Department of Anaesthesia and Department of Pharmacology & Therapeutics, Faculty of Medicine, The University of British Columbia Vancouver, British Columbia V6T 1Z3 Canada
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ABSTRACT |
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Ries, Craig R. and Ernest Puil. Ionic mechanism of isoflurane's actions on thalamocortical neurons. We studied the actions of isoflurane (IFL) applied in aqueous solutions on ventrobasal neurons from thalamic brain slices of juvenile rats. By using the whole cell, patch-clamp method with current- and voltage-clamp recording techniques, we found that IFL increased a noninactivating membrane conductance in a concentration-dependent reversible manner. In an eightfold concentration range that extended into equivalent in vivo lethal concentrations, IFL did not produce a maximal effect on the conductance; this is consistent with a nonreceptor-mediated mechanism of action. TTX eliminated action potential activity but did not alter IFL effects. The effects on the membrane potential and current induced by IFL were voltage independent but depended on the external [K+], reversing near the equilibrium potential for K+. External Ba2+ or internal Cs+ applications, which block K+ channels, suppressed the conductance increase caused by IFL. External applications of the Ca2+ channel blockers Co2+ or Cd2+ or internal application of the Ca2+ chelator 1,2-bis-(2-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid did not prevent the effects of IFL, implying little involvement of Ca2+-dependent K+ currents. A contribution of inwardly rectifying K+ channels to the increased steady-state conductance seemed unlikely because IFL decreased inward rectification. An involvement of ATP-mediated K+ channels also was unlikely because application of the ATP-mediated K+ channel blocker glibenclamide (1-80 µM) did not prevent IFL's actions. In contrast to spiking cells, IFL depolarized presumed glial cells, consistent with an efflux of K+ from thalamocortical neurons. The results imply that a leak K+ channel mediated the IFL-induced increase in postsynaptic membrane conductance in thalamic relay neurons. Thus a single nonreceptor-mediated mechanism of IFL action was responsible for the hyperpolarization and conductance shunt of voltage-dependent Na+ and Ca2+ spikes, as reported in the preceding paper. Although anesthetics influence various neurological systems, an enhanced K+ leak generalized in thalamocortical neurons alone could account for anesthesia in vivo.
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INTRODUCTION |
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The actions of inhalational anesthetics on neurons
enhance endogenous inhibition in the CNS. This augmentation probably
results from an increase in ionic conductance through receptor- and
nonreceptor-mediated processes in the neuronal membrane (see reviews by
Krnjevi and Puil 1997
; Tanelian et al.
1993
). To account for the anesthetic state in vivo, the
increased conductance should be observed in neurons that are major
participants in generating and maintaining consciousness.
Although anesthetics influence various neurological systems, effects on
the corticothalamocortical system alone could account for anesthesia.
In the preceding paper (Ries and Puil 1999) we reported
that isoflurane (IFL) decreased the excitability of thalamocortical neurons in vitro through a conductance shunt. We now describe the
specific changes in conductance, predominantly a rise in K+
leak, that are responsible for the shunt. The ionic mechanism of action
appears generalized because of IFL's hydrophobicity and the ubiquity
of the leak conductance in neurons. This generalized mechanism together
with the importance of corticothalamocortical excitability in
generating conscious states (cf. Ries and Puil 1999
)
raises the likelihood that IFL's actions at unified sites widely
distributed along somatic and dendritic membranes produce the
anesthetic state.
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METHODS |
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Experiments were performed on neurons in slices of the ventral
posterior thalamic nucleus at room temperature (~22°C) from juvenile Sprague-Dawley rats (aged P9-P22) of either gender. The preparation and maintenance of the slices as well as
electrophysiological and pharmacological techniques are described in
the preceding paper (Ries and Puil 1999).
