The Journal of General Physiology最新文献

{"title":"Interactions of external K+ and internal blockers in a weak inward-rectifier K+ channel.","authors":"Lei Yang,&nbsp;Johan Edvinsson,&nbsp;Lawrence G Palmer","doi":"10.1085/jgp.201210835","DOIUrl":"https://doi.org/10.1085/jgp.201210835","url":null,"abstract":"<p><p>We investigated the effects of changing extracellular K(+) concentrations on block of the weak inward-rectifier K(+) channel Kir1.1b (ROMK2) by the three intracellular cations Mg(2+), Na(+), and TEA(+). Single-channel currents were monitored in inside-out patches made from Xenopus laevis oocytes expressing the channels. With 110 mM K(+) in the inside (cytoplasmic) solution and 11 mM K(+) in the outside (extracellular) solution, these three cations blocked K(+) currents with a range of apparent affinities (K(i) (0) = 1.6 mM for Mg(2+), 160 mM for Na(+), and 1.8 mM for TEA(+)) but with similar voltage dependence (zδ = 0.58 for Mg(2+), 0.71 for Na(+), and 0.61 for TEA(+)) despite having different valences. When external K(+) was increased to 110 mM, the apparent affinity of all three blockers was decreased approximately threefold with no significant change in the voltage dependence of block. The possibility that the transmembrane cavity is the site of block was explored by making mutations at the N152 residue, a position previously shown to affect rectification in Kir channels. N152D increased the affinity for block by Mg(2+) but not for Na(+) or TEA(+). In contrast, the N152Y mutation increased the affinity for block by TEA(+) but not for Na(+) or Mg(2+). Replacing the C terminus of the channel with that of the strong inward-rectifier Kir2.1 increased the affinity of block by Mg(2+) but had a small effect on that by Na(+). TEA(+) block was enhanced and had a larger voltage dependence. We used an eight-state kinetic model to simulate these results. The effects of voltage and external K(+) could be explained by a model in which the blockers occupy a site, presumably in the transmembrane cavity, at a position that is largely unaffected by changes in the electric field. The effects of voltage and extracellular K(+) are explained by shifts in the occupancy of sites within the selectivity filter by K(+) ions.</p>","PeriodicalId":173753,"journal":{"name":"The Journal of General Physiology","volume":" ","pages":"529-40"},"PeriodicalIF":3.8,"publicationDate":"2012-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1085/jgp.201210835","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"31012704","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Taking a translational turn.","authors":"Elizabeth M Adler","doi":"10.1085/jgp.201210911","DOIUrl":"https://doi.org/10.1085/jgp.201210911","url":null,"abstract":"This month’s installment of Generally Physiological takes a translational turn, discussing functional analyses of two proteins whose mutation is associated with disease (heart failure and cystic fibrosis, respectively) and the mechanism underlying the painful response to cold found in ciguatera. \u0000 \u0000 \u0000 \u0000 \u0000 \u0000 \u0000 \u0000Image 1. Model for how β-adrenergic signaling could modulate cardiac contraction through phosphorylation of MyBP-C. (From Burghardt and Ajtai. 2012. Science. 337:1182–1183. Reprinted with permission from AAAS.) \u0000 \u0000 \u0000 \u0000A tunable brake on cardiac contractility \u0000Cardiac contraction, like that of skeletal muscle, depends on the interaction of myosin thick filaments with actin thin filaments, which slide past each other to shorten the sarcomeres that make up muscle fibers. Although mutations in cardiac myosin–binding protein C (cMyBP-C, which binds to thick filaments) have been linked to heart disease, its precise physiological role—and thus the mechanisms underlying the pathophysiological consequences of its malfunction—has been unclear (see Burghardt and Ajtai, 2012). Previs et al. (2012) developed an in vitro sarcomere model in which they visualized the movement of fluorescently labeled actin filaments along thick filaments immobilized on coverslips. Analyses of actin movement along thick filaments isolated from the hearts of wild-type mice or mice lacking cMyBP-C revealed that cMyBP-C, which localizes