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Channel Subunits (channel + subunit)
Kinds of Channel Subunits Selected AbstractsReduced Expression of the KATP Channel Subunit, Kir6.2, is Associated with Decreased Expression of Neuropeptide Y and Agouti-Related Protein in the Hypothalami of Zucker Diabetic Fatty RatsJOURNAL OF NEUROENDOCRINOLOGY, Issue 12 2007A. Gyte The link between obesity and diabetes is not fully understood but there is evidence to suggest that hypothalamic signalling pathways may be involved. The hypothalamic neuropeptides, pro-opiomelanocortin (POMC), neuropeptide Y (NPY) and agouti-related protein (AGRP) are central to the regulation of food intake and have been implicated in glucose homeostasis. Therefore, the expression of these genes was quantified in hypothalami from diabetic Zucker fatty (ZDF) rats and nondiabetic Zucker fatty (ZF) rats at 6, 8, 10 and 14 weeks of age. Although both strains are obese, only ZDF rats develop pancreatic degeneration and diabetes over this time period. In both ZF and ZDF rats, POMC gene expression was decreased in obese versus lean rats at all ages. By contrast, although there was the expected increase in both NPY and AGRP expression in obese 14-week-old ZF rats, the expression of NPY and AGRP was decreased in 6-week-old obese ZDF rats with hyperinsulinaemia and in 14-week-old rats with the additional hyperglycaemia. Therefore, candidate genes involved in glucose, and insulin signalling pathways were examined in obese ZDF rats over this age range. We found that expression of the ATP-sensitive potassium (KATP) channel component, Kir6.2, was decreased in obese ZDF rats and was lower compared to ZF rats in each age group tested. Furthermore, immunofluorescence analysis showed that Kir6.2 protein expression was reduced in the dorsomedial and ventromedial hypothalamic nuclei of 6-week-old prediabetic ZDF rats compared to ZF rats. The Kir6.2 immunofluorescence colocalised with NPY throughout the hypothalamus. The differences in Kir6.2 expression in ZF and ZDF rats mimic those of NPY and AGRP, which could infer that the changes occur in the same neurones. Overall, these data suggest that chronic changes in hypothalamic Kir6.2 expression may be associated with the development of hyperinsulinaemia and hyperglycaemia in ZDF rats. [source] Thyroid hormone receptor , can control action potential duration in mouse ventricular myocytes through the KCNE1 ion channel subunitACTA PHYSIOLOGICA, Issue 2 2010A. Mansén Abstract Aims:, The reduced heart rate and prolonged QTend duration in mice deficient in thyroid hormone receptor (TR) ,1 may involve aberrant expression of the K+ channel ,-subunit KCNQ1 and its regulatory ,-subunit KCNE1. Here we focus on KCNE1 and study whether increased KCNE1 expression can explain changes in cardiac function observed in TR,1-deficient mice. Methods:, TR-deficient, KCNE1-overexpressing and their respective wildtype (wt) mice were used. mRNA and protein expression were assessed with Northern and Western blot respectively. Telemetry was used to record electrocardiogram and temperature in freely moving mice. Patch-clamp was used to measure action potentials (APs) in isolated cardiomyocytes and ion currents in Chinese hamster ovary (CHO) cells. Results:, KCNE1 was four to 10-fold overexpressed in mice deficient in TR,1. Overexpression of KCNE1 with a heart-specific promoter in transgenic mice resulted in a cardiac phenotype similar to that in TR,1-deficient mice, including a lower heart rate and prolonged QTend time. Cardiomyocytes from KCNE1-overexpressing mice displayed increased AP duration. CHO cells transfected with expression plasmids for KCNQ1 and KCNE1 showed an outward rectifying current that was maximal at equimolar plasmids for KCNQ1-KCNE1 and decreased at higher KCNE1 levels. Conclusion:, The bradycardia and prolonged QTend time in hypothyroid states can be explained by altered K+ channel function due to decreased TR,1-dependent repression of KCNE1 expression. [source] Developmental expression of potassium-channel subunit Kv3.2 within subpopulations of mouse hippocampal inhibitory interneurons,HIPPOCAMPUS, Issue 2 2002Emily Phillips Tansey Abstract The developmental expression of the voltage-gated potassium channel subunit, Kv3.2, and its localization within specific mouse hippocampal inhibitory interneuron populations were determined using immunoblotting and immunohistochemical techniques. Using immunoblotting techniques, the Kv3.2 protein was weakly detected at postnatal age day 7 (P7), and full expression was attained at P21 in tissue extracts from homogenized hippocampal preparations. A similar developmental profile was observed using immunohistochemical techniques in hippocampal tissue