Near steady-state, voltage-current relationships were obtained by
applying ramp commands during voltage-clamp conditions. Such commands
could be completed within a few seconds and were applied in a
hyperpolarizing direction to avoid activation of the
Ca2+-mediated T-type current. Slow voltage ramps (10 mV/s)
started from a holding potential (Vh) of either
approximately 30 mV during application of TTX or approximately
60
mV in the absence of TTX. After reaching a final potential as low as
120 mV, the ramp application was repeated such that the currents
could be averaged on-line to reduce "noise." These responses are
shown in horizontally mirrored form. In addition, steady-state,
voltage-current relationships were obtained by applying hyperpolarizing
step voltage commands. Whole cell capacitance was not compensated as
currents were measured after the capacitive transient. The conductance
of the neuron was calculated from the current response evoked near the
resting membrane potential (RMP).
The ionic composition of the external solution, used in conjunction
with a gluconate-based internal pipette solution (Ries and Puil
1999), was changed in some experiments. Normally, the calculated equilibrium potentials were as follows (in mV at 25°C): ENa = +46; EK =
85;
ECa = +159 (based on a calculated internal [Ca2+] of 10
8 M); and
ECl =
54. The external [K+] was
changed from 5.25 to 1.25, 2.45, and 9.25 mM by adjusting the [KCl].
This change shifted the calculated EK from
85
mV to
123,
106, and
71 mV, respectively. The NaCl also was
adjusted in the different K+ solutions to maintain the same
[Cl
]. This procedure avoided changes in the junction
potential at the Ag-AgCl reference electrode while changing
ENa by only + 0.9, +0.7, and
0.5 mV,
respectively. In other experiments, choline chloride was partly
substituted for NaCl in the artificial cerebrospinal fluid (ACSF). This
substitution changed the external [Na+] from 124 to 26 mM
and shifted ENa from +46 to 0 mV. On occasion sodium isethionate partly replaced NaCl in the ACSF, changing the
external [Cl
] from 136 to 12 mM, which shifted
ECl from
54 to +10 mV. When this low
[Cl
] was applied the resulting change in junction
potential was compensated in part by adjusting the junction-null
control to display the same membrane potential; this added voltage to
the internal circuitry of the amplifier. During analysis a final
correction for the junction potential was made with, as guides, the
action-potential threshold and K+-mediated persistent
inward rectifier (IKir) (Ries and Puil
1999
).
Ionic channel blockers as well as drugs with known ionic actions
sometimes were applied in the external solutions. Their concentrations were as follows: TTX, 300 nM; tetraethylammonium (TEA), 10 mM; 4-aminopyridine (4-AP), 3 mM; Cs+, 5 mM; Ba2+,
100 µM; Cd2+, 50 µM; and Ni2+, 0.5 mM. In
other experiments, substitutions for Ca2+ were made with
Ba2+, Co2+, and Mg2+. The
GABAB agonist baclofen (racemate, 10 µM) and the
anticholinesterase agent tacrine (100 µM), were administered
occasionally to verify EK. In addition the
ATP-sensitive K+ channel blocker glibenclamide (in 0.5%
dimethylsulfoxide) (Erdemli and Krnjevi 1994
),
was used to test involvement of IK(ATP).
The internal pipette solutions also were altered in three types of
experiments: 1) CsOH was substituted for KOH; 2)
internal Cl was increased from 12 to 50 mM by
substituting KCl for some of the KOH, which shifted
ECl from
54 to
26 mV; and 3)
1,2-bis-(2-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid (BAPTA, 10 mM) was substituted for EGTA. In the BAPTA experiments CaCl2 was decreased to 0.5 mM to maintain the same internal
[Ca2+]. All substances were obtained from Sigma-Aldrich
Canada, except for IFL (Abbott Laboratories, Montreal, Canada) and
glibenclamide (Research Biochemicals International, Natick, USA).
Statistical analysis was performed with GraphPad Prism software (version 2.0, San Diego, CA). Results were expressed as means ± SD. Regression lines were plotted with the method of least squares.
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RESULTS |
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The data were obtained from 66 ventral posterior nucleus neurons,
exhibiting overshooting action potentials on current pulse injection
and the characteristic low-threshold Ca2+ spike (LTS) when
depolarized from potentials more negative than 65 mV. The average
RMPs and input resistances were
66 ± 4 mV and 240 ± 83 M
, respectively.
