| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Journal of Physiology (2002), 544.1, pp. 19-27
© Copyright 2002 The Physiological Society
DOI: 10.1113/jphysiol.2001.013557
 |
ABSTRACT |
|---|
 TopAbstract IntroductionMethodsResultsDiscussionReferences |
|---|
The objective for this paper was to characterize the transient outward current (Ito) present in smooth muscle cells of the intestinal external muscularis. Two populations of cells were identified, one with a fast rate of Ito inactivation (< 100 ms) and another with a slow rate of Ito inactivating (
(Resubmitted 13 November 2001; accepted after revision 7 June 2002; first published online 2 August 2002)
Corresponding author J. D. Huizinga: McMaster University, HSC-3N5C, 1200 Main Street West, Hamilton, ON L8N 3Z5, Canada. Email: huizinga{at}mcmaster.ca
|
INTRODUCTION |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
A transient potassium outward current was first identified by Connor & Stevens (1971) in neurons of the sea slug Anisodoris. This current inactivates rapidly and has been named the rapidly inactivating K+ current, or KA. According to inactivation kinetics, various types of transient K+ currents appear to exist. In some neurons, activation of these currents is fast enough to participate in the repolarization phase of the action potential. These currents might also serve as a damper in the interspike interval to space successive action potentials (Bielefeld et al. 1990). Similar transient K+ currents have been observed in smooth muscle preparations from artery (James et al. 1995), vein (Beech & Bolton, 1989), bladder (Smith et al. 1998) and intestine (Ohya et al. 1987; Hollywood et al. 2000).
In the mouse small intestine, the musculature generates two major oscillatory electrical activities: the slow wave and the superimposed action potentials (Malysz et al. 1995). The slow wave occurs at a frequency of 40-60 cycles min-1 at 37 °C with a average rate of rise of the upstroke of ~100 mV s-1. Action potentials are superimposed on the plateau of the slow waves and occur at a frequency of ~5 Hz and have an average rate of rise of ~5 V s-1. Both the slow waves and action potentials occur within the voltage activation range of transient outward currents, suggesting that transient outward currents may play a role in influencing these major electrical activities.
Our objective was to characterize transient outward currents in the smooth muscle cells from the mouse small intestine as part of our general aim to elucidate the cellular and electrophysiological basis of excitation in this tissue (Huizinga et al. 1997). Our observations indicate the presence of at least two types of transient outward currents with different kinetic properties, occurring in different cells. These outward currents will take part in shaping the slow waves and action potentials of the intestinal musculature.
|
METHODS |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
Isolation of single smooth muscle cells
Adult female mice (20-25 g) were killed by cervical dislocation and an ileal segment was removed. The Animal Ethics Committee of McMaster University approved animal handling procedures. After washing, the segment was placed in pre-equilibrated M199 cell media and the muscularis externa was removed by separation along the deep muscular plexus margin. The dissected muscularis was cut into small pieces (~1-2 mm2) and placed for 15 min at 37 °C in dissociation solution A (see below) to which was added 1 mg ml-1 trypsin. Thereafter, the supernatant was removed and the tissue was incubated for another 30 min in fresh dissociation solution A with trypsin. The supernatant was carefully removed and the preparation was further incubated for 35 min in dissociation solution B (see below), containing 3 mg ml-1 collagenase and 1 mg ml-1 BSA. The cells were released by shaking, thereafter the cell suspension was carefully layered on top of 20 % (w/v) Ficoll and spun at 15 g for 15 min. The cell band located at the interface was removed and plated onto collagen-coated cover slips. The cells were maintained in 5 % CO2 at 37 °C until needed.
