed from zinc-containing HAp (Zn-HAp) and Zn-HAp calcined at 400 degrees C, were used as carriers of EPO, and five Zn-HAp formulation samples incorporating EPO were prepared; no formulation, poly-L-lactic acid (PLA) formulation, zinc (Zn) formulation, Zn/PLA formulation, and calcined/Zn/PLA formulation. ICR mice were administered these samples or commercial rHuEPO (Epogin) as a control from dorsal neck subcutaneous, and hematological and histopathological analyses, including enzyme-linked immunosorbent assay for plasma EPO concentration, were performed. An increase in the blood EPO level was detected on days 3 and 8 post-administration. Peak hematopoiesis was delayed and higher hematological values were obtained on day 14 post-administration with no serious adverse reactions compared with the control. The Zn/PLA formulation sample was found to be most effective in reducing the initial peak while sustaining the delayed release of EPO. In conclusion, the Zn-HAp formulation samples were considered to be useful carriers for the sustained release of EPO, and the Zn/PLA formulation appears to be the most effective of five Zn-HAp formulation samples in sustaining EPO release.

[Case of optic neuritis associated with cerebral autosomal recessive arteriopathy with subcortical infarcts and leukoencephalopathy (CARASIL)].

PURPOSE: To report a case of optic neuritis associated with cerebral autosomal recessive arteriopathy with subcortical infarcts and leukoencephalopathy (CARASIL). CASE: A 38-year-old woman presented with visual field defects, ocular pain and relative afferent papillary defect (RAPD). Fluorescein angiography demonstrated peripapillary hyperfluorescence of the optic nerve. Optic neuritis was diagnosed soon after. Routine laboratory data were unremarkable. Brain MRI showed a remarkable enhanced region of white matter and abnormal spots in the capsula externa. Multiple sclerosis was outruled by neurological findings and therefore CARASIL was diagnosed according to the criteria. The patient's vision and inflammation of the optic nerve head gradually improved with systemic steroid therapy, but arterial sheathing was observed during the follow-up period. CONCLUSIONS: Clinical manifestations such as optic neuritis and retinal vascular change might be caused by the pathological changes of CARASIL. This is the first report of ophthalmic findings of clinicopathological events of CARASIL.

Non-destructive evaluation of tomato based on optical scattering.

Equipment for the non-destructive evaluations of a tomato has been developed based on the scattering spectra, the angular distributions of the scattering intensities, and the scattering images from the specimens. The sugar concentrations of tomato can be estimated optically based on the scattering intensities using a halogen lamp, a white light-emitting diode (white LED), a yellow LED, a red LED, and near infrared LEDs (λ = 850 nm and 940 nm) as the incident lights. The scattering intensity is found to vary linearly with sugar concentrations of tomato with any wavelengths at which the scattering light can be observed. The gradient of calibration lines of the sugar concentrations against the scattering intensities varies with scattering angles, θ. Evaluations at around θ = 160° are found to be suitable because of the large gradient and good linearity with the correlation coefficient, R, close to 1.

Real-World Effectiveness of Ramelteon and Suvorexant for Delirium Prevention in 948 Patients With Delirium Risk Factors.

OBJECTIVE: The aim of this study was to examine the effectiveness of ramelteon and suvorexant for delirium prevention in real-world practice. It explored whether ramelteon and/or suvorexant would affect delirium prevention among both patients at risk for but without delirium (patients at risk) and those with delirium the night before a consultation. METHODS: This multicenter, prospective, observational study was conducted by trained psychiatrists at consultation-liaison psychiatric services from October 1, 2017, to October 7, 2018. Patients who were aged 65 years or older and hospitalized because of acute diseases or elective surgery, had risk factors for delirium, and had insomnia or delirium on the night before the consultation were prescribed ramelteon and/or suvorexant. The decision to take medication was left to the discretion of each patient. The primary outcome was incidence of delirium based on the Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition, during the first 7 days. RESULTS: Among 526 patients at risk, those taking ramelteon and/or suvorexant developed delirium significantly less frequently than those who did not, after control for the effects of risk factors on the estimate of an independent association between the effects of ramelteon and/or suvorexant and the outcome of developing delirium (15.7% vs 24.0%; odds ratio [OR] = 0.48;, 95% CI, 0.29-0.80; P = .005). Similar results were found among 422 patients with delirium (39.9% vs 66.3%; OR = 0.36; 95% CI, 0.22-0.59; P < .0001). CONCLUSIONS: Ramelteon and suvorexant appear to be effective for delirium prevention in real-world practice.