IFL induced a postsynaptic, net outward current
By using TTX application (300 nM) we first abolished
Na+-dependent rectification and action potentials. We then
determined voltage-current relationships by applying hyperpolarizing
ramp commands in neurons held at a Vh of
approximately 30 mV (see METHODS). A high concentration
(4%) of IFL was applied for 1-2 min to obtain a rapid, yet submaximal
effect. This led to an increased conductance, enhancing the outward
current at voltages >RMP (n = 12, Fig.
1A). The magnitude and
relative linearity of the IFL-induced outward current were evident from
subtraction of the control and IFL-induced currents (Fig.
1B). It seemed likely then that postsynaptic voltage-dependent Na+ currents did not distort the outward
current.
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We performed experiments to assess possible Ca2+ contributions to the net outward current. First, we applied external solutions with nominally zero [Ca2+] and high [Mg2+] (4 mM MgCl2, 0 mM CaCl2), which also would diminish Ca2+-dependent transmitter release. This procedure eliminated the postsynaptic LTS but did not cause detectable changes in the IFL-induced hyperpolarization and conductance increase (n = 5). Second, coapplication of IFL with the Ca2+ channel blockers Cd2+ (CdCl2 50 µM, n = 2) or Co2+ (1 mM CoCl2, 0 mM CaCl2 and 3 mM MgCl2, n = 2, Fig. 1C) had no effect on the rate of onset and peak amplitude of the conductance increase induced by IFL. Although we have not completely dismissed a presynaptic contribution, a postsynaptic influx of Ca2+ did not distort the IFL-induced outward current.
Concentration-effect relationship
We assessed the concentration-dependence of IFL effects with
hyperpolarizing step commands of long duration (1 s) in a voltage range between
70 mV and
100 mV from a Vh of
approximately
60 mV. In addition to TTX, coapplications of TEA (10 mM) and 4-AP (3 mM) were used to block rectifying currents. The
magnitudes of the conductance increase were approximately the same in
the absence of the Na+ and K+ blockers
(Ries and Puil 1999
). In a concentration-dependent
manner, cumulative applications of IFL (0.5-2%) produced
noninactivating currents (n = 8, Fig.
2A). As with ramp commands,
the voltage-current plots demonstrated that the increases in
conductance caused by IFL were voltage independent (Fig.
2B). The results, pooled with single 4% IFL applications,
were used to construct a concentration-effect relationship for the
conductance increase (Fig. 2C). This relationship showed a
linearity over an eightfold concentration range (IFL 0.5 to 4%).
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Principal ionic conductance in IFL actions
By changing the K+ driving force with different external K+ concentrations (1.25, 2.45, 5.25, and 9.25 mM), we assessed a K+ involvement in the IFL response. From a Vh at or near RMP, a change in K+ driving force was verified by the increase in outward holding current with low [K+] perfusion and an inward shift in the holding current with high [K+] perfusion (Fig. 3A). Application of IFL then induced outward currents during all changed external [K+] (Fig. 3A). However, the current magnitude was larger during low [K+] and smaller during high [K+] conditions. These observations were consistent with a K+ involvement in IFL actions.
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By using hyperpolarizing voltage ramps we next determined the reversal potential for IFL actions (EIFL) at various external K+ concentrations. For example, when the extracellular [K+] was decreased, EIFL shifted with the change in EK to a more negative value (Fig. 3, B and C). At all external K+ concentrations, however, EIFL was at a positive value relative to the calculated EK. Figure 3D shows EIFL as a function of log Kout/Kin, which behaves like the Nernst potential for K+. A slope of 39 mV for the experimental data compares approximately to a slope of 60 mV for calculated EK at 25°C. The increasing disparity of the relationship of EIFL to EK as external [K+] decreases implies a systematic experimental error or a contribution of ions other than K+ to IFL's effects.