to a particular region of the thick filament (the C-zone), acts in this region to slow actin movement. β-adrenergic stimulation of the heart promotes cMyBP-C phosphorylation, and manipulation of the degree of phosphorylation revealed that increasing phosphorylation of cMyBP-C was associated with a graded reduction in its inhibition of actin filament velocity. Thus, the authors propose that cMyBP-C enables the fine-tuning of cardiac contraction, providing a potential mechanism to link its loss or dysfunction to aberrant cardiac contractility. \u0000 \u0000 \u0000 \u0000 \u0000 \u0000Image 2. Model for the energetic coupling of CFTR gating; ATP hydrolysis provides a shortcut from open state O1 to open state O2. (from Jih et al., 2012). \u0000 \u0000 \u0000 \u0000 \u0000A one-way cycle for CFTR gating \u0000 \u0000The cystic fibrosis transmembrane reporter (CFTR, mutation of which leads to cystic fibrosis) is unusual in being an ATP-gated chloride channel in a family of active transporters (the ATP-binding cassette [ABC] protein superfamily). Unlike active transporters, which couple ATP hydrolysis to substrate movement against a concentration gradient, channels mediate the passive transmembrane diffusion of ions down their concentration gradients. Thus, there is no clear requirement for the energy derived from ATP hydrolysis in CFTR function (see Tsai, 2012). Noting that CFTR retains key structural elements common to other members of the ABC family, Jih et al. (2012) exploited a mutant form of the CFTR that exhibits two different open states (characterized by distinct conductances) to explore the role of ATP hy","PeriodicalId":173753,"journal":{"name":"The Journal of General Physiology","volume":" ","pages":"455-6"},"PeriodicalIF":3.8,"publicationDate":"2012-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1085/jgp.201210911","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"31012702","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Modeling-independent elucidation of inactivation pathways in recombinant and native A-type Kv channels.","authors":"Jeffrey D Fineberg,&nbsp;David M Ritter,&nbsp;Manuel Covarrubias","doi":"10.1085/jgp.201210869","DOIUrl":"https://doi.org/10.1085/jgp.201210869","url":null,"abstract":"<p><p>A-type voltage-gated K(+) (Kv) channels self-regulate their activity by inactivating directly from the open state (open-state inactivation [OSI]) or by inactivating before they open (closed-state inactivation [CSI]). To determine the inactivation pathways, it is often necessary to apply several pulse protocols, pore blockers, single-channel recording, and kinetic modeling. However, intrinsic hurdles may preclude the standardized application of these methods. Here, we implemented a simple method inspired by earlier studies of Na(+) channels to analyze macroscopic inactivation and conclusively deduce the pathways of inactivation of recombinant and native A-type Kv channels. We investigated two distinct A-type Kv channels expressed heterologously (Kv3.4 and Kv4.2 with accessory subunits) and their native counterparts in dorsal root ganglion and cerebellar granule neurons. This approach applies two conventional pulse protocols to examine inactivation induced by (a) a simple step (single-pulse inactivation) and (b) a conditioning step (double-pulse inactivation). Consistent with OSI, the rate of Kv3.4 inactivation (i.e., the negative first derivative of double-pulse inactivation) precisely superimposes on the profile of the Kv3.4 current evoked by a single pulse because the channels must open to inactivate. In contrast, the rate of Kv4.2 inactivation is asynchronous, already changing at earlier times relative to the profile of the Kv4.2 current evoked by a single pulse. Thus, Kv4.2 inactivation occurs uncoupled from channel opening, indicating CSI. Furthermore, the inactivation time constant versus voltage relation of Kv3.4 decreases monotonically with depolarization and levels off, whereas that of Kv4.2 exhibits a J-shape profile. We also manipulated the inactivation phenotype by changing the subunit composition and show how CSI and CSI combined with OSI might affect spiking properties in a full computational model of the hippocampal CA1 neuron. This work unambiguously elucidates contrasting inactivation pathways in neuronal A-type Kv channels and demonstrates how distinct pathways might impact neurophysiological activity.