Abstract 1522: Local Ca Releases And Spontaneous Beating Rate Of Rabbit Sinoatrial Node Cells Are Controlled By Interaction Of Basal Phosphodiesterase 3 And 4 Activity

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Tatiana M Vinogradova ◽  
Alexey E Lyashkov ◽  
Harold Spurgeon ◽  
Edward G Lakatta
Keyword(s):  
Sinoatrial Node ◽  
Pde4 Inhibitor ◽  
Phosphodiesterase 3 ◽  
Whole Cell Patch Clamp ◽  
Diastolic Depolarization ◽  
Spontaneous Action ◽  
Beating Rate ◽  
Sinoatrial Node Cells ◽  
Spontaneous Action Potentials

A high basal phosphodiesterase (PDE) activity in sinoatrial node cells (SANC) regulates cAMP and modulates cAMP-mediated PKA-dependent phosphorylation of Ca 2+ cycling proteins that determines the characteristics of local subsarcolemmal Ca 2+ releases (LCR). LCRs occur during the terminal diastolic depolarization and activate the inward Na + -Ca 2+ exchange current, thus, regulating the SANC basal spontaneous beating rate. Though both PDE3 and PDE4 are present in rabbit SANC, a role of PDE4 or its interaction with PDE3 is not clear. To determine to what extent PDE4 controls LCRs and the SANC spontaneous beating rate, we used perforated patch to record spontaneous action potentials (APs) in conjunction with confocal linescan images or whole-cell patch clamp to measure L-type Ca 2+ current (I Ca,L ) at 35 o C. A specific PDE3 inhibitor, cilostamide (Cil, 0.3 μmol/L), increased the spontaneous beating rate by 28% (from 154 ± 21 to 191 ± 17 beat/min); in contrast a specific PDE4 inhibitor, rolipram (Rol, 2 μmol/L), had no effect on the firing rate. The substantial role of PDE4 in the control of basal spontaneous SANC firing was unmasked when PDE3 and PDE4 were simultaneously inhibited: the combination of Rol and Cil produced ~57% acceleration of the beating rate (from129 ± 10 to 200 ± 10 beat/min) equivalent to that produced by the broad-spectrum PDE inhibitor, IBMX (100μM), i.e. ~55% (from 145 ± 8 to 221 ± 6 beat/min). The combination of Cil and Rol increased LCR amplitude, size and decreased the LCR period similar to IBMX. Both IBMX and the combination of Cil and Rol produced a substantial increase in I Ca,L ~135% (from 10 ± 2 to 22 ± 3 pA/pF) and ~125% (from 6 ± 1 to 13 ± 3 pA/pF) respectively. The efficiency of PDE3 inhibition alone on spontaneous beating rate might be ascribed to its lower K m for cAMP, than that of PDE4. When PDE3 is suppressed level of cAMP is sufficiently elevated to activate PDE4. Therefore, in the presence of PDE3 inhibition, PDE4 inhibition contributes to the increase in the spontaneous beating rate. Thus, an interaction of basal PDE3 and PDE4 activities in SANC modulates cAMP level and I Ca,L which controls Ca 2+ influx, cell Ca 2+ load, determining LCR characteristics. When both PDE3 and PDE4 are inhibited the maximum spontaneous beating rate of SANC is achieved.

Regional differences in the role of the Ca2+ and Na+ currents in pacemaker activity in the sinoatrial node

1997 ◽  
Vol 272 (6) ◽  
pp. H2793-H2806 ◽  
Author(s):  
I. Kodama ◽  
M. R. Nikmaram ◽  
M. R. Boyett ◽  
R. Suzuki ◽  
H. Honjo ◽  
...  
Keyword(s):  
Action Potentials ◽  
Regional Differences ◽  
Sinoatrial Node ◽  
Pacemaker Activity ◽  
Na Current ◽  
Na Currents ◽  
Spontaneous Action ◽  
Spontaneous Action Potentials ◽  
Rabbit Sinoatrial Node

The effect of block of the L-type Ca2+ current by 2 microM nifedipine and of the Na+ current by 20 microM tetrodotoxin on the center (normally the leading pacemaker site) and periphery (latent pacemaker tissue) of the rabbit sinoatrial node was investigated. Spontaneous action potentials were recorded with microelectrodes from either an isolated right atrium containing the whole node or small balls of tissue (approximately 0.3-0.4 mm in diameter) from different regions of the node. Nifedipine abolished the action potential in the center, but not usually in the periphery, in both the intact sinoatrial node and the small balls. Tetrodotoxin had no effect, on electrical activity in small balls from the center, but it decreased the takeoff potential and upstroke velocity and slowed the spontaneous activity (by 49 +/- 10%; n = 11) in small balls from the periphery. It is concluded that whereas the L-type Ca2- current plays an obligatory role in pacemaking in the center, the Na+ current plays a major role in pacemaking in the periphery.

