scholarly journals Acute Detubulation of Ventricular Myocytes Amplifies the Inhibitory Effect of Cholinergic Agonist on Intracellular Ca2+ Transients

2021 ◽  
Vol 12 ◽  
Author(s):  
Andriy E. Belevych ◽  
Vladimir Bogdanov ◽  
Dmitry A. Terentyev ◽  
Sandor Gyorke
Keyword(s):  
Muscarinic Receptor ◽  
Cardiac Myocytes ◽  
Muscarinic Receptors ◽  
Parasympathetic Nervous System ◽  
Ventricular Myocytes ◽  
Heart Function ◽  
Critical Role ◽  
Inhibitory Effect ◽  
Intracellular Ca ◽  
Image Pixels

Muscarinic receptors expressed in cardiac myocytes play a critical role in the regulation of heart function by the parasympathetic nervous system. How the structural organization of cardiac myocytes affects the regulation of Ca2+ handling by muscarinic receptors is not well-defined. Using confocal Ca2+ imaging, patch-clamp techniques, and immunocytochemistry, the relationship between t-tubule density and cholinergic regulation of intracellular Ca2+ in normal murine ventricular myocytes and myocytes with acute disruption of the t-tubule system caused by formamide treatment was studied. The inhibitory effect of muscarinic receptor agonist carbachol (CCh, 10 μM) on the amplitude of Ca2+ transients, evoked by field-stimulation in the presence of 100 nM isoproterenol (Iso), a β-adrenergic agonist, was directly proportional to the level of myocyte detubulation. The timing of the maximal rate of fluorescence increase of fluo-4, a Ca2+-sensitive dye, was used to classify image pixels into the regions functionally coupled or uncoupled to the sarcolemmal Ca2+ influx (ICa). CCh decreased the fraction of coupled regions and suppressed Ca2+ propagation from sarcolemma inside the cell. Formamide treatment reduced ICa density and decreased sarcoplasmic reticulum (SR) Ca2+ content. CCh did not change SR Ca2+ content in Iso-stimulated control and formamide-treated myocytes. CCh inhibited peak ICa recorded in the presence of Iso by ∼20% in both the control and detubulated myocytes. Reducing ICa amplitude up to 40% by changing the voltage step levels from 0 to –25 mV decreased Ca2+ transients in formamide-treated but not in control myocytes in the presence of Iso. CCh inhibited CaMKII activity, whereas CaMKII inhibition with KN93 mimicked the effect of CCh on Ca2+ transients in formamide-treated myocytes. It was concluded that the downregulation of t-tubules coupled with the diminished efficiency of excitation–contraction coupling, increases the sensitivity of Ca2+ release and propagation to muscarinic receptor-mediated inhibition of both ICa and CaMKII activity.

Fluid pressure modulates L-type Ca2+ channel via enhancement of Ca2+-induced Ca2+ release in rat ventricular myocytes

2008 ◽  
Vol 294 (4) ◽  
pp. C966-C976 ◽  
Author(s):  
Sunwoo Lee ◽  
Joon-Chul Kim ◽  
Yuhua Li ◽  
Min-Jeong Son ◽  
Sun-Hee Woo
Keyword(s):  
Ventricular Myocytes ◽  
Fluid Pressure ◽  
Inhibitory Effect ◽  
Cellular Mechanism ◽  
Confocal Imaging ◽  
Rat Ventricular Myocytes ◽  
Whole Cell Patch Clamp ◽  
Current Voltage ◽  
Intracellular Ca ◽  
High Concentrations