</p>","PeriodicalId":173753,"journal":{"name":"The Journal of General Physiology","volume":" ","pages":"513-27"},"PeriodicalIF":3.8,"publicationDate":"2012-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1085/jgp.201210869","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"31012703","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The intrinsic energy of the gating isomerization of a neuromuscular acetylcholine receptor channel.","authors":"Tapan K Nayak,&nbsp;Prasad G Purohit,&nbsp;Anthony Auerbach","doi":"10.1085/jgp.201110752","DOIUrl":"https://doi.org/10.1085/jgp.201110752","url":null,"abstract":"<p><p>Nicotinic acetylcholine receptor (AChR) channels at neuromuscular synapses rarely open in the absence of agonists, but many different mutations increase the unliganded gating equilibrium constant (E0) to generate AChRs that are active constitutively. We measured E0 for two different sets of mutant combinations and by extrapolation estimated E0 for wild-type AChRs. The estimates were 7.6 and 7.8×10(-7) in adult-type mouse AChRs (-100 mV at 23°C). The values are in excellent agreement with one obtained previously by using a completely different method (6.5×10(-7), from monoliganded gating). E0 decreases with depolarization to the same extent as does the diliganded gating equilibrium constant, e-fold with ∼60 mV. We estimate that at -100 mV the intrinsic energy of the unliganded gating isomerization is +8.4 kcal/mol (35 kJ/mol), and that in the absence of a membrane potential, the intrinsic chemical energy of this global conformational change is +9.4 kcal/mol (39 kJ/mol). Na+ and K+ in the extracellular solution have no measureable effect on E0, which suggests that unliganded gating occurs with only water occupying the transmitter binding sites. The results are discussed with regard to the energy changes in receptor activation and the competitive antagonism of ions in agonist binding.</p>","PeriodicalId":173753,"journal":{"name":"The Journal of General Physiology","volume":" ","pages":"349-58"},"PeriodicalIF":3.8,"publicationDate":"2012-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1085/jgp.201110752","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40559563","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Inorganic polyphosphate is a potent activator of the mitochondrial permeability transition pore in cardiac myocytes.","authors":"Lea K Seidlmayer,&nbsp;Maria R Gomez-Garcia,&nbsp;Lothar A Blatter,&nbsp;Evgeny Pavlov,&nbsp;Elena N Dedkova","doi":"10.1085/jgp.201210788","DOIUrl":"https://doi.org/10.1085/jgp.201210788","url":null,"abstract":"<p><p>Mitochondrial dysfunction caused by excessive Ca2+ accumulation is a major contributor to cardiac cell and tissue damage during myocardial infarction and ischemia-reperfusion injury (IRI). At the molecular level, mitochondrial dysfunction is induced by Ca2+-dependent opening of the mitochondrial permeability transition pore (mPTP) in the inner mitochondrial membrane, which leads to the dissipation of mitochondrial membrane potential (ΔΨm), disruption of adenosine triphosphate production, and ultimately cell death. Although the role of Ca2+ for induction of mPTP opening is established, the exact molecular mechanism of this process is not understood. The aim of the present study was to test the hypothesis that the adverse effect of mitochondrial Ca2+ accumulation is mediated by its interaction with inorganic polyphosphate (polyP), a polymer of orthophosphates linked by phosphoanhydride bonds. We found that cardiac mitochondria contained significant amounts (280±60 pmol/mg of protein) of short-chain polyP with an average length of 25 orthophosphates. To test the role of polyP for mPTP activity, we investigated kinetics of Ca2+ uptake and release, ΔΨm and Ca2+-induced mPTP opening in polyP-depleted mitochondria. polyP depletion was achieved by mitochondria-targeted expression of a polyP-hydrolyzing enzyme. Depletion of polyP in mitochondria of rabbit ventricular myocytes led to significant inhibition of mPTP opening without affecting mitochondrial Ca2+ concentration by itself. This effect was observed when mitochondrial Ca2+ uptake was stimulated by increasing cytosolic [Ca2+] in permeabilized myocytes mimicking mitochondrial Ca2+ overload observed during IRI. Our findings suggest that inorganic polyP is a previously unrecognized major activator of mPTP. We propose that the adverse effect of polyphosphate might be caused by its ability to form stable complexes with Ca2+ and directly contribute to inner mitochondrial membrane permeabilization.