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L-type Calcium Channels (l-type + calcium_channel)
Terms modified by L-type Calcium Channels Selected AbstractsAngiotensin II regulates endothelial cell migration through calcium influx via T-type calcium channel in human umbilical vein endothelial cellsACTA PHYSIOLOGICA, Issue 4 2010A. Martini Abstract Aim:, The T-type calcium channel is expressed in vascular endothelial cells, but its role in endothelial cell function is yet to be elucidated. We analysed the endothelial functional role of T-type calcium channel-dependent calcium under angiotensin II (Ang II) stimulation. Methods:, Human umbilical vein endothelial cells were co-incubated with hormone at 10,7 m and either Efonidipine 10,5 m or Verapamil 10,5 m or Mibefradil 10,5 m or Wortmannin 10,6 m. The contribution of Ang II receptors was evaluated using PD123319 10,7 m and ZD 7155 10,7 m. The calcium ion concentration was observed using Fluo-3 acetossimetil ester. The cells were observed after 3, 6, 9 and 12 h. Results:, The microfluorescence method points out that Ang II induces intracellular calcium modulation in time by distinct mechanisms. AT2 receptor blockade is necessary to observe significant increase in [Ca2+]i levels. Pre-treatment with Mibefradil abolishes Ang II -induced cell migration. Conclusions:, Our data show that Ang II, via AT1 receptor, modulates calcium concentration involving T-type calcium channel and L-type calcium channel but only the calcium influx via T-type calcium channels regulates endothelial cell migration which is essential for angiogenesis. [source] Bepridil Reverses Atrial Electrical Remodeling and L-Type Calcium Channel Downregulation in a Canine Model of Persistent Atrial TachycardiaJOURNAL OF CARDIOVASCULAR ELECTROPHYSIOLOGY, Issue 7 2007KUNIHIRO NISHIDA M.D. Introduction: This study tested whether bepridil, a multichannel blocker, would reverse electrical remodeling induced by persistent atrial tachycardia. Methods and Results: Fourteen dogs were subjected to rapid atrial pacing at 400 bpm for 6 weeks after atrioventricular block was created to control the ventricular rate. During the study period, seven dogs were given placebo for 6 weeks (Control group), and seven were given placebo for 3 weeks, followed by 3 weeks of bepridil (10 mg/kg/day, Bepridil group). The atrial effective refractory period (ERP) and the inducibility and duration of atrial fibrillation (AF) were determined on a weekly basis. After 6 weeks, expression of L-type calcium channel ,1C messenger ribonucleic acid (mRNA) was quantified by real-time reverse transcription-polymerase chain reaction. In the Control group, ERP was shortened and the inducibility and duration of AF increased through the 6-week period. In the Bepridil group, the same changes occurred during the first 3 weeks, but were gradually reversed with bepridil. After 6 weeks, ERP was longer, AF inducibility was lower, and AF duration was shorter in Bepridil group than in the Control group. Expression of ,1C mRNA was decreased by 64% in the Control group (P < 0.05 vs sham), but in the Bepridil group, it was not different compared with the sham dogs. As a whole group of dogs, ERP was positively correlated with ,1C mRNA expression. Conclusion: Bepridil reverses the electrophysiological consequences of atrial remodeling to some extent and L-type calcium channel downregulation in a canine model of atrial tachycardia. [source] Highly efficient synthesis of [11C]S12968 and [11C]S12967, for the in vivo imaging of the cardiac calcium channels using PETJOURNAL OF LABELLED COMPOUNDS AND RADIOPHARMACEUTICALS, Issue 7 2001Frédéric Dolle Abstract The dihydrophyridines S12968 ((,)-S11568, absolute configuration S) and S12967 ((+)-S11568, absolute configuration R), 3-ethyl 5-methyl (,/+)-2-[(2-(2-aminoethoxy)ethoxy)methyl]-4-(2,3-dichlorophenyl)-6-methyl-1,4-dihydropyridine-3,5-dicarboxylate, have both an in vitro profile of high potency and of high selectivity for the low-voltage-dependent L-type calcium channel. In this paper, the radiosynthesis of both enantiomers, S12968 and S12967, with carbon-11, a positron-emitting isotope (half-life : 20.4 min) was investigated and oriented towards the preparation of multi milliCuries of radiotracer. Typically, 130,250 mCi (4.81,9.25 GBq) of [11C]S12968 and [11C]S12967 were obtained within 30 min of radiosynthesis (HPLC purification included) with specific radioactivities ranging from 500 to 1000 mCi/,mol (18.5,37.0 GBq/,mol) using no-carrier-added [11C]methyl triflate as the alkylating agent and the appropriate, enantiomerically pure carboxylic acid precursor at 100°C for 1 min. Based on preliminary PET experiments, only the levo enantiomer S12968 ((,)-[11C]-1) appears to be suitable for myocardial PET imaging as demonstrated in vivo in beagle dogs: with S12968, 85% of the uptake of [11C]S12968 could be inhibited in pretreatment experiments and up to 70% of [11C]S12968 could be displaced. Further investigations are currently underway in order to provide an absolute quantification of ventricular calcium channels with PET. Copyright © 2001 John Wiley & Sons, Ltd. [source] EXCITATION,CONTRACTION COUPLING FROM THE 1950s INTO THE NEW MILLENNIUMCLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY, Issue 9 2006AF Dulhunty SUMMARY 1Excitation,contraction coupling is broadly defined as the process linking the action potential to contraction in striated muscle or, more narrowly, as the process coupling surface membrane depolarization to Ca2+ release from the sarcoplasmic reticulum. 