This study examines whether fluid pressure (FP) modulates the L-type Ca2+ channel in cardiomyocytes and investigates the underlying cellular mechanism(s) involved. A flow of pressurized (∼16 dyn/cm2) fluid, identical to that bathing the myocytes, was applied onto single rat ventricular myocytes using a microperfusion method. The Ca2+ current ( ICa) and cytosolic Ca2+ signals were measured using a whole cell patch-clamp and confocal imaging, respectively. It was found that the FP reversibly suppressed ICa (by 25%) without altering the current-voltage relationships, and it accelerated the inactivation of ICa. The level of ICa suppression by FP depended on the level and duration of pressure. The Ba2+ current through the Ca2+ channel was only slightly decreased by the FP (5%), suggesting an indirect inhibition of the Ca2+ channel during FP stimulation. The cytosolic Ca2+ transients and the basal Ca2+ in field-stimulated ventricular myocytes were significantly increased by the FP. The effects of the FP on the ICa and on the Ca2+ transient were resistant to the stretch-activated channel inhibitors, GsMTx-4 and streptomycin. Dialysis of myocytes with high concentrations of BAPTA, the Ca2+ buffer, eliminated the FP-induced acceleration of ICa inactivation and reduced the inhibitory effect of the FP on ICa by ≈80%. Ryanodine and thapsigargin, abolishing sarcoplasmic reticulum Ca2+ release, eliminated the accelerating effect of FP on the ICa inactivation, and they reduced the inhibitory effect of FP on the ICa. These results suggest that the fluid pressure indirectly suppresses the Ca2+ channel by enhancing the Ca2+-induced intracellular Ca2+ release in rat ventricular myocytes.

Voltage-independent changes in L-type Ca2+current uncoupled from SR Ca2+ release in cardiac myocytes

2000 ◽  
Vol 279 (4) ◽  
pp. H2024-H2031 ◽  
Author(s):  
Andrzej M. Janczewski ◽  
Edward G. Lakatta ◽  
Michael D. Stern
Keyword(s):  
Cardiac Myocytes ◽  
Ventricular Myocytes ◽  
Abrupt Increase ◽  
Time To Peak ◽  
Time Integral ◽  
Marked Disparity ◽  
Release Flux ◽  
Intracellular Ca ◽  
Combined Actions ◽  
Continuous Presence

To determine the effect of voltage-independent alterations of L-type Ca2+ current ( I Ca) on the sarcoplasmic reticular (SR) Ca2+ release in cardiac myocytes, we measured I Ca and cytosolic Ca2+ transients (Cai 2+; intracellular Ca2+ concentration) in voltage-clamped rat ventricular myocytes during 1) an abrupt increase of extracellular [Ca2+] (Cao 2+) or 2) application of 1 μM FPL-64176, a Ca2+channel agonist, to selectively alter I Ca in the absence of changes in SR Ca2+ loading. On the first depolarization in higher Cao 2+, peak I Ca was increased by 46 ± 6% ( P < 0.001), but the increases in the maximal rate of rise of Cai 2+(dCai 2+/d t max, where t is time; an index of SR Ca2+ release flux) and the Cai 2+ transient amplitude were not significant. Rapid exposure to FPL-64176 greatly slowed inactivation of I Ca, increasing its time integral by 117 ± 8% ( P < 0.001) without significantly increasing peak I Ca, dCai 2+/d t max, or amplitude of the corresponding Cai 2+ transient. Prolongation of exposure to higher Cao 2+ or FPL-64176 did not further increase peak I Ca but greatly increased dCai 2+/d t max, Cai 2+ transient amplitude, and the gain of Ca2+ release (dCai 2+/d t max/ I Ca), evidently due to augmentation of the SR Ca2+ loading. Also, the time to peak dCai 2+/d t maxwas significantly increased in the continuous presence of higher Cao 2+ (by 37 ± 5%, P < 0.001) or FPL-64176 (by 63 ± 5%, P < 0.002). Our experiments provide the first evidence of a marked disparity between an increased peak I Ca and the corresponding SR Ca2+ release. We attribute this to saturation of the SR Ca2+ release flux as predicted by local control theory. Prolongation of the SR Ca2+ release flux, caused by combined actions of a larger I Ca and maximally augmented SR Ca2+ loading, might reflect additional Ca2+ release from corbular SR.

