kt-5720 and Disease-Models--Animal

γ-Aminobutyric acid (GABA) is the major inhibitory neurotransmitter in adult central nervous system, and profound alterations of GABA receptor functions are linked to temporal lobe epilepsy (TLE). Here we describe the functional relationships between GABA receptors type B (GABA. Miniature inhibitory postsynaptic currents (mIPSCs) were recorded by patch-clamp techniques from human L5 pyramidal neurons in slices from temporal cortex tissue obtained from surgery.. We describe a constitutive functional crosstalk between GABA. Our results highlight the association of TLE with functional changes of GABAergic signaling that may be related to seizure propagation, and suggest that the selective activation of a definite subset of nonpresynaptic GABA

Topics: 8-Bromo Cyclic Adenosine Monophosphate; Adolescent; Adult; Animals; Baclofen; Carbazoles; Cyclic AMP-Dependent Protein Kinases; Disease Models, Animal; Drug Resistant Epilepsy; Enzyme Inhibitors; Epilepsy; Epilepsy, Temporal Lobe; Female; GABA-B Receptor Agonists; Humans; Inhibitory Postsynaptic Potentials; Male; Middle Aged; Muscarinic Agonists; Neocortex; Patch-Clamp Techniques; Pertussis Toxin; Pilocarpine; Protein Kinase C; Pyramidal Cells; Pyrroles; Rats; Receptors, GABA-A; Receptors, GABA-B; Temporal Lobe

Present study was performed to examine whether ADH was implicated in psychological stress asthma and to explore the underlying molecular mechanism.. We not only examined ADH levels in the cerebrospinal fluid (CSF) via radioimmunoassay, but also measured ADH receptor (ADHR) expression in airway-related vagal preganglionic neurons (AVPNs) through real-time PCR in all experimental mice. Western blotting was performed to evaluate the relationship between ADH and PKA/PKC in psychological stress asthma. Finally, the role of PKA/PKC in psychological stress asthma was analyzed.. Marked asthma exacerbations were noted owing to significantly elevated levels of ADH and ADHR after psychological stress induction as compared to OVA alone (asthma group). ADHR antagonists (SR-49095 or SR-121463A) dramatically lowered higher protein levels of PKAα and PKCα induced by psychological stress as compared to OVA alone, suggesting the correlation between ADH and PKA/PKC in psychological stress asthma. KT-5720 (PKA inhibitor) and Go-7874 (PKC inhibitor) further directly revealed the involvement of PKA/PKC in psychological stress asthma. Some notable changes were also noted after employing PKA and PKC inhibitors in psychological stress asthma, including reduced asthmatic inflammation (lower eosinophil peroxidase (EPO) activity, myeloperoxidase (MPO) activity, immunoglobulin E (IgE) level, and histamine release), substantial decrements in inflammatory cell counts (eosinophils and lymphocytes), and decreased cytokine secretion (IL-6, IL-10, and IFN-γ), indicating the involvement of PKA/PKC in asthma exacerbations induced by psychological stress.. Our results strongly suggested that ADH participated in psychological stress-induced asthma exacerbations via PKA/PKC signal pathway in AVPNs.

Topics: Animals; Antidiuretic Hormone Receptor Antagonists; Asthma; Carbazoles; Cyclic AMP-Dependent Protein Kinases; Cytokines; Disease Models, Animal; Eosinophils; Female; Mice; Mice, Inbred BALB C; Morpholines; Neurons; Ovalbumin; Protein Kinase C; Protein Kinase Inhibitors; Pyrroles; Receptors, Vasopressin; Signal Transduction; Spiro Compounds; Stress, Psychological; Vasopressins

β2-AR activation increases the risk of sudden cardiac death (SCD) in heart failure (HF) patients. Non-selective β-AR blockers have greater benefits on survival than selective β1-AR blockers in chronic HF patients, indicating that β2-AR activation contributes to SCD in HF. This study investigated the role of β2-AR activation on repolarization and ventricular arrhythmia (VA) in the experimental HF model. The guinea pig HF was induced by descending aortic banding. The effective refractoriness period (ERP), corrected QT (QTc) and the incidence of VA were examined using Langendorff and programmed electrical stimulation. Ikr and APD were recorded by the whole cell patch clamp. Selective β2-AR agonist salbutamol significantly increased the incidence of VA, prolonged QTc and shortened ERP. These effects could be prevented by the selective β2-AR antagonist, ICI118551. Salbutamol prolonged APD90 and reduced Ikr in guinea pig HF myocytes. The antagonists of cAMP (Rp-cAMP) and PKA (KT5720) attenuated Ikr inhibition and APD prolongation induced by salbutamol. However, the antagonists of Gi protein (PTX) and PDE III (amrinone) showed opposite effects. This study indicates that β2-AR activation increases the incidence of VA in the experimental HF model via activation of Gs/cAMP/PKA and/or inhibition of Gi/PDE pathways.