2We now know that excitation,contraction coupling depends on a macromolecular protein complex or ,calcium release unit'. The complex extends the extracellular space within the transverse tubule invaginations of the surface membrane, across the transverse tubule membrane into the cytoplasm and then across the sarcoplasmic reticulum membrane and into the lumen of the sarcoplasmic reticulum. 3The central element of the macromolecular complex is the ryanodine receptor calcium release channel in the sarcoplasmic reticulum membrane. The ryanodine receptor has recruited a surface membrane L-type calcium channel as a ,voltage sensor' to detect the action potential and the calcium-binding protein calsequestrin to detect in the environment within the sarcoplasmic reticulum. Consequently, the calcium release channel is able to respond to surface depolarization in a manner that depends on the Ca2+ load within the calcium store. 4The molecular components of the ,calcium release unit' are the same in skeletal and cardiac muscle. However, the mechanism of excitation,contraction coupling is different. The signal from the voltage sensor to ryanodine receptor is chemical in the heart, depending on an influx of external Ca2+ through the surface calcium channel. In contrast, conformational coupling links the voltage sensor and the ryanodine receptor in skeletal muscle. 5Our current understanding of this amazingly efficient molecular signal transduction machine has evolved over the past 50 years. None of the proteins had been identified in the 1950s; indeed, there was debate about whether the molecules involved were, in fact, protein. Nevertheless, a multitude of questions about the molecular interactions and structures of the proteins and their interaction sites remain to be answered and provide a challenge for the next 50 years. [source] BMP-2 and FGF-2 Synergistically Facilitate Adoption of a Cardiac Phenotype in Somatic Bone Marrow c-kit+/Sca-1+ Stem CellsCLINICAL AND TRANSLATIONAL SCIENCE, Issue 2 2008Brent R. DeGeorge, Jr. B.S. Abstract The aim of this study was to explore the effect of bone morphogenetic protein-2 (BMP-2) and fibroblast growth factor-2 (FGF-2), paracrine factors implicated in both cardiac embryogenesis and cardiac repair following myocardial infarction (MI),on murine bone marrow stem cell (mBMSC) differentiation in an ex vivo cardiac microenvironment. For this purpose, green fluorescent protein (GFP) expressing hematopoietic lineage negative (lin-) c-kit ligand (c-kit) and stem cell antigen-1 (Sca-1) positive (GFP-lin-/c-kit+/sca+) mBMSC were co-cultured with neonatal rat ventricular cardiomyocytes (NVCMs). GFP+ mBMSC significantly induced the expression of BMP-2 and FGF-2 in NVCMs, and approximately 4% GFP+ mBMSCs could be recovered from the co-culture at day 10. The addition of BMP-2 in concert with FGF-2 significantly enhanced the amount of integrated GFP+ mBMSCs by 5-fold (,20%), whereas the addition of anti-BMP-2 and/or anti-FGF-2 antibodies completely abolished this effect. An analysis of calcium cycling revealed robust calcium transients in GFP+ mBMSCs treated with BMP-2/FGF-2 compared to untreated co-cultures. BMP-2 and FGF-2 addition led to a significant induction of early (NK2 transcription factor related, locus 5; Nkx2.5, GATA binding protein 4; GATA-4) and late (myosin light chain kinase [MLC-2v], connexin 43 [Cx43]) cardiac marker mRNA expression in mBMSCs following co-culture. In addition, re-cultured fluorescence-activated cell sorting (FACS)-purified BMP-2/FGF-2-treated mBMSCs revealed robust calcium transients in response to electrical field stimulation which were inhibited by the L-type calcium channel (LTCC) inhibitor, nifedipine, and displayed caffeine-sensitive intracellular calcium stores. In summary, our results show that mBMSCs can adopt a functional cardiac phenotype through treatment with factors essential to embryonic cardiogenesis that are induced after cardiac ischemia. This study provides the first evidence that mBMSCs with long-term self-renewal potential possess the capability to serve as a functional cardiomyocyte precursor through the appropriate paracrine input and cross-talk within an appropriate cardiac microenvironment. [source] Influx of calcium through L-type calcium channels in early postnatal regulation of chloride transporters in the