Department of Physiology and Cell Biology, University of Nevada School of Medicine, Reno, Nevada 89557
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Whole cell patch-clamp
recordings were made from cultured myenteric neurons taken from murine
proximal colon. The micropipette contained Cs+ to remove
K+ currents. Depolarization elicited a slowly activating
time-dependent outward current (Itdo), whereas
repolarization was followed by a slowly deactivating tail current
(Itail). Itdo and
Itail were present in ~70% of neurons. We
identified these currents as Cl currents
(ICl), because changing the transmembrane
Cl
gradient altered the measured reversal potential
(Erev) of both Itdo and
Itail with that for Itail
shifted close to the calculated Cl
equilibrium potential
(ECl). ICl are
Ca2+-activated Cl
current
[ICl(Ca)] because they were Ca2+
dependent. ECl, which was measured from the
Erev of ICl(Ca) using a
gramicidin perforated patch, was
33 mV. This value is more positive
than the resting membrane potential (
56.3 ± 2.7 mV), suggesting
myenteric neurons accumulate intracellular Cl
.
-Conotoxin GIVA [0.3 µM; N-type Ca2+ channel
blocker] and niflumic acid [10 µM; known
ICl(Ca) blocker], decreased the
ICl(Ca). In conclusion, these neurons have
ICl(Ca) that are activated by Ca2+
entry through N-type Ca2+ channels. These currents likely
regulate postspike frequency adaptation.
myenteric neurons; chloride currents; cell culture; murine large intestine
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
THE PROXIMAL COLON RECEIVES the liquefied waste products of digestion and reabsorbs the remaining water and electrolytes. These functions are largely dependent upon the integrated activities of the enteric nervous system (ENS; see Ref. 46). In small mammals, the ENS consists of two ganglionated neural networks called the myenteric plexus, which is between the longitudinal and circular muscle layers, and the submucous plexus, which lies in the submucosa on the surface of the inner circular muscle layer. The neurons in the myenteric plexus largely regulate motility reflexes, whereas those in the submucous plexus regulate secretomotor reflexes. The myenteric plexus, which is studied here, contains a number of functionally different neurons that include sensory neurons, interneurons, and excitatory and inhibitory motor neurons supplying the longitudinal and circular muscle layers (31, 34, 47).
Intracellular microelectrode recordings from myenteric neurons in guinea pig small intestine have revealed two broad electrophysiological classes of myenteric neurons, S/type I and AH/type II neurons (23, 36). S neurons are uniaxonal, lack a prolonged slow afterhyperpolarization (AHP, up to 20 s), have prominent fast synaptic input (7, 23), and comprise both interneurons and motor neurons (44). AH neurons, which comprise ~25% of all neurons in the small intestine, are generally multipolar and named for their characteristic AHP (4-20 s) that follows action potential firing in these neurons (7, 21, 23, 36, 49, 50). Many AH neurons appear to be intrinsic primary afferent neurons (17). Ca2+-activated K+ channels underlie the AHP (22, 32) that involves the opening of small-to-intermediate conductance channels that are tetraethylammonium (TEA) and apamin resistant (55).
Myenteric neurons in the large intestine of a number of species, such as human (8), guinea pig (30, 31, 33, 43, 49, 58, 59), rat (9), and mouse (18), appear to be electrically more heterogeneous than those in the small intestine since they exhibit more diverse firing patterns in response to current injection. However, like those in the small intestine, AH neurons are characterized by a prolonged AHP (30, 31, 33, 43, 49, 58), are usually multipolar (30, 31, 33, 43, 49), and project to the mucosa (33, 43). Some ascending interneurons in the colon are also rapidly adapting and exhibit an intermediate AHP (30). However, in the mouse colon, the AH neuronal population (~8%) appears to be only about one-third that in the guinea pig small intestine (18).
Patch-clamp techniques have been used to study Na+,
Ca2+, and K+ currents in myenteric neurons in
guinea pig small intestine (3, 41, 48, 54, 61) and to a
lesser extent in the large intestine (56, 57). However, to
date, Cl channels have not been characterized in
myenteric neurons using patch-clamp studies. Indirect evidence,
however, suggests that Cl
channels participate in slow
synaptic transmission (6) and the depolarizing responses
to exogenous
-aminobutyric acid (GABA; see Ref. 10) and
glycine (35). In addition, the presence of afterdepolarizing responses in some tonic S-type neurons
(45) and transient inward currents in AH neurons
(54) suggests the possible involvement of
Ca2+-activated Cl
(ClCa) channels
in these events. Robust action potential-dependent increases in
Ca2+ are observed in both of these types of neuron
(21, 42, 50, 53, 56, 61), and both the
after-depolarization (unpublished observations) and the transient
inward current (54) are reduced by niflumic acid.