GABAA/Benzodiazepine receptor binding in patients with schizophrenia using [11C]Ro15-4513, a radioligand with relatively high affinity for alpha5 subunit.

Dysfunction of the GABA system is considered to play a role in the pathology of schizophrenia. Individual subunits of GABA(A)/Benzodiazepine (BZ) receptor complex have been revealed to have different functional properties. alpha5 subunit was reported to be related to learning and memory. Changes of alpha5 subunit in schizophrenia were reported in postmortem studies, but the results were inconsistent. In this study, we examined GABA(A)/BZ receptor using [(11)C]Ro15-4513, which has relatively high affinity for alpha5 subunit, and its relation to clinical symptoms in patients with schizophrenia. [(11)C]Ro15-4513 bindings of 11 patients with schizophrenia (6 drug-naïve and 5 drug-free) were compared with those of 12 age-matched healthy control subjects using positron emission tomography. Symptoms were assessed using the Positive and Negative Syndrome Scale. [(11)C]Ro15-4513 binding was quantified by binding potential (BP) obtained by the reference tissue model. [(11)C]Ro15-4513 binding in the prefrontal cortex and hippocampus was negatively correlated with negative symptom scores in patients with schizophrenia, although there was no significant difference in BP between patients and controls. GABA(A)/BZ receptor including alpha5 subunit in the prefrontal cortex and hippocampus might be involved in the pathophysiology of negative symptoms of schizophrenia.

Voltage-gated channel properties of epithelial cells in porcine vomeronasal organ.

Bipolar vomeronasal sensory neurons (VSNs) in the vomeronasal organ (VNO) are believed to detect pheromones in most mammals. The vomeronasal sensory epithelium (VSE) is composed of VSNs and supporting cells. There are morphological differences in VNOs between species. Many electrophysiological experiments have been performed on rodent VSEs but few on other mammals. We therefore investigated voltage-gated channel properties of cells in the porcine VSE using slice whole-cell voltage-clamp techniques. In immunohistochemical study of the porcine VSE, most PGP9.5-immunoreactive cells were found between the middle and basal region, and negative cells were distributed in the apical to middle region. Depolarizing pulses to epithelial cells from -90mV produced transient inward Na+ channel currents and sustained outward K+ channel currents with various amplitudes. The distribution of cells having high and low Na+ current densities was mostly consistent with the histological distribution of VSNs and supporting cells, respectively. The half-inactivation voltage of voltage-gated Na+ channels in supporting cells was 26mV more negative than that in VSNs. Voltage-gated K+ channel currents in both cell types were suppressed by tetraethylammonium to the same extent. VSNs possessed TTX-sensitive voltage-gated Na+ channels and Ni2+ -sensitive T-type Ca2+ channels. These results suggest that the histological distribution of porcine vomeronasal epithelial cells is more similar to the dog and goat than to rodents, and that the electrophysiological characteristics of porcine vomeronasal epithelial cells are similar to those of rodents. It is also suggested that porcine VSNs detecting pheromones generate action potentials through these channels.

Measurement and simulation of the effect of snowfall on free-space optical propagation.

We measured the time variation of a received laser signal level during snowfall over a distance of 72 m. The signal level dropped sharply for up to 10 ms when a snowflake crossed the laser beam. The probability distribution of the variation due to snowfall was calculated by assuming it to be the linear superposition of the light diffracted by snowflakes. The measured distributions could be reproduced by assuming reasonable snowflake size distributions. Furthermore, the probability distributions due to snowfall over a 1 km distance were calculated, and the expected bit errors during snowfall and the transmitted beam sizes were evaluated.

IL-1ß augments H2S-induced increase in intracellular Ca2+ through polysulfides generated from H2S/NO interaction.