rat hippocampusDEVELOPMENTAL NEUROBIOLOGY, Issue 13 2009Jennifer G. Bray Abstract During the early postnatal period, GABAB receptor activation facilitates L-type calcium current in rat hippocampus. One developmental process that L-type current may regulate is the change in expression of the K+Cl, co-transporter (KCC2) and N+K+2Cl, co-transporter (NKCC1), which are involved in the maturation of the GABAergic system. The present study investigated the connection between L-type current, GABAB receptors, and expression of chloride transporters during development. The facilitation of L-type current by GABAB receptors is more prominent in the second week of development, with the highest percentage of cells exhibiting facilitation in cultures isolated from 7 day old rats (37.5%). The protein levels of KCC2 and NKCC1 were investigated to determine the developmental timecourse of expression as well as expression following treatment with an L-type channel antagonist and a GABAB receptor agonist. The time course of both chloride transporters in culture mimics that seen in hippocampal tissue isolated from various ages. KCC2 levels increased drastically in the first two postnatal weeks while NKCC1 remained relatively stable, suggesting that the ratio of the chloride transporters is important in mediating the developmental change in chloride reversal potential. Treatment of cultures with the L-type antagonist nimodipine did not affect protein levels of NKCC1, but significantly decreased the upregulation of KCC2 during the first postnatal week. In addition, calcium current facilitation occurs slightly before the large increase in KCC2 expression. These results suggest that the expression of KCC2 is regulated by calcium influx through L-type channels in the early postnatal period in hippocampal neurons. © 2009 Wiley Periodicals, Inc. Develop Neurobiol 2009 [source] A role for acetylcholine receptors in their own aggregation on muscle cellsDEVELOPMENTAL NEUROBIOLOGY, Issue 8 2007Rebecca B.R. Milholland Abstract Both neurotrophic factors and activity regulate synaptogenesis. At neuromuscular synapses, the neural factor agrin released from motor neuron terminals stimulates postsynaptic specialization by way of the muscle specific kinase MuSK. In addition, activity through acetylcholine receptors (AChRs) has been implicated in the stabilization of pre- and postsynaptic contacts on muscle at various stages of development. We show here that activation of AChRs with specific concentrations of nicotine is sufficient to induce AChR aggregation and that this induction requires the function of L-type calcium channels (L-CaChs). Furthermore, AChR function is required for agrin induced AChR aggregation in C2 muscle cells. The same concentrations of nicotine did not induce observable tyrosine phosphorylation on either MuSK or the AChR , subunit, suggesting significant differences between the mechanisms of agrin and activity induced aggregation. The AChR/L-CaCh pathway provides a mechanism by which neuromuscular signal transmission can act in concert with the agrin-MuSK signaling cascade to regulate NMJ formation. © 2007 Wiley Periodicals, Inc. Develop Neurobiol, 2007. [source] Effects of oestrogen replacement therapy on pattern reversal visual evoked potentialsEUROPEAN JOURNAL OF NEUROLOGY, Issue 2 2000H. Yilmaz As a result of a regression in the ovarian functions, oestrogen level in circulation during the menopause drops to 1/50 of its value in the normal reproductive cycle. Excitatory oestrogen increases the sensitivity of the central nervous system to catecholamines by changing the opening frequency of voltage-related L-type calcium channels and augmenting the effect of glutamate; in addition it inhibits the formation of gamma-amino butyric acid (GABA) by the inhibition of glutamate decarboxylase enzyme. It is argued that oestrogen increases transmission in the optic pathways and that oestrogen is responsible for the shorter latency values and higher amplitudes of visual evoked potentials in women. We recorded the monocular pattern reversal visual evoked potentials (PRVEP) of both eyes of 54 post-menopausal women before treatment and of 30 of them after replacement therapy with Tibolon, and of 24 women receiving placebo treatment. The explicit values of P100 latency of right and left eyes before treatment were 98.8 ± 3.5 and 99.0 ± 3.3 ms, respectively. The explicit values of P100 latency of right and left eyes after placebo treatment were 98.6 ± 3.7 and 98.8 ± 4.0, respectively. The explicit values of P100 latency of right and left eyes after replacement treatment were 94.6 ± 3.7 and 94.8 ± 4.0, respectively. We found a statistically significant decrease in the mean PRVEP latencies and a statistically significant increase in mean amplitudes after replacement treatment (P < 0.001) compared with those before treatment and those after placebo treatment. We attributed the changes in PRVEP values after