scholarly journals Activation of Ca2+-activated Cl- current by depolarizing steps in rabbit urethral interstitial cells

2003 ◽  
Vol 285 (2) ◽  
pp. C327-C333 ◽  
Author(s):  
M. A. Hollywood ◽  
G. P. Sergeant ◽  
N. G. McHale ◽  
K. D. Thornbury
Keyword(s):  
Amphotericin B ◽  
Cyclopiazonic Acid ◽  
Interstitial Cells ◽  
Bath Solution ◽  
Perforated Patch ◽  
Inositol 1,4,5 Trisphosphate ◽  
Rabbit Urethra ◽  
Patch Technique ◽  
Aminoethoxydiphenyl Borate

Interstitial cells were isolated from strips of rabbit urethra for study using the amphotericin B perforated-patch technique. Depolarizing steps to -30 mV or greater activated a Ca2+ current ( ICa), followed by a Ca2+-activated Cl- current, and, on stepping back to -80 mV, large Cl- tail currents were observed. Both currents were abolished when the cells were superfused with Ca2+-free bath solution, suggesting that Ca2+ influx was necessary for activation of the Cl- current. The Cl- current was also abolished when Ba2+ was substituted for Ca2+ in the bath or the cell was dialyzed with EGTA (2 mM). The Cl- current was also reduced by cyclopiazonic acid, ryanodine, 2-aminoethoxydiphenyl borate (2-APB), and xestospongin C, suggesting that Ca2+-induced Ca2+ release (CICR) involving both ryanodine and inositol 1,4,5-trisphosphate receptors contributes to its activation.

scholarly journals Characterization of norepinephrine-evoked inward currents in interstitial cells isolated from the rabbit urethra

2002 ◽  
Vol 283 (3) ◽  
pp. C885-C894 ◽  
Author(s):  
G. P. Sergeant ◽  
K. D. Thornbury ◽  
N. G. McHale ◽  
M. A. Hollywood
Keyword(s):  
Reversal Potential ◽  
Cyclopiazonic Acid ◽  
Interstitial Cells ◽  
Equilibrium Potential ◽  
Inward Currents ◽  
Rabbit Urethra ◽  
Patch Technique ◽  
Dependent Mechanism ◽  
Stimulation Of

Freshly dispersed interstitial cells from the rabbit urethra were studied by using the perforated-patch technique. When cells were voltage clamped at −60 mV and exposed to 10 μM norepinephrine (NE) at 80-s intervals, either large single inward currents or a series of oscillatory inward currents of diminishing amplitude were evoked. These currents were blocked by either phentolamine (1 μM) or prazosin (1 μM), suggesting that the effects of NE were mediated via α1-adrenoceptors. NE-evoked currents were depressed by the blockers of Ca2+-activated Cl− currents, niflumic acid (10 μM), and 9-anthracenecarboxylic acid (9-AC, 1 mM). The reversal potential of the above currents changed in a predictable manner when the Cl− equilibrium potential was altered, again suggesting that they were due to activation of a Cl−conductance. NE-evoked currents were decreased by 10 μM cyclopiazonic acid, suggesting that they were dependent on store-released Ca2+. Inhibition of NE-evoked currents by the phospholipase C inhibitor 2-nitro-4-carboxyphenyl- N, N-diphenylcarbamate (100 μM) suggested that NE releases Ca2+ via an inositol 1,4,5-trisphosphate (IP3)-dependent mechanism. These results support the idea that stimulation of α1-adrenoceptors releases Ca2+ from an IP3-sensitive store, which in turn activates Ca2+-activated Cl− current in freshly dispersed interstitial cells of the rabbit urethra. This elevates slow wave frequency in these cells and may underlie the mechanism responsible for increased urethral tone during nerve stimulation.

scholarly journals Role of IP3 in modulation of spontaneous activity in pacemaker cells of rabbit urethra

2001 ◽  
Vol 280 (5) ◽  
pp. C1349-C1356 ◽  
Author(s):  
G. P. Sergeant ◽  
M. A. Hollywood ◽  
K. D. McCloskey ◽  
N. G. McHale ◽  
K. D. Thornbury
Keyword(s):  
Cyclopiazonic Acid ◽  
Pacemaker Activity ◽  
Pacemaker Cells ◽  
Outward Currents ◽  
Inositol 1,4,5 Trisphosphate ◽  
Inward Currents ◽  
Rabbit Urethra ◽  
Patch Technique ◽  
Spontaneous Transient Outward Currents