Effect of calcium and cAMP on Goα expression in neonatal rat cardiac myocytes

1991 ◽  
Vol 261 (4) ◽  
pp. 15-20
Author(s):  
Karen A. Foster ◽  
Janet D. Robishaw
Keyword(s):  
Cardiac Myocytes ◽  
Inhibitory Effect ◽  
Neonatal Rat ◽  
Intracellular Camp ◽  
Dibutyryl Camp ◽  
High K ◽  
Neonatal Rat Cardiac Myocytes ◽  
Intracellular Ca ◽  
The Mean ◽  
Rat Cardiac Myocytes

Culturing neonatal rat cardiac myocytes in 50 mM KCl inhibits the accumulation of Go that occurs when myocytes are placed in culture. The mechanism by which high extracellular K+ inhibits Go accumulation in myocytes was investigated by measurement of the concentration of intracellular Ca2+ ([Ca2+]) and adenosine 3',5'-cyclic monophosphate concentration ([cAMP]) of control and K+-depolarized myocytes. Although intracellular [Ca2++] in K+-depolarized myocytes was twofold higher than basal intracellular [Ca2+] in control cells, the mean intracellular [Ca2+] in contracting control myocytes was comparable to that of K+-depolarized myocytes. Furthermore, myocytes cultured in low Ca2+ plus high K+ exhibited an inhibition of Go accumulation, even though intracellular [Ca2+] was 10-fold lower than that of cells cultured in normal Ca2+ plus high K+. In addition, intracellular [cAMP] of K+-depolarized myocytes was comparable to that of control cells. Moreover, dibutyryl cAMP inhibited Go accumulation in myocytes to the same extent as high K+, even though intracellular [cAMP] differed 10-fold. Thus neither intracellular Ca2+ nor cAMP appear to mediate the inhibitory effect of high K+ on Go accumulation. However, cAMP has an inhibitory effect on Goα expression that is independent of K+. dibutyryl cAMP; fura-2; immunoblotting

scholarly journals inhibitor 2-[(4-methylphenyl)sulfonylcarbamido]-1-(4-nitrobenzyl)pyridinium bromide (2-AP27) is a muscarinic M2 receptor antagonist

Medicina ◽  
2009 ◽  
Vol 45 (7) ◽  
pp. 516
Author(s):  
Vytenis Skeberdis ◽  
Vida Gendvilienė ◽  
Danguolė Zablockaitė ◽  
Irma Martišienė ◽  
Antanas Stankevičius
Keyword(s):  
Cardiac Myocytes ◽  
Ventricular Myocytes ◽  
Inhibitory Effect ◽  
Patch Clamp Technique ◽  
Pyridinium Bromide ◽  
K Current ◽  
Isolated Cardiac Myocytes ◽  
Series Of Experiments ◽  
Muscarinic M2 Receptors ◽  
M2 Muscarinic Receptors

Aminopyridines are known to inhibit acetylcholine-activated K+ current (IKACh) in cardiac myocytes. The aim of this study was to examine the effect of 2-aminopyridine sulfonylcarbamide derivative 2-AP27 on isoprenaline-stimulated L-type Ca2+ current (ICaL) and to identify whether 2-AP27 acts via blocking of muscarinic M2-receptors in frog cardiomyocytes. The whole-cell configuration of the patch-clamp technique was used to record ICaL in enzymatically isolated cardiac myocytes. Isoprenaline (0.1 μM), an agonist of β1-β2-adrenoreceptors, stimulated the ICaL up to 475±61% (n=4) (P<0.05) vs. control. Then, in the first series of experiments, carbachol (0.01 μM), an agonist of M2 muscarinic receptors, reduced the stimulatory effect of isoprenaline to 42±15% vs. isoprenaline alone. 2- AP27 (100 μM) alone completely abolished the inhibitory effect of carbachol on isoprenaline-stimulated ICaL, which recovered to 95±5.8% of the effect of isoprenaline. In the second series of experiments, adenosine (1 μM), an agonist of A1-adenosine receptors, reduced the stimulatory effect of isoprenaline on ICaL to 56±10% (n=3) (P<0.05). Then 2-AP27 (100 μM) applied in the presence of adenosine, had no effect on ICaL, which remained at 51±7.9% (n=3) (P<0.05) of the effect of isoprenaline. These results suggest that 2-AP27, a new derivative of 2-AP, containing 4-toluolsulfonylcarbamide instead of amino group and quaternizated nitrogen by 4-nitrobenzylbromide in pyridine ring, is acting as an antagonist of muscarinic M2 receptors in frog ventricular myocytes.