Electrophysiology
Cells were not used beyond 5 days post-isolation because stable tight seals were more difficult to obtain. The cells were continuously perfused with extracellular solution (ECS). Whole cell electrical activity was recorded using the nystatin perforated patch. This was achieved by adding 2 mg ml-1 nystatin in the intracellular solution (ICS). Pipettes were made using a Sutter micropipette puller, they had a resistance of 3-5 M
when filled with the recording solution. Electrical activity was recorded using the Axopatch 1B amplifier (Axon Instruments, CA, USA) and pCLAMP v5.2 acquisition software. Different voltage protocols were utilized to assess different kinetic aspects of whole cell currents. The sampling frequency was 4 kHz, filtering was done at 2 kHz, the cell capacitance was measured after capacitance compensation using the Axopatch 200A amplifier. Membrane potentials were corrected for the liquid junction potentials between the pipette and the external solutions.
Analysis
Data analysis was done by the Axon pCLAMP v6.1 utilities and by Microcal Origin graphing/fitting program. Fitting was done by 
2 reduction by Simplex and Levenberg-Marquardt algorithms. Goodness of fit was assessed both by 2 minimization and by visual inspection. Average values are followed by S.E.M.
Solutions, with concentrations in millimolar
ECS (extracellular solution): NaCl 125.0, KCl 5.0, MgCl2 1.2, NaH2PO4 1.2, glucose 11.0, NaHCO3 4.0, CaCl2 2.0, Hepes 10.0, pH 7.35. ICS (intracellular solution): potassium methanesulfonate 129.0, NaCl 5.0, magnesium acetate 2.0, CaCl2 1.0, EGTA 11.0, Hepes 10.0, pH 7.25. Solution A: NaCl 134.0, KCl 3.0, Taurine 5.0, EDTA 5.0, MnCl2 2.0, Hepes 10.0, pH 7.4. Solution B: NaCl 133.0, KCl 3.0, taurine 5.0, CaCl2 0.1, MgCl2 2.0, glucose 10.0, Hepes 10.0, pH 7.35. M199 media: 1X M199 media, NaHCO3 26.0, glutamine 2.0, penicillin 0.25 mg ml-1, fetal bovine serum (FBS) 10 %.
Chemicals and drugs were purchased from Sigma-Aldrich Canada Ltd. Nifedipine (2 
10-7 M) was added as indicated. When 60 mM TEA was used, 60 mM NaCl was omitted and the pH was adjusted. Collagenase used was Sigma no. C0130, trypsin was Sigma no. T8253. Media was obtained from Gibco-Life Sciences, Canada.
|
RESULTS |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
General properties of whole cell outward currents
Outward potassium currents were recorded from 104 intestinal smooth muscle cells by using step depolarizations from -70 to +60 mV. In 49 % of the cells, a transient outward current (Ito) was recorded as a component of the total outward currents. The other outward currents which showed either slower or no inactivation were completely inhibited by the K+ channel blocker TEA (60 mM; Fig. 1A and B). The only outward current remaining after the application of TEA was Ito, an outward current that activated quickly to peak amplitude (in less than 10 ms) and inactivated to 24 ± 16 % (n = 51) of the peak currents, assessed by a test pulse of +30 mV. Nifedipine (5 10-7 M) did not affect transient outward currents (n = 18).
 |
View larger version [in this window] [in a new window] |
|
|
Figure 1. Transient outward currents isolated by 60 mM TEA Whole cell currents were evoked by a voltage step to +30 mV from a holding potential of -50 mV. With the addition of 60 mM TEA, transient outward currents were revealed. The transient outward currents were either A, fast inactivating ( | ||
The peak Ito was always prominent when the holding potential was set to -100 mV, but was small when the holding potential was set to -30 mV (Fig. 2). Therefore, to fully express Ito, a prepulse of -100 mV for at least 500 ms was applied before the test pulses from -70 to +60 mV were executed in order to remove any inactivation imposed at the typical holding potentials of -50 to -70 mV. The transient outward currents expressed using these voltage protocols had either 'fast' inactivation (Fig. 2A), in that the Ito inactivated to a steady state in less than 150 ms, or 'slow' inactivation where Ito needed greater than 150 ms to inactivate to steady state (Fig. 2B). The transient outward currents showing 'fast' inactivation were designated as 'fast Ito'. Similarly, the transient outward currents showing 'slow' inactivation were designated as 'slow Ito'. Another method to classify the transient outward current was by using a -30 mV pre-pulse instead of the -100 mV pre-pulse for the standard voltage protocol. The fast Ito completely inactivated, while the slow Ito, peak currents were still seen, indicating incomplete inactivation by the -30 mV pre-pulse.