Baclofen and tacrine effects serve as markers for EK
Independently from IFL application, we obtained measurements of
reversal potentials for two agents that have actions that involve
mostly K+ conductances. First, the effects of the
GABAB agonist baclofen (Crunelli et al.
1988) and its reversal potential were compared with those of
IFL. As with IFL baclofen application (10 µM) induced a persistent
outward current when Vh was near rest (Fig.
4A). Unlike IFL, however, the
outward current caused by baclofen reversed polarity at
84 ± 1 mV (n = 3; Fig. 4B and C), i.e.,
close to the calculated EK (
85 mV). In another
test for EK in two neurons, we applied the
anticholinesterase tacrine (100 µM) (Stevens and Cotman
1987
). Tacrine decreased the resting outward current when Vh was near rest with a reversal potential
identical to EK. We concluded then that errors
such as junction potentials were probably not responsible for the
difference between EIFL and
EK. Because RMP also differed from
EK, it seemed reasonable to explore a
relationship between EIFL and RMP. Indeed the
variation in EIFL correlated to the variation in
RMP, measured before IFL application (r = 0.74, n = 20).
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Contributions of ions other than K+ to IFL actions
Manipulations in the ionic environment were performed to
investigate a secondary role for ions other than K+.
EIFL did not change during blockade of a
theoretical steady-state window T current (Hutcheon et al.
1994) by either a nominally zero external [Ca2+]
with replacement of external Ca2+ with Mg2+ (4 mM, n = 4) or with Ni2+ (0.5 mM,
n = 3). In other experiments we reduced the external [Na+] from 124 to 26 mM by replacement with the
impermeant cation choline (n = 3) and external
[Cl
] from 136 to 12 mM by replacement with the
impermeant anion isethionate (n = 3). We also raised
the internal [Cl
] from 12 to 50 mM (n = 3). Although liquid junction potentials could still be a complicating
factor in some of these experiments (see METHODS), we did
not obtain evidence for a contribution of transmembrane
Ca2+, Na+, or Cl
fluxes to
EIFL and hence to IFL's effects (see also
previous results on Ca2+ channel blockers).
Contributions of K+ conductances to IFL
Although the IFL-induced current appeared resistant to blockade by
4-AP and TEA, other K+ channel blockers were used to
identify the increase in K+ conductance. External
Ba2+ (2 and 0 mM Ca2+) depolarized neurons held
near rest by decreasing conductance and inward rectification; a
subsequent application of IFL (1%) produced little or no change in
membrane conductance and potential. In these neurons, this total
blockade was surmountable by application of a higher concentration of
IFL (4%, n = 4). The reversal potential for
Ba2+ application (77 ± 3 mV) was similar to
EIFL (
76 ± 3 mV) under control
K+ conditions. In another set of experiments the usual
effects of IFL (1 or 2%) were not observed when internal
Cs+ was used to replace K+ in the internal
pipette solution. Like the external Ba2+ blockade, this
apparently total blockade by internal Cs+ was surmountable
in three neurons (n = 4) with higher concentrations of
IFL (4%). These results also were consistent with an IFL-induced increase in K+ conductance.
We investigated a possible involvement of a Ca2+-dependent
K+ current (IK(Ca)). Consistent with
the results obtained previously with nominally zero external
[Ca2+], TEA (10 mM, n = 8) blockade of
IK(Ca) or blockade of Ca2+ channels
with Co2+ (1 mM, n = 2) or Cd2+
(50 µM, n = 2) did not affect either the increase in
conductance induced by IFL application or EIFL.
The influence of internal Ca2+ buffering on IFL actions
also was tested by applying internal BAPTA (10 mM, same internal
[Ca2+]), a faster Ca2+ chelator than EGTA. In
the BAPTA-applied neurons, the increase in conductance during IFL
application was unchanged in magnitude (n = 4) compared
with other neurons recorded with EGTA in the internal pipette solution.
With the caution that internal gluconate may inhibit
IK(Ca) (Velumian et al. 1997), we
tentatively concluded that IFL did not greatly activate
IK(Ca).
