Perforated-patch recording does not enhance effect of 3-isobutyl-1-methylxanthine on cardiac calcium current

1994 ◽  
Vol 266 (6) ◽  
pp. C1619-C1627 ◽  
Author(s):  
A. Kawamura ◽  
G. M. Wahler
Keyword(s):  
Ventricular Myocytes ◽  
Cell Voltage ◽  
Rapid Decline ◽  
Response Curves ◽  
Whole Cell ◽  
Perforated Patch ◽  
Cell Responses ◽  
Whole Cell Recording ◽  
Cell Recording ◽  
Beta Adrenergic Agonist

Conventional whole cell voltage-clamp recording results in washout of the cardiac Ca2+ current (ICa) response to the beta-adrenergic agonist isoproterenol (Iso), for reasons which are not clear. When dose-response curves for the phosphodiesterase (PDE) inhibitor 3-isobutyl-1-methylxanthine (IBMX) were compared using perforated-patch vs. conventional whole cell recording in guinea pig ventricular myocytes, the conventional whole cell IBMX responses were unexpectedly larger than the perforated-patch responses. Furthermore, during conventional whole cell recording the response to repeated application of Iso declined rapidly, whereas the IBMX response initially increased and then declined. When pipette [Ca2+] was increased to 10(-7) M, conventional whole cell responses to 300 microM IBMX and 10(-9) M Iso were identical to perforated-patch responses. Thus loss of the Iso response during conventional whole cell recording seems to not be solely due to a washout of some constituent of the adenosine 3',5'-cyclic monophosphate pathway. We suggest that unphysiological intracellular [Ca2+] enhances the relative PDE activity and that this contributes to the rapid decline of the Iso response and the initial enhancement of the IBMX response.

scholarly journals Muscarinic activation of ionic currents measured by a new whole-cell recording method.

1988 ◽  
Vol 92 (2) ◽  
pp. 145-159 ◽  
Author(s):  
R Horn ◽  
A Marty
Keyword(s):  
Membrane Conductance ◽  
Ionic Currents ◽  
Response Curves ◽  
Whole Cell ◽  
Whole Cell Recording ◽  
Muscarinic Response ◽  
Cell Recording ◽  
K Current ◽  
A Cell ◽  
K Currents

A new method is described as an alternative to whole-cell recording in order to prevent "wash-out" of the muscarinic response to acetylcholine (ACh) in rat lacrimal gland cells. The membrane of a cell-attached patch is permeabilized by nystatin in the patch pipette, thus providing electrical continuity between the pipette and the cytoplasm of the cell without the loss or alteration of cytoplasmic compounds necessary for the maintenance of the response to ACh. With normal whole-cell recording in these cells, the response to ACh, seen as the activation of Ca-activated K and Cl currents, lasts for approximately 5 min. With the nystatin method, the response is not diminished after 1 h. Nystatin, applied extracellularly, is shown to cause a rapid and reversible increase of membrane conductance to cations. In the absence of wash-out, we were able to obtain dose-response curves for the effect of ACh on Ca-activated K currents. An increase of [ACh] caused an increase in the K current, with apparent saturation at concentrations above approximately 1 microM ACh. The delay between ACh application and the activation of K current was inversely related to [ACh] and reached a minimum value of 0.7-1.0 s at high [ACh].

α1-Adrenergic activation of L-type Ca current in rat ventricular myocytes: perforated patch-clamp recordings

1998 ◽  
Vol 274 (6) ◽  
pp. H2203-H2207 ◽  
Author(s):  
Shi J. Liu ◽  
Richard H. Kennedy
Keyword(s):  
Ventricular Myocytes ◽  
Cell Voltage ◽  
Adrenergic Stimulation ◽  
Whole Cell ◽  
Rat Ventricular Myocytes ◽  
Perforated Patch ◽  
Adult Rat ◽  
Transient Decrease ◽  
Adrenergic Antagonist ◽  
Patch Configuration