replacement treatment to the action of Tibolon, which acted as a natural sex steroid and speeded the visual transmission time via the widespread receptors in the central nervous system. It is concluded that PRVEP is an objective electrophysiological assessment method in evaluating the efficiency of hormone replacement therapy in post-menopausal women. [source] L-type calcium channels are involved in fast endocytosis at the mouse neuromuscular junctionEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 6 2008Paula P. Perissinotti Abstract We used fluorescence microscopy of FM dyes-labeled synaptic vesicles and electrophysiological recordings to examine the functional characteristics of vesicle recycling and study how different types of voltage-dependent Ca2+ channels (VDCCs) regulate the coupling of exocytosis and endocytosis at mouse neuromuscular junction. Our results demonstrate the presence of at least two different pools of recycling vesicles: a high-probability release pool (i.e. a fast destaining vesicle pool), which is preferentially loaded during the first 5 s (250 action potentials) at 50 Hz; and a low-probability release pool (i.e. a slow destaining vesicle pool), which is loaded during prolonged stimulation and keeps on refilling after end of stimulation. Our results suggest that a fast recycling pool mediates neurotransmitter release when vesicle use is minimal (i.e. during brief high-frequency stimulation), while vesicle mobilization from a reserve pool is the prevailing mechanism when the level of synaptic activity increases. We observed that specific N- and L -type VDCC blockers had no effect on evoked transmitter release upon low-frequency stimulation (5 Hz). However, at high-frequency stimulation (50 Hz), L -type Ca2+ channel blocker increased FM2-10 destaining and at the same time diminished quantal release. Furthermore, when L -type channels were blocked, FM2-10 loading during stimulation was diminished, while the amount of endocytosis after stimulation was increased. Our experiments suggest that L -type VDCCs promote endocytosis of synaptic vesicles, directing the newly formed vesicles to a high-probability release pool where they compete against unused vesicles. [source] Distribution and regulation of L-type calcium channels in deep dorsal horn neurons after sciatic nerve injury in ratsEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 12 2005E. Dobremez Abstract Deep dorsal horn neurons are involved in the processing of nociceptive information in the spinal cord. Previous studies revealed a role of the intrinsic bioelectrical properties (plateau potentials) of deep dorsal horn neuron in neuronal hyperexcitability, indicating their function in pain sensitization. These properties were considered to rely on L -type calcium currents. Two different isotypes of L -type calcium channel alpha 1 subunit have been cloned (CaV1.2 and CaV1.3). Both are known to be expressed in the spinal cord. However, no data were available on their subcellular localization. Moreover, possible changes in CaV1.2 and CaV1.3 expression had never been investigated in nerve injury models. Our study provides evidence for a differential expression of CaV1.2 and CaV1.3 subunits in the somato-dendritic compartment of deep dorsal horn neurons. CaV1.2 immunoreactivity is restricted to the soma and proximal dendrites whereas CaV1.3 immunoreactivity is found in the whole somato-dendritic compartment, up to distal dendritic segments. Moreover, these specific immunoreactive patterns are also found in electrophysiologically identified deep dorsal horn neurons expressing plateau potentials. After nerve injury, namely total axotomy or partial nerve ligation, CaV1.2 and CaV1.3 expression undergo differential changes, showing up- and down-regulation, respectively, both at the protein and at the mRNA levels. Taken together, our data support the role of L-type calcium channels in the control of intrinsic biolectrical regenerative properties. Furthermore, CaV1.2 and CaV1.3 subunits may have distinct and specific roles in sensory processing in the dorsal horn of the spinal cord, the former being most likely involved in long-term changes after nerve injury. [source] Dendritic L-type calcium currents in mouse spinal motoneurons: implications for bistabilityEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 5 2000K. P. Carlin Abstract The intrinsic properties of mammalian spinal motoneurons provide them with the capability to produce high rates of sustained firing in response to transient inputs (bistability). Even though it has been suggested that a persistent dendritic calcium current is responsible for the depolarizing drive underlying this firing property, such a current has not been demonstrated in these cells. In this study, calcium currents are recorded from functionally mature mouse spinal motoneurons using somatic whole-cell patch-clamp techniques. Under these conditions a component of the current