However, Cl
channels cannot be identified definitively by
pharmacological means, since there are no specific blockers of these
channels (16).
ICl(Ca) have been identified in a variety of
peripheral and central neurons, including spinal cord
(38), dorsal root ganglia (11), pelvic
parasympathetic ganglia (37), trigeminal sensory and
parasympathetic neurons (2), rod and cone photoreceptors (4), and olfactory receptor neurons (28). The
physiological role of ICl(Ca) is variable,
depending on the Cl equilibrium potential in different
types of neurons.
Because there is no clear proof for the presence of ClCa
channels in enteric neurons, we determined whether
ICl(Ca) could be found in cultured myenteric
neurons of the murine proximal colon. We also attempted to directly
measure the Cl equilibrium potential, since this might
disclose a physiological role for ICl(Ca). We
chose to characterize the ICl(Ca) in the murine
intestine in view of the advantages offered by future studies using
transgenic mouse models.
A preliminary account of our findings has been published in abstract form (26).
![]() |
MATERIALS AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Dissociation of the myenteric plexus. Adult mice (C57BL/6) were killed by isofluorane inhalation and cervical dislocation in compliance with the requirements of the Animal Ethics Committee at the University of Nevada. A 2.5-cm length of proximal colon was removed, opened longitudinally, and pinned out flat in a dish lined with Sylgard containing Krebs solution. The mucosa and submucosa were completely dissected away, the remaining muscle layer preparation was turned upside down, and the longitudinal muscle layer was peeled away. The remaining myenteric plexus-circular muscle preparation (see Ref. 18) was cut into small pieces and transferred to a test tube containing 0.2% collagenase (type II; Worthington) and 0.12% protease (type IX; Sigma, St. Louis, MO) dissolved in Ca2+-free Hanks' solution. After 30 min of incubation at 37°C, the tissues were washed four times with enzyme-free Ca2+-free Hanks' solution and gently triturated through a fire-polished glass Pasteur pipette for 10-15 min. The suspension was then centrifuged at 200 rpm for 5 min, after which the supernatant was discarded, and the pellet was resuspended in 2 ml Ca2+-free Hanks' solution. Aliquots of this solution were added to 35-mm plastic dishes. Each of the dishes contained 2.5 ml cell culture medium consisting of medium-199 (GIBCO) plus 10% FBS. The medium was also supplemented with 10 mM glucose (GIBCO), 20 µM 5-fluoro-2-deoxyuridine (Sigma), and 1.5% antibiotic/antimycotic solution (10,000 U/ml penicillin, 10 mg/ml streptomycin, and 0.5 mg/ml amphotericin B). The dishes were maintained in a humidified incubator (gassed 5% CO2) at 37°C for 2-5 days before use. The culture medium in the dishes was changed every 2 days.
Patch-clamp recording.
Whole cell currents were recorded at room temperature (20-22°C)
using a perforated patch configuration with a patch-clamp amplifier
(EPC-9; HEKA Instruments, Lambrecht, Germany) and Pulse software.
Currents were filtered on-line at 3 kHz and digitized at 0.5-20
kHz. Patch pipettes were drawn from thin-walled borosilicate capillary
glass (Sutter Instrument, Novato, Canada) to have resistances of
1.5-3.0 M. An Ag-AgCl reference electrode was connected to the
bath through an agar bridge saturated with KCl solution. To obtain a
perforated patch, the pipette solution contained gramicidin dissolved
in DMSO to a final concentration of 10-60 µg/ml. To measure the
reversal potential (Erev) of Cl
,
we used nystatin (250 µg/ml) perforated patches to equilibrate the
intracellular Cl
with that of the pipette solution
(20).
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Depolarizing voltage-activated currents in myenteric neurons.
Whole cell currents were recorded from murine colonic myenteric neurons
using a Cs+-containing pipette solution to block
K+ currents (see MATERIALS AND METHODS).
Gramicidin (10-60 µg/ml) was used to perforate cell-attached
membrane patches with cation-selective channels to preserve the
intracellular Cl concentration (14, 29).