 |
View larger version [in this window] [in a new window] |
|
|
Figure 2. Voltage dependent transient outward currents have different inactivation kinetics In the presence of TEA (60 mM), transient outward currents (Ito) were evoked. Inward Ca2+ currents were seen at the beginning of the test pulse when L-type calcium channel blockers were not applied. Because Ito shows voltage dependent inactivation, a -100 mV pre-pulse was given for 500 ms before the test pulses from -70 to +60 mV were applied, as seen for the top figures. The bottom figures were the result of changing the pre-pulse to -30 mV, and demonstrating the sensitivity of the transient outward currents to voltage dependent inactivation. A, in 56 % of cells, the transient outward current exhibited fast inactivation (peak | ||
Voltage dependent activation and inactivation of Ito
The two different transient outward currents were distinguished according to their electrophysiological properties. Fast inactivation was seen in 28 cells (55 %), and slow inactivation was seen in 23 cells (43 %) out of a total of 51 cells. Both transient outward currents were very fast activating (activation to peak current < 10 ms). Current-voltage (I-V) plots of elicited peak currents against imposed membrane voltages gave similar thresholds for the fast and slow inactivating Ito at -43 ± 13 and -45 ± 7 mV, respectively (Fig. 3). From the I-V plots, the peak cord conductances were 0.5 ± 0.1 nS pF-1 for the fast Ito, and 1.3 ± 0.1 nS pF-1 for the slow Ito. The steady-state currents had similar threshold points.
 |
View larger version [in this window] [in a new window] |
|
|
Figure 3. Current-voltage (I-V) plots of the different transient outward currents I-V curves are expressed as peak currents (Pk) as well as the sustained, steady-state currents (SS) for A, fast Ito, and B, slow Ito. Both the peak and steady-state currents showed similar thresholds for activation and can be seen to develop monophasically. The slow Ito had a higher peak chord conductance than the fast Ito (1.31 ± 0.06 vs. 0.54 ± 0.13 nS pF-1, respectively). C, a plot of the steady-state current to peak- current ratio (SS/Pk) showed that the SS/Pk ratio increased proportionally with voltage for the fast Ito but that it levelled off at ~-10 mV for the slow Ito. | ||
The relative amount of steady-state currents (SS) remaining after time dependent inactivation was compared to the peak currents (Pk) evoked at the same potentials. As shown in Fig. 3C, the SS/Pk ratios were always higher for the slow Ito. Hence, the slow Ito shows a lesser degree of inactivation than the fast Ito.
The voltage dependency of inactivation
The time dependent inactivation from peak current to steady state was best described for both populations by mono-exponential fits (Fig. 4A and B). The time constant for the inactivation, (time when 66 % of inactivation is completed), was plotted against the imposed voltages (Fig. 4C). The fast Ito had an average = 44 ms while the slow Ito had an average = 229 ms over all voltages. The wide margin between the values validates the grouping of the Ito into two populations of fast and slow inactivating currents.