sections. Kv3.2 protein expression was clustered on the somata and proximal dendrites of presumed inhibitory interneurons. Using double immunofluorescence, Kv3.2 subunit expression was detected on subpopulations of GABAergic inhibitory interneurons. Kv3.2 was detected in ,100% of parvalbumin-positive interneurons, 86% of interneurons expressing nitric oxide synthase, and ,50% of somatostatin-immunoreactive cells. Kv3.2 expression was absent from both calbindin- and calretinin-containing interneurons. Using immunoprecipitation, we further demonstrate that Kv3.2 and its related subunit Kv3.1b are coexpressed within the same protein complexes in the hippocampus. These data demonstrate that potassium channel subunit Kv3.2 expression is developmentally regulated in a specific set of interneurons. The vast majority of these interneuron subpopulations possess a "fast-spiking" phenotype, consistent with a role for currents through Kv3.2 containing channels in determining action potential kinetics in these cells. Hippocampus 2002;12:137,148. Published 2002 Wiley-Liss, Inc. [source] Transcription of rat TRPV1 utilizes a dual promoter system that is positively regulated by nerve growth factorJOURNAL OF NEUROCHEMISTRY, Issue 1 2007Qing Xue Abstract The capsaicin receptor, also known as ,transient receptor potential vanilloid receptor subtype 1' (TRPV1, VR1), is an ion channel subunit expressed in primary afferent nociceptors, which plays a critical role in pain transduction and thermal hyperalgesia. Increases in nociceptor TRPV1 mRNA and protein are associated with tissue injury,inflammation. As little is understood about what controls TRPV1 RNA transcription in nociceptors, we functionally characterized the upstream portion of the rat TRPV1 gene. Two functional rTRPV1 promoter regions and their transcription initiation sites were identified. Although both promoter regions directed transcriptional activity in nerve growth factor (NGF) treated rat sensory neurons, the upstream Core promoter was the most active in cultures enriched in sensory neurons. Because NGF is a key modulator of inflammatory pain, we examined the effect of NGF on rTRPV1 transcription in PC12 cells. NGF positively regulated transcriptional activity of both rTRPV1 promoter regions in PC12 cells. We propose that the upstream regulatory region of the rTRPV1 gene is composed of a dual promoter system that is regulated by NGF. These findings support the hypothesis that NGF produced under conditions of tissue injury and/or inflammation directs an increase of TRPV1 expression in nociceptors in part through a transcription-dependent mechanism. [source] A Patient Suffering from Hypokalemic Periodic Paralysis Is Deficient in Skeletal Muscle ATP-sensitive K+ channelsCLINICAL AND TRANSLATIONAL SCIENCE, Issue 1 2008Sofija Jovanovi Abstract Hypokalemic periodic paralysis (HOPP) is a rare disease associated with attacks of muscle weakness and hypokalemia. In the present study, immunoprecipitation/Western blotting has shown that a HOPP patient was deficient in sarcolemmal KATP channels. Real-time RT-PCR has revealed that HOPP has decreased mRNA levels of Kir6.2, a pore-forming KATP channel subunit, without affecting the expression of other KATP channel-forming proteins. Based on these findings, we conclude that HOPP could be associated with impaired expression of Kir6.2 which leads to deficiency in skeletal muscle KATP channels, which may explain the symptoms and clinical signs of this disease. [source] Glucose-induced inhibition: how many ionic mechanisms?ACTA PHYSIOLOGICA, Issue 3 2010D. Burdakov Abstract Sensing of sugar by specialized ,glucose-inhibited' cells helps organisms to counteract swings in their internal energy levels. Evidence from several cell types in both vertebrates and invertebrates suggests that this process involves sugar-induced stimulation of plasma membrane K+ currents. However, the molecular composition and the mechanism of activation of the underlying channel(s) remain controversial. In mouse hypothalamic neurones and neurosecretory cells of the crab Cancer borealis, glucose stimulates K+ currents displaying leak-like properties. Yet knockout of some of the candidate ,leak' channel subunits encoded by the KCNK gene family so far failed to abolish glucose inhibition of hypothalamic cells. Moreover, in other tissues, such as the carotid body, glucose-stimulated K+ channels appear to be not leak-like but voltage-gated, suggesting that glucose-induced inhibition may engage multiple types of K+ channels. Other mechanisms of glucose-induced inhibition, such as hyperpolarization mediated by opening of Cl, channels and depolarization block caused by closure of KATP channels have also been proposed. Here we review known ionic