demonstrated kinetics consistent with a current originating at a site spatially segregated from the soma. In response to step commands this component was seen as a late-onset, low amplitude persistent current whilst in response to depolarizing,repolarizing ramp commands a low voltage clockwise current hysteresis was recorded. Simulations using a neuromorphic motoneuron model could reproduce these currents only if a noninactivating calcium conductance was placed in the dendritic compartments. Pharmacological studies demonstrated that both the late-onset and hysteretic currents demonstrated sensitivity to both dihydropyridines and the L-channel activator FPL-64176. Furthermore, the ,1D subunits of L-type calcium channels were immunohistochemically demonstrated on motoneuronal dendrites. It is concluded that there are dendritically located L-type channels in mammalian motoneurons capable of mediating a persistent depolarizing drive to the soma and which probably mediate the bistable behaviour of these cells. [source] Actions of Arachidonic Acid on Contractions and Associated Electrical Activity in Guinea-Pig Isolated Ventricular MyocytesEXPERIMENTAL PHYSIOLOGY, Issue 4 2001M. A. Mamas The actions of arachidonic acid (AA) were investigated in guinea-pig isolated ventricular myocytes. Exposure of myocytes to 10 ,M AA reduced the amplitude of contractions and calcium transients accompanying action potentials at a frequency of 1 Hz. AA (10 ,M) also reduced the amplitude of calcium currents recorded under voltage-clamp conditions. The suppression of contraction by AA was not prevented by either 10 ,M trihydroindomethicin (to inhibit cyclo-oxygenase) or 10 ,M ETYA (5,8,11,14-eicosatetraynoic acid, to inhibit AA metabolising enzymes), showing that the actions of AA appeared not to be mediated by these metabolites. The reduction of contraction by 10 ,M AA was also not prevented by the protein kinase C inhibitor, Ro31-8220 (1 ,M), showing that this pathway appeared not to be required for the observed effect. Direct effects of AA may be involved. A further action of 10 ,M AA was to suppress spontaneous electrical activity induced by either the ,-adrenergic agonist isoprenaline or the Na+ pump inhibitor, ouabain. This effect of AA on spontaneous activity might be associated with the observed reduction of calcium entry through L-type calcium channels, although additional effects of AA on calcium release from the sarcoplasmic reticulum might also be involved. [source] The triakontatetraneuropeptide TTN increases [Ca2+]i in rat astrocytes through activation of peripheral-type benzodiazepine receptorsGLIA, Issue 2 2001Pierrick Gandolfo Abstract Astrocytes synthesize a series of regulatory peptides called endozepines, which act as endogenous ligands of benzodiazepine receptors. We have recently shown that one of these endozepines, the triakontatetraneuropeptide TTN, stimulates DNA synthesis in astroglial cells. The purpose of the present study was to determine the mechanism of action of TTN on cultured rat astrocytes. Binding of the peripheral-type benzodiazepine receptor ligand [3H]Ro5-4864 to intact astrocytes was displaced by TTN, whereas its C-terminal fragment (TTN[17,34], the octadecaneuropeptide ODN) did not compete for [3H]Ro5-4864 binding. Microfluorimetric measurement of cytosolic calcium concentrations ([Ca2+]i) with the fluorescent probe indo-1 showed that TTN (10,10 to 10,6 M) provokes a concentration-dependent increase in [Ca2+]i in cultured astrocytes. Simultaneous administration of TTN (10,8 M) and Ro5-4864 (10,5 M) induced an increase in [Ca2+]i similar to that obtained with Ro5-4864 alone. In contrast, the effects of TTN (10,8 M) and ODN (10,8 M) on [Ca2+]i were strictly additive. Chelation of extracellular Ca2+ by EGTA (6 mM) or blockage of Ca2+ channels with Ni2+ (2 mM) abrogated the stimulatory effect of TTN. The calcium influx evoked by TTN (10,7 M) or by Ro5-4864 (10,5 M) was not affected by the N- and T-type calcium channel blockers ,-conotoxin (10,6 M) and mibefradil (10,6 M), but was significantly reduced by the L-type calcium channel blocker nifedipine (10,7 M). Patch-clamp studies showed that, at negative potentials, TTN (10,7 M) induced a sustained depolarization. Reduction of the chloride concentration in the extracellular solution shifted the reversal potential from 0 mV to a positive potential. These data show that TTN, acting through peripheral-type benzodiazepine receptors, provokes chloride efflux, which in turn induces calcium influx via L-type calcium channels in rat astrocytes. GLIA 35:90,100, 2001. © 2001 Wiley-Liss, Inc. [source] Correlation Between Atrial ZnT-1 Expression and Atrial Fibrillation in Humans: A Pilot StudyJOURNAL OF CARDIOVASCULAR ELECTROPHYSIOLOGY, Issue 2 2008Ph.D. , YORAM ETZION M.D. Background: Until recently, the membrane protein ZnT-1 was studied mainly in the context of zinc homeostasis. However, new findings indicate that it acts as