 |
View larger version [in this window] [in a new window] |
|
|
Figure 4. Inactivation rates of the fast Ito and slow Ito Inactivation rates were fitted as mono-exponentials over the voltage ranges at which time dependent inactivation can be seen. Representative mono-exponential fits were superimposed on data points obtained from voltage steps from 0 mV to +60 mV for A, fast Ito and B, slow Ito. C, although the inactivation process was clearly voltage dependent, the rate of inactivation did not change significantly over the observed voltage range. The slow Ito inactivation rate ( | ||
The voltage dependency of inactivation was explored using the double pulse (pre-pulse) protocol (Fig. 5). The membrane potential was allowed to settle to steady-state inactivation before a test pulse was given in order to assess the level of inactivation that had occurred. With increasing levels of depolarization in the first pulse, the inactivation decreased the peak current according to a Boltzmann distribution. Two distinct voltage ranges for inactivation were observed (Fig. 5B and C). The values for 5 and 95 % thresholds were estimated from values derived from successful fits of the data (n = 5). The fast inactivating Ito has a half-inactivation potential (Vh) of -78 mV and slope factor (k) of 14, and started inactivating (5 % threshold) at -110 mV and completely inactivated (95 % threshold) at -36 mV (Fig. 5B). The slower inactivating Ito had a Vh of +33 mV and a k of 13, and displayed the 5 % threshold at -70 mV and the 95 % threshold at +5 mV (Fig. 5C).
 |
View larger version [in this window] [in a new window] |
|
|
Figure 5. Voltage dependent inactivation process as assessed by a double-voltage-pulse protocol A, from a holding potential of -50 mV, voltage pulses from -120 to 0 mV were given for 500 ms before the test pulse of +50 mV was applied. As seen with the currents elicited in the first pulse, the inactivation process has settled to steady state. The peak amplitude from the test pulse can be seen to decrease with increasingly positive first pulses, as expected with voltage dependent inactivation. This voltage dependent inactivation of the transient outward currents is shown as plots of normalized peak currents (I/Imax) to the corresponding voltages of the first pulse. The data were fitted with the Boltzmann equation to obtain the half-inactivation voltage (Vh) and slope factor (k). B, the fast Ito had Vh = -78 mV and k = 14. C, the slow Ito had Vh = -33 mV and k = 13. | ||
Simultaneous voltage dependent activation and inactivation of Ito
Conversion of peak currents to conductances corresponded to Boltzmann distributions with a half-activation potential (Vh) of -12.1 mV, slope factor k = 15.0 for the fast Ito, and Vh = 6.8 mV, k = 16.7 for the slower inactivating Ito. Considering the 5 and 95 % activation points to full conductance, the fast inactivating Ito started activating (5 %) at -56.3 mV and reached full conductance (95 % complete) at +32.1 mV. Similarly, the slow Ito started activating (5 %) at -45.1 mV and reached full activation (95 %) at +56.1 mV.
Both the activation and inactivation processes were plotted together against the imposed membrane potential. The degree of overlap between activation and inactivation processes determines the amount of 'window' Ito available at any membrane potential. Figure 6 shows the ranges for 'window' currents for both populations of Ito using 5 % threshold of activation and 95 % threshold for inactivation (n = 5). Since both fast and slow Ito activated in approximately the same ranges, the differences in inactivation were the main determinants of the differences in window currents. The voltage range for the fast Ito window currents were from -56 to -36 mV (Fig. 6A). The slow Ito had a larger voltage range for windows currents that extended from -42 to +5 mV (Fig. 6B). The fast Ito had little overlap between the activation and inactivation processes. At the activation threshold (-56 mV), the inactivation curve indicates 72 % inactivation. For the slow Ito, there was substantial overlap such that at threshold for activation (-45 mV), only 28 % of the total current had been inactivated. Hence, at the voltages where slow waves and action potentials occur, the slow Ito can deliver more standing current.