During the recording, the series resistance was <10 M
and
compensated (~80%). Under these conditions, depolarization (+10 mV,
400 ms) evoked an initial transient inward current and the slowly
activating time-dependent outward current (Itdo), whereas repolarizing the membrane
potential to
80 mV induced a slowly deactivating tail current
(Itail; Fig.
1A). This Itdo and Itail was
present in ~70% of recorded neurons (65 out of 94 patched neurons).
Initial transient inward currents were composed of fast-activating and
fast-inactivating current and sustained inward currents (Fig.
1B). The fast-activating and fast-inactivating current was
an Na+ current because it was almost blocked by TTX (0.7 µM, n = 7; Fig. 1B). Myenteric neurons
were identified as those cells generating fast Na+ current
at the onset of the depolarizing voltage step. The sustained inward
current was Ca2+ current because it was abolished by
Cd2+ (0.4 mM; see Fig. 7A2).
|
|
|
Measurement of resting membrane potential and cell capacitance.
The resting membrane potential of these cultured myenteric neurons
measured using a K+-rich (without TEA) pipette solution was
56.3 ± 2.7 mV (n = 15).
Itdo and Itail are carried by
Cl.
To test whether Itdo and
Itail are ICl, we
measured the Erev of
Itail before and after changing the
transmembrane Cl
gradient (Fig.
4). Whole cell currents were recorded in
a nystatin-perforated patch configuration to equilibrate the
intracellular Cl
concentration with that of pipette
solution (20). The extracellular Cl
concentration was reduced from 145 to 46 mM, thereby shifting the
calculated ECl from ~0 mV to approximately +29
mV. These changes shifted the measured Erev of
Itail from 1.4 ± 0.6 to 19.2 ± 0.7 mV (n = 5; Fig. 4, A and B). From
these measured Erev, we can calculate the
relative permeability of methanesulfonic acid to Cl
(0.23 ± 0.12).
|
Myenteric neurons accumulate intracellular
Cl.
Using the gramicidin-perforated patch, we measured the
ICl without disturbing the intracellular
Cl
homeostasis (14, 29).
Itail reversed at
33.4 ± 1.0 mV
(n = 17; Fig. 5), which
was more positive than the resting membrane potential. Using the Nernst
equation, we could approximate the intracellular Cl
concentration to be ~39 mM. This implies that, in myenteric neurons, the intracellular Cl
concentration is maintained above
that expected for passive Cl
distribution.
|
Does ECl change during long depolarizing pulses?
Figure 3 shows that Itdo appears to grow after
Itail has saturated; we therefore examined
whether such a difference may be attributable to changes in
ECl in response to Cl accumulation
during long-duration depolarizing pulses. We therefore measured the
Erev of Itdo and
Itail using a ramp protocol. Increasing the
depolarization duration from 300 to 1,100 ms in increments of 200 ms
produced incremental shifts in ECl from
32.8 ± 1.0 mV (300 ms) to
27.8 ± 0.9 mV (1,100 ms;
n = 7, P < 0.01; Fig 6).
|
ICl are Ca2+ dependent. To explore the Ca2+ dependence of the ICl, we compared ICl recorded before and after exposure to Cd2+ (0.4 mM) in the bath solution and after changes in the extracellular Ca2+ concentration.
Cd2+ (0.4 mM) abolished ICl (Fig. 7A1), indicating that Ca2+ influx is a prerequisite for the activation of this current.
|
Role of N-type Ca2+ channel on
ICl(Ca).
In the guinea pig myenteric neurons, Ca2+-activated
K+ current, which is responsible for the prolonged AHP in
AH neurons and the intermediate AHP in tonic S neurons, is reported to
be dependent on Ca2+ entry through N-type Ca2+
channels (42, 54). To test the role of N-type channel on ICl(Ca), we have used the -conotoxin GIVA
(0.3 µM). Bath application of
-conotoxin GIVA decreased
Itdo from 1,080 ± 180 to 550 ± 66 pA
(n = 6, P < 0.01) and decreased
Itail from
990 ± 130 to
480 ± 77 pA (n = 6, P < 0.01). Also, it
increased (but not significantly) the time constant of
Itdo from 90 ± 9.3 to 210 ± 60 ms
(n = 6, P > 0.05) and decreased the
time constant of Itail from 470 ± 65 to
355 ± 43 ms (n = 6, P < 0.01;
Fig. 8).
|
Blockade of ICl(Ca) by niflumic acid.