and pharmacological features of glucose-induced inhibition in different cell types, which may help to identify its molecular correlates. [source] Localization of KCNC1 (Kv3.1) potassium channel subunits in the avian auditory nucleus magnocellularis and nucleus laminaris during developmentDEVELOPMENTAL NEUROBIOLOGY, Issue 2 2003Suchitra Parameshwaran-Iyer Abstract The KCNC1 (previously Kv3.1) potassium channel, a delayed rectifier with a high threshold of activation, is highly expressed in the time coding nuclei of the adult chicken and barn owl auditory brainstem. The proposed role of KCNC1 currents in auditory neurons is to reduce the width of the action potential and enable neurons to transmit high frequency temporal information with little jitter. Because developmental changes in potassium currents are critical for the maturation of the shape of the action potential, we used immunohistochemical methods to examine the developmental expression of KCNC1 subunits in the avian auditory brainstem. The KCNC1 gene gives rise to two splice variants, a longer KCNC1b and a shorter KCNC1a that differ at the carboxy termini. Two antibodies were used: an antibody to the N-terminus that does not distinguish between KCNC1a and b isoforms, denoted as panKCNC1, and another antibody that specifically recognizes the C terminus of KCNC1b. A comparison of the staining patterns observed with the panKCNC1 and the KCNC1b specific antibodies suggests that KCNC1a and KCNC1b splice variants are differentially regulated during development. Although panKCNC1 immunoreactivity is observed from the earliest time examined in the chicken (E10), a subcellular redistribution of the immunoproduct was apparent over the course of development. KCNC1b specific staining has a late onset with immunostaining first appearing in the regions that map high frequencies in nucleus magnocellularis (NM) and nucleus laminaris (NL). The expression of KCNC1b protein begins around E14 in the chicken and after E21 in the barn owl, relatively late during ontogeny and at the time that synaptic connections mature morphologically and functionally. © 2003 Wiley Periodicals, Inc. J Neurobiol 55: 165,178, 2003 [source] Genetic Architecture of Idiopathic Generalized Epilepsy: Clinical Genetic Analysis of 55 Multiplex FamiliesEPILEPSIA, Issue 5 2004Carla Marini Summary: Purpose: In families with idiopathic generalized epilepsy (IGE), multiple IGE subsyndromes may occur. We performed a genetic study of IGE families to clarify the genetic relation of the IGE subsyndromes and to improve understanding of the mode(s) of inheritance. Methods: Clinical and genealogic data were obtained on probands with IGE and family members with a history of seizures. Families were grouped according to the probands' IGE subsyndrome: childhood absence epilepsy (CAE), juvenile absence epilepsy (JAE), juvenile myoclonic epilepsy (JME), and IGE with tonic,clonic seizures only (IGE-TCS). The subsyndromes in the relatives were analyzed. Mutations in genes encoding ,1 and ,2 ,-aminobutyric acid (GABA)-receptor subunits, ,1 and ,1 sodium channel subunits, and the chloride channel CLC-2 were sought. Results: Fifty-five families were studied. 122 (13%) of 937 first- and second-degree relatives had seizures. Phenotypic concordance within families of CAE and JME probands was 28 and 27%, respectively. JAE and IGE-TCS families had a much lower concordance (10 and 13%), and in the JAE group, 31% of relatives had CAE. JME was rare among affected relatives of CAE and JAE probands and vice versa. Mothers were more frequently affected than fathers. No GABA-receptor or sodium or chloride channel gene mutations were identified. Conclusions: The clinical genetic analysis of this set of families suggests that CAE and JAE share a close genetic relation, whereas JME is a more distinct entity. Febrile seizures and epilepsy with unclassified tonic,clonic seizures were frequent in affected relatives of all IGE individuals, perhaps representing a nonspecific susceptibility to seizures. A maternal effect also was seen. Our findings are consistent with an oligogenic model of inheritance. [source] Environmental manipulations early in development alter seizure activity, Ih and HCN1 protein expression later in lifeEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 12 2006Ulrich Schridde Abstract Although absence epilepsy has a genetic origin, evidence from an animal model (Wistar Albino Glaxo/Rijswijk; WAG/Rij) suggests that seizures are sensitive to environmental manipulations. Here, we show that manipulations of the early rearing environment (neonatal handling, maternal deprivation) of WAG/Rij rats leads to a pronounced decrease in seizure activity later in life. Recent observations link seizure