 |
View larger version [in this window] [in a new window] |
|
|
Figure 6. Simultaneous plots of activation and inactivation to reveal differences in the window currents Both activation and inactivation processes were normalized as conductances and expressed as ratios relative to the maximum values. Since both the fast and slow Ito show similar thresholds for activation, the main difference in the voltage behaviour between the two transient outward currents was the inactivation voltage dependency. The voltage range for window currents was expressed as the threshold for activation (5 % point) to completion of inactivation (95 % point). A, the voltage range for window currents in the fast Ito was -56 to -36 mV. B, for the slow Ito, the window currents were between -42 to +5 mV. Therefore, the slow Ito has significantly more window current relative to the fast Ito. | ||
Latencies for recovery from inactivation
To assess the time needed for recovery from inactivation, a three-pulse latency protocol was used (Fig. 7A). The first voltage pulse was used to maximally inactivate the current, followed by a hyperpolarizing pulse of varying durations to remove the inactivation, and the last pulse was used to elicit the test current in which the peak amplitude would reflect the degree of inactivation still present. A plot of the peak currents from the test pulse against the duration (or latency) spent recovering from inactivation (duration of the second pulse, 4-200 ms) revealed a mono-exponential relationship (Fig. 7B and C). The currents obtained using varying times to remove inactivation were compared to the currents obtained when a time of 200 ms was allowed to completely remove inactivation. The fast Ito recovered more quickly from inactivation with = 11 ms and was complete (95 %) at 32 ms. In contrast, the slow Ito had a = 42 ms with complete recovery (95 %) at 126 ms. Therefore, the slow Ito required approximately four times longer to recover from inactivation than the fast Ito and would be unlikely to recover completely from fast events with very high frequency (> 8 Hz).
 |
View larger version [in this window] [in a new window] |
|
|
Figure 7. The latency for recovery from inactivation as assessed by the three pulse experiment A, from a holding potential of -50 mV, the first inactivating pulse (0 mV, 500 ms) was given, followed by a second recovery pulse of -100 mV with increasing durations ( | ||
The effect of depolarization rate on Ito activation
Although the above experiments examined activation and inactivation processes in detail, they may not reflect activity during continuously changing potentials. This is important since in vivo cellular membrane potential changes are not instantaneous and step-wise, but have definite rates of depolarization. To examine how Ito would behave under different rates of change in membrane potential (dV/dt), voltage ramps with different velocities were used (Fig. 8A). Fast rates of depolarization (> 10 mV ms-1) evoked peak currents that approached the maxima, limited only by the rate of activation of Ito itself. A plot of the peak current evoked by the ramps of the same voltage ranges (-150 to +60 mV) against ramp velocity revealed a mono-exponential relationship which was different between the fast and slow Ito (Fig. 8B and C). The fast Ito demonstrated a significantly faster response to depolarization with = 2 ± 1 mV ms-1 and approached its maximum (95 %) at 5 mV ms (Fig. 8B). The response to depolarization was markedly slower with the slow Ito; it had not reached its maximum after 10 ms. Assuming a mono-exponential relationship similar to the fast Ito, the of the slow Ito was 14 ± 9 mV ms-1 with maximal activation (95 %) at 42 mV ms-1 (Fig. 8C). Therefore, the slow Ito requires very fast events (approximately nine times faster than the fast Ito) in order to be maximally activated.
 |
View larger version [in this window] [in a new window] |
|
|
Figure 8. The response of Ito to different rates of depolarization as assessed by voltage ramps A, ramps from -150 to +60 mV with increasing durations ( | ||
Since a depolarization rate of 42 mV ms-1 may not be reached in vivo, the slow Ito may not be fully activated under normal rates of depolarization. Considering the rates of depolarization of slow waves and spiking activity to be approximately 1 and 5 mV ms-1, respectively, in the mouse small intestine (J. Malysz & J. D. Huizinga unpublished data), only the fast Ito will maximally respond to spiking events while both currents will respond to slow wave type depolarizations.