Isolated interstitial (“pacemaker”) cells from rabbit urethra were examined using the perforated-patch technique. Under voltage clamp at −60 mV, these cells fired large spontaneous transient inward currents (STICs), averaging −860 pA and >1 s in duration, which could account for urethral pacemaker activity. Spontaneous transient outward currents (STOCs) were also observed and fell into two categories, “fast” (<100 ms in duration) and “slow” (>1 s in duration). The latter were coupled to STICs, suggesting that they shared the same mechanism, while the former occurred independently at faster rates. All of these currents were abolished by cyclopiazonic acid, caffeine, or ryanodine, suggesting that they were activated by Ca2+ release. Whend- myo-inositol 1,4,5-trisphosphate (IP3)-sensitive stores were blocked with 2-aminoethoxydiphenyl borate, the STICs and slow STOCs were abolished, but the fast STOCs remained. In contrast, the fast STOCs were more nifedipine sensitive than the STICs or the slow STOCs. These results suggest that while fast STOCs are mediated by a mechanism similar to STOCs in smooth muscle, STICs and slow STOCs are driven by IP3. These results support the hypothesis that pacemaker activity in the urethra is driven by the IP3-sensitive store.

Differential modulation of caffeine- and IP3-induced calcium release in cultured arterial tissue

1999 ◽  
Vol 276 (5) ◽  
pp. C1115-C1120 ◽  
Author(s):  
Karl Dreja ◽  
Per Hellstrand
Keyword(s):  
Calcium Release ◽  
Cyclopiazonic Acid ◽  
Differential Modulation ◽  
Transient Responses ◽  
Inositol 1,4,5 Trisphosphate ◽  
Arterial Tissue ◽  
Intracellular Ca ◽  
Rat Tail

To investigate the Ca2+-dependent plasticity of sarcoplasmic reticulum (SR) function in vascular smooth muscle, transient responses to agents releasing intracellular Ca2+ by either ryanodine (caffeine) ord- myo-inositol 1,4,5-trisphosphate [IP3; produced in response to norepinephrine (NE), 5-hydroxytryptamine (5-HT), arginine vasopressin (AVP)] receptors in rat tail arterial rings were evaluated after 4 days of organ culture. Force transients induced by all agents were increased compared with those induced in fresh rings. Stimulation by 10% FCS during culture further potentiated the force and Ca2+ responses to caffeine (20 mM) but not to NE (10 μM), 5-HT (10 μM), or AVP (0.1 μM). The effect was persistent, and SR capacity was not altered after reversible depletion of stores with cyclopiazonic acid. The effects of serum could be mimicked by culture in depolarizing medium (30 mM K+) and blocked by the addition of verapamil (1 μM) or EGTA (1 mM) to the medium, lowering intracellular Ca2+ concentration ([Ca2+]i) during culture. These results show that modulation of SR function can occur in vitro by a mechanism dependent on long-term levels of basal [Ca2+]iand involving ryanodine- but not IP3 receptor-mediated Ca2+release.

Neuropeptide galanin inhibits omega-conotoxin GVIA-sensitive calcium channels in parasympathetic neurons

1995 ◽  
Vol 73 (4) ◽  
pp. 1374-1382 ◽  
Author(s):  
L. A. Merriam ◽  
R. L. Parsons
Keyword(s):  
Calcium Channels ◽  
Patch Clamp Technique ◽  
Maximal Inhibition ◽  
Whole Cell ◽  
Recording Technique ◽  
Perforated Patch ◽  
Cell Recording ◽  
Parasympathetic Neurons ◽  
Current Activation ◽  
Patch Technique

1. We determined the effect of the neuropeptide galanin on barium currents (IBa) flowing through voltage-gated calcium channels. We voltage clamped parasympathetic neurons dissociated from mudpuppy cardiac ganglia using both the standard whole cell and the perforated-patch variations of the patch-clamp technique. 2. Galanin produced a concentration-dependent inhibition of IBa. The maximal inhibition was 50-60% and the concentration that produced half-maximal inhibition (IC50) was 0.42 nM. In mud-puppy parasympathetic neurons, omega-conotoxin-GVIA (CTX)-sensitive channels are the predominant type of calcium channels, and only a small portion of IBa is contributed by dihydropyridine-sensitive channels. Galanin preferentially inhibited a portion of the CTX-sensitive current. 3. In currents recorded with the standard whole cell technique, activation of IBa was slowed in the presence of galanin. In contrast, in the majority of neurons studied with the perforated-patch technique, galanin decreased IBa without altering the kinetics of current activation. With both recording methods, the decrease in IBa was greatest with voltage steps to 0 mV and persisted with steps to +50 mV. For control currents, large depolarizing voltage steps (+70 to +120 mV) did not markedly facilitate IBa when either recording technique was used. However, the degree of facilitation in galanin was significantly greater with the standard whole cell recording technique. 4. IBa exhibited inactivation under the conditions of these experiments. Inactivation of IBa recorded during a 900-ms depolarizing voltage step was fitted to a double exponential. Galanin decreased the amplitude of IBa but did not alter the time constants of inactivation.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Inositol 1,4,5-trisphosphate–induced Calcium Release Is Necessary for Generating the Entire Light Response of Limulus Ventral Photoreceptors