Inhibition of transient outward K+ current by DHP Ca2+ antagonists and agonists in rabbit cardiac myocytes

1991 ◽  
Vol 260 (5) ◽  
pp. H1737-H1742 ◽  
Author(s):  
Y. Gotoh ◽  
Y. Imaizumi ◽  
M. Watanabe ◽  
E. F. Shibata ◽  
R. B. Clark ◽  
...  
Keyword(s):  
Cardiac Myocytes ◽  
Rank Order ◽  
Inhibitory Effect ◽  
Bay K 8644 ◽  
Voltage Dependent ◽  
Intracellular Ca ◽  
K Current ◽  
Rabbit Atrium

The 1,4-dihydropyridine (DHP) Ca2+ antagonists and agonists can inhibit a time- and voltage-dependent, but intracellular Ca(2+)-independent transient outward K+ current (It), in myocytes from rabbit atrium. In the presence of 0.3 mM CdCl2, DHPs decreased the peak It slightly and markedly accelerated its apparent rate of inactivation. When the inhibition of It was measured from integrated It records, the 50% inhibitory concentrations (IC50) of nicardipine and BAY K 8644 were 630 nM and 7 microM, respectively, and the IC50 of nicardipine for inhibition of the Ca2+ current (ICa) was only approximately fourfold lower (160 nM). The inhibition of It by nicardipine was not affected by changing holding potential from -55 to -100 mV; in contrast, the inhibitory effect on ICa was significantly reduced by this hyperpolarization. We conclude that the DHP Ca2+ antagonist nicardipine blocks It at similar doses to those that block ICa and that nicardipine blocks this K+ current by mechanism different from that for ICa inhibition. This inhibitory effect on It is shared by other DHP compounds; the rank order for potency of It inhibition is nicardipine greater than benidipine greater than nisoldipine greater than BAY K 8644 greater than nitrendipine greater than nifedipine.

scholarly journals cADP ribose and [Ca2+]iregulation in rat cardiac myocytes

2000 ◽  
Vol 279 (4) ◽  
pp. H1482-H1489 ◽  
Author(s):  
Y. S. Prakash ◽  
Mathur S. Kannan ◽  
Timothy F. Walseth ◽  
Gary C. Sieck
Keyword(s):  
Electrical Stimulation ◽  
Sarcoplasmic Reticulum ◽  
Confocal Microscopy ◽  
Real Time ◽  
Cardiac Myocytes ◽  
Ryanodine Receptor ◽  
Ventricular Myocytes ◽  
Adult Rat ◽  
Intracellular Ca ◽  
Rat Cardiac Myocytes

cADP ribose (cADPR)-induced intracellular Ca2+ concentration ([Ca2+]i) responses were assessed in acutely dissociated adult rat ventricular myocytes using real-time confocal microscopy. In quiescent single myocytes, injection of cADPR (0.1–10 μM) induced sustained, concentration-dependent [Ca2+]i responses ranging from 50 to 500 nM, which were completely inhibited by 20 μM 8-amino-cADPR, a specific blocker of the cADPR receptor. In myocytes displaying spontaneous [Ca2+]i waves, increasing concentrations of cADPR increased wave frequency up to ∼250% of control. In electrically paced myocytes (0.5 Hz, 5-ms duration), cADPR increased the amplitude of [Ca2+]i transients in a concentration-dependent fashion, up to 150% of control. Administration of 8-amino-cADPR inhibited both spontaneous waves as well as [Ca2+]i responses to electrical stimulation, even in the absence of exogenous cADPR. However, subsequent [Ca2+]i responses to 5 mM caffeine were only partially inhibited by 8-amino-cADPR. In contrast, even under conditions where ryanodine receptor (RyR) channels were blocked with ryanodine, high cADPR concentrations still induced an [Ca2+]i response. These results indicate that in cardiac myocytes, cADPR induces Ca2+ release from the sarcoplasmic reticulum through both RyR channels and via mechanisms independent of RyR channels.

scholarly journals Effects of β-adrenoceptor stimulation on contractility, [Ca2+]i, and Ca2+ current in diabetic rat cardiomyocytes