Pharmacology and effects of time in culture
Both currents were revealed by their insensitivity to 60 mM TEA (n = 32; Fig. 9A). The amplitude of the slow Ito was not significantly different in the presence or absence of 2 or 5 mM 4-AP (n = 7), The presence of 4-AP reduced the fast Ito by only 15.5 ± 2.1 % (n = 8). Both transient outward currents were blocked by Cs+ (n = 7) or by 1 mM quinine or quinidine (n = 3; Fig. 9B).
 |
View larger version [in this window] [in a new window] |
|
|
Figure 9. Pharmacology of Ito A, after blocking the other outward currents with 5 mM TEA to reveal Ito, additional application with 2 mM 4-AP did not block Ito. B, after the addition of 5 mM TEA, 1 mM quinine completely abolishes Ito. | ||
Differences in electrophysiological properties could potentially be related to days in culture. Since changes in cell morphology were observed over time in culture (Lee et al. 1996), the number of days in culture were noted for each experiment. No relationship was found between kinetic properties of the transient outward currents and days in culture.
|
DISCUSSION |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
The study of the transient outward currents revealed in the presence of TEA demonstrated the existence of two cell populations, which expressed two distinct types of Ito, markedly different in their electrophysiological properties. Both transient outward currents activated in approximately the same voltage range (~-50 mV) and showed similar evoked peak currents but different chord conductances. The inactivation profiles showed the fast Ito to be inactivated at more hyperpolarized potentials than the slow Ito. The fast Ito had relatively little overlap between the activation/inactivation curves while the slow Ito had considerable overlap. Hence at a given voltage, the slow Ito current may contribute more compared to the fast Ito. The latencies for recovery from inactivation revealed that the slow Ito required approximately four times longer to recover than the fast Ito. To illustrate the importance of the differences in latencies, the fast Ito can act on depolarizing events with frequencies up to 30 Hz and still be able to recover from inactivation while the slow Ito can only express itself with frequencies up to 8 Hz. The voltage ramp analysis showed that the fast Ito generated more current in response to fast rates of depolarization and was attenuated sharply by slow rates in comparison with the slow Ito. All results are consistent with the existence of two distinct populations of transient outward currents.
Functional consequences of Ito expression
The amount of Ito that can be evoked is dependent on both the voltage and the rate of depolarization; and hence the two major electrical activities in the intestine, slow waves and action potentials, will be affected differently. We predict that the Ito in the mouse small intestine will, similar to IA, provide transient inhibition to excitatory electrical activity. From their kinetic properties, they can modulate only specific excitatory events. For example, the ramp analysis demonstrated that the Ito would be effective in inhibiting relatively fast depolarizing events. If the depolarizing event outlasts the time for inactivation, the excitatory event would only be opposed by the steady-state current remaining. The response of Ito to fast velocities of depolarization and its relatively short latencies for recovery from inactivation suggests that it could modulate the frequency of fast electrical events. Thus, in essence, the Ito provides a frequency filter such that only events that are large enough (high amplitude), fast enough (high rate of depolarization), (long duration) and frequent enough (high frequency) would overcome the inhibitory influence of Ito. Fast events with high frequency would essentially be dampened up to the point at which Ito can no longer recover from inactivation. Action potentials superimposed on slow waves can easily reach 7 Hz so that the slow Ito may indeed set the limit for this frequency.
It is probable that the fast Ito would be more involved with inhibiting action potential generation while both the fast and slow transient outward currents would influence the slow wave type depolarizations. The slow waves are special in that they have a fast upstroke and a relatively long duration of depolarization (plateau) before repolarizing. The long duration will inactivate the transient outward currents, and thus allow for easier occurrences of action potentials on the plateau of the slow waves. But, since the plateau typically occurs at -40 to -30 mV, there will still be significant contributions from 'window' currents. Therefore, transient outward currents, and especially fast Ito, will modulate the threshold for action potential generation.