Topics: Adrenergic beta-Agonists; Adrenergic beta-Antagonists; Albuterol; Amrinone; Animals; Arrhythmias, Cardiac; Carbazoles; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Disease Models, Animal; Electric Stimulation; Electrocardiography; GTP-Binding Protein alpha Subunits, Gi-Go; GTP-Binding Protein alpha Subunits, Gs; Guinea Pigs; Heart Failure; In Vitro Techniques; Male; Myocytes, Cardiac; Patch-Clamp Techniques; Pyrroles; Receptors, Adrenergic, beta-2; Thionucleotides; Ventricular Function

Paclitaxel chemotherapy is limited by a long-lasting painful neuropathy that lacks an effective therapy. In this study, we tested the hypothesis that paclitaxel may release mast cell tryptase, which activates protease-activated receptor 2 (PAR2) and, subsequently, protein kinases A and C, resulting in mechanical and thermal (both heat and cold) hypersensitivity. Correlating with the development of neuropathy after repeated administration of paclitaxel, mast cell tryptase activity was found to be increased in the spinal cord, dorsal root ganglia, and peripheral tissues in mice. FSLLRY-amide, a selective PAR2 antagonist, blocked paclitaxel-induced neuropathic pain behaviors in a dose- and time-dependent manner. In addition, blocking downstream signaling pathways of PAR2, including phospholipase C (PLC), protein kinase A (PKA), and protein kinase Cε (PKC), effectively attenuated paclitaxel-induced mechanical, heat, or cold hypersensitivity. Furthermore, sensitized pain response was selectively inhibited by antagonists of transient receptor potential (TRP) V1, TRPV4, or TRPA1. These results revealed specific cellular signaling pathways leading to paclitaxel-induced neuropathy, including the activation of PAR2 and downstream enzymes PLC, PKCε, and PKA and resultant sensitization of TRPV1, TRPV4, and TRPA1. Targeting one or more of these signaling molecules may present new opportunities for the treatment of paclitaxel-induced neuropathy.

Topics: Analysis of Variance; Anilides; Animals; Ankyrins; Antineoplastic Agents, Phytogenic; Capsaicin; Carbazoles; Central Nervous System; Cinnamates; Cyclic AMP-Dependent Protein Kinases; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Interactions; Enzyme Inhibitors; Estrenes; Gene Expression Regulation; Hyperalgesia; Male; Mice; Mice, Inbred ICR; Neuralgia; Oligopeptides; Paclitaxel; Pain Measurement; Physical Stimulation; Protein Kinase C; Pyrroles; Pyrrolidinones; Receptor, PAR-2; Sulfonamides; Time Factors; TRPV Cation Channels; Tryptases; Type C Phospholipases

The laterocapsular division of the central nucleus of the amygdala (CeLC) has emerged as an important site of pain-related plasticity and pain modulation. Glutamate and neuropeptide receptors in the CeLC contribute to synaptic and behavioral changes in the arthritis pain model, but the intracellular signaling pathways remain to be determined. This study addressed the role of PKA, PKC, and ERK in the CeLC. Adult male Sprague-Dawley rats were used in all experiments. Whole-cell patch-clamp recordings of CeLC neurons were made in brain slices from normal rats and from rats with a kaolin/carrageenan-induced monoarthritis in the knee (6 h postinduction). Membrane-permeable inhibitors of PKA (KT5720, 1 microM; cAMPS-Rp, 10 microM) and ERK (U0126, 1 microM) activation inhibited synaptic plasticity in slices from arthritic rats but had no effect on normal transmission in control slices. A PKC inhibitor (GF109203x, 1 microM) and an inactive structural analogue of U0126 (U0124, 1 microM) had no effect. The NMDA receptor-mediated synaptic component was inhibited by KT5720 or U0126; their combined application had additive effects. U0126 did not inhibit synaptic facilitation by forskolin-induced PKA-activation. Administration of KT5720 (100 microM, concentration in microdialysis probe) or U0126 (100 microM) into the CeLC, but not striatum (placement control), inhibited audible and ultrasonic vocalizations and spinal reflexes of arthritic rats but had no effect in normal animals. GF109203x (100 microM) and U0124 (100 microM) did not affect pain behavior. The data suggest that in the amygdala PKA and ERK, but not PKC, contribute to pain-related synaptic facilitation and behavior by increasing NMDA receptor function through independent signaling pathways.