1998 ◽  
Vol 274 (6) ◽  
pp. H1849-H1857 ◽  
Author(s):  
Atsushi Tamada ◽  
Yuichi Hattori ◽  
Hideki Houzen ◽  
Yoichi Yamada ◽  
Ichiro Sakuma ◽  
...  
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Cardiac Myocytes ◽  
Ventricular Myocytes ◽  
Diabetic Rat ◽  
Inotropic Response ◽  
Dibutyryl Camp ◽  
Rat Cardiomyocytes ◽  
Adrenergic Response ◽  
Significant Difference ◽  
Intracellular Ca

The mechanism of the diminished inotropic response to β-adrenoceptor stimulation in diabetic hearts was studied in enzymatically isolated diabetic rat ventricular myocytes in comparison with age-matched controls. The increases in contractions and intracellular Ca2+ concentration ([Ca2+]i) transients produced by isoproterenol were markedly diminished in diabetic myocytes. The inotropic and [Ca2+]iresponses to forskolin and dibutyryl cAMP (DBcAMP) were also reduced. No significant difference was found in the stimulating effects of isoproterenol, forskolin, and DBcAMP on the L-type Ca2+ current ( I Ca) between control and diabetic myocytes. The rise of [Ca2+]iin response to rapid caffeine application, an index of sarcoplasmic reticulum (SR) Ca2+ content, was significantly decreased in diabetic myocytes. Isoproterenol, forskolin, and DBcAMP enhanced this [Ca2+]iresponse to caffeine in control myocytes more markedly than in diabetic myocytes. The changes in the isoproterenol responses observed in diabetic myocytes were prevented by insulin therapy. We conclude that 1) diabetes causes an impairment of the contractile and [Ca2+]iresponses of cardiac myocytes when stimulated at both β-adrenoceptors and the postreceptor level without affecting the I Ca response and 2) altered SR functions of uptake and/or release of Ca2+ may primarily contribute to the diminished β-adrenergic response.

Myofilament response to Ca2+ and Na+/H+ exchanger activity in sex hormone-related protection of cardiac myocytes from deactivation in hypercapnic acidosis

2007 ◽  
Vol 292 (2) ◽  
pp. R837-R843 ◽  
Author(s):  
Tepmanas Bupha-Intr ◽  
Jonggonnee Wattanapermpool ◽  
James R. Peña ◽  
Beata M. Wolska ◽  
R. J. Solaro
Keyword(s):  
Cardiac Myocytes ◽  
Ventricular Myocytes ◽  
Specific Inhibitor ◽  
Hypercapnic Acidosis ◽  
Intact Cells ◽  
Cell Shortening ◽  
Female Sex Hormones ◽  
Ovx Rats ◽  
Intracellular Ca ◽  
Myofilament Activation

Compared to sham-operated controls, myofilaments from hearts of ovariectomized (OVX) rats demonstrate an increase in Ca2+ sensitivity with no change in maximum tension (Wattanapermpool J and Reiser PJ. Am J Physiol 277: H467–H473, 1999). To test the significance of this modification in intact cells, we compared intracellular Ca2+ transients and shortening of ventricular myocytes isolated from sham and 10-wk OVX rats. There was a decrease in the peak Ca2+ transient with prolonged 50% decay time in OVX cardiac myocytes without changes in the resting intracellular Ca2+ concentration. Percent cell shortening was also depressed, and relaxation was prolonged in cardiac myocytes from OVX rats compared with shams. Ovariectomy induced a sensitization of the myofilaments to Ca2+. Hypercapnic acidosis suppressed the shortening of OVX myocytes to a lesser extent than that detected in shams. Moreover, a larger compensatory increase in %cell shortening was obtained in OVX myocytes during prolonged acidosis. The elevated compensation in cell shortening was related to a higher amount of increase in the amplitude of the Ca2+ transient in OVX myocytes. However, these differences in Ca2+ transients and %cell shortening were no longer evident in the presence of 1 μM cariporide, a specific inhibitor of Na+/H+ exchanger type 1 (NHE1). Our results indicate that deprivation of female sex hormones modulates the intracellular Ca2+ concentration in cardiac myocytes, possibly via an increased NHE1 activity, which may act in concert with Ca2+ hypersensitivity of myofilament activation as a determinant of sex differences in cardiac function.