Protein kinase A mediates activation of ATP-sensitive K+ currents by CGRP in gallbladder smooth muscle

1994 ◽  
Vol 267 (3) ◽  
pp. G494-G499 ◽  
Author(s):  
L. Zhang ◽  
A. D. Bonev ◽  
G. M. Mawe ◽  
M. T. Nelson
Keyword(s):  
Smooth Muscle ◽  
Protein Kinase ◽  
Protein Kinase A ◽  
Whole Cell Patch Clamp ◽  
Camp Analogue ◽  
Calcitonin Gene ◽  
Transduction Mechanisms ◽  
Spontaneous Action ◽  
Spontaneous Action Potentials ◽  
Kinase A

The signal transduction mechanisms underlying the activation of ATP-sensitive potassium (KATP) current by calcitonin gene-related peptide (CGRP) in gallbladder smooth muscle were examined with intracellular microelectrode recording and whole cell patch-clamp techniques. In the intact gallbladder preparation, the adenylyl cyclase activator forskolin hyperpolarized the membrane potential and abolished spontaneous action potentials. This response was inhibited by the KATP channel blocker glibenclamide. CGRP (10 nM), forskolin (10 microM), the membrane-permeable adenosine 3',5'-cyclic monophosphate (cAMP) analogue adenosine 3',5'-cyclic monophosphothioate (Sp-cAMP[S]; 500 microM), and the catalytic subunit of protein kinase A (100 U/ml) activated glibenclamide-sensitive currents in enzymatically dissociated gallbladder smooth muscle cells. CGRP activation of potassium currents was prevented by dialysis of the cell cytoplasm with guanosine 5'-O-(2-thiodiphosphate) (5 mM) or a specific peptide inhibitor of protein kinase A (2.3 microM). Okadaic acid (5 microM), a phosphatase inhibitor, slowed the deactivation of the KATP current, following removal of CGRP. The results of this study indicate that CGRP hyperpolarizes gallbladder smooth muscle by elevation of cAMP and subsequent stimulation of protein kinase A.

A role for a background sodium current in spontaneous action potentials and secretion from rat lactotrophs

1996 ◽  
Vol 271 (6) ◽  
pp. C1927-C1934 ◽  
Author(s):  
S. Sankaranarayanan ◽  
S. M. Simasko
Keyword(s):  
Action Potentials ◽  
Sodium Current ◽  
Plaque Assay ◽  
Input Resistance ◽  
Cytosolic Calcium ◽  
Calcium Dependent ◽  
Whole Cell Patch Clamp ◽  
Spontaneous Action ◽  
Spontaneous Action Potentials ◽  
Spontaneous Secretion

We have used the perforated-patch variation of whole cell patch-clamp techniques, measurements of cytosolic calcium with use of fura 2, and secretion measurements with use of the reverse-hemolytic plaque assay to address the role of depolarizing background currents in maintaining spontaneous action potentials and spontaneous secretion from rat lactotrophs in primary culture. Replacement of bath sodium with tris(hydroxymethyl)aminomethane or N-methyl-D-glucamine caused a dramatic hyperpolarization of the cells, a cessation of spontaneous action potentials, and an increase in input resistance of cells. Tetrodotoxin had no effect on spontaneous action potentials, and removal of bath calcium stopped spiking but did not hyperpolarize the cells. The hyperpolarization in response to removal of bath sodium was associated with a decrease in cytosolic calcium levels. Finally, removal of bath sodium caused a decrease in spontaneous secretion of prolactin from lactotrophs. These data suggest that a background sodium current is essential to drive the membrane to threshold for firing spontaneous calcium-dependent action potentials in lactotrophs. This, in turn, results in elevated intracellular calcium, which supports spontaneous secretion of prolactin from these cells.

Desensitization to acetylcholine in single sinoatrial node cells isolated from rabbit hearts

1992 ◽  
Vol 263 (6) ◽  
pp. H1779-H1789 ◽  
Author(s):  
H. Honjo ◽  
I. Kodama ◽  
W. J. Zang ◽  
M. R. Boyett
Keyword(s):  
Potassium Current ◽  
Sinoatrial Node ◽  
Chronotropic Effect ◽  
Patch Clamp Technique ◽  
Membrane Hyperpolarization ◽  
Chronotropic Response ◽  
Negative Chronotropic Effect ◽  
Whole Cell Patch Clamp ◽  
Sinoatrial Node Cells ◽  
Continuous Presence