We then investigated a possible activation of
IK(ATP) by IFL. For example, glibenclamide, a
specific blocker of IK(ATP), prevents much of
the conductance change caused by anoxia (Politi and Rogawski 1991). During current-clamp recordings and manual clamping at Vh of approximately
60 mV, we monitored input
conductance on injection of small intermittent hyperpolarizing current
pulses. A standardized 2-min application of IFL (4%) was applied
before, during, and after glibenclamide application (1-80 µM,
n = 7). Application of glibenclamide at 40 and 80 µM
(n = 4) tended to increase input resistance and
slightly decreased a positive holding current (implicating a
depolarizing action). However, glibenclamide application did not
significantly change the IFL-induced increase in conductance (Fig.
5A). We concluded from these
results that IFL did not likely activate
IK(ATP).
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The voltage-current relationships obtained with voltage-ramp commands
demonstrated that inward rectification activated on hyperpolarization
just beyond EK (Fig. 5B,
top). In voltage-step experiments in the absence of
K+ channel blockers, the current traces showed that the
inward rectification was fast in onset. In addition, application of low
concentrations of Ba2+ (100 µM) (Sutor and Hablitz
1993) blocked the inward rectification, confirming that it was
due to IKir. Although IFL application increased conductance at potentials positive to EK, we
also observed that IFL decreased fast inward rectification at
potentials negative to EK (Fig. 5B,
top). Subtraction of current obtained under control conditions from the current induced by IFL (4%) resulted in either no
change or a decreased conductance at potentials negative to EK. When fast inward rectification was blocked
by TEA (10 mM), however, the increase in conductance caused by IFL
application was voltage independent at potentials above and below
EK (n = 8, Fig. 2B).
The IFL-evoked decrease in inward rectification likely resulted from an
interaction with channels mediating IKir.
Effects of IFL on nonspiking cells, consistent with K+ efflux from neurons
Five cells were presumed to be glia because of the following
characteristics: 1) relatively negative RMP (79 ± 1 mV), 2) low input resistance (4 ± 3 M
),
3) short membrane time constant (1.5 ± 1.1 ms), and
4) absence of action potentials despite injections of
depolarizing pulses of large amplitude. During voltage clamping of
these "unresponsive" cells at RMP, applications of a high IFL concentration (4%) produced an inward current that was both reversible and reproducible on several repeated applications (n = 5, Fig. 6A). The application
of a high concentration of K+ (9.25 mM) similarly evoked an
inward current (n = 1, Fig. 6A). Because
glial cells behave as a K+-selective electrode, the inward
current may have resulted from a decreased steady-state outward
current. Consistent with a decrease in K+ efflux, both the
IFL- and the K+-induced inward currents were associated
with depolarization.
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Voltage ramps were used during IFL applications to investigate
conductance changes in nonspiking cells. Along with the depolarization, IFL application decreased conductance in a voltage-independent manner
throughout a hyperpolarized voltage range (from 65 to
95 mV,
n = 5, Fig. 6B). This action had a reversal
potential of
83 ± 1 mV, close to EK. A
small positive separation in these values was consistent with a
postulated increase in external [K+] and a corresponding
depolarizing shift in the equilibrium potential. These results were
consistent then with a hypothesis that the IFL-induced depolarization
of nonspiking cells was due to K+ efflux from neurons.
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DISCUSSION |
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These investigations have shown that, in a concentration-dependent
manner, IFL application to thalamocortical neurons increased a
postsynaptic conductance that was linear throughout a range of membrane
voltages that occur during wakefulness and sleep. As described in the
preceding paper (Ries and Puil 1999), the increased
conductance produced a hyperpolarization and inhibited neurons by
shunting voltage-dependent Na+ and Ca2+
currents. If IFL were to have similar actions on the nerve terminal membranes, i.e., a possibility that seems likely, we would expect an
additional decrease in synaptic activity.
