Suppression of Central Nervous System Sodium Channels by Propofol

Background: Previous studies have provided evidence that clinical levels of propofol alter the functions of voltage-dependent sodium channels, thereby inhibiting synaptic release of glutamate. However, most of these experiments were conducted in the presence of sodium-channel activators, which alter channel inactivation. This study electrophysiologically characterized the interactions of propofol with unmodified sodium channels.

Methods: Sodium currents were measured using whole-cell patch-clamp recordings of rat brain IIa sodium channels expressed in a stably transfected Chinese hamster ovary cell line. Standard electrophysiologic protocols were used to record sodium currents in the presence or absence of externally applied propofol.

Results: Propofol, at concentrations achieved clinically in the brain, significantly altered sodium channel currents by two mechanisms: a voltage-independent block of peak currents and a concentration-dependent shift in steady-state inactivation to hyperpolarized potentials, leading to a voltage dependence of current suppression. The two effects combined to give an apparent concentration yielding a half-maximal inhibitory effect of 10 [micro sign]M near the threshold potential of action potential firing (about -60 mV). Propofol inhibition was also use-dependent, causing a further block of sodium currents at these anesthetic concentrations.     

Interaction of Volatile Anesthetics with Human Kv Channels in Relation to Clinical Concentrations

Background: Recent evidence shows that inhibition of human Kv3 channels by intravenous anesthetics occurs at clinical concentrations. The effects of volatile anesthetics on these human ion channels are unknown. This study was designed to establish whether minimum alveolar concentrations (MAC) of halothane, enflurane, isoflurane, and desflurane exhibit effects on Kv3 channels. To obtain an indication whether these findings may be specific to Kv3 channels, the effects of enflurane and isoflurane on human Kv1.1 channels were also investigated.

Methods: Kv3 channels natively expressed in SH-SY5Y cells and Kv1.1 channels expressed in HEK293 cells were measured with the whole cell patch clamp technique by standard protocols. Concentrations of volatile anesthetics were determined by gas chromatography.

Results: Halothane, enflurane, isoflurane, and desflurane reversibly inhibited Kv3 channels in a concentration-dependent manner. Concentrations at half-maximal effect (IC50 values) ranged between 1,800 and 4,600 [mu]m. Hill coefficients were between 1.7 and 2.5. IC50 values for inhibition of Kv1.1 channels were 2,800 and 5,200 [mu]m, and Hill coefficients were 3.9 and 5.6 for enflurane and isoflurane, respectively.     

Inhibition of Sodium/Calcium Exchange and Calcium Channels of Heart Cells by Volatile Anesthetics

Background: Volatile anesthetics exert profound effects on the heart, probably through their effect on Calcium2+ movements during the cardiac cycle. Calcium2+ movements across the sarcolemma are thought to involve mainly Calcium2+ channels and the Sodium sup +/Calcium2+ exchanger. We have therefore investigated the action of halothane, isoflurane, and enflurane on Sodium sup +/Calcium2+ exchange and Calcium2+ channel activity to assess the contribution of these pathways to the observed effect of the anesthetics on the myocardium.

Methods: Sarcolemmal ion fluxes were investigated using radioisotope uptake by isolated adult rat heart cells in suspension. Sodium sup +/Calcium2+ exchange activity was measured from45 Calcium2+ uptake by Sodium sup + -loaded cells. Calcium2+ channel activity was measured from verapamil-sensitive trace54 Manganese2+ uptake during electric stimulation.

Results: Halothane, isoflurane, and enflurane inhibited Sodium sup +/Calcium2+ exchange completely, with similar potency when concentrations were expressed in millimolar units in aqueous medium but not when expressed as minimum alveolar concentration (MAC). The inhibition by enflurane was particularly strong, > 50%, at 2 MAC. In contrast, the three anesthetics inhibited Calcium2+ channels with similar potency when concentrations were expressed as MAC but not when expressed in millimolar units in aqueous medium. Hill plots of pooled data with all three anesthetics showed a slope of 3.87 plus/minus 0.50 for inhibition of Sodium sup +/Calcium2+ exchange and 1.73 plus/minus 0.19 for inhibition of Calcium2+ channels.     