an inhibitor of L-type calcium channels. We recently found that acute rapid pacing of the rat atria in vivo augments the expression of ZnT-1, while knockdown of ZnT-1 in culture can oppose the inhibition of L-type calcium channels following rapid pacing. This pilot study, the first to assess cardiac ZnT-1 in humans, was designed to look for possible correlation between the atrial expression of ZnT-1 and atrial fibrillation. Methods: Right atrial appendage tissue was collected from 39 patients (27 with sinus rhythm and 12 with atrial fibrillation; 6-permanent, 6- paroxysmal or persistent) undergoing open-heart surgery. The expression of ZnT-1 was analyzed by Western blot utilizing ,-actin as an internal loading control and a standard rat heart sample (STD) for inter-blot comparison. Results: Overall atrial fibrillation patients (n = 12) had median ZnT-1/,-actin of 1.80 STD (inter-quartile range 1.26 to 2.85) versus 0.73 STD (0.24 to 1.64) in the sinus rhythm group (P = 0.002). No association was found between ZnT-1 level and most other clinical parameters tested. Multivariate analysis determined that atrial fibrillation and increased body mass index were the only independent variables clearly associated with higher ZnT-1 levels (Standardized coefficients Beta = 0.62, 0.31; P = 0.002, P = 0.04, respectively). Conclusions: This pilot study provides evidence for increased ZnT-1 expression in the atria of patients with atrial fibrillation. [source] Phosphate and calcium are required for TGF,-mediated stimulation of ANK expression and function during chondrogenesisJOURNAL OF CELLULAR PHYSIOLOGY, Issue 2 2010Paulina Oca The expression of ANK, a key player in biomineralization, is stimulated by treatment with TGF,. The purpose of this study was to determine whether TGF, stimulation of ANK expression during chondrogenesis was dependent upon the influx of calcium and phosphate into cells. Treatment of ATDC5 cells with TGF, increased ANK expression during all phases of chondrogenic differentiation, particularly at day 14 (proliferation) and day 32 (mineralizing hypertrophy) of culture. Phosphate uptake studies in the presence and absence of phosphonoformic acid (PFA), a competitive inhibitor of the type III Na+/Pi channels Pit-1 and Pit-2, indicated that the stimulation of ANK expression by TGF, required the influx of phosphate, specifically by the Pit-1 transporter, at all phases of differentiation. At hypertrophy, when alkaline phosphatase is highly expressed, inhibition of its activity with levamisole also abrogated the stimulatory effect of TGF, on ANK expression, further illustrating that Pi availability and uptake by the cells is necessary for stimulation of ANK expression in response to TGF,. Since previous studies of endochondral ossification in the growth plate have shown that L-type calcium channels are essential for chondrogenesis, we investigated their role in the TGF,-stimulated ANK response in ATDC5 cells. Treatment with nifedipine to inhibit calcium influx via the L-type channel Cav1.2 (,1C) inhibited the TGF, stimulated increase in ANK expression at all phases of chondrogenesis. Our findings indicate that TGF, stimulation of ANK expression is dependent upon the influx of phosphate and calcium into ATDC5 cells at all stages of differentiation. J. Cell. Physiol. 224: 540,548, 2010. © 2010 Wiley-Liss, Inc. [source] The Role of K+ Channels in Determining Pulmonary Vascular Tone, Oxygen Sensing, Cell Proliferation, and Apoptosis: Implications in Hypoxic Pulmonary Vasoconstriction and Pulmonary Arterial HypertensionMICROCIRCULATION, Issue 8 2006ROHIT MOUDGIL ABSTRACT Potassium channels are tetrameric, membrane-spanning proteins that selectively conduct K+ at near diffusion-limited rates. Their remarkable ionic selectivity results from a highly-conserved K+ recognition sequence in the pore. The classical function of K+ channels is regulation of membrane potential (EM) and thence vascular tone. In pulmonary artery smooth muscle cells (PASMC), tonic K+ egress, driven by a 145/5 mM intracellular/extracellular concentration gradient, contributes to a EM of about ,60 mV. It has been recently discovered that K+ channels also participate in vascular remodeling by regulating cell proliferation and apoptosis. PASMC express voltage-gated (Kv), inward rectifier (Kir), calcium-sensitive (KCa), and two-pore (K2P) channels. Certain K+ channels are subject to rapid redox regulation by reactive oxygen species (ROS) derived from the PASMC's oxygen-sensor (mitochondria and/or NADPH oxidase). Acute hypoxic inhibition of ROS production inhibits Kv1.5, which depolarizes EM, opens voltage-sensitive, L-type calcium channels, elevates cytosolic calcium, and initiates hypoxic pulmonary vasoconstriction (HPV). Hypoxia-inhibited K+ currents are not seen in systemic arterial SMCs. Kv expression is also transcriptionally regulated by HIF-1, and NFAT. Loss of PASMC Kv1.5 and Kv2.1 contributes to the pathogenesis