The negative chronotropic effect of acetylcholine (ACh) on the sinoatrial node fades in the continuous presence of ACh as a result of desensitization. We have investigated the mechanism underlying desensitization in single rabbit sinoatrial node cells using the whole cell patch clamp technique. The negative chronotropic effect resulting from the injection of a constant hyperpolarizing current faded. ACh activated an inwardly rectifying potassium current (iK,ACh), which faded in the continuous presence of ACh. ACh had no effect on “basal” L-type calcium current (iCa), but ACh decreased iCa, which had been potentiated by isoprenaline. This effect did not fade during a 2-min exposure to ACh. ACh decreased the hyperpolarization-activated current (i(f)). This effect again did not fade. These results suggest that desensitization of the negative chronotropic response to ACh is, in part, the result of the membrane hyperpolarization and, in part, the result of the fade of iK,ACh. These results also suggest that, whereas the activation of potassium current by ACh rapidly fades, the effects resulting from the inhibition of adenylate cyclase do not.

scholarly journals Hierarchical clustering of ryanodine receptors enables emergence of a calcium clock in sinoatrial node cells

2014 ◽  
Vol 143 (5) ◽  
pp. 577-604 ◽  
Author(s):  
Michael D. Stern ◽  
Larissa A. Maltseva ◽  
Magdalena Juhaszova ◽  
Steven J. Sollott ◽  
Edward G. Lakatta ◽  
...  
Keyword(s):  
Hierarchical Clustering ◽  
Calcium Release ◽  
Sinoatrial Node ◽  
Sinus Node ◽  
Ryanodine Receptors ◽  
Membrane Currents ◽  
Immunofluorescent Staining ◽  
Adrenergic Stimulation ◽  
Beating Rate ◽  
Sinoatrial Node Cells

The sinoatrial node, whose cells (sinoatrial node cells [SANCs]) generate rhythmic action potentials, is the primary pacemaker of the heart. During diastole, calcium released from the sarcoplasmic reticulum (SR) via ryanodine receptors (RyRs) interacts with membrane currents to control the rate of the heartbeat. This “calcium clock” takes the form of stochastic, partially periodic, localized calcium release (LCR) events that propagate, wave-like, for limited distances. The detailed mechanisms controlling the calcium clock are not understood. We constructed a computational model of SANCs, including three-dimensional diffusion and buffering of calcium in the cytosol and SR; explicit, stochastic gating of individual RyRs and L-type calcium channels; and a full complement of voltage- and calcium-dependent membrane currents. We did not include an anatomical submembrane space or inactivation of RyRs, the two heuristic components that have been used in prior models but are not observed experimentally. When RyRs were distributed in discrete clusters separated by >1 µm, only isolated sparks were produced in this model and LCR events did not form. However, immunofluorescent staining of SANCs for RyR revealed the presence of bridging RyR groups between large clusters, forming an irregular network. Incorporation of this architecture into the model led to the generation of propagating LCR events. Partial periodicity emerged from the interaction of LCR events, as observed experimentally. This calcium clock becomes entrained with membrane currents to accelerate the beating rate, which therefore was controlled by the activity of the SERCA pump, RyR sensitivity, and L-type current amplitude, all of which are targets of β-adrenergic–mediated phosphorylation. Unexpectedly, simulations revealed the existence of a pathological mode at high RyR sensitivity to calcium, in which the calcium clock loses synchronization with the membrane, resulting in a paradoxical decrease in beating rate in response to β-adrenergic stimulation. The model indicates that the hierarchical clustering of surface RyRs in SANCs may be a crucial adaptive mechanism. Pathological desynchronization of the clocks may explain sinus node dysfunction in heart failure and RyR mutations.

Role of mitochondria in spontaneous rhythmic activity and intracellular calcium waves in the guinea pig gallbladder smooth muscle

2008 ◽  
Vol 294 (2) ◽  
pp. G467-G476 ◽  
Author(s):  
Onesmo B. Balemba ◽  
Aaron C. Bartoo ◽  
Mark T. Nelson ◽  
Gary M. Mawe
Keyword(s):  
Smooth Muscle ◽  
Rhythmic Activity ◽  
Atp Production ◽  
Mitochondrial Inhibitors ◽  
Carbonyl Cyanide ◽  
Spontaneous Action ◽  
Cells Of Cajal ◽  
Spontaneous Action Potentials ◽  
Spontaneous Rhythmic Activity