Bath application of 10 µM niflumic acid, an
ICl(Ca) blocker (16), significantly
decreased Itdo from 950 ± 170 to 420 ± 91 pA (n = 7, P < 0.01) and
Itail from 600 ± 130 to
250 ± 50 pA (n = 7, P < 0.01). Although
niflumic acid increased the deactivation time constant of
Itail from 560 ± 62 to 970 ± 98 ms
(n = 6, P < 0.01), it had no effect on
the activation time constant of Itdo [from
100 ± 9.1 to 110 ± 4.0 ms (n = 7, P > 0.05); Fig. 9].
Niflumic acid did not appear to inhibit Ca2+ entry (Fig.
9A2) but decreased the peak of the
ICl(Ca). Furthermore, niflumic acid did not
affect the activation time constant of Itdo, suggesting it to be an open channel blocker, as reported previously (24). However, the deactivation time constant of
Itail was significantly slowed, even though the
Ca2+ load to be buffered was the same.
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
We present, for the first time, direct evidence for
ICl(Ca) in myenteric neurons cultured from
murine proximal colon. We identified these currents by showing that,
when the transmembrane Cl gradient was changed, the
Erev of this Ca2+-dependent current
was shifted close to the calculated value for ECl. Using a gramicidin perforated patch so as
not to perturb the intracellular Cl
concentration, we
found ECl to be
33 mV. This
Erev, which is more positive than the resting
membrane potential of these neurons (
56.3 ± 2.7 mV), suggests
that ICl(Ca) regulates postspike excitability.
ICl(Ca) in murine colonic myenteric neurons have
similar characteristics to those described in other cells: a slow rate
of activation, little inactivation during sustained depolarization, and
very long deactivation kinetics (39, 60). The
characteristics of ICl(Ca) were closely related
to Ca2+ entry in the cell. ICl(Ca),
which started to activate around 20 mV, was dependent upon
Ca2+ entry through voltage-dependent Ca2+
channels (blocked by Cd2+). A significant amount of this
Ca2+ entry is through voltage-gated N-type Ca2+
channels, since ICl(Ca) was reduced by
-conotoxin GIVA, as are K+ currents underlying the
postspike afterhypolarization in both AH neurons (54) and
some tonic S neurons (42). Itail of
ICl(Ca) were maximal at the activation voltage
for peak Ca2+ current (~10 mV; Fig. 2B). The
changes in the deactivation time constant were compared at the same
voltage (
80 mV), and, in contrast to
of
Itdo, are unlikely to be contaminated by other
underlying currents. When the Itail was maximal,
the deactivation time constant was also maximal (Fig. 2, B
and C). Increasing the duration of the depolarizing step
prolongs the activation of Ca2+ channels, which promotes
sustained Ca2+ entry into the cell. Increasing the duration
of the depolarizing step caused an increase in
ICl(Ca) and prolonged the deactivation time
constant (
of Itail; Fig. 3C).
Moreover, increased Ca2+ entry because of raised
extracellular Ca2+ concentration (from 2 to 6 mM) prolonged
the deactivation time constant, whereas decreased Ca2+
entry resulting from lowered extracellular Ca2+
concentration (from 2 to 0.7 mM) or N-type Ca2+ channel
blocker (
-conotoxin GIVA) shortened the deactivation time constant.
These data suggest that increased Ca2+ entry increases the
intracellular Ca2+ load to be buffered, which reflects
slower deactivation of ICl(Ca) and vice versa
for decreased Ca2+ entry to the cell. Therefore, the
deactivation time constant likely reflects how fast cells remove excess
Ca2+. However, increased Ca2+ entry resulting
from raised extracellular Ca2+ concentration (from 2 to 6 mM) shortened the activation time constant, whereas decreased
Ca2+ entry resulting from lowered extracellular
Ca2+ concentration (from 2 to 0.7 mM) or N-type
Ca2+ channel blocker (
-conotoxin GIVA) prolonged the
activation time constant. This supports the link between
Ca2+ influx and activation of the
ICl(Ca). Therefore, the activation time constant
may reflect the level of available intracellular Ca2+
needed to activate ClCa channels.
So far, specific blockers for pharmacological identification of
ClCa channels are not available. Niflumic acid, a
nonsteroidal anti-inflammatory agent, has been used widely. However,
its actions can be complex, such as blocking outward currents and
enhancing inward currents, depending on the intracellular
Ca2+ level (see Ref. 40 for results in rat
pulmonary artery). Also, niflumic acid has been shown to have both
open-channel and voltage-dependent blocking effects (24).
