cAMP-independent effects of 8-(4-parachlorophenylthio)-cyclic AMP on spike duration and membrane currents in pleural sensory neurons of Aplysia

1994 ◽  
Vol 72 (3) ◽  
pp. 1250-1259 ◽  
Author(s):  
S. Sugita ◽  
D. A. Baxter ◽  
J. H. Byrne
Keyword(s):  
Protein Kinase ◽  
Action Potential ◽  
Voltage Clamp ◽  
Sensory Neurons ◽  
Membrane Current ◽  
Membrane Currents ◽  
Cyclic Monophosphate ◽  
Subsequent Application ◽  
Independent Effects

1. The serotonergic modulation of pleural sensory neurons in Aplysia is mediated via two second messenger systems: the adenosine cyclic monophosphate/protein kinase A (cAMP/PKA) and diacylglycerol/protein kinase C systems. Often membrane permeable derivatives of cAMP, such as 8-(4-parachlorophenylthio)-cAMP (pcpt-cAMP), have been used to investigate the role of cAMP/PKA in modulating sensory neurons. In light of recent findings that pcpt-cAMP may have cAMP-independent actions, we have reexamined the effects of pcpt-cAMP on the action potential and membrane currents of the sensory neurons. 2. Although pcpt-cAMP (500 microM to 1 mM) and serotonin (5-HT; 10 microM) induced comparable measures of spike broadening (an average increase above baseline of 29 and 40%, respectively), the broadening produced by the two was qualitatively different. Serotonin-induced broadening developed slowly over 9-12 min, was most prominent during later phases of the spike repolarization, and reduced the spike afterhyperpolarization. In contrast, pcpt-cAMP-induced broadening developed rapidly, was rather uniform throughout the repolarization phase of the spike, delayed the peak of the action potential, and increased the afterhyperpolarization. 3. Preexposure of sensory neurons to 5-HT did not occlude further spike broaden by subsequent application of pcpt-cAMP. Indeed the effects of the two were additive. In addition, the effects of pcpt-cAMP were not mimicked by another analogue of cAMP, 8-bromo-cAMP. Interestingly, most of the effects of pcpt-cAMP on the action potential were mimicked by 8-(4-parachlorophenyl-thio)-guanosine cyclic monophosphate (pcpt-cGMP), but not by 8-bromo-cGMP. 4. During voltage-clamp pulses to 20 mV, pcpt-cAMP reduced the membrane current throughout the voltage-clamp pulse, which was qualitatively different from the modulation of the membrane current by 5-HT. In addition, the pcpt-cAMP-induced reduction in the membrane current at the beginning of the pulse was much greater than that induced by 5-HT. Moreover, preexposure of sensory neurons to 5-HT did not occlude further reduction in the membrane current by subsequent application of pcpt-cAMP. 5. These results suggest that pcpt-cAMP has some mechanisms of action that are not shared by 5-HT or cAMP but are shared by pcpt-cGMP. In addition, these findings provide further evidence that results obtained with this compound should be interpreted with caution.

Activators of protein kinase C mimic serotonin-induced modulation of a voltage-dependent potassium current in pleural sensory neurons of Aplysia

1994 ◽  
Vol 72 (3) ◽  
pp. 1240-1249 ◽  
Author(s):  
S. Sugita ◽  
D. A. Baxter ◽  
J. H. Byrne
Keyword(s):  
Protein Kinase ◽  
Voltage Clamp ◽  
Sensory Neurons ◽  
Phorbol Esters ◽  
Outward Current ◽  
Membrane Currents ◽  
Kinase C ◽  
Voltage Dependent ◽  
K Current ◽  
Kinetics Of