of pulmonary arterial hypertension (PAH) by causing a sustained depolarization, which increases intracellular calcium and K+, thereby stimulating cell proliferation and inhibiting apoptosis, respectively. Restoring Kv expression (via Kv1.5 gene therapy, dichloroacetate, or anti-survivin therapy) reduces experimental PAH. Electrophysiological diversity exists within the pulmonary circulation. Resistance PASMC have a homogeneous Kv current (including an oxygen-sensitive component), whereas conduit PASMC current is a Kv/KCa mosaic. This reflects regional differences in expression of channel isoforms, heterotetramers, splice variants, and regulatory subunits as well as mitochondrial diversity. In conclusion, K+ channels regulate pulmonary vascular tone and remodeling and constitute potential therapeutic targets in the regression of PAH. [source] Effect of otilonium bromide on contractile patterns in the human sigmoid colonNEUROGASTROENTEROLOGY & MOTILITY, Issue 6 2010D. Gallego Abstract Background, The mechanism of action of the spasmolytic compound otilonium bromide (OB) on human colonic motility is not understood. The aim of our study was to characterize the pharmacological effects of OB on contractile patterns in the human sigmoid colon. Methods, Circular sigmoid strips were studied in organ baths. Isolated smooth muscle cells from human sigmoid colon were examined using the calcium imaging technique. Key Results, Otilonium bromide inhibited by 85% spontaneous non-neural rhythmic phasic contractions (RPCs), (IC50 = 49.9 nmol L,1) and stretch-induced tone (IC50 = 10.7 nmol L,1) with maximum effects at micromolar range. OB also inhibited by 50% both on- (IC50 = 38.0 nmol L,1) and off- contractions induced by electrical stimulation of excitatory motor neurons. In contrast, the inhibitory latency period prior to off -contractions was unaffected by OB. OB inhibited acetylcholine-, substance P-, and neurokinin A-induced contractions. The L-type Ca2+ channel agonist BayK8644 reversed the effects of OB on RPCs, on- and off -contractions. Hexamethonium, atropine, the NK2 antagonist, or depletion of intracellular Ca2+ stores by thapsigargin did not prevent the inhibitory effect of OB on RPCs and electrical contractions. KCl-induced calcium transients in isolated smooth muscle cells were also inhibited by OB (IC50 = 0.2 ,mol L,1). Conclusions & Inferences, Otilonium bromide strongly inhibited the main patterns of human sigmoid motility in vitro by blocking calcium influx through L-type calcium channels on smooth muscle cells. This pharmacological profile may mediate the clinically observed effects of the drug in patients with irritable bowel syndrome. [source] Nifedipine trials: effectiveness and safety aspectsBJOG : AN INTERNATIONAL JOURNAL OF OBSTETRICS & GYNAECOLOGY, Issue 2005Herman P. van Geijn Nifedipine (Adalat) is marketed as an anti-hypertensive agent. Nifedipine inhibits voltage-dependent L-type calcium channels, which leads to vascular (and other) smooth muscle relaxation and negative inotropic and chronotropic effects on the heart. Vasodilation, followed by a baroreceptor-mediated increase in sympathetic tone then results in indirect cardiostimulation. Nifedipine was introduced as a tocolytic agent at a time when ,-agonists and magnesium sulphate dominated the arena for the prevention of preterm birth. The oral administration route, the availability of immediate and slow-release preparations, the low incidence of (mild) side effects, and its limited costs explain the attraction to this medication from the obstetric field and its rapid and widespread distribution. Currently, over 40 studies have been published on nifedipine's tocolytic effectiveness, including seven meta-analyses. The quality of the studies suffers particularly from performance bias because the majority of them failed to ensure adequate blinding to treatment both for providers and patients. Concerns about other methodological flaws include measurements, outcome assessment and attrition bias. In particular, the safety aspects of nifedipine for tocolysis have been underassessed. Conclusions from the meta-analyses, favouring the use of nifedipine as a tocolytic agent, are not supported by close examination of the data. The tocolytic effectiveness and ,safety' of nifedipine has been studied primarily in normal pregnancies. Based on its pharmacological properties, one should be cautious to administer nifedipine when the maternal cardiovascular condition is compromised, such as with intrauterine infection, twin pregnancy, maternal hypertension, cardiac disease, etc. Life-threatening pulmonary oedema and/or cardiac failure are definite risks and have been reported. Under such circumstances, the baroreceptor-mediated increase in sympathetic tone may not balance the cardiac-depressant activity of nifedipine. [source] Effects of angiotensin II blockade on inflammation-induced alterations of pharmacokinetics and pharmacodynamics