Mitochondrial Ca2+ handling has been implicated in spontaneous rhythmic activity in smooth muscle and interstitial cells of Cajal. In this investigation we evaluated the effect of mitochondrial inhibitors on spontaneous action potentials (APs), Ca2+ flashes, and Ca2+ waves in gallbladder smooth muscle (GBSM). Disruption of the mitochondrial membrane potential with carbonyl cyanide 3-chlorophenylhydrazone, carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazone, rotenone, and antimycin A significantly reduced or eliminated APs, Ca2+ flashes, and Ca2+ waves in GBSM. Blockade of ATP production with oligomycin did not alter APs or Ca2+ flashes but significantly reduced Ca2+ wave frequency. Inhibition of mitochondrial Ca2+ uptake and Ca2+ release with Ru360 and CGP-37157, respectively, reduced the frequency of Ca2+ flashes and Ca2+ waves in GBSM. Similar to oligomycin, cyclosporin A did not alter AP and Ca2+ flash frequency but significantly reduced Ca2+ wave activity. These data suggest that mitochondrial Ca2+ handling is necessary for the generation of spontaneous electrical activity and may therefore play an important role in gallbladder tone and motility.

scholarly journals SK Current, Expressed During the Development and Regeneration of Chick Hair Cells, Contributes to the Patterning of Spontaneous Action Potentials

2022 ◽  
Vol 15 ◽  
Author(s):  
Snezana Levic
Keyword(s):  
Electrical Activity ◽  
Hair Cells ◽  
Action Potentials ◽  
Functional Expression ◽  
Basilar Papilla ◽  
Spontaneous Electrical Activity ◽  
Intracellular Ca ◽  
Spontaneous Action ◽  
Spontaneous Action Potentials

Chick hair cells display calcium (Ca2+)-sensitive spontaneous action potentials during development and regeneration. The role of this activity is unclear but thought to be involved in establishing proper synaptic connections and tonotopic maps, both of which are instrumental to normal hearing. Using an electrophysiological approach, this work investigated the functional expression of Ca2+-sensitive potassium [IK(Ca)] currents and their role in spontaneous electrical activity in the developing and regenerating hair cells (HCs) in the chick basilar papilla. The main IK(Ca) in developing and regenerating chick HCs is an SK current, based on its sensitivity to apamin. Analysis of the functional expression of SK current showed that most dramatic changes occurred between E8 and E16. Specifically, there is a developmental downregulation of the SK current after E16. The SK current gating was very sensitive to the availability of intracellular Ca2+ but showed very little sensitivity to T-type voltage-gated Ca2+ channels, which are one of the hallmarks of developing and regenerating hair cells. Additionally, apamin reduced the frequency of spontaneous electrical activity in HCs, suggesting that SK current participates in patterning the spontaneous electrical activity of HCs.

scholarly journals Unique Ca2+-Cycling Protein Abundance and Regulation Sustains Local Ca2+ Releases and Spontaneous Firing of Rabbit Sinoatrial Node Cells

2018 ◽  
Vol 19 (8) ◽  
pp. 2173 ◽  
Author(s):  
Tatiana Vinogradova ◽  
Syevda Tagirova (Sirenko) ◽  
Edward Lakatta
Keyword(s):  
Sinoatrial Node ◽  
Ventricular Myocytes ◽  
Ryanodine Receptors ◽  
Cell Types ◽  
Protein Abundance ◽  
Gradual Change ◽  
Spontaneous Firing ◽  
Diastolic Depolarization ◽  
Sinoatrial Node Cells ◽  
Rabbit Sinoatrial Node

Spontaneous beating of the heart pacemaker, the sinoatrial node, is generated by sinoatrial node cells (SANC) and caused by gradual change of the membrane potential called diastolic depolarization (DD). Submembrane local Ca2+ releases (LCR) from sarcoplasmic reticulum (SR) occur during late DD and activate an inward Na+/Ca2+ exchange current, which accelerates the DD rate leading to earlier occurrence of an action potential. A comparison of intrinsic SR Ca2+ cycling revealed that, at similar physiological Ca2+ concentrations, LCRs are large and rhythmic in permeabilized SANC, but small and random in permeabilized ventricular myocytes (VM). Permeabilized SANC spontaneously released more Ca2+ from SR than VM, despite comparable SR Ca2+ content in both cell types. In this review we discuss specific patterns of expression and distribution of SR Ca2+ cycling proteins (SR Ca2+ ATPase (SERCA2), phospholamban (PLB) and ryanodine receptors (RyR)) in SANC and ventricular myocytes. We link ability of SANC to generate larger and rhythmic LCRs with increased abundance of SERCA2, reduced abundance of the SERCA inhibitor PLB. In addition, an increase in intracellular [Ca2+] increases phosphorylation of both PLB and RyR exclusively in SANC. The differences in SR Ca2+ cycling protein expression between SANC and VM provide insights into diverse regulation of intrinsic SR Ca2+ cycling that drives automaticity of SANC.

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