activity in WAG/Rij rats to the hyperpolarization-activated cation current (Ih) in the somatosensory cortex, the site of seizure generation. Therefore, we investigated whether the alterations in seizure activity between rats reared differently might be correlated with changes in Ih and its channel subunits hyperpolarization-activated cation channel HCN1, 2 and 4. Whole-cell recordings from layer 5 pyramidal neurons, in situ hybridization and Western blot of the somatosensory cortex revealed an increase in Ih and HCN1 in neonatal handled and maternal deprived, compared to control rats. The increase was specific to HCN1 protein expression and did not involve HCN2/4 protein expression, or mRNA expression of any of the subunits (HCN1, 2, 4). Our findings provide the first evidence that relatively mild changes in the neonatal environment have a long-term impact of absence seizures, Ih and HCN1, and suggest that an increase of Ih and HCN1 is associated with absence seizure reduction. Our findings shed new light on the role of Ih and HCN in brain functioning and development and demonstrate that genetically determined absence seizures are quite sensitive for early interventions. [source] Auxiliary subunit regulation of high-voltage activated calcium channels expressed in mammalian cellsEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 1 2004Takahiro Yasuda Abstract The effects of auxiliary calcium channel subunits on the expression and functional properties of high-voltage activated (HVA) calcium channels have been studied extensively in the Xenopus oocyte expression system, but are less completely characterized in a mammalian cellular environment. Here, we provide the first systematic analysis of the effects of calcium channel , and ,2,, subunits on expression levels and biophysical properties of three different types (Cav1.2, Cav2.1 and Cav2.3) of HVA calcium channels expressed in tsA-201 cells. Our data show that Cav1.2 and Cav2.3 channels yield significant barium current in the absence of any auxiliary subunits. Although calcium channel , subunits were in principle capable of increasing whole cell conductance, this effect was dependent on the type of calcium channel ,1 subunit, and ,3 subunits altogether failed to enhance current amplitude irrespective of channel subtype. Moreover, the ,2,, subunit alone is capable of increasing current amplitude of each channel type examined, and at least for members of the Cav2 channel family, appears to act synergistically with , subunits. In general agreement with previous studies, channel activation and inactivation gating was regulated both by , and by ,2,, subunits. However, whereas pronounced regulation of inactivation characteristics was seen with the majority of the auxiliary subunits, effects on voltage dependence of activation were only small (< 5 mV). Overall, through a systematic approach, we have elucidated a previously underestimated role of the ,2,,1 subunit with regard to current enhancement and kinetics. Moreover, the effects of each auxiliary subunit on whole cell conductance and channel gating appear to be specifically tailored to subsets of calcium channel subtypes. [source] Contribution of Kir3.1, Kir3.2A and Kir3.2C subunits to native G protein-gated inwardly rectifying potassium currents in cultured hippocampal neuronsEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 8 2003Joanne L. Leaney Abstract G protein-gated inwardly rectifying potassium (GIRK) channels are found in neurons, atrial myocytes and neuroendocrine cells. A characteristic feature is their activation by stimulation of Gi/o -coupled receptors. In central neurons, for example, they are activated by adenosine and GABA and, as such, they play an important role in neurotransmitter-mediated regulation of membrane excitability. The channels are tetrameric assemblies of Kir3.x subunits (Kir3.1,3.4 plus splice variants). In this study I have attempted to identify the channel subunits which contribute to the native GIRK current recorded from primary cultured rat hippocampal pyramidal neurons. Reverse transcriptase,polymerase chain reaction revealed the expression of mRNA for Kir3.1, 3.2A, 3.2C and 3.3 subunits and confocal immunofluorescence microscopy was used to investigate their expression patterns. Diffuse staining was observed on both cell somata and dendrites for Kir3.1 and Kir3.2A yet that for Kir3.2C was weaker and punctate. Whole-cell patch clamp recordings were used to record GIRK currents from hippocampal pyramidal neurons which were identified on the basis of inward rectification, dependence of reversal potential on external potassium concentration and sensitivity to tertiapin. The GIRK currents were enhanced by the stimulation of a number of Gi/o -coupled receptors and were inhibited by pertussis toxin. In order to ascertain which Kir3.x subunits were responsible for the native GIRK current I compared the properties with those of the cloned Kir3.1 + 3.2A and Kir3.1 + 3.2C channels heterologously expressed in HEK293 cells. [source] Inhibition of human ether à go-go potassium channels by Ca2+/calmodulin binding to the cytosolic N- and C-terminiFEBS JOURNAL, Issue 5 2006Ulrike Ziechner Human ether à go-go potassium channels (hEAG1) open in response to membrane depolarization and they are inhibited by Ca2+/calmodulin (CaM), presumably binding to the C-terminal domain of the channel subunits. Deletion of the cytosolic N-terminal domain resulted in complete abolition of Ca2+/CaM sensitivity suggesting the existence of further CaM binding sites. A peptide array-based screen of the entire cytosolic protein of hEAG1 identified three putative CaM-binding domains, two in the C-terminus (BD-C1: 674,683, BD-C2: 711,721) and one in the N-terminus (BD-N: 151,165). Binding of GST-fusion proteins to Ca2+/CaM was assayed with fluorescence correlation spectroscopy, surface plasmon resonance spectroscopy and precipitation assays. In the presence of Ca2+, BD-N and BD-C2 provided dissociation constants in the nanomolar range, BD-C1 bound with lower affinity. Mutations in the binding domains reduced inhibition of the functional channels by Ca2+/CaM. Employment of CaM-EF-hand mutants showed that CaM binding to the N- and C-terminus are primarily dependent on EF-hand motifs 3 and 4. Hence, closure of EAG channels presumably requires the binding of multiple CaM molecules in a manner more complex than previously assumed. [source] Automated mutation screening using dideoxy fingerprinting and capillary array electrophoresisHUMAN MUTATION, Issue 5 2001Lars Allan Larsen Abstract The rapid progress in the isolation of genes associated with human disease has resulted in an increasing demand for mutation screening methods. The molecular diagnosis of the long QT syndrome (LQTS), a cardiac disorder characterized by prolongation of the QTc interval in the ECG, syncopes, and sudden death, requires mutation screening of all exons in at least five genes, encoding cardiac Na+ and K+ channel subunits. A method for automated dideoxy fingerprinting (ddF) using capillary array electrophoresis (CAE) was developed and the efficiency of the method was tested by analyzing 24 DNA samples with mutations in one of the genes KCNQ1 and KCNH2, which are involved in 50% of LQTS cases. One of these mutations, 362insQK in KCNQ1, is novel. The sensitivity was 100% using a single electrophoresis temperature of 18°C or 25°C. However, analysis of the samples in both the sense and anti-sense direction were required for high sensitivity. Analysis in a single direction resulted in a decrease of the sensitivity to 74% and 70%, respectively. The throughput of the ddF method, if performed with a 16 capillary CAE instrument, is 288 samples per seven hr if each sample is analyzed on both strands. Hum Mutat 18:451,457, 2001. © 2001 Wiley-Liss, Inc. [source] Melatonin ameliorates hippocampal nitric oxide production and large conductance calcium-activated potassium channel activity in chronic intermittent hypoxiaJOURNAL OF PINEAL RESEARCH, Issue 3 2008Y. W. Tjong Abstract:, Melatonin protects against hippocampal injury induced by intermittent hypoxia (IH). IH-induced oxidative stress is associated with decreases in constitutive production of nitric oxide (NO) and in the activity of large conductance calcium-activated potassium (BK) channels in hippocampal neurons. We tested the hypothesis that administration of melatonin alleviates the NO deficit and impaired BK channel activity in the hippocampus of IH rats. Sprague,Dawley rats were injected with melatonin (10 mg/kg, i.p.) or vehicle before daily IH exposure for 8 hr for 7 days. The NO and intracellular calcium ([Ca2+]i) levels in the CA1 region of hippocampal slices were measured by electrochemical microsenor and spectrofluorometry, respectively. The activity of BK channels was recorded by patch-clamping electrophysiology in dissociated CA1 neurons. Malondialdehyde levels were increased in the hippocampus of hypoxic rats and were lowered by the melatonin treatment. Levels of NO under resting and hypoxic conditions, and the protein expression of neuronal NO synthase (nNOS) were significantly reduced in the CA1 neurons of hypoxic animals compared with the normoxic controls. These deficits were mitigated in the melatonin-treated hypoxic rats with an improved [Ca2+]i response to acute hypoxia. The open probability of BK channels was decreased in the hypoxic rats and was partially restored in the melatonin-treated animals, without alterations in the