Comparisons with other transient outward currents in smooth muscle
Transient outward currents have been described in other tissues, but the remarkable variability in properties makes it imperative that they be examined carefully within each tissue of interest. Within the gastrointestinal system, transient outward currents were identified in opossum oesophageal circular muscle (Akbarali et al. 1995), newborn rat ileum (Smirnov et al. 1992), guinea-pig proximal colon (Vogalis & Sanders, 1991), dog colon (Farrugia et al. 1993) and human jejunal circular muscle (Duridanova et al. 1997). These currents differed from the currents described here in that they were sensitive to 4 mM 4-AP, whereas in our preparations, 5 mM 4-AP did not affect the mouse intestinal slow Ito with minor effects on the fast Ito. Insensitivity to 4-AP was also reported for transient outward currents in guinea-pig gastric antrum (Noack et al. 1992). Most reported high concentrations (up to 10 mM) of 4-AP to block Ito (Bielefeld et al. 1990; Hisada et al. 1990), while some show selective block with micromolar concentrations of quinine (Farrugia et al. 1993; Duridanova et al. 1997). It is clear that characteristics as well as the pharmacology of transient outward current is diverse in the various tissues (Hisada et al. 1990; Imaizumi et al. 1990; McFadzean & England, 1992).
Our findings support the hypothesis that the population of smooth muscle cells of the intestinal external muscularis is heterogeneous with respect to presence of transient outward currents. At least three observations indicate that this heterogeneity exists within both muscle layers and is not due to the two muscle layers having different ion channel populations. First, there are three type of cells according to the Ito that they express, one type (~50 % of the studied cells) without any transient outward current, a second type with slow Ito, and a third type with fast Ito. Second, the ratio of the number of smooth muscle cells in the longitudinal muscle layer vs. the circular muscle layer in the mouse small intestine is 1:4 (X.-Y. Wang & J. D. Huizinga, unpublished observations), whereas the ratio of smooth muscle cells with the slow and the fast transient outward current is 1:1.2. Third, in hundreds of intracellular recordings where one of the objectives was to find differences between the smooth muscle cells in the different muscle layers, no difference in resting membrane potential or characteristics of slow waves and action potentials was found (J. Malysz & J. D. Huizinga, unpublished observations). Currently, the lack of specific inhibitors for Ito prevents further characterization of the effects of fast and slow Ito on the electrical activity of the muscle layers of the small intestine.
|
REFERENCES |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
| AKBARALI, H. I., HATAKEYAMA, N., WANG, Q. & GOYAL, R. K. (1995). Transient outward current in opossum esophageal circular muscle. American Journal of Physiology 268, G979-987 | [Medline] |
| BEECH, D. J. & BOLTON, T. B. (1989). A voltage-dependent outward current with fast kinetics in single smooth muscle cells isolated from rabbit portal vein. Journal of Physiology 412, 397-414 | [Abstract] |
| BIELEFELD, D. R., HUME, J. R. & KRIER, J. (1990). Action potentials and membrane currents of isolated single smooth muscle cells of cat and rabbit colon. Pflügers Archiv 415, 678-687 | [Medline] |
| CONNOR, J. A. & STEVENS, C. F. (1971). Voltage clamp studies of a transient outward membrane current in gastropod neural somata. Journal of Physiology 213, 21-30 | [Medline] |
| DURIDANOVA, D. B., GAGOV, H. S., DAMYANOV, D. & BOEV, K. K. (1997). Two components of potassium outward current in smooth muscle cells from the circular layer of human jejunum. General Physiology and Biophysics 16, 49-58 | [Medline] |
| FARRUGIA, G., RAE, J. L., SARR, M. G. & SZURSZEWSKI, J. H. (1993). Potassium current in circular smooth muscle of human jejunum activated by fenamates. American Journal of Physiology 265, G873-879 | [Medline] |