H2S has excitatory and inhibitory effects on Ca2+ signals via transient receptor potential ankyrin 1 (TRPA1) and ATP-sensitive K+ channels, respectively. H2S converts intracellularly to polysulfides, which are more potent agonists for TRPA1 than H2S. Under inflammatory conditions, changes in the expression and activity of these H2S target channels and/or the conversion of H2S to polysulfides may modulate H2S effects. Effects of proinflammatory cytokines on H2S-induced Ca2+ signals and polysulfide production in RIN14B cells were examined using fluorescence imaging with fura-2 and SSP4, respectively. Na2S, a H2S donor, induced 1) the inhibition of spontaneous Ca2+ signals, 2) inhibition followed by [Ca2+]i increase, and 3) rapid [Ca2+]i increase without inhibition in 50% (23/46), 22% (10/46), and 17% (8/46) of cells tested, respectively. IL-1ß augmented H2S-induced [Ca2+]i increases, which were inhibited by TRPA1 and voltage-dependent L-type Ca2+ channel blockers. However, IL-1ß treatment did not affect [Ca2+]i increases evoked by a TRPA1 agonist or high concentration of KCl. Na2S increased intracellular polysulfide levels, which were enhanced by IL-1ß treatment. A NOS inhibitor suppressed the increased polysulfide production and [Ca2+]i increase in IL-1ß-treated cells. These results suggest that IL-1ß augments H2S-induced [Ca2+]i increases via the conversion of H2S to polysulfides through NO synthesis, but not via changes in the activity and expression of target channels. Polysulfides may play an important role in the effects of H2S during inflammation.

Novel agonistic action of mustard oil on recombinant and endogenous porcine transient receptor potential V1 (pTRPV1) channels.

Neurogenic components play a crucial role in inflammation and nociception. Mustard oil (MO) is a pungent plant extract from mustard seed, horseradish and wasabi, the main constituent of which is allylisothiocyanate. We have characterized the action of MO on transient receptor potential V1 (TRPV1), a key receptor of signal transduction pathways in the nociceptive system, using fura-2-based [Ca(2+)](i) imaging and the patch-clamp technique in a heterologous expression system and sensory neurons. In human embryonic kidney (HEK) 293 cells expressing porcine TRPV1 (pTRPV1), MO evoked increases of [Ca(2+)](i) in a concentration-dependent manner. A high concentration of MO elicited irreversible cell swelling. Capsazepine, ruthenium red and iodoresiniferatoxin dose-dependently suppressed the MO-induced [Ca(2+)](i) increase. MO elicited outward rectified currents in pTRPV1-expressing HEK 293 cells with a reversal potential similar to that of capsaicin. [Ca(2+)](i) responses to MO were completely abolished by the removal of external Ca(2+). MO simultaneously elicited an inward current and increase of [Ca(2+)](i) in the same cells, indicating that MO promoted Ca(2+) influx through TRPV1 channels. In cultured porcine dorsal root ganglion (DRG) neurons, MO elicited a [Ca(2+)](i) increase and inward current. Among DRG neurons responding to MO, 85% were also sensitive to capsaicin. The present data indicate that MO is a novel agonist of TRPV1 channels, and suggest that the action of MO in vivo may be partly mediated via TRPV1. These results provide an insight into the TRPV1-mediated effects of MO on inflammation and hyperalgesia.

T-type Ca2+ channels contribute to IBMX/forskolin- and K(+)-induced Ca(2+) transients in porcine olfactory receptor neurons.

T-type Ca(2+) channels are low-voltage-activated Ca(2+) channels that control Ca(2+) entry in excitable cells during small depolarization above resting potentials. Using Ca(2+) imaging with a laser scanning confocal microscope we investigated the involvement of T-type Ca(2+) channels in IBMX/forskolin- and sparingly elevated extracellular K(+)-induced Ca(2+) transients in freshly isolated porcine olfactory receptor neurons (ORNs). In the presence of mibefradil (10microM) or Ni(2+) (100microM), the selective T-type Ca(2+) channel inhibitors, IBMX/forskolin-induced Ca(2+) transients in the soma were either strongly (>60%) inhibited or abolished completely. However, the Ca(2+) transients in the knob were only partially (<60%) inhibited. Ca(2+) transients induced by 30mM K(+) were also partially ( approximately 60%) inhibited at both the knob and soma. Furthermore, ORNs responded to as little as a 2.5mM increase in the extracellular K(+) concentration (7.5mM K(+)), and such responses were completely inhibited by mibefradil or Ni(2+). These results reveal functional expression of T-type Ca(2+) channels in porcine ORNs, and suggest a role for these channels in the spread Ca(2+) transients from the knob to the soma during activation of the cAMP cascade following odorant binding to G-protein-coupled receptors on the cilia/knob of ORNs.