Consistent with its open-channel blocking effects, we found that
niflumic acid had no effect on the activation time constant but
prolonged the deactivation time constant. However, there was no
evidence for a voltage-dependent blocking effect by niflumic acid,
since it decreased both the outward currents (Itdo; activated by stepping to +10 mV) and the
inward currents (Itail; elicited by stepping to
80 mV) by a similar amount (decreased by 55.6 and 58.3%, respectively).
Although the specific functional classes of neurons possessing ICl(Ca) were not identified in our study, we found the majority (~70%) of cultured myenteric neurons in murine proximal colon to have ICl(Ca). Also, we did not find any relation between the cell capacitance (and therefore cell size) and the presence or absence of this current.
The physiological role of ClCa channels in neurons is
determined by the ECl and resting membrane
potential. Opening ClCa channels hyperpolarizes cultured
spinal neurons (mouse; see Ref. 39) and taste cells
(Necturus; see Ref. 51). Also, inhibitory postsynaptic potentials generated by activation of GABAA and glycine
receptors are also reported to be the result of activation of
Cl conductances in neuronal cells (12, 25).
For Cl
conductances to hyperpolarize or stabilize the
membrane potential, the ECl must be maintained
at a value equal to or higher than the resting membrane potential. In
this case, ECl results from passive distribution
of Cl
across the membrane (the Donnan equilibrium;
ECl would be the same as the resting membrane
potential) and active extrusion of Cl
from the cytoplasm
(ECl would be higher than the resting membrane potential; see Ref. 52). On the other hand,
ICl(Ca) can be responsible for posttetanic
afterdepolarizing potentials in rabbit sympathetic ganglia
(1). Furthermore, the fact that GABA and glycine can also
depolarize neurons through activation of Cl
channels
(13, 19, 35) reflects an ECl lower
than the resting membrane potential. These depolarization responses
resulting from an increase in Cl
conductance result from
intracellular Cl
accumulation (27). Hence,
our results (ECl =
33 mV) imply that
myenteric neurons actively accumulate intracellular Cl
,
which are responsible for membrane depolarization. In myenteric neurons, there is some evidence for membrane depolarization resulting from an increase in Cl
conductance, which is consistent
with our results. Previous studies have suggested the possibility of
ICl(Ca) in the afterdepolarizing response
observed in some tonic S neurons (45) and poststimulus transient inward currents in AH neurons (54). However,
these studies relied on either the estimates of the
Erev (
34 to
40 mV) of these events or the
sensitivity of inward currents to high concentrations of niflumic acid,
which is not selective for ICl(Ca) (16). Other studies that examined the membrane potential
dependence of depolarizing responses to agonists estimated the
Cl
reversal potential in AH neurons to be around
18 mV
with sharp KCl electrodes (10, 35) and
39 mV when
potassium acetate, citrate, or sulfate electrodes were used
(10).
In the present study, the ECl in murine colonic
myenteric neurons was found to be approximately 33 mV, which is more
positive than the resting membrane potential of these neurons
(
56.3 ± 2.7 mV); therefore, they would accumulate
Cl
. Activation of ICl(Ca) is
likely to depolarize the neuron (up to ECl) and
decrease the threshold for neuronal firing, thereby regulating both
postspike excitability and spike frequency adaptation.
![]() |
ACKNOWLEDGEMENTS |
---|
We thank Drs. James Kenyon and Sang Don Koh for helpful suggestions.
![]() |
FOOTNOTES |
---|
This study was supported by National Institute of Diabetes and Digestive and Kidney Diseases Grant PO1 DK-41315. P. Vanden Berghe is a Postdoctoral Fellow of the Fund for Scientific Research, Flanders, Belgium.
Address for reprint requests and other correspondence: T. K. Smith, Dept. of Physiology and Cell Biology, Univ. of Nevada School of Medicine, Reno, Nevada 89557 (E-mail: tks{at}physio.unr.edu).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
First published November 27, 2002;10.1152/ajpcell.00437.2002
Received 23 September 2002; accepted in final form 25 November 2002.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Akasu, T,
Nishimura T,
and
Tokimasa T.
Calcium-dependent chloride current in neurones of the rabbit pelvic parasympathetic ganglia.
J Physiol
422:
303-320,
1990[Abstract].
2.
Bader, CR,
Bertrand D,
and
Schlichter R.
Calcium-activated chloride current in cultured sensory and parasympathetic quail neurones.
J Physiol
394:
125-148,
1987[Abstract].