expression of channel subunits and unitary conductance. Acute treatment of melatonin had no significant effects on the BK channel activity or on the [Ca2+]i response to hypoxia. Collectively, these results suggest that melatonin ameliorates the constitutive NO production and BK channel activity via an antioxidant mechanism against an IH-induced down-regulation of nNOS expression in hippocampal neurons. [source] KATP channel openers: Structure-activity relationships and therapeutic potentialMEDICINAL RESEARCH REVIEWS, Issue 2 2004Raimund Mannhold Abstract ATP-sensitive potassium channels (KATP channels) are heteromeric complexes of pore-forming inwardly rectifying potassium channel subunits and regulatory sulfonylurea receptor subunits. KATP channels were identified in a variety of tissues including muscle cells, pancreatic ,-cells, and various neurons. They are regulated by the intracellular ATP/ADP ratio; ATP induces channel inhibition and MgADP induces channel opening. Functionally, KATP channels provide a means of linking the electrical activity of a cell to its metabolic state. Shortening of the cardiac action potential, smooth muscle relaxation, inhibition of both insulin secretion, and neurotransmitter release are mediated via KATP channels. Given their many physiological functions, KATP channels represent promising drug targets. Sulfonylureas like glibenclamide block KATP channels; they are used in the therapy of type 2 diabetes. Openers of KATP channels (KCOs), for example, relax smooth muscle and induce hypotension. KCOs are chemically heterogeneous and include as different classes as the benzopyrans, cyanoguanidines, thioformamides, thiadiazines, and pyridyl nitrates. Examples for new chemical entities more recently developed as KCOs include cyclobutenediones, dihydropyridine related structures, and tertiary carbinols. © 2003 Wiley Periodicals, Inc. Med Res Rev, 24, No. 2, 213,266, 2004 [source] Protein distribution of Kcnq1, Kcnh2, and Kcne3 potassium channel subunits during mouse embryonic developmentTHE ANATOMICAL RECORD : ADVANCES IN INTEGRATIVE ANATOMY AND EVOLUTIONARY BIOLOGY, Issue 3 2006María Pilar de Castro Abstract Voltage-dependent potassium channels consist of a pore-forming ,-subunit, which is modulated by additional ,-ancillary or regulatory subunits. Kcnq1 and Kcnh2 ,-channel subunits play pivotal roles in the developing and adult heart. However, Kcnq1 and Kcnh2 have a much wider expression profile than strictly confined to the myocardium, similar to their putative regulatory Kcne1-5 ,-subunits. At present, the distribution of distinct potassium channel subunits has been partially mapped in adult tissues, whereas almost no information is available during embryonic development. In this study, we report a detailed analysis of Kcnq1, Kcnh2, and Kcne3 protein expression during mouse embryogenesis. Our results demonstrate that Kcnq1 and Kcnh2 are widely distributed. Coexpression of both ,-subunits is observed in a wide variety of organs, such as heart and the skeletal muscle, whereas others display unique Kcnq1 or Knch2 expression. Interestingly, Kcne3 expression is also widely observed in distinct tissue layers during embryogenesis, supporting the notion that an exquisite balance of ,- and ,-subunit expression is required for modulating potassium conductance in distinct organs and tissue layers. © 2006 Wiley-Liss, Inc. [source] Immunohistochemical localization of Ih channel subunits, HCN1,4, in the rat brainTHE JOURNAL OF COMPARATIVE NEUROLOGY, Issue 3 2004Takuya Notomi Abstract Hyperpolarization-activated cation currents (Ih) contribute to various physiological properties and functions in the brain, including neuronal pacemaker activity, setting of resting membrane potential, and dendritic integration of synaptic input. Four subunits of the Hyperpolarization-activated and Cyclic-Nucleotide-gated nonselective cation channels (HCN1,4), which generate Ih, have been cloned recently. To better understand the functional diversity of Ih in the brain, we examined precise immunohistochemical localization of four HCNs in the rat brain. Immunoreactivity for HCN1 showed predominantly cortical distribution, being intense in the neocortex, hippocampus, superior colliculus, and cerebellum, whereas those for HCN3 and HCN4 exhibited subcortical distribution mainly concentrated in the hypothalamus and thalamus, respectively. Immunoreactivity for HCN2 had a widespread distribution throughout the brain. Double immunofluorescence revealed colocalization of immunoreactivity for HCN1 and HCN2 in distal dendrites of pyramidal cells in the hippocampus and neocortex. At the electron microscopic level, immunogold particles for HCN1 and HCN2 had similar distribution patterns along plasma