1. In the pleural mechanoafferent sensory neurons of Aplysia, serotonin (5-HT)-induced spike broadening consists of at least two components: a cAMP and protein kinase A (PKA)-dependent, rapidly developing component and a protein kinase C (PKC)-dependent, slowly developing component. Voltage-clamp experiments were conducted to identify currents that are modulated by PKC and thus may contribute to the slowly developing component of 5-HT-induced spike broadening. 2. We compared the effects of phorbol esters, activators of PKC, on membrane currents with those of 5-HT. Bath application of 5-HT had complex modulatory effects on currents elicited by voltage-clamp pulses to potentials > 0 mV. The kinetics of both activation and inactivation of the membrane currents were slowed by 5-HT. This led to a decrease in an outward current at the beginning of the voltage-clamp pulse and an increase at the end of the pulse. Previous work has shown that these effects represent, in part, the modulation of a large, voltage-dependent K+ current (IK,V) by 5-HT. 3. Active phorbol esters mimicked some of the actions of 5-HT on membrane currents in that they slowed activation and inactivation kinetics of current responses to voltage-clamp pulses more positive than 0 mV. This led to a decrease in an outward current at the beginning of the pulse and an increase at the end of the pulse. Because inactive phorbols did not mimic the actions of 5-HT, the effects of active phorbol esters appeared to be PKC specific. In addition, preexposure of the sensory neurons to active phorbol esters appeared to occlude the modulatory actions of 5-HT on IK,V. Thus it is likely that modulation of IK,V by 5-HT is mediated, at lease in part, by PKC. 4. To further characterize which currents were modulated by PKC, low concentrations of tetraethylammonium (TEA, 2 mM) were used to block Ca(2+)-activated K+ current (IK,Ca). Low TEA partially blocked the phorbol ester-induced increase of the outward current at the end of voltage-clamp pulses. These results agreed with previous reports that activation of PKC enhanced a fast component of IK,Ca in these sensory neurons. Such an enhancement would lead to an increase in outward current that should be blocked by low TEA. Low TEA, however, did not affect phorbol ester-induced decrease of the outward current at the beginning of pulse, where the predominant current is IK,V, which is less sensitive to TEA.(ABSTRACT TRUNCATED AT 400 WORDS)

A re-examination of late outward plateau currents of cardiac Purkinje fibers

1985 ◽  
Vol 249 (1) ◽  
pp. H108-H121
Author(s):  
J. M. Jaeger ◽  
W. R. Gibbons
Keyword(s):  
Action Potential ◽  
Voltage Clamp ◽  
Critical Role ◽  
Membrane Current ◽  
Purkinje Fiber ◽  
Purkinje Fibers ◽  
Outward Currents ◽  
Cardiac Purkinje Fibers

Two outward currents, IX1 and IX2, are thought to be activated by depolarization of the Purkinje fiber. One of these, IX1, is presently believed to play a critical role in repolarization of the action potential. The IX currents were originally analyzed in voltage-clamp experiments in sheep Purkinje fibers. These experiments were designed to minimize interference by other currents, and it was assumed that changes of the net current were produced entirely by the IX currents. We have tried to repeat the original experiments and the analysis that led to acceptance of the existence and roles of the IX currents, without success. Moreover, tests of how membrane current should behave if the IX current hypothesis is correct did not give satisfactory results. Our data suggest the original conclusions about IX1 and IX2 may need substantial revision.

Regression of LV hypertrophy with captopril normalizes membrane currents in rabbits

1998 ◽  
Vol 275 (4) ◽  
pp. H1216-H1224 ◽  
Author(s):  
Seth J. Rials ◽  
Xiaoping Xu ◽  
Ying Wu ◽  
Roger A. Marinchak ◽  
Peter R. Kowey
Keyword(s):  
Cell Membrane ◽  
Action Potential ◽  
Renal Artery ◽  
Current Density ◽  
Action Potential Duration ◽  
Left Ventricular ◽  
Membrane Current ◽  
Membrane Capacitance ◽  
Membrane Currents ◽  
Cell Membrane Capacitance

Recent studies indicate that regression of left ventricular hypertrophy (LVH) normalizes the in situ electrophysiological abnormalities of the left ventricle. This study was designed to determine whether regression of LVH also normalizes the abnormalities of individual membrane currents. LVH was induced in rabbits by renal artery banding. Single ventricular myocytes from rabbits with LVH at 3 mo after renal artery banding demonstrated increased cell membrane capacitance, prolonged action potential duration, decreased inward rectifier K+ current density, and increased transient outward K+ current density compared with myocytes from age-matched controls. Additional rabbits were randomized at 3 mo after banding to treatment with either vehicle or captopril for an additional 3 mo. Myocytes from LVH rabbits treated with vehicle showed persistent membrane current abnormalities. However, myocytes isolated from LVH rabbits treated with captopril had normal cell membrane capacitance, action potential duration, and membrane current densities. Captopril had no direct effect on membrane currents of either control or LVH myocytes. These data support the hypothesis that the action potential prolongation and membrane current abnormalities of LVH are reversed by regression. Normalization of membrane currents probably explains the reduced vulnerability to ventricular arrhythmia observed in this LVH model after treatment with captopril.