Along with the increased conductance, IFL application to neurons at the RMP increased the positive holding current, implicating an outward K+ current. Several observations confirmed a major involvement of a K+ conductance in the IFL actions: 1) the magnitude of the steady-state outward current increased during low [K+] perfusion and decreased during high [K+] perfusion; 2) EIFL was close to the EK calculated for different external [K+] conditions; 3) during application of IFL at high concentrations glia cells depolarized in association with a decrease in membrane conductance, presumably caused by IFL-induced K+ leakage from thalamocortical neurons; and 4) applications of external Ba2+ or internal Cs+, which block leak and other K+ channels but not external TEA or 4-AP, which block voltage-dependent K+ channels, suppressed the increased conductance caused by application of IFL. It seemed likely then that the IFL-induced increase in conductance involved K+ leak channels. The ability of IFL to influence electrical signaling and produce an anesthetic state likely relates to the abundance of nonligand gated K+ leak channels and their important role in determining the RMP.
Is an ionic conductance other than K+ involved in IFL actions?
The increased disparity of EIFL and
EK with decreases in extracellular
[K+] and the correlation of EIFL
to the initial RMP implied a minor contribution of ionic currents other
than K+. Yet the results of experiments where we altered
Na+, Cl, or Ca2+ concentrations
did not implicate an involvement of ions other than K+. A
lack of Cl
involvement in IFL actions may relate to a
higher concentration dependence for Cl
channel activation
as well as conflicting reports of anesthetic-induced potentiation
(Jones et al. 1992
) and depression (El-Beheiry
and Puil 1989a
) of inhibitory postsynaptic potentials (mediated
by GABA). Although experimental limitations may have contributed to an
underestimation of EIFL, the accuracy of
voltage-clamp estimations should be greater for a steady-state
conductance evoked throughout the soma and dendrites than for a
transient dendritic conductance (Spruston et al. 1993
;
see Ries and Puil 1999
). By acting unspecifically on
both somatic and dendritic membranes, however, IFL application would
have caused spatial control to diminish during the voltage-clamp recording. In such a case, an electrotonically distant K+
conductance would have shifted the apparent EIFL
to values more negative than EK and resulted in
an overestimation of EIFL negativity. By using
baclofen and tacrine to verify the calculated
EK, it was apparent that errors resulting from a
reduced external [K+] or a junction potential were not
responsible for the difference between EIFL and
EK. Thus the results are consistent with a major role for an increased K+ leak conductance, with or without
a role for an increased conductance to other ions.
Nonleak K+ conductances
Another possibility is that anesthetics affect a
Ca2+-mediated K+ conductance
(Krnjevi 1974
). Steroid anesthetics such as
althesin or IFL block slow afterhyperpolarizations (AHPs) and
spike-train AHPs activated by Ca2+ entry into neocortical
neurons (El-Beheiry and Puil 1989b
). This contrasts with
observations on hippocampal and cerebellar neurons where ethanol,
pentobarbital, and the benzodiazepines midazolam and clonazepam
increase such Ca2+-mediated K+ conductances
(Carlen et al. 1985
). We found that external
IK(Ca) blockers and chelation of internal
Ca2+ by BAPTA did not suppress IFL actions. Considering
these results together with the experiments on altered Ca2+
concentrations, it does not seem likely that Ca2+ release
or entry contributed significantly to the actions of IFL.
Other noninactivating K+ currents did not contribute to the
IFL-induced increase in steady-state conductance in our studies. In
fact, IFL decreased inward rectification caused by
IKir, probably by shunting this
voltage-dependent current. In addition an action of IFL on
GABAB-activated K+ channels seemed unlikely
here because 4-AP, which reportedly blocks baclofen effects
(Inoue et al. 1985; see also Sugiyama et al.
1992
), did not reduce the hyperpolarization induced by IFL.
More likely are possiblities that anesthetics act on ligand-gated, voltage-independent K+ channels, especially the
muscarine-sensitive K+ leak conductance (Puil and
El-Beheiry 1990
) or the serotonin-sensitive K+
channel (Winegar et al. 1996
).