Subcellular properties of [Ca2+]itransients in phospholamban-deficient mouse ventricular cells

1998 ◽  
Vol 274 (5) ◽  
pp. H1800-H1811 ◽  
Author(s):  
Jörg Hüser ◽  
Donald M. Bers ◽  
Lothar A. Blatter
Keyword(s):  
Ventricular Myocytes ◽  
Marked Decrease ◽  
Regulatory Protein ◽  
Inhibitory Effect ◽  
Deficient Mouse ◽  
Wild Type ◽  
Contraction Coupling ◽  
Ventricular Cells ◽  
Pump Rate ◽  
Intracellular Ca

The regulatory protein phospholamban exerts a physiological inhibitory effect on the sarcoplasmic reticulum (SR) Ca2+ pump that is relieved with phosphorylation. We have studied the subcellular properties of intracellular Ca2+([Ca2+]i) transients in ventricular myocytes isolated from wild-type (WT) and phospholamban-deficient (PLB-KO) mice. In PLB-KO myocytes, steady-state twitch [Ca2+]itransients revealed an accelerated relaxation and the occurrence of highly localized failures of Ca2+release. The acceleration of SR Ca2+ uptake caused an increase in SR Ca2+ load with the frequent occurrence of spontaneous [Ca2+]iwaves and Ca2+ sparks. [Ca2+]iwaves in PLB-KO cells showed a marked decrease in spatial width and more frequently appeared to abort. Local Ca2+ release events (Ca2+ sparks) were larger and more variable in amplitude and [Ca2+]ideclined faster in PLB-KO myocytes. Increased local buffering and reduction in the refractoriness of SR Ca2+ release caused by the increased SR pump rate led to an overall enhancement of local [Ca2+]igradients and inhomogeneities in the [Ca2+]idistribution during spontaneous Ca2+ release, [Ca2+]iwaves, and excitation-contraction coupling.

Muscarinic Receptors Differentially Modulate the Persistent Potassium Current in Striatal Spiny Projection Neurons

1999 ◽  
Vol 81 (3) ◽  
pp. 1418-1423 ◽  
Author(s):  
Lisa A. Gabel ◽  
Eric S. Nisenbaum
Keyword(s):  
Muscarinic Receptor ◽  
Muscarinic Receptors ◽  
Potassium Current ◽  
Critical Role ◽  
Neuron Activity ◽  
Projection Neurons ◽  
Maximal Conductance ◽  
Negative Shift ◽  
Receptor Modulation ◽  
Spiny Neurons

Muscarinic receptors differentially modulate the persistent potassium current in striatal spiny projection neurons. Cholinergic regulation of striatal spiny projection neuron activity is predominantly mediated through muscarinic receptor modulation of several subclasses of ion channels. Because of its critical role in governing the recurring episodes of hyperpolarization and depolarization characteristic of spiny neurons in vivo, the 4-aminopyridine–resistant, persistent potassium (K+) current, I Krp, would be a strategic target for modulation. The present results show that I Krp can be either suppressed or enhanced by muscarinic receptor stimulation. Biophysical analysis demonstrated that the depression of I Krp was associated with a hyperpolarizing shift in the voltage dependence of inactivation and a reduction in maximal conductance. By contrast, the enhancement of I Krp was linked to hyperpolarizing shifts in both activation and inactivation voltage dependencies. Viewed in the context of the natural activity of spiny neurons, muscarinic depression of I Krp should uniformly increase excitability in both hyperpolarized and depolarized states. In the hyperpolarized state, the reduction in maximal conductance should bolster the efficacy of impending excitatory input. Likewise, in the depolarized state, the decreased availability of I Krp produced by the shift in inactivation should enhance ongoing synaptic input. The alterations associated with enhancement of I Krpare predicted to have a more dynamic influence on spiny cell excitability. In the hyperpolarized state, the negative shift in activation should increase the flow of I Krp and attenuate subsequent excitatory synpatic input; whereas once the cell has traversed into the depolarized state, the negative shift in inactivation should reduce the availability of this current and diminish its influence on the existing excitatory barrage.

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