α1-Adrenergic stimulation has little effect on L-type Ca2+channel current ( I Ca,L) in adult cardiac myocytes measured using conventional whole cell voltage-clamp techniques. In this study using perforated-patch techniques, we reevaluated the effect of α1-adrenergic stimulation on I Ca,L in adult rat ventricular myocytes. Action potentials and I Ca,L were examined in the presence of 1 μM nadolol, a β-adrenergic antagonist, in myocytes internally dialyzed with Na+- and K+-free solutions (Cs+ and tetraethylammonium as substitutes). Phenylephrine (PE; 30 μM) increased the action potential duration measured at 25 and 70% of repolarization by 104 and 86%, respectively. In the perforated-patch configuration, PE elicited a transient decrease followed by a ∼60% increase in I Ca,L, whereas only the transient decrease in I Ca,L was observed in myocytes when the conventional whole cell configuration was used. The PE-induced increase in I Ca,L was reversibly blocked by 1 μM prazosin, an α1-adrenergic antagonist. These results suggest that α1-adrenergic stimulation enhances cardiac I Ca,L and that obligatory intracellular mediators for this action are lost during whole cell recordings.

Antagonistic Modulation of a Hyperpolarization-Activated Cl– Current in Aplysia Sensory Neurons by SCPB and FMRFamide

2003 ◽  
Vol 90 (2) ◽  
pp. 586-598 ◽  
Author(s):  
Ned Buttner ◽  
Steven A. Siegelbaum
Keyword(s):  
Sensory Neurons ◽  
Reversal Potential ◽  
Outward Current ◽  
Cell Voltage ◽  
High Concentration ◽  
Pipette Solution ◽  
Whole Cell ◽  
Whole Cell Recording ◽  
Type K ◽  
Cell Recording

Whole cell voltage-clamp recordings from Aplysia mechanosensory neurons obtained from the pleural ganglion were used to investigate the actions on membrane currents of the neuropeptides SCPB and FMRFamide. At the start of whole cell recording, SCPB typically evoked an inward current at a holding potential of –40 mV, due to the cAMP-mediated closure of the S-type K+ channel, whereas FMRFamide evoked an outward current, due to the opening of the S-type K+ channels mediated by 12-lipoxygenase metabolites of arachidonic acid. However, after several minutes of whole cell recording with a high concentration of chloride in the whole cell patch pipette solution, the responses to SCPB and FMRF-amide at –40 mV were inverted; SCPB evoked an outward current, whereas FMRFamide and YGGFMRFamide evoked inward currents. Ion substitution experiments and reversal potential measurements revealed that these responses were due to the opposing regulation of a Cl– current, whose magnitude was greatly enhanced by dialysis with the high Cl–-containing pipette solution. SCPB inhibited this Cl– current through production of cAMP and activation of PKA. YGGFMRFamide activated this Cl– current by stimulating a cGMP-activated phosphodiesterase that hydrolyzed cAMP. Thus a cAMP-dependent Cl– current undergoes antagonistic modulation by two neuropeptides in Aplysia sensory neurons.

Relationship between changes of glomus cell current and neural response of rat carotid body

1995 ◽  
Vol 74 (5) ◽  
pp. 2077-2086 ◽  
Author(s):  
P. M. Cheng ◽  
D. F. Donnelly
Keyword(s):  
Carotid Body ◽  
Outward Current ◽  
Membrane Resistance ◽  
Glomus Cell ◽  
Nerve Activity ◽  
Whole Cell ◽  
Glomus Cells ◽  
Perforated Patch ◽  
Whole Cell Recording ◽  
Cell Recording