Effects of cAMP on serotonin evoked calcium transients in cultured rat airway smooth muscle cells

1997 ◽  
Vol 272 (5) ◽  
pp. L865-L871 ◽  
Author(s):  
B. Tolloczko ◽  
Y. L. Jia ◽  
J. G. Martin
Keyword(s):  
Smooth Muscle ◽  
Protein Kinase ◽  
Airway Smooth Muscle ◽  
Airway Smooth Muscle Cells ◽  
Intracellular Camp ◽  
Dependent Protein Kinase ◽  
Cyclic Monophosphate ◽  
Dependent Protein ◽  
High Concentrations

Agents increasing intracellular adenosine 3',5'-cyclic monophosphate (cAMP) cause relaxation of airway smooth muscle. However, the mechanisms of their action are not fully understood. We investigated the role of cAMP in the modulation of intracellular Ca2+ concentration ([Ca2+]i) transients evoked by serotonin (5-HT) in cultured rat tracheal smooth muscle (TSM) cells. Forskolin (10(-7) M) caused a significant elevation of intracellular cAMP and a 60% relaxation of tracheal rings contracted with 5-HT but did not affect [Ca2+]i in TSM cells. Forskolin (10(-5) M) completely relaxed tracheal rings and significantly decreased [Ca2+]i during the sustained phase of the 5-HT response. Forskolin-induced relaxation was attenuated by the cAMP-dependent protein kinase A (PKA) inhibitor Rp diastereomer of cAMP (Rp-cAMPS; 10(-4) M) and by the guanosine 3',5'-cyclic monophosphate (cGMP)-dependent protein kinase (PKG) inhibitor [Rp isomer of 8-(4-chlorophenylthio)-guanosine 3',5'-cyclic monophosphorothioate, 10(-4) M]. The effects of forskolin on [Ca2+]i were not altered by the PKA inhibitor but were abolished by the PKG inhibitor and thapsigargin. These results indicate that, in rat TSM, the relaxant effects of high concentrations of cAMP may be mediated, at least in part, by facilitating the sequestration of Ca2+ into intracellular stores by a mechanism involving PKG.

scholarly journals Ionic Relations of Cells of Chara Australis VIII. Membrane Currents During a Voltage Clamp

1964 ◽  
Vol 17 (2) ◽  
pp. 388 ◽  
Author(s):  
GP Findlay
Keyword(s):  
Membrane Potential ◽  
Action Potential ◽  
Detailed Analysis ◽  
Voltage Clamp ◽  
Membrane Currents ◽  
Voltage Clamps ◽  
Transient Activity ◽  
Predetermined Level ◽  
Chara Australis ◽  
Ionic Relations

Experiments are described in which "voltage clamps" were applied to the "membrane" of O. australis cells comprising tonoplast and plasmalemma and also to the plasmalemma alone. The voltage-clamp system maintained the membrane potential at a predetermined level, and enabled a detailed analysis to be made of the transient electrical phenomena occurring during the action potential. A scarming technique is also described, by means of which the membrane currentpotential characteristics could be determined at any particular time during the transient activity of the membrane.

Role of protein kinase A in the regulation of cytosolic free calcium in human osteoblast-like SaOS-2 cells

1993 ◽  
Vol 264 (2) ◽  
pp. C464-C470 ◽  
Author(s):  
S. Fukayama ◽  
A. H. Tashjian ◽  
F. R. Bringhurst
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Resistant Mutant ◽  
Free Calcium ◽  
Wild Type ◽  
Cyclic Monophosphate ◽  
Type Pattern ◽  
High Concentrations ◽  
Kinase A