</p>","PeriodicalId":173753,"journal":{"name":"The Journal of General Physiology","volume":" ","pages":"321-31"},"PeriodicalIF":3.8,"publicationDate":"2012-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1085/jgp.201210788","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40571994","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Gating properties of the P2X2a and P2X2b receptor channels: experiments and mathematical modeling.","authors":"Anmar Khadra,&nbsp;Zonghe Yan,&nbsp;Claudio Coddou,&nbsp;Melanija Tomić,&nbsp;Arthur Sherman,&nbsp;Stanko S Stojilkovic","doi":"10.1085/jgp.201110716","DOIUrl":"https://doi.org/10.1085/jgp.201110716","url":null,"abstract":"<p><p>Adenosine triphosphate (ATP)-gated P2X2 receptors exhibit two opposite activation-dependent changes, pore dilation and pore closing (desensitization), through a process that is incompletely understood. To address this issue and to clarify the roles of calcium and the C-terminal domain in gating, we combined biophysical and mathematical approaches using two splice forms of receptors: the full-size form (P2X2aR) and the shorter form missing 69 residues in the C-terminal domain (P2X2bR). Both receptors developed conductivity for N-methyl-D-glucamine within 2-6 s of ATP application. However, pore dilation was accompanied with a decrease rather than an increase in the total conductance, which temporally coincided with rapid and partial desensitization. During sustained agonist application, receptors continued to desensitize in calcium-independent and calcium-dependent modes. Calcium-independent desensitization was more pronounced in P2X2bR, and calcium-dependent desensitization was more pronounced in P2X2aR. In whole cell recording, we also observed use-dependent facilitation of desensitization of both receptors. Such behavior was accounted for by a 16-state Markov kinetic model describing ATP binding/unbinding and activation/desensitization. The model assumes that naive receptors open when two to three ATP molecules bind and undergo calcium-independent desensitization, causing a decrease in the total conductance, or pore dilation, causing a shift in the reversal potential. In calcium-containing media, receptor desensitization is facilitated and the use-dependent desensitization can be modeled by a calcium-dependent toggle switch. The experiments and the model together provide a rationale for the lack of sustained current growth in dilating P2X2Rs and show that receptors in the dilated state can also desensitize in the presence of calcium.</p>","PeriodicalId":173753,"journal":{"name":"The Journal of General Physiology","volume":" ","pages":"333-48"},"PeriodicalIF":3.8,"publicationDate":"2012-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1085/jgp.201110716","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40571463","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Selective disruption of high sensitivity heat activation but not capsaicin activation of TRPV1 channels by pore turret mutations.","authors":"Yuanyuan Cui,&nbsp;Fan Yang,&nbsp;Xu Cao,&nbsp;Vladimir Yarov-Yarovoy,&nbsp;KeWei Wang,&nbsp;Jie Zheng","doi":"10.1085/jgp.201110724","DOIUrl":"https://doi.org/10.1085/jgp.201110724","url":null,"abstract":"<p><p>The capsaicin receptor transient receptor potential vanilloid (TRPV)1 is a highly heat-sensitive ion channel. Although chemical activation and heat activation of TRPV1 elicit similar pungent, painful sensation, the molecular mechanism underlying synergistic activation remains mysterious. In particular, where the temperature sensor is located and whether heat and capsaicin share a common activation pathway are debated. To address these fundamental issues, we searched for channel