membrane of dendritic shafts in layer I of the neocortex and stratum lacunosum moleculare of the hippocampal CA1 area, suggesting that these subunits could form heteromeric channels. Our results further indicate that HCNs are localized not only in somato-dendritic compartments but also in axonal compartments of neurons. Immunoreactivity for HCNs often occurred in preterminal rather than terminal portions of axons and in specific populations of myelinated axons. We also found HCN2-immunopositive oligodendrocytes including perineuronal oligodendrocytes throughout the brain. These results support previous electrophysiological findings and further suggest unexpected roles of Ih channels in the brain. J. Comp. Neurol. 471:241,276, 2004. © 2004 Wiley-Liss, Inc. [source] Characteristics and physiological role of hyperpolarization activated currents in mouse cold thermoreceptorsTHE JOURNAL OF PHYSIOLOGY, Issue 9 2009Patricio Orio Hyperpolarization-activated currents (Ih) are mediated by the expression of combinations of hyperpolarization-activated, cyclic nucleotide-gated (HCN) channel subunits (HCN1,4). These cation currents are key regulators of cellular excitability in the heart and many neurons in the nervous system. Subunit composition determines the gating properties and cAMP sensitivity of native Ih currents. We investigated the functional properties of Ih in adult mouse cold thermoreceptor neurons from the trigeminal ganglion, identified by their high sensitivity to moderate cooling and responsiveness to menthol. All cultured cold-sensitive (CS) neurons expressed a fast activating Ih, which was fully blocked by extracellular Cs+ or ZD7288 and had biophysical properties consistent with those of heteromeric HCN1,HCN2 channels. In CS neurons from HCN1(,/,) animals, Ih was greatly reduced but not abolished. We find that Ih activity is not essential for the transduction of cold stimuli in CS neurons. Nevertheless, Ih has the potential to shape the excitability of CS neurons. First, Ih blockade caused a membrane hyperpolarization in CS neurons of about 5 mV. Furthermore, impedance power analysis showed that all CS neurons had a prominent subthreshold membrane resonance in the 5,7 Hz range, completely abolished upon blockade of Ih and absent in HCN1 null mice. This frequency range matches the spontaneous firing frequency of cold thermoreceptor terminals in vivo. Behavioural responses to cooling were reduced in HCN1 null mice and after peripheral pharmacological blockade of Ih with ZD7288, suggesting that Ih plays an important role in peripheral sensitivity to cold. [source] De novo expression of Kv6.3 contributes to changes in vascular smooth muscle cell excitability in a hypertensive mice strainTHE JOURNAL OF PHYSIOLOGY, Issue 3 2009Alejandro Moreno-Domínguez Essential hypertension involves a gradual and sustained increase in total peripheral resistance, reflecting an increased vascular tone. This change associates with a depolarization of vascular myocytes, and relies on a change in the expression profile of voltage-dependent ion channels (mainly Ca2+ and K+ channels) that promotes arterial contraction. However, changes in expression and/or modulation of voltage-dependent K+ channels (Kv channels) are poorly defined, due to their large molecular diversity and their vascular bed-specific expression. Here we endeavor to characterize the molecular and functional expression of Kv channels in vascular smooth muscle cells (VSMCs) and their regulation in essential hypertension, by using VSMCs from resistance (mesenteric) or conduit (aortic) arteries obtained from a hypertensive inbred mice strain, BPH, and the corresponding normotensive strain, BPN. Real-time PCR reveals a differential distribution of Kv channel subunits in the different vascular beds as well as arterial bed-specific changes under hypertension. In mesenteric arteries, the most conspicuous change was the de novo expression of Kv6.3 (Kcng3) mRNA in hypertensive animals. The functional relevance of this change was studied by using patch-clamp techniques. VSMCs from BPH arteries were more depolarized than BPN ones, and showed significantly larger capacitance values. Moreover, Kv current density in BPH VSMCs is decreased mainly due to the diminished contribution of the Kv2 component. The kinetic and pharmacological profile of Kv2 currents suggests that the expression of Kv6.3 could contribute to the natural development of hypertension. [source] Molecular determinants of inactivation in voltage-gated Ca2+ channelsTHE JOURNAL OF PHYSIOLOGY, Issue 2 2000Steffen Hering Evolution has created a large family of different classes of voltage-gated Ca2+ channels and a variety of additional splice variants with different inactivation