Monophasic action potential in anaesthetized guinea pigs as a biomarker for prediction of liability for drug-induced delayed ventricular repolarization.

INTRODUCTION: Drug-induced QT interval prolongation has been one of the critical issues for developing new chemical entities and pharmaceutical companies need to evaluate the risk early in the development stage. At such stage, guinea pigs are appropriate due to their small size requiring only small amounts of test drugs. The purpose of this study was to determine the utility of guinea pig monophasic action potential (MAP) using 12 reference drugs in order to clarify prediction of the QT interval prolonging risk. METHODS: Male guinea pigs were anaesthetized with pentobarbital (40 mg/kg, i.p.). Parameters analyzed were epicardial MAP duration (MAP(90)) at sinus rhythm (MAP(90(sinus))) and MAP(90) during atrial pacing (MAP(90(pacing))). Test drugs were administered to animals intravenously and cumulatively. RESULTS: Vehicle control did not affect the parameters tested. All 8 QT-prolonging drugs prolonged MAP(90(sinus)) and MAP(90(pacing)) dose-dependently, whereas all 4 non-QT-prolonging drugs showed no or very slight prolongations of these MAP(90) parameters. Rank order potency of MAP(90(pacing)) prolongations by the QT-prolonging drugs tended to correspond to clinical plasma concentrations associated with QT interval prolongations or Torsades de Pointes but showed less of a link with hERG inhibition activities. CONCLUSION: The present study demonstrates that the MAP model using anaesthetized guinea pigs could predict the liability of drugs for QT interval prolongation with high accuracy. QT assessment using the combination of the hERG assay with high sensitivity and the current in vivo assay would be desirable for early risk assessment within drug development.

Changes in central 5-HT(1A) receptor binding in mesial temporal epilepsy measured by positron emission tomography with [(11)C]WAY100635.

OBJECTIVE: The possible involvement of the brain 5-HT(1A) receptor in epilepsy has been indicated in animal seizure models. Recent in vivo neuroimaging studies demonstrated decreased 5-HT(1A) receptor binding in epilepsy. Using positron emission tomography (PET) with [(11)C]WAY100635, we investigated 5-HT(1A) receptor binding in patients with mesial temporal lobe epilepsy and aimed to clarify the involvement of the brain 5-HT(1A) receptor system in epilepsy. METHOD: PET measurements with [(11)C]WAY100635 were performed on 23 healthy volunteers and 13 patients who were diagnosed with mesial temporal lobe epilepsy based on clinical symptoms and electroencephalogram (EEG) findings. They had non-lesional mesial temporal lobe epilepsy with unilateral EEG foci and no hippocampal atrophy on magnetic resonance imaging. The binding potential (BP) of [(11)C]WAY100635 was calculated by the reference tissue model method. Data were analyzed for each region of interest (ROI) and on a voxel-by-voxel basis by statistical parametric mapping (SPM) system. RESULTS: ROI and voxel-based analyses consistently demonstrated that 5-HT(1A) receptor BP was significantly decreased in the temporal lobe, hippocampus and amygdala on the ipsilateral side of the EEG focus compared to controls. In addition, decreased 5-HT(1A) receptor BP was also observed on the contralateral side of the amygdala. CONCLUSION: 5-HT(1A) receptor binding in patients with mesial temporal lobe epilepsy decreased predominantly in the ipsilateral mesial temporal lobe structures but also in the contralateral side. The imaging of 5-HT(1A) receptor binding by PET detects functional changes of the limbic system in mesial temporal lobe epilepsy, proving to be a sensitive and useful method.