We have used wild-type and adenosine 3',5'-cyclic monophosphate (cAMP)-resistant mutant osteoblast-like SaOS-2 cells to investigate the role of protein kinase A (PKA) in the regulation of cytosolic free Ca2+ concentration ([Ca2+]i). Basal levels of [Ca2+]i were the same in wild-type (127 +/- 6.1 nM) and transfected (117 +/- 6.8 nM) SaOS-2 cells, although 45Ca2+ efflux was slower in the transfected cells. In wild-type cells, thapsigargin (TG, > or = 200 nM), an inhibitor of the Ca(2+)-ATPase activity of the endoplasmic reticulum, acutely increased [Ca2+]i (by up to 2-fold), which then returned promptly to basal [Ca2+]i. In cAMP-resistant cells, TG elicited a significantly greater acute rise in [Ca2+]i, which then decayed to an elevated plateau level. In mutant cells, high concentrations of dibutyryladenosine 3',5'-cyclic monophosphate, which overcome the PKA blockade, restored the changes in [Ca2+]i to the wild-type pattern. In cAMP-resistant, TG-blocked cells, ionomycin (or alpha-thrombin) induced a further elevation in [Ca2+]i, which then declined rapidly to the original basal level. We conclude that basal PKA activity is involved actively in regulation of [Ca2+]i in SaOS-2 cells by promoting Ca2+ efflux from the cell and, possibly, by inhibiting Ca2+ release from or stimulating net Ca2+ sequestration into the ER. We have also obtained evidence for an alternate Ca(2+)-triggered Ca2+ reuptake mechanism in SaOS-2 cells that is not dependent on either Ca(2+)-ATPase or PKA.

scholarly journals Featured Article: AMPKα2 deficiency enhanced susceptibility to ventricular arrhythmias in mice by the role of β-adrenoceptor signaling

2018 ◽  
Vol 243 (8) ◽  
pp. 708-714
Author(s):  
Hong Cao ◽  
Xin Wang ◽  
Shaozheng Ying ◽  
Congxin Huang
Keyword(s):  
Protein Kinase ◽  
Action Potential ◽  
Clinical Significance ◽  
Ventricular Arrhythmias ◽  
Cardiac Electrophysiology ◽  
Monophasic Action Potential ◽  
Wild Type ◽  
Adrenoceptor Antagonist ◽  
Amp Activated Protein Kinase

AMP-activated protein kinase-α2 is the main catalytic subunit of the heart, which is mainly located in cardiac myocytes. The effect of AMP-activated protein kinase-α2 on the cardiac electrophysiology is barely studied. From the previous study, it is possible that AMP-activated protein kinase-α2 may have some effect on the electrophysiology of the heart. To prove the hypothesis, we used the AMP-activated protein kinase-α2 knockout (AMPKα2−/−) mice to estimate the electrophysiological characteristics of AMPKα2−/− mice and try to find the mechanism between them. We used AMP-activated protein kinase-α2 gene knockout (AMPKα2−/−) mice and control wild-type mice as the experimental animals. In the experiment, we measured the monophasic action potential duration and test the inducibility to ventricular arrhythmia in isolated mice heart with and without β-adrenoceptor antagonist metoprolol. Meanwhile, plasma concentration of catecholamine was collected. We found that AMPKα2−/− significantly shortened 90% repolarization of monophasic action potential (MAP) (MAPD90) than wild-type (47.4 ± 2.6 ms vs. 55.5 ± 2.4 ms, n = 10, P < 0.05) and were more vulnerable to be induced to ventricular arrhythmias (70% (7/10) vs. 10% (1/10), P < 0.05), accompanied by the higher concentration of catecholamine (epinephrine: 1.75 ± 0.18 nmol/L vs. 0.68 ± 0.10 nmol/L n = 10, P < 0.05; norepinephrine: 9.56 ± 0.71 nmol/L vs. 2.52 ± 0.31 nmol/L n = 10, P < 0.05). The shortening of MAPD90 and increased inducibility to ventricular arrhythmias of AMPKα2−/− could almost be abolished when perfusion with β-adrenoceptor antagonist metoprolol. It indicated that the β-adrenoceptor activation resulting from catecholamine release was mainly responsible for the relating changes of electrophysiology of AMPKα2−/−. It had great clinical significance, as in patients who had problem with AMP-activated protein kinase-α2 gene, we might use β-adrenoceptor antagonists as the prevention of arrhythmias in future. Impact statement As far as we know, this is the first time the role of AMP-activated protein kinase-α2 (AMPKα2) on the cardiac electrophysiology is explored, and we found that the β-adrenoceptor activation resulting from catecholamine release was mainly responsible for the changes of electrophysiology related to the absence of AMPKα2. This has great clinical significance, as in patients who have problems with AMPKα2 gene, we may use β-adrenoceptor antagonists for the prevention of arrhythmias in future.