3.
Baidan, LV,
Zholos AV,
and
Wood JD.
Modulation of calcium currents by G-proteins and adenosine receptors in myenteric neurones cultured from adult guinea-pig small intestine.
Br J Pharmacol
116:
1882-1886,
1995[Abstract].
4.
Barnes, S,
and
Bui Q.
Modulation of calcium-activated chloride current via pH-induced changes of calcium channel properties in cone photoreceptors.
J Neurosci
11:
4015-4023,
1991[Abstract].
5.
Barry, PH.
JPCalc, a software package for calculating liquid junction potential corrections in patch-clamp, intracellular, epithelial and bilayer measurements and for correcting junction potential measurements.
J Neurosci Methods
51:
107-116,
1994[ISI][Medline].
6.
Bertrand, PP,
and
Galligan JJ.
Contribution of chloride conductance increase to slow EPSC and tachykinin current in guinea-pig myenteric neurones.
J Physiol
481:
47-60,
1994[Abstract].
7.
Bornstein, JC,
Furness JB,
and
Kunze WA.
Electrophysiological characterization of myenteric neurons: how do classification schemes relate?
J Auton Nerv Syst
48:
1-15,
1994[ISI][Medline].
8.
Brookes, SJ,
Ewart WR,
and
Wingate DL.
Intracellular recordings from myenteric neurones in the human colon.
J Physiol
390:
305-318,
1987[Abstract].
9.
Browning, KN,
and
Lees GM.
Myenteric neurons of the rat descending colon: electrophysiological and correlated morphological properties.
Neuroscience
73:
1029-1047,
1996[ISI][Medline].
10.
Cherubini, E,
and
North RA.
Actions of gamma-aminobutyric acid on neurones of guinea-pig myenteric plexus.
Br J Pharmacol
82:
93-100,
1984[Abstract].
11.
Currie, KP,
and
Scott RH.
Calcium-activated currents in cultured neurones from rat dorsal root ganglia.
Br J Pharmacol
106:
593-602,
1992[Abstract].
12.
Deisz, RA,
and
Lux HD.
The role of intracellular chloride in hyperpolarizing post-synaptic inhibition of crayfish stretch receptor neurones.
J Physiol
326:
123-138,
1982[ISI][Medline].
13.
Deschenes, M,
Feltz P,
and
Lamour Y.
A model for an estimate in vivo of the ionic basis of presynaptic inhibition: an intracellular analysis of the GABA-induced depolarization in rat dorsal root ganglia.
Brain Res
118:
486-493,
1976[ISI][Medline].
14.
Ebihara, S,
Shirato K,
Harata N,
and
Akaike N.
Gramicidin-perforated patch recording: GABA response in mammalian neurones with intact intracellular chloride.
J Physiol
484:
77-86,
1995[Abstract].
15.
Evans, MG,
and
Marty A.
Calcium-dependent chloride currents in isolated cells from rat lacrimal glands.
J Physiol
378:
437-460,
1986[Abstract].
16.
Frings, S,
Reuter D,
and
Kleene SJ.
Neuronal Ca2+-activated Cl channels: homing in on an elusive channel species.
Prog Neurobiol
60:
247-289,
2000[ISI][Medline].
17.
Furness, JB,
Kunze WA,
Bertrand PP,
Clerc N,
and
Bornstein JC.
Intrinsic primary afferent neurons of the intestine.
Prog Neurobiol
54:
1-18,
1998[ISI][Medline].
18.
Furukawa, K,
Taylor GS,
and
Bywater RA.
An intracellular study of myenteric neurons in the mouse colon.
J Neurophysiol
55:
1395-1406,
1986
19.
Gallagher, JP,
Higashi H,
and
Nishi S.
Characterization and ionic basis of GABA-induced depolarizations recorded in vitro from cat primary afferent neurones.
J Physiol
275:
263-282,
1978[Abstract].
20.
Hille, B.
Counting channels and measuring fluctuations.
In: Ion Channels of Excitable Membranes. Sunderland, MA: Sinauer, 2001, p. 402.
21.
Hillsley, K,
Kenyon JL,
and
Smith TK.
Ryanodine-sensitive stores regulate the excitability of AH neurons in the myenteric plexus of guinea-pig ileum.
J Neurophysiol
84:
2777-2785,
2000
22.
Hirst, GD,
Johnson SM,
and
van Helden DF.
The slow calcium-dependent potassium current in a myenteric neurone of the guinea-pig ileum.