properties. Inactivation controls the amount of Ca2+ entry during an action potential and is, therefore, believed to play an important role in tissue-specific Ca2+ signalling. Furthermore, mutations in a neuronal Ca2+ channel (Cav2.1) that are associated with the aetiology of neurological disorders such as familial hemiplegic migraine and ataxia cause significant changes in the process of channel inactivation. Ca2+ channels of a given subtype may inactivate by three different conformational changes: a fast and a slow voltage-dependent inactivation process and in some channel types by an additional Ca2+ -dependent inactivation mechanism. Inactivation kinetics of Ca2+ channels are determined by the intrinsic properties of their pore-forming ,1 -subunits and by interactions with other channel subunits. This review focuses on structural determinants of Ca2+ channel inactivation in different parts of Ca2+ channel ,1 -subunits, including pore-forming transmembrane segments and loops, intracellular domain linkers and the carboxyl terminus. Inactivation is also affected by the interaction of the ,1 -subunits with auxiliary ,-subunits and intracellular regulator proteins. The evidence shows that pore-forming S6 segments and conformational changes in extra- (pore loop) and intracellular linkers connected to pore-forming segments may play a principal role in the modulation of Ca2+ channel inactivation. Structural concepts of Ca2+ channel inactivation are discussed. [source] AtKC1, a conditionally targeted Shaker-type subunit, regulates the activity of plant K+ channelsTHE PLANT JOURNAL, Issue 1 2008Geoffrey Duby Summary Amongst the nine voltage-gated K+ channel (Kv) subunits expressed in Arabidopsis, AtKC1 does not seem to form functional Kv channels on its own, and is therefore said to be silent. It has been proposed to be a regulatory subunit, and to significantly influence the functional properties of heteromeric channels in which it participates, along with other Kv channel subunits. The mechanisms underlying these properties of AtKC1 remain unknown. Here, the transient (co-)expression of AtKC1, AKT1 and/or KAT1 genes was obtained in tobacco mesophyll protoplasts, which lack endogenous inward Kv channel activity. Our experimental conditions allowed both localization of expressed polypeptides (GFP-tagging) and recording of heterologously expressed Kv channel activity (untagged polypeptides). It is shown that AtKC1 remains in the endoplasmic reticulum unless it is co-expressed with AKT1. In these conditions heteromeric AtKC1-AKT1 channels are obtained, and display functional properties different from those of homomeric AKT1 channels in the same context. In particular, the activation threshold voltage of the former channels is more negative than that of the latter ones. Also, it is proposed that AtKC1-AKT1 heterodimers are preferred to AKT1-AKT1 homodimers during the process of tetramer assembly. Similar results are obtained upon co-expression of AtKC1 with KAT1. The whole set of data provides evidence that AtKC1 is a conditionally-targeted Kv subunit, which probably downregulates the physiological activity of other Kv channel subunits in Arabidopsis. [source] Functional Roles Of KATP Channels In Vascular Smooth MuscleCLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY, Issue 4 2002Joseph E Brayden SUMMARY 1. ATP-sensitive potassium channels (KATP) are present in vascular smooth muscle cells and play important roles in the vascular responses to a variety of pharmacological and endogenous vasodilators. 2. The KATP channels are composed of four inwardly rectifying K+ channel subunits and four regulatory sulphonylurea receptors. The KATP channels are inhibited by intracellular ATP and by sulphonylurea agents. 3. Pharmacological vasodilators such as cromakalim, pinacidil and diazoxide directly activate KATP channels. The associated membrane hyperpolarization closes voltage-dependent Ca2+ channels, which leads to a reduction in intracellular Ca2+ and vasodilation. 4. Endogenous vasodilators such as calcitonin gene-related peptide, vasoactive intestinal polypeptide, prostacylin and adenosine activate KATP by stimulating the formation of cAMP and increasing the activity of protein kinase A. Part of the mechanism of contraction of endogenous vasoconstrictors is due to inhibition of KATP channels. 5. The KATP channels appear to be tonically active in some vascular beds and contribute to the physiological regulation of vascular tone and blood flow. These channels also are activated under pathophysiological conditions, such as hypoxia, ischaemia, acidosis and septic shock, and, in these disease states, may play an important role in the regulation of tissue perfusion. [source] | ||||||||||||||||||||||||||||