mutations that selectively affected one form of activation. We found that deletion of the first 10 amino acids of the pore turret significantly reduced the heat response amplitude and shifted the heat activation threshold, whereas capsaicin activation remained unchanged. Removing larger portions of the turret disrupted channel function. Introducing an artificial sequence to replace the deleted region restored sensitive capsaicin activation in these nonfunctional channels. The heat activation, however, remained significantly impaired, with the current exhibiting diminishing heat sensitivity to a level indistinguishable from that of a voltage-gated potassium channel, Kv7.4. Our results demonstrate that heat and capsaicin activation of TRPV1 are structurally and mechanistically distinct processes, and the pore turret is an indispensible channel structure involved in the heat activation process but is not part of the capsaicin activation pathway. Synergistic effect of heat and capsaicin on TRPV1 activation may originate from convergence of the two pathways on a common activation gate.</p>","PeriodicalId":173753,"journal":{"name":"The Journal of General Physiology","volume":" ","pages":"273-83"},"PeriodicalIF":3.8,"publicationDate":"2012-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1085/jgp.201110724","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40157956","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The voltage dependence of the TMEM16B/anoctamin2 calcium-activated chloride channel is modified by mutations in the first putative intracellular loop.","authors":"Valentina Cenedese,&nbsp;Giulia Betto,&nbsp;Fulvio Celsi,&nbsp;O Lijo Cherian,&nbsp;Simone Pifferi,&nbsp;Anna Menini","doi":"10.1085/jgp.201110764","DOIUrl":"https://doi.org/10.1085/jgp.201110764","url":null,"abstract":"<p><p>Ca(2+)-activated Cl(-) channels (CaCCs) are involved in several physiological processes. Recently, TMEM16A/anoctamin1 and TMEM16B/anoctamin2 have been shown to function as CaCCs, but very little information is available on the structure-function relations of these channels. TMEM16B is expressed in the cilia of olfactory sensory neurons, in microvilli of vomeronasal sensory neurons, and in the synaptic terminals of retinal photoreceptors. Here, we have performed the first site-directed mutagenesis study on TMEM16B to understand the molecular mechanisms of voltage and Ca(2+) dependence. We have mutated amino acids in the first putative intracellular loop and measured the properties of the wild-type and mutant TMEM16B channels expressed in HEK 293T cells using the whole cell voltage-clamp technique in the presence of various intracellular Ca(2+) concentrations. We mutated E367 into glutamine or deleted the five consecutive glutamates (386)EEEEE(390) and (399)EYE(401). The EYE deletion did not significantly modify the apparent Ca(2+) dependence nor the voltage dependence of channel activation. E367Q and deletion of the five glutamates did not greatly affect the apparent Ca(2+) affinity but modified the voltage dependence, shifting the conductance-voltage relations toward more positive voltages. These findings indicate that glutamates E367 and (386)EEEEE(390) in the first intracellular putative loop play an important role in the voltage dependence of TMEM16B, thus providing an initial structure-function study for this channel.</p>","PeriodicalId":173753,"journal":{"name":"The Journal of General Physiology","volume":" ","pages":"285-94"},"PeriodicalIF":3.8,"publicationDate":"2012-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1085/jgp.201110764","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40157957","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Structural basis for the channel function of a degraded ABC transporter, CFTR (ABCC7).","authors":"Yonghong Bai,&nbsp;Min Li,&nbsp;Tzyh-Chang Hwang","doi":"10.1085/jgp.201110705","DOIUrl":"https://doi.org/10.1085/jgp.201110705","url":null,"abstract":"<p><p>Cystic fibrosis transmembrane conductance regulator (CFTR) is a member of the ATP-binding cassette (ABC) transporter superfamily, but little is known about how this ion channel that harbors an uninterrupted ion permeation pathway evolves from a transporter that works