1. Mature rat carotid bodies were harvested and sinus nerve activity was recorded in vitro during superfusion with Ringer saline. Membrane currents of glomus cells were simultaneously recorded using conventional whole cell or perforated-patch whole cell recording. Presumptive glomus cells were identified by the presence of a rapidly activated, voltage-dependent outward current above a threshold of -20 mV. 2. Outward current of presumptive glomus cells was inhibited by tetraethylammonium chloride (TEA) (20 mM) and by verapamil (5-10 microM), consistent with previous studies in which isolated glomus cells were used. Somal capacitance, calculated from the current transient following a step hyperpolarization, was 7.47 +/- 0.54 (SE) pF (n = 52). Membrane resistance for perforated-patch recordings was 820 +/- 187 M omega. 3. In perforated-patch recordings, brief periods of hypoxia (30-45 s) caused a marked increase in nerve activity to 21.6 +/- 2.7 times baseline spiking frequency (n = 59) but no significant change in membrane resistance or outward current. No change in holding current was detected, although the low amplifier gain precluded high-resolution measurement. Similar results were obtained using conventional whole cell recording, except that outward current significantly decreased during hypoxia but failed to recover in the immediate posthypoxia period. 4. TEA (20 mM) rapidly inhibited outward current to 55 +/- 7% (n = 15) of predrug current, but nerve activity only slightly increased to 2.0 +/- 0.3 times baseline spike frequency (n = 15). Brief anoxia (40 s in duration) in the presence of TEA evoked a brisk increase in nerve activity to 30 +/- 13 times baseline frequency (n = 3), demonstrating that organ function was not blocked by TEA. 5. Charybdotoxin (10 nM) significantly reduced outward current by 12.1 +/- 3.0% (n = 11) but did not significantly alter nerve activity, holding current, or membrane resistance. Apamin (100 nM) did not significantly affect nerve activity, membrane resistance, or holding current. Outward current decreased by 11.4 +/- 6.1% (n = 13). 6. These results show a dissociation between changes in glomus cell voltage-gated outward currents and changes in afferent nerve activity. This suggests that modulation of glomus cell K+ current by hypoxia is not the primary step in initiating the nerve response to hypoxia in the rat carotid body.

scholarly journals Altered Na/Ca exchange distribution in ventricular myocytes from failing hearts

2016 ◽  
Vol 310 (2) ◽  
pp. H262-H268 ◽  
Author(s):  
Hanne C. Gadeberg ◽  
Simon M. Bryant ◽  
Andrew F. James ◽  
Clive H. Orchard
Keyword(s):  
Heart Failure ◽  
Ventricular Myocytes ◽  
Cell Voltage ◽  
Coronary Artery Ligation ◽  
Sham Operation ◽  
Ca Current ◽  
Artery Ligation ◽  
Whole Cell ◽  
Rat Hearts ◽  
T Tubules

In mammalian cardiac ventricular myocytes, Ca efflux via Na/Ca exchange (NCX) occurs predominantly at T tubules. Heart failure is associated with disrupted t-tubular structure, but its effect on t-tubular function is less clear. We therefore investigated t-tubular NCX activity in ventricular myocytes isolated from rat hearts ∼18 wk after coronary artery ligation (CAL) or corresponding sham operation (Sham). NCX current ( INCX) and l-type Ca current ( ICa) were recorded using the whole cell, voltage-clamp technique in intact and detubulated (DT) myocytes; intracellular free Ca concentration ([Ca]i) was monitored simultaneously using fluo-4. INCX was activated and measured during application of caffeine to release Ca from sarcoplasmic reticulum (SR). Whole cell INCX was not significantly different in Sham and CAL myocytes and occurred predominantly in the T tubules in Sham myocytes. CAL was associated with redistribution of INCX and ICa away from the T tubules to the cell surface and an increase in t-tubular INCX/ ICa density from 0.12 in Sham to 0.30 in CAL myocytes. The decrease in t-tubular INCX in CAL myocytes was accompanied by an increase in the fraction of Ca sequestered by SR. However, SR Ca content was not significantly different in Sham, Sham DT, and CAL myocytes but was significantly increased by DT of CAL myocytes. In Sham myocytes, there was hysteresis between INCX and [Ca]i, which was absent in DT Sham but present in CAL and DT CAL myocytes. These data suggest altered distribution of NCX in CAL myocytes.

Whole cell recording from neurons in slices of reptilian and mammalian cerebral cortex

1989 ◽  
Vol 30 (3) ◽  
pp. 203-210 ◽  
Author(s):  
Mark G. Blanton ◽  
Joseph J. Lo Turco ◽  
Arnold R. Kriegstein
Keyword(s):  
Cerebral Cortex ◽  
Whole Cell ◽  
Whole Cell Recording ◽  
Cell Recording
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