J Physiol
361:
315-337,
1985[Abstract].
23.
Hirst, GDS,
Holman ME,
and
Spence I.
Two types of neurones in the myenteric plexus of duodenum in the guinea-pig.
J Physiol
236:
303-326,
1974[ISI].
24.
Hogg, RC,
Wang Q,
and
Large WA.
Action of niflumic acid on evoked and spontaneous calcium-activated chloride and potassium currents in smooth muscle cells from rabbit portal vein.
Br J Pharmacol
112:
977-984,
1994[Abstract].
25.
Kaila, K.
Ionic basis of GABAA receptor channel function in the nervous system.
Prog Neurobiol
42:
489-537,
1994[ISI][Medline].
26.
Kang, SH,
Vanden Berghe P,
and
Smith TK.
Calcium-activated chloride current in cultured myenteric neurons from mouse colon (Abstract).
Gastroenterolgy
122:
A85,
2002.
27.
Kenyon, JL.
The reversal potential of Ca(2+)-activated Cl(-) currents indicates that chick sensory neurons accumulate intracellular Cl(-).
Neurosci Lett
296:
9-12,
2000[ISI][Medline].
28.
Kleene, SJ,
and
Gesteland RC.
Calcium-activated chloride conductance in frog olfactory cilia.
J Neurosci
11:
3624-3629,
1991[Abstract].
29.
Kyrozis, A,
and
Reichling DB.
Perforated-patch recording with gramicidin avoids artifactual changes in intracellular chloride concentration.
J Neurosci Methods
57:
27-35,
1995[ISI][Medline].
30.
Lomax, AE,
Sharkey KA,
Bertrand PP,
Low AM,
Bornstein JC,
and
Furness JB.
Correlation of morphology, electrophysiology and chemistry of neurons in the myenteric plexus of the guinea-pig distal colon.
J Auton Nerv Syst
76:
45-61,
1999[ISI][Medline].
31.
Messenger, JP,
Bornstein JC,
and
Furness JB.
Electrophysiological and morphological classification of myenteric neurons in the proximal colon of the guinea-pig.
Neuroscience
60:
227-244,
1994[ISI][Medline].
32.
Morita, K,
and
North RA.
Significance of slow synaptic potentials for transmission of excitation in guinea-pig myenteric plexus.
Neuroscience
14:
661-672,
1985[ISI][Medline].
33.
Neunlist, M,
Dobreva G,
and
Schemann M.
Characteristics of mucosally projecting myenteric neurones in the guinea-pig proximal colon.
J Physiol
517:
533-546,
1999
34.
Neunlist, M,
Michel K,
Aube AC,
Galmiche JP,
and
Schemann M.
Projections of excitatory and inhibitory motor neurones to the circular and longitudinal muscle of the guinea pig colon.
Cell Tissue Res
305:
325-330,
2001[ISI][Medline].
35.
Neunlist, M,
Michel K,
Reiche D,
Dobreva G,
Huber K,
and
Schemann M.
Glycine activates myenteric neurones in adult guinea-pigs.
J Physiol
536:
727-739,
2001
36.
Nishi, S,
and
North RA.
Intracellular recording from the myenteric plexus of the guinea-pig ileum.
J Physiol
231:
471-491,
1973[ISI][Medline].
37.
Nishimura, T,
Akasu T,
and
Tokimasa T.
A slow calcium-dependent chloride current in rhythmic hyperpolarization in neurones of the rabbit vesical pelvic ganglia.
J Physiol
437:
673-690,
1991[Abstract].
38.
Owen, DG,
Segal M,
and
Barker JL.
A Ca-dependent Cl conductance in cultured mouse spinal neurones.
Nature
311:
567-570,
1984[ISI][Medline].
39.
Owen, DG,
Segal M,
and
Barker JL.
Voltage-clamp analysis of a Ca2+- and voltage-dependent chloride conductance in cultured mouse spinal neurons.
J Neurophysiol
55:
1115-1135,
1986
40.
Piper, AS,
Greenwood IA,
and
Large WA.
Dual effect of blocking agents on Ca2+-activated Cl.
J Physiol
539:
119-131,
2002
41.
Rugiero, F,
Gola M,
Kunze WA,
Reynaud JC,
Furness JB,
and
Clerc N.
Analysis of whole-cell currents by patch clamp of guinea-pig myenteric neurones in intact ganglia.
J Physiol
538:
447-463,
2002
42.
Shuttleworth, CW,
and
Smith TK.