of calcium channel blockersBRITISH JOURNAL OF PHARMACOLOGY, Issue 1 2008S Hanafy Background and purpose: Inflammation elevates plasma verapamil concentrations but diminishes pharmacological response. Angiotensin II is a pro-inflammatory mediator. We examined the effect of angiotensin II receptor blockade on the pharmacokinetics and pharmacodynamics of verapamil, as well as the binding properties and amounts of its target protein in calcium channels, in a rat model of inflammation. Experimental approach: We used 4 groups of male Sprague,Dawley rats (220,280 g): inflamed-placebo, inflamed-treated, control-placebo and control-treated. Inflammation as pre-adjuvant arthritis was induced by injecting Mycobacterium butyricum on day 0. From day 6 to 12, 30 mg kg,1 oral valsartan or placebo was administered twice daily. On day 12, a single oral dose of 25 mg kg,1 verapamil was administered and prolongation of the PR interval measured and plasma samples collected for verapamil and nor-verapamil analysis. The amounts of the target protein Cav1.2 subunit of L-type calcium channels in heart was measured by Western blotting and ligand binding with 3H-nitrendipine. Key results: Inflammation reduced effects of verapamil, although plasma drug concentrations were increased. This was associated with a reduction in ligand binding capacity and amount of the calcium channel target protein in heart extracts. Valsartan significantly reversed the down-regulating effect of inflammation on verapamil's effects on the PR interval, and the lower level of protein binding and the decreased target protein. Conclusions and implications: Reduced responses to calcium channel blockers in inflammatory conditions appeared to be due to a reduced amount of target protein that was reversed by the angiotensin II antagonist, valsartan. British Journal of Pharmacology (2008) 153, 90,99; doi:10.1038/sj.bjp.0707538; published online 29 October 2007 [source] ,1 -Adrenoceptor effects mediated by protein kinase C , in human cultured prostatic stromal cellsBRITISH JOURNAL OF PHARMACOLOGY, Issue 1 2003A Preston We have investigated the effects of ,1 -adrenoceptor stimulation upon contractility, Ca2+ influx, inositol phosphate production, and protein kinase C (PKC) translocation in human cultured prostatic stromal cells (HCPSC). The ,1 -adrenoceptor selective agonist phenylephrine elicited contractile responses of HCPSC, i.e. a maximal cell shortening of 45±6% of initial cell length, with an EC50 of 1.6±0.1 ,M. The ,1 -adrenoceptor selective antagonists prazosin (1 ,M) and terazosin (1 ,M) both blocked contractions to phenylephrine (10 ,M). The L-type calcium channel blocker nifedipine (10 ,M), and the PKC inhibitors Gö 6976 (1 ,M) and bisindolylmaleimide (1 ,M) also inhibited phenylephrine-induced contractions. Phenylephrine caused a concentration dependent increase in inositol phosphate production (EC50 119±67 nM). This response was blocked by terazosin (1 ,M). Phenylephrine caused the translocation of the PKC , isoform, but not the ,, ,, ,, , or , isoforms, from the cytosolic to the particulate fraction of HCPSC, with an EC50 of 5.7±0.5 ,M. In FURA-2AM (5 ,M) loaded cells, phenylephrine elicited concentration dependent increases in [Ca2+]i, with an EC50 of 3.9±0.4 ,M. The response to phenylephrine (10 ,M) was blocked by prazosin (1 ,M), bisindolymaleimide (1 ,M), and nifedipine (10 ,M). In conclusion, this study has shown that HCPSC express functional ,1 -adrenoceptors, and that the intracellular pathways responsible for contractility may be largely dependent upon protein kinase C activation and subsequent opening of L-type calcium channels. British Journal of Pharmacology (2003) 138, 218,224. doi:10.1038/sj.bjp.0705021 [source] Localization of voltage-sensitive L-type calcium channels in the chicken retinaCLINICAL & EXPERIMENTAL OPHTHALMOLOGY, Issue 3 2001Sally I Firth PhD ABSTRACT L-type calcium channels have been associated with synaptic transmission in the retina, and are a potential site for modulation of the release of neurotransmitters. The present study documents the immunohistochemical localization of neuronal ,1 subunits of L-type calcium channels in chicken retina, using antibodies to the ,1c, ,1d and ,1f subunits of L-type calcium channels. The ,1c-like subunits were localized to Müller cells, with predominantly radial processes, and a prominent band of horizontal processes in the outer plexiform layer. The antibody to ,1d subunits labelled most, if not all, cell bodies. The antibody to a human ,1f subunit strongly labelled photoreceptor terminals. Fainter immunoreactivity was detected in the inner segments of the photoreceptors, a subset of amacrine cells, two bands of labelling in the inner plexiform layer and many ganglion cells. The differential cellular distributions of these ,1-subunits suggests subtle functional differences in their roles at different cellular locations. [source] | ||||||||||||||||||||||