by alternately exposing its substrate conduit to the two sides of the membrane. Here, we assessed reactivity of intracellularly applied thiol-specific probes with cysteine residues substituted into the 12th transmembrane segment (TM12) of CFTR. Our experimental data showing high reaction rates of substituted cysteines toward the probes, strong blocker protection of cysteines against reaction, and reaction-induced alterations in channel conductance support the idea that TM12 of CFTR contributes to the lining of the ion permeation pathway. Together with previous work, these findings raise the possibility that pore-lining elements of CFTR involve structural components resembling those that form the substrate translocation pathway of ABC transporters. In addition, comparison of reaction rates in the open and closed states of the CFTR channel leads us to propose that upon channel opening, the wide cytoplasmic vestibule tightens and the pore-lining TM12 rotates along its helical axis. This simple model for gating conformational changes in the inner pore domain of CFTR argues that the gating transition of CFTR and the transport cycle of ABC proteins share analogous conformational changes. Collectively, our data corroborate the popular hypothesis that degradation of the cytoplasmic-side gate turned an ABC transporter into the CFTR channel.</p>","PeriodicalId":173753,"journal":{"name":"The Journal of General Physiology","volume":" ","pages":"495-507"},"PeriodicalIF":3.8,"publicationDate":"2011-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1085/jgp.201110705","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40119917","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Influence of pH on Ca²⁺ current and its control of electrical and Ca²⁺ signaling in ventricular myocytes.","authors":"Noriko Saegusa, Emma Moorhouse, Richard D Vaughan-Jones, Kenneth W Spitzer","doi":"10.1085/jgp.201110658","DOIUrl":"10.1085/jgp.201110658","url":null,"abstract":"<p><p>Modulation of L-type Ca(2+) current (I(Ca,L)) by H(+) ions in cardiac myocytes is controversial, with widely discrepant responses reported. The pH sensitivity of I(Ca,L) was investigated (whole cell voltage clamp) while measuring intracellular Ca(2+) (Ca(2+)(i)) or pH(i) (epifluorescence microscopy) in rabbit and guinea pig ventricular myocytes. Selectively reducing extracellular or intracellular pH (pH(o) 6.5 and pH(i) 6.7) had opposite effects on I(Ca,L) gating, shifting the steady-state activation and inactivation curves to the right and left, respectively, along the voltage axis. At low pH(o), this decreased I(Ca,L), whereas at low pH(i), it increased I(Ca,L) at clamp potentials negative to 0 mV, although the current decreased at more positive potentials. When Ca(2+)(i) was buffered with BAPTA, the stimulatory effect of low pH(i) was even more marked, with essentially no inhibition. We conclude that extracellular H(+) ions inhibit whereas intracellular H(+) ions can stimulate I(Ca,L). Low pH(i) and pH(o) effects on I(Ca,L) were additive, tending to cancel when appropriately combined. They persisted after inhibition of calmodulin kinase II (with KN-93). Effects are consistent with H(+) ion screening of fixed negative charge at the sarcolemma, with additional channel block by H(+)(o) and Ca(2+)(i). Action potential duration (APD) was also strongly H(+) sensitive, being shortened by low pH(o), but lengthened by low pH(i), caused mainly by H(+)-induced changes in late Ca(2+) entry through the L-type Ca(2+) channel. Kinetic analyses of pH-sensitive channel gating, when combined with whole cell modeling, successfully predicted the APD changes, plus many of the accompanying changes in Ca(2+) signaling. We conclude that the pH(i)-versus-pH(o) control of I(Ca,L) will exert a major influence on electrical and Ca(2+)-dependent signaling during acid-base disturbances in the heart.</p>","PeriodicalId":173753,"journal":{"name":"The Journal of General Physiology","volume":" ","pages":"537-59"},"PeriodicalIF":0.0,"publicationDate":"2011-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/0c/ea/JGP_201110658.PMC3206307.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40120392","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}