Action potential-dependent calcium transients in myenteric S neurons of the guinea-pig ileum.
Neuroscience
92:
751-762,
1999[ISI][Medline].
43.
Smith, TK.
An electrophysiological identification of intrinsic sensory neurons responsive to 5-HT applied to the mucosa that underly peristalsis in the guinea-pig proximal colon (Abstract).
J Physiol
495:
102P,
1996.
44.
Smith, TK,
Bornstein JC,
and
Furness JB.
Convergence of reflex pathways excited by distension and mechanical stimulation of the mucosa onto the same myenteric neurons of the guinea pig small intestine.
J Neurosci
12:
1502-1510,
1992[Abstract].
45.
Smith, TK,
Burke EP,
and
Shuttleworth CW.
Topographical and electrophysiological characteristics of highly excitable S neurones in the myenteric plexus of the guinea-pig ileum.
J Physiol
517:
817-830,
1999
46.
Smith, TK,
and
Sanders KM.
Motility of the large intestine.
In: Textbook of Gastroenterology I, edited by Yamada T,
Owyang C,
Powell DW,
and Silverstein JB.. Philadelphia, PA: Lippincott, 1995, p. 234-260.
47.
Spencer, NJ,
and
Smith TK.
Simultaneous intracellular recordings from longitudinal and circular muscle during the peristaltic reflex in guinea-pig distal colon.
J Physiol
533:
787-799,
2001
48.
Starodub, AM,
and
Wood JD.
Selectivity of omega-CgTx-MVIIC toxin from Conus magus on calcium currents in enteric neurons.
Life Sci
64:
L305-L310,
1999.
49.
Tamura, K,
Ito H,
and
Wade PR.
Morphology, electrophysiology, and calbindin immunoreactivity of myenteric neurons in the guinea pig distal colon.
J Comp Neurol
437:
423-437,
2001[ISI][Medline].
50.
Tatsumi, H,
Hirai K,
and
Katayama Y.
Measurement of the intracellular calcium concentration in guinea-pig myenteric neurons by using fura-2.
Brain Res
451:
371-375,
1988[ISI][Medline].
51.
Taylor, R,
and
Roper S.
Ca2+-dependent Cl conductance in taste cells from Necturus.
J Neurophysiol
72:
475-478,
1994
52.
Thompson, SM,
Deisz RA,
and
Prince DA.
Outward chloride/cation co-transport in mammalian cortical neurons.
Neurosci Lett
89:
49-54,
1988[ISI][Medline].
53.
Vanden Berghe, P,
Kenyon JL,
and
Smith TK.
Mitochondrial Ca2+ uptake regulates the excitability of myenteric neurons.
J Neurosci
22:
6962-6971,
2002
54.
Vogalis, F,
Furness JB,
and
Kunze WA.
Afterhyperpolarization current in myenteric neurons of the guinea pig duodenum.
J Neurophysiol
85:
1941-1951,
2001
55.
Vogalis, F,
Harvey JR,
and
Furness JB.
TEA- and apamin-resistant K(Ca) channels in guinea-pig myenteric neurons; slow AHP channels.
J Physiol
538:
421-433,
2002
56.
Vogalis, F,
Hillsley K,
and
Smith TK.
Recording ionic events from cultured, DiI-labelled myenteric neurons in the guinea-pig proximal colon.
J Neurosci Methods
96:
25-34,
2000[ISI][Medline].
57.
Vogalis, F,
Hillsley K,
and
Smith TK.
Diverse ionic currents and electrical activity of cultured myenteric neurons from the guinea pig proximal colon.
J Neurophysiol
83:
1253-1263,
2000
58.
Wade, PR,
and
Wood JD.
Electrical behavior of myenteric neurons in guinea pig distal colon.
Am J Physiol Gastrointest Liver Physiol
254:
G522-G530,
1988
59.
Wade, PR,
and
Wood JD.
Synaptic behavior of myenteric neurons in guinea pig distal colon.
Am J Physiol Gastrointest Liver Physiol
255:
G184-G190,
1988
60.
Yuan, X.
Role of calcium-activated chloride current in regulating pulmonary vasomotor tone.
Am J Physiol Lung Cell Mol Physiol
272:
L959-L968,
1997
61.
Zholos, AV,
Baidan LV,
and
Wood JD.
Sodium conductance in cultured myenteric AH-type neurons from guinea-pig small intestine.
Auton Neurosci
96:
93-102,
2002[ISI][Medline].