| FARRUGIA, G., RAE, J. L. & SZURSZEWSKI, J. H. (1993). Characterization of an outward potassium current in canine jejunal circular smooth muscle and its activation by fenamates. Journal of Physiology 468, 297-310 | [Abstract] |
| HISADA, T., KURACHI, Y. & SUGIMOTO, T. (1990). Properties of membrane currents in isolated smooth muscle cells from guinea-pig trachea. Pflügers Archiv 416, 151-161 | [Medline] |
| HOLLYWOOD, M. A., MCCLOSKEY, K. D., MCHALE, N. G. & THORNBURY, K. D. (2000). Characterization of outward K+. currents in isolated smooth muscle cells from sheep urethra. American Journal of Physiology 279, C420-428 | |
| HUIZINGA, J. D., THUNEBERG, L., VANDERWINDEN, J. M. & RUMESSEN, J. J. (1997). Interstitial cells of Cajal as pharmacological targets for gastrointestinal motility disorders. Trends in Pharmacological Sciences 18, 393-403 | [Medline] |
| IMAIZUMI, Y., MURAKI, K. & WATANABE, M. (1990). Characteristics of transient outward currents in single smooth muscle cells from the ureter of the guinea-pig. Journal of Physiology 427, 301-324 | [Abstract] |
| JAMES, A. F., OKADA, T. & HORIE, M. (1995). A fast transient outward current in cultured cells from human pulmonary artery smooth muscle. American Journal of Physiology 268, H2358-2365 | [Medline] |
| LEE, J. C. F., THUNEBERG, L., FARRAWAY, L. & HUIZINGA, J. D. (1996). Plasticity in ICC isolated from the mouse small intestine. Digestive Diseases and Sciences 41, 1904 | |
| MCFADZEAN, I. & ENGLAND, S. (1992). Properties of the inactivating outward current in single smooth muscle cells isolated from the rat anococcygeus. Pflügers Archiv 421, 117-124 | [Medline] |
| MALYSZ, J., RICHARDSON, D., FARRAWAY, L., CHRISTEN, M. O. & HUIZINGA, J. D. (1995). Generation of slow wave type action potentials in the mouse small intestine involves a non-L-type calcium channel. Canadian Journal of Physiology and Pharmacology 73, 1502-1511 | [Medline] |
| NOACK, T., DEITMER, P. & LAMMEL, E. (1992). Characterization of membrane currents in single smooth muscle cells from the guinea-pig gastric antrum. Journal of Physiology 451, 387-417 | [Abstract] |
| OHYA, Y., KITAMURA, K. & KURIYAMA, H. (1987). Cellular calcium regulates outward currents in rabbit intestinal smooth muscle cell. American Journal of Physiology 252, C401-410 | [Medline] |
| SMIRNOV, S. V., ZHOLOS, A. V. & SHUBA, M. F. (1992). A potential-dependent fast outward current in single smooth muscle cells isolated from the newborn rat ileum. Journal of Physiology 454, 573-589 | [Abstract] |
| SMITH, R. D., EISNER, D. A. & WRAY, S. (1998). The effects of changing intracellular pH on calcium and potassium currents in smooth muscle cells from the guinea-pig ureter. Pflügers Archiv 435, 518-522 | [Medline] |
| VOGALIS, F. & SANDERS, K. M. (1991). Characterization of ionic currents of circular smooth muscle cells of the canine pyloric sphincter. Journal of Physiology 436, 75-92 | [Abstract] |
Acknowledgements
The authors wish to thank Laura Farraway for preparation of the cells. The Canadian Institutes for Health Research provided salary support to JCFL and JDH and operating grants.
Author's present address
C. Barajas-López: Department of Anatomy and Cell Biology, Queen's University, Kingston, Ontario, Canada.
This article has been cited by other articles:
|
|
 |
|
  Y. Zhu, C. M. Golden, J. Ye, X.-Y. Wang, H. I. Akbarali, and J. D. Huizinga ERG K+ currents regulate pacemaker activity in ICC Am J Physiol Gastrointest Liver Physiol, December 1, 2003; 285(6): G1249 - G1258. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
t = 4 ms). The final third test pulse of +50 mV was administered to assess the amount of recovery during that duration. As the duration increases, the peak current elicited by the test pulse is seen to increase in a monophasic manner (inset). The peak currents from the test pulses were normalized and plotted against the time spent recovering from 0 mV inactivation (duration of second pulse, 4-200 ms). The data were fitted according to an inverse exponential relationship to obtain the