2003 ◽  
Vol 121 (5) ◽  
pp. 441-449 ◽  
Author(s):  
Alan Fein
Keyword(s):  
Calcium Release ◽  
Cyclopiazonic Acid ◽  
Light Response ◽  
Calcium Stores ◽  
Ip3 Receptor ◽  
Treatment Results ◽  
Light Responses ◽  
Calcium Buffer ◽  
Pressure Injection ◽  
Inositol 1,4,5 Trisphosphate

The experiments reported here were designed to answer the question of whether inositol 1,4,5-trisphosphate (IP3)-induced calcium release is necessary for generating the entire light response of Limulus ventral photoreceptors. For this purpose the membrane-permeable IP3 receptor antagonist 2-aminoethoxydiphenyl borate (2APB) (Maruyama, T., T. Kanaji, S. Nakade, T. Kanno, and K. Mikoshiba. 1997. J. Biochem. (Tokyo). 122:498–505) was used. Previously, 2APB was found to inhibit the light activated current of Limulus ventral photoreceptors and reversibly inhibit both light and IP3 induced calcium release as well as the current activated by pressure injection of calcium into the light sensitive lobe of the photoreceptor (Wang, Y., M. Deshpande, and R. Payne. 2002. Cell Calcium. 32:209). In this study 2APB was found to inhibit the response to a flash of light at all light intensities and to inhibit the entire light response to a step of light, that is, both the initial transient and the steady-state components of the response to a step of light were inhibited. The light response in cells injected with the calcium buffer 1,2-bis(o-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid (BAPTA) was reversibly inhibited by 2APB, indicating that these light responses result from IP3-mediated calcium release giving rise to an increase in Cai. The light response obtained from cells after treatment with 100 μM cyclopiazonic acid (CPA), which acts to empty intracellular calcium stores, was reversibly inhibited by 2APB, indicating that the light response after CPA treatment results from IP3-mediated calcium release and a consequent rise in Cai. Together these findings imply that IP3-induced calcium release is necessary for generating the entire light response of Limulus ventral photoreceptors.

scholarly journals Calcium oscillations in interstitial cells of the rabbit urethra

2005 ◽  
Vol 565 (2) ◽  
pp. 449-461 ◽  
Author(s):  
L. Johnston ◽  
G. P. Sergeant ◽  
M. A. Hollywood ◽  
K. D. Thornbury ◽  
N. G. McHale
Keyword(s):  
Interstitial Cells ◽  
Calcium Oscillations ◽  
Rabbit Urethra

Gramicidin perforated patch-clamp technique reveals glycine-gated outward chloride current in dissociated nucleus solitarii neurons of the rat

1994 ◽  
Vol 72 (3) ◽  
pp. 1103-1108 ◽  
Author(s):  
J. S. Rhee ◽  
S. Ebihara ◽  
N. Akaike
Keyword(s):  
Patch Clamp ◽  
Hill Coefficient ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Concentration Dependent Manner ◽  
Perforated Patch ◽  
Clamp Technique ◽  
Outward Currents ◽  
Patch Technique ◽  
Perforated Patch Clamp

1. The inhibitory response of exogenously applied glycine was investigated in freshly dissociated rat nucleus tractus solitarii neurons under whole cell configuration using new perforated patch-clamp technique termed "gramicidin perforated patch technique," which maintains intact intracellular Cl- concentrations. 2. Using the gramicidin perforated patch technique, at a holding potential (VH) of -45 mV, glycine induced outward currents in a concentration-dependent manner with a EC50 of 4.0 x 10(-5) M and at a Hill coefficient of 1.5. In contrast, using the nystatin perforated patch technique, glycine induced inward currents at the same VH in a concentration-dependent manner with an EC50 of 4.9 x 10(-5) M and at a Hill coefficient of 1.2. 3. The glycine-induced outward currents were blocked by strychnine in a concentration dependent manner with an IC50 of 2.2 x 10(-8) M. The blockade was competitive. 4. The current-voltage relationship for the 10(-5) M glycine response showed a clear outward rectification. 5. Ten-fold change of extracellular Cl- with a large impermeable anion resulted in a 65 mV shift of the reversal potential of glycine-induced currents (EGly), indicating that the membrane behaves like a Cl- electrode in the presence of glycine. 6. The intracellular Cl- activity calculated from the EGly ranged from 7.3 to 18.2 mM, with a mean value of 13.3 mM. 7. The values of EGly in the individual neurons were significantly negative to the resting membrane potentials, suggesting the existence of active transport of Cl-.

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