Topics: Amygdala; Animals; Arthritis; Behavior; Butadienes; Carbazoles; Colforsin; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Disease Models, Animal; Enzyme Activation; Extracellular Signal-Regulated MAP Kinases; Indoles; Male; Maleimides; Neuronal Plasticity; Neurons; Nitriles; Pain; Protein Kinase C; Protein Kinase Inhibitors; Pyrroles; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Synaptic Transmission; Thionucleotides

The highly dynamic nature of voltage-gated Na+ channel (VGSC) expression and its controlling mechanism(s) are not well understood. In this study, we investigated the possible involvement of nerve growth factor (NGF) in regulating VGSC activity in the strongly metastatic Mat-LyLu cell model of rat prostate cancer (PCa). NGF increased peak VGSC current density in a time- and dose-dependent manner. NGF also shifted voltage to peak and the half-activation voltage to more positive potentials, and produced currents with faster kinetics of activation; sensitivity to the VGSC blocker tetrodotoxin (TTX) was not affected. The NGF-induced increase in peak VGSC current density was suppressed by both the pan-trk antagonist K252a, and the protein kinase A (PKA) inhibitor KT5720. NGF did not affect the Nav1.7 mRNA level, but the total VGSC alpha-subunit protein level was upregulated. NGF potentiated the cells' migration in Transwell assays, and this was not affected by TTX. We concluded that NGF upregulated functional VGSC expression in Mat-LyLu cells, with PKA as a signaling intermediate, but enhancement of migration by NGF was independent of VGSC activity.

Topics: Animals; Carbazoles; Cell Line, Tumor; Cell Movement; Cyclic AMP-Dependent Protein Kinases; Disease Models, Animal; Dose-Response Relationship, Drug; Electrophysiology; Enzyme Inhibitors; Gene Expression Regulation, Neoplastic; Indole Alkaloids; Indoles; Male; NAV1.7 Voltage-Gated Sodium Channel; Nerve Growth Factor; Patch-Clamp Techniques; Prostatic Neoplasms; Pyrroles; Rats; Receptors, Nerve Growth Factor; RNA, Messenger; Sodium Channels; Time Factors; Up-Regulation

We recently demonstrated that activation of the pulmonary sensory neurons plays a critical role in prevention of endotoxin-induced shock by releasing calcitonin gene-related peptide (CGRP) in rats. CGRP increased the endothelial production of prostacyclin (PGI(2)) in the lungs, thereby preventing endotoxin-induced shock response by inhibiting tumor necrosis factor-alpha (TNF-alpha) production. Since antithrombin (AT) enhances sensory neuron activation, we hypothesized that AT might reduce endotoxin-induced hypotension by enhancing the activation of pulmonary sensory neurons in rats. We examined this possibility using a rat model of endotoxin shock. AT-induced effects including reduction of hypotension (n = 5) and inhibition of induction of iNOS (n = 4 or 5) and TNF- alpha (n = 5) in the lungs of endotoxin-treated animals were completely reversed by pretreatment with capsazepine (CPZ) (n = 4 or 5), a vanilloid receptor antagonist, or CGRP(8-37), a CGRP receptor antagonist (n = 4 or 5). AT enhanced endotoxin-induced increases in lung tissue levels of CGRP (n = 4), but this effect of AT was not seen in animals pretreated with CPZ (n = 4). CGRP produced therapeutic effects (n = 5) similar to those induced by AT, and such therapeutic effects were completely abrogated by pretreatment with indomethacin (n = 4). AT increased CGRP release from cultured dorsal root ganglion neurons only in the presence of anandamide (n = 5), and AT-induced increase in CGRP release was not observed in the presence KT5720, an inhibitor of protein kinase A (n = 5). AT markedly increased intracellular levels of cAMP in the presence of anandamide (n = 5). These results strongly suggested that AT might reduce endotoxin-induced hypotension in rats by enhancing activation of sensory neurons via activation of protein kinase A.

Topics: Animals; Antithrombins; Arachidonic Acids; Blood Pressure; Calcitonin Gene-Related Peptide; Calcitonin Gene-Related Peptide Receptor Antagonists; Capsaicin; Carbazoles; Cells, Cultured; Cyclic AMP; Cyclic AMP-Dependent Protein Kinase Type II; Cyclic AMP-Dependent Protein Kinases; Cyclooxygenase Inhibitors; Disease Models, Animal; Endocannabinoids; Endotoxins; Ganglia, Spinal; Gene Expression Regulation; Hypotension; Indoles; Indomethacin; Lung; Male; Neurons, Afferent; Nitrates; Nitric Oxide Synthase Type II; Nitrites; Peptide Fragments; Polyunsaturated Alkamides; Protein Kinase Inhibitors; Pyrroles; Rats; Rats, Wistar; Receptors, Calcitonin Gene-Related Peptide; RNA, Messenger; TRPV Cation Channels; Tumor Necrosis Factor-alpha