kn-93 and chelerythrine

Prazosin is an α1 adrenoceptor antagonist used in pharmacotherapy for the treatment of hypertension. Prazosin alters lipid metabolism in vivo, but the involved mechanism is not fully understood. In this study, we investigated the mechanism underlying the alteration of lipid metabolism. We show that the prazosin-stimulated release of hepatic triacylglyceride lipase (HTGL) from primary cultured rat hepatocytes involved Ca(2+)/calmodulin-dependent protein kinase II (CaMK-II) activation.. Primary cultured rat hepatocytes were incubated with prazosin and other agents. The hepatocytes were used in the CaMK-II and protein kinase A (PKA) activity assay. The supernatant was used in the HTGL activity assay and western blotting.. Prazosin-stimulated HTGL release was suppressed by the inositol triphosphate receptor inhibitor xestospongin C and by the calmodulin inhibitor trifluoperazine but not by the protein kinase C inhibitor chelerythrine chloride or a diacylglycerol kinase inhibitor (R59949). Furthermore, the calmodulin-dependent protein kinase II (CaMK-II) activity in prazosin-treated hepatocytes increased in a time- and dose-dependent manner. The cAMP-dependent PKA activity of prazosin-stimulated hepatocytes was suppressed by a phospholipase C (PLC) inhibitor (U-73122), trifluoperazine, and a CaMK-II inhibitor (KN-93).. These results suggested that prazosin-stimulated HTGL release from hepatocytes was caused by activation of PKA associated with stimulation of CaMK-II activity through a signal cascade from PLC.

Topics: Animals; Benzophenanthridines; Benzylamines; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cyclic AMP-Dependent Protein Kinases; Dose-Response Relationship, Drug; Estrenes; Hepatocytes; Lipoprotein Lipase; Macrocyclic Compounds; Male; Oxazoles; Piperidines; Prazosin; Primary Cell Culture; Pyrrolidinones; Quinazolinones; Rats; Sulfonamides; Time Factors; Trifluoperazine

Fibroblast growth factor 23 (FGF23), elevated in chronic renal failure, increases atrial arrhythmogenesis and dysregulates calcium homeostasis. Late sodium currents (INa-Late) critically induces ectopic activity of pulmoanry vein (the most important atrial fibrillation trigger). This study was to investigate whether FGF23 activates the INa-Late leading to calcium dysregulation and increases PV arrhythmogenesis. Patch clamp, western blot, and confocal microscopy were used to evaluate the electrical activities, calcium homeostasis, and mitochondrial reactive oxygen species (ROS) in PV cardiomyocytes with or without FGF23 (0.1 or 1 ng/mL) incubation for 4~6 h. Compared to the control, FGF23 (1 ng/mL, but not 0.1 ng/mL)-treated PV cardiomyocytes had a faster beating rate. FGF23 (1 ng/mL)-treated PV cardiomyocytes had larger INa-Late, calcium transients, and mitochondrial ROS than controls. However, ranolazine (an inhibitor of INa-Late) attenuated FGF23 (1 ng/mL)-increased beating rates, calcium transients and mitochondrial ROS. FGF23 (1 ng/mL)-treated PV cardiomyocytes exhibited larger phosphorylation of calcium/calmodulin-dependent protein kinase II (CaMKII). Chelerythrine chloride (an inhibitor of protein kinase C) decreased INa-Late in FGF23 (1 ng/mL)-treated PV cardiomyocytes. However, KN93 (a selective CaMKII blocker) decreased INa-Late in control and FGF23 (1 ng/mL)-treated PV cardiomyocytes to a similar extent. In conclusion, FGF23 increased PV arrhythmogenesis through sodium and calcium dysregulation by acting protein kinase C signaling.

Topics: Action Potentials; Animals; Benzophenanthridines; Benzylamines; Calcium; Fibroblast Growth Factor-23; Fibroblast Growth Factors; Homeostasis; Male; Myocytes, Cardiac; Patch-Clamp Techniques; Protein Kinase Inhibitors; Pulmonary Veins; Rabbits; Sodium; Sulfonamides

Although the effects of mechanical loading on chondrocyte metabolic activities have been extensively characterized, the sequence of events through which extracellular mechanical signals are transduced into chondrocytes and ultimately modulate cell activities is not well understood. Here, studies were performed to map out the sequential intracellular signalling pathways through which compression-induced signals modulate aggrecan mRNA levels in bovine articular chondrocytes. Bovine articular cartilage explants were subjected to a compressive stress of 0.1 MPa for 1 h in the presence or absence of inhibitors or antagonists of the phosphoinositol and Ca(2+)/calmodulin signalling pathways in order to determine the roles of second messengers and effector molecules of these pathways in transducing the compression-induced signals. In the absence of the inhibitors, aggrecan mRNA levels were stimulated by compression 2-4-fold relative to levels in tare-loaded (see below) explants. Treatment of the explants with graded levels of the protein kinase C inhibitor chelerythrine or bisindolylmaleimide I, followed by 1 h compressive loading, did not significantly alter the load-induced elevation of aggrecan mRNA levels. In contrast, thapsigargin, which depletes the Ins(1,4,5)P3-sensitive intracellular Ca(2+) stores, completely blocked the load response without significantly altering aggrecan mRNA levels in tare-loaded explants. Similarly, antagonists of the Ca(2+)/calmodulin signalling pathway dose-dependently or completely blocked the load-response. The results obtained demonstrate that transduction of the compression-induced aggrecan mRNA-regulating signals requires Ins(1,4,5)P3- and Ca(2+)/calmodulin-dependent signalling processes in bovine articular chondrocytes.

Topics: Aggrecans; Alkaloids; Animals; Benzophenanthridines; Benzylamines; Calcineurin; Calcium-Calmodulin-Dependent Protein Kinases; Calmodulin; Cartilage; Cattle; Chondrocytes; Cyclic AMP; Cyclosporine; Dose-Response Relationship, Drug; Egtazic Acid; Enzyme Inhibitors; Extracellular Matrix Proteins; Indoles; Inositol 1,4,5-Trisphosphate; Lectins, C-Type; Maleimides; Models, Biological; Phenanthridines; Protein Kinase C; Proteoglycans; RNA, Messenger; Signal Transduction; Sulfonamides; Thapsigargin; Time Factors

Intracellular recordings were made from pyramidal neurons in layers V and VI of the rat medial prefrontal cortex in slice preparations to investigate the effect of the serotonin 5-HT2A,2C receptor agonist (-)-1-2,5-dimethoxy-4-bromophenol-2-aminopropane (DOB) and 5-hydroxytryptamine (5-HT) on N-methyl-D-aspartate (NMDA)-induced responses. Bath application of either DOB or 5-HT [in the presence of antagonists to 5-HT1A, 5-HT3 and gamma-aminobutytric acid (GABA) receptors] produced a concentration-dependent biphasic modulation of the NMDA responses. They facilitated and inhibited NMDA responses at low (

Topics: Alkaloids; Animals; Benzophenanthridines; Benzylamines; Electrophysiology; Enzyme Inhibitors; Excitatory Amino Acid Agonists; In Vitro Techniques; Male; N-Methylaspartate; Phenanthridines; Phenols; Prefrontal Cortex; Presynaptic Terminals; Propane; Pyramidal Cells; Rats; Rats, Sprague-Dawley; Serotonin; Serotonin Antagonists; Serotonin Receptor Agonists; Sulfonamides; Synapses