A model of the action potential and underlying membrane currents in a rabbit atrial cell

1996 ◽  
Vol 271 (4) ◽  
pp. H1666-H1696 ◽  
Author(s):  
D. S. Lindblad ◽  
C. R. Murphey ◽  
J. W. Clark ◽  
W. R. Giles
Keyword(s):  
Action Potential ◽  
Voltage Clamp ◽  
Cell Voltage ◽  
Ionic Currents ◽  
Membrane Currents ◽  
Whole Cell ◽  
Whole Cell Voltage Clamp ◽  
Voltage Dependent ◽  
Atrial Myocyte ◽  
Atrial Cell

We have developed a mathematical model of the rabbit atrial myocyte and have used it in an examination of the ionic basis of the atrial action potential. Available biophysical data have been incorporated into the model to quantify the specific ultrastructural morphology, intracellular ion buffering, and time- and voltage-dependent currents and transport mechanisms of the rabbit atrial cell. When possible, mathematical expressions describing ionic currents identified in rabbit atrium are based on whole cell voltage-clamp data from enzymatically isolated rabbit atrial myocytes. This membrane model is coupled to equations describing Na+, K+, and Ca2+ homeostasis, including the uptake and release of Ca2+ by the sarcoplasmic reticulum and Ca2+ buffering. The resulting formulation can accurately simulate the whole cell voltage-clamp data on which it is based and provides fits to a family of rabbit atrial cell action potentials obtained at 35 degrees C over a range of stimulus rates (0.2–3.0 Hz). The model is utilized to provide a qualitative prediction of the intracellular Ca2+ concentration transient during the action potential and to illustrate the interactions between membrane currents that underlie repolarization in the rabbit atrial myocyte.

scholarly journals The Ca2+-activated Cl− channel TMEM16B regulates action potential firing and axonal targeting in olfactory sensory neurons

2016 ◽  
Vol 148 (4) ◽  
pp. 293-311 ◽  
Author(s):  
Gianluca Pietra ◽  
Michele Dibattista ◽  
Anna Menini ◽  
Johannes Reisert ◽  
Anna Boccaccio
Keyword(s):  
Action Potential ◽  
Sensory Neurons ◽  
Major Change ◽  
Electrophysiological Recording ◽  
Olfactory Sensory Neurons ◽  
Firing Activity ◽  
Action Potential Firing ◽  
Cl Channel ◽  
Axonal Targeting

The Ca2+-activated Cl− channel TMEM16B is highly expressed in the cilia of olfactory sensory neurons (OSNs). Although a large portion of the odor-evoked transduction current is carried by Ca2+-activated Cl− channels, their role in olfaction is still controversial. A previous report (Billig et al. 2011. Nat. Neurosci. http://dx.doi.org/10.1038/nn.2821) showed that disruption of the TMEM16b/Ano2 gene in mice abolished Ca2+-activated Cl− currents in OSNs but did not produce any major change in olfactory behavior. Here we readdress the role of TMEM16B in olfaction and show that TMEM16B knockout (KO) mice have behavioral deficits in odor-guided food-finding ability. Moreover, as the role of TMEM16B in action potential (AP) firing has not yet been studied, we use electrophysiological recording methods to measure the firing activity of OSNs. Suction electrode recordings from isolated olfactory neurons and on-cell loose-patch recordings from dendritic knobs of neurons in the olfactory epithelium show that randomly selected neurons from TMEM16B KO mice respond to stimulation with increased firing activity than those from wild-type (WT) mice. Because OSNs express different odorant receptors (ORs), we restrict variability by using a mouse line that expresses a GFP-tagged I7 OR, which is known to be activated by heptanal. In response to heptanal, we measure dramatic changes in the firing pattern of I7-expressing neurons from TMEM16B KO mice compared with WT: responses are prolonged and display a higher number of APs. Moreover, lack of TMEM16B causes a markedly reduced basal spiking activity in I7-expressing neurons, together with an alteration of axonal targeting to the olfactory bulb, leading to the appearance of supernumerary I7 glomeruli. Thus, TMEM16B controls AP firing and ensures correct glomerular targeting of OSNs expressing I7. Altogether, these results show that TMEM16B does have a relevant role in normal olfaction.

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