Comparison with anoxic effects
We considered the analogy that unconsciousness rapidly occurs
during induction of either anesthesia or anoxia in vivo. Like IFL
application to thalamocortical neurons, anoxic insults hyperpolarize CNS neurons by increasing conductance to K+
(Krnjevi and Leblond 1989
). In part
IK(ATP) is responsible for these effects during
energy depletion, as demonstrated by the effects of applications of
specific ATP-sensitive K+ channel blockers. Glibenclamide,
for example, prevents much of the conductance change during acute
anoxia (Politi and Rogawski 1991
). Although
glibenclamide inhibits IFL-induced coronary artery vasodilation
(Cason et al. 1994
) and myocardial protection
(Kersten et al. 1996
), glibenclamide does not affect the
minimum alveolar concentration for IFL in rats (Zucker
1992
). Whereas glibenclamide occasionally is ineffective in
some hippocampal neurons (Erdemli and Krnjevi
1994
), TEA application blocks their anoxic responses (Krnjevi
and Leblond 1989
). In contrast we
observed intact IFL responses during glibenclamide or TEA application
and hence found no evidence for an involvement of
IK(ATP).
Previous studies on inhalational anesthetics
The pioneering studies of Nicoll and Madison (1982) showed that
anesthetics other than IFL increase membrane conductance, probably to
K+, and hyperpolarize hippocampal and spinal neurons of in
vitro preparations. These effects occur despite bicuculline blockade of
GABAA-activated Cl
channels and under low or
high external Cl
conditions. Their observations included
the effects of the inhalational agents diethylether and halothane
during TTX blockade of action potentials. More recent studies confirmed
such pharmacological properties for IFL and halothane in neocortical,
hippocampal, and intralaminar thalamic neurons (Berg-Johnsen and
Langmoen 1990
; El-Beheiry and Puil 1989b
;
Sugiyama et al. 1992
). There are some inconsistencies,
however, because inhalational agents can have insignificant effects on
membrane potential of hippocampal neurons (Miu and Puil
1989
), depending on the concentration or type of agent
(MacIver and Kendig 1991
). While hippocampal neurons
probably participate in consciousness, coherent activity of neurons in the corticothalamocortical system is essential for conscious behavior. Hence the blunted membrane excitability (Ries and Puil
1999
) and disrupted oscillatory activity (Tennigkeit et
al. 1997
) caused by an increased K+ leak
conductance in thalamocortical neurons provide a likely mechanism of
IFL anesthesia in humans.
Significance
The IFL actions are consistent with the historical hydrophobic mechanism involving the lipid membrane and hydrophobic channel proteins. In the current investigations, an eightfold range of IFL concentrations increased the K+ leak conductance without producing a maximal effect. From a pharmacological viewpoint this suggests a nonspecific (i.e., nonreceptor-mediated) mechanism for IFL.
Likely generalized, the enhanced leak conductance caused by IFL shunted the effectiveness of somatic injected current for eliciting voltage transfer to the axon hillock and dendrites. The leak enhancement mechanism accounted for the ability of IFL to annihilate action potentials as well as low- and high-threshold Ca2+ spikes in thalamocortical neurons. The apparently widespread reduction in their membrane excitability, the hydrophobicity of IFL, and its K+ mechanism of action are consistent with the classical viewpoint of "general" (i.e., nonreceptor-mediated) actions for anesthetics.
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ACKNOWLEDGMENTS |
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We thank L. Corey for technical assistance.
The authors acknowledge the financial support of the Medical Research Council of Canada for a fellowship to C. R. Ries, for research grants to E. Puil, and the Department of Anaesthesia of The University of British Columbia for supporting C. R. Ries.
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FOOTNOTES |
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Address for reprint requests: C. R. Ries, Dept. of Pharmacology and Therapeutics, The University of British Columbia, 2176 Health Sciences Mall, Vancouver, British Columbia V6T 1Z3, Canada.
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.
Received 1 June 1998; accepted in final form 14 December 1998.
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REFERENCES |
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