Central Nervous System Concentrations of Cyclooxygenase-2 Inhibitors in Humans

Background: Cyclooxygenase-2 (COX-2)-selective inhibitors (coxibs) are under investigation for the potential therapy, attenuation, or prevention of neuroinflammatory and neurodegenerative disorders. Coxibs are also a significant advance in pain therapy and are traditionally considered to achieve analgesia via peripheral effects. However, in animals, central nervous system (CNS) COX-2 activity and prostanoid concentrations are increased by peripheral inflammation, central sensitization has been proposed to account for long-term pain-related phenomena, and coxibs achieve significant cerebrospinal fluid (CSF) concentrations and may cause analgesia via CNS action. Nevertheless, it remains unknown whether or which coxibs reach the CNS in humans. This investigation determined whether coxibs can reach the CNS in humans, based on CSF concentrations.

Methods: Ten healthy human volunteers simultaneously received a single oral dose of celecoxib (200 mg), rofecoxib (50 mg), and valdecoxib (40 mg). Blood and CSF were serially sampled for 10 h, and plasma total and unbound and CSF coxib concentrations were quantified by mass spectrometry.

Results: Total plasma concentrations and time to maximum plasma concentration were similar among the three coxibs. In contrast, unbound (free) plasma concentrations differed significantly. Maximum unbound plasma concentrations were 1.4 +/- 0.5, 42 +/- 17, and 6.0 +/- 2.9 ng/ml, respectively, for celecoxib, rofecoxib, and valdecoxib. COX-2 inhibitors rapidly penetrated the CNS. Maximum CSF concentrations were 2 +/- 2, 57 +/- 25, and 10 +/- 4 ng/ml, respectively, for celecoxib, rofecoxib, and valdecoxib. CSF concentrations exceeding the median inhibitory concentration for COX-2 were achieved by rofecoxib and valdecoxib but not celecoxib.     

Modulation of Cardiac Sodium Current by alpha sub 1 -stimulation and Volatile Anesthetics

Background: alpha1 -adrenoceptor stimulation is known to produce electrophysiologic changes in cardiac tissues, which may involve modulations of the fast inward Na sup + current (INa). A direct prodysrhythmic alpha1 -mediated interaction between catecholamines and halothane has been demonstrated, supporting the hypothesis that generation of halothane-epinephrine dysrhythmias may involve slowed conduction, leading to reentry. In this study, we examined the effects of a selective alpha1 -adrenergic receptor agonist, methoxamine, on cardiac INa in the absence and presence of equianesthetic concentrations of halothane and isoflurane in single ventricular myocytes from adult guinea pig hearts.

Methods: INa was recorded using the standard whole-cell configuration of the patch-clamp technique. Voltage clamp protocols initiated from two different holding potentials (VH) were applied to examine state-dependent effects of methoxamine in the presence of anesthetics. Steady state activation and inactivation and recovery from inactivation were characterized using standard protocols.

Results: Methoxamine decreased INa in a concentration- and voltage-dependent manner, being more potent at the depolarized VH. Halothane and isoflurane interacted synergistically with methoxamine to suppress INa near the physiologic cardiac resting potential of -80 mV. The effect of methoxamine with anesthetics appeared to be additive when using a VH of -110 mV, a potential where no Na sup + channels are in the inactivated state. Methoxamine in the absence and presence of anesthetics significantly shifted the half maximal inactivation voltage in the hyperpolarizing direction but had no effect on steady-state activation.     

Effects of Small Concentrations of Volatile Anesthetics on Action Potential Firing of Neocortical Neurons In Vitro

Background: Volatile general anesthetics depress neuronal activity in the mammalian central nervous system and enhance inhibitory Cl- currents flowing across the [Greek small letter gamma]-aminobutyric acidA (GABA (A)) receptor - ion channel complex. The extent to which an increase in GABAA-mediated synaptic inhibition contributes to the decrease in neuronal firing must be determined, because many further effects of these agents have been reported on the molecular level.

Methods: The actions of halothane, isoflurane, and enflurane on the firing patterns of single neurons were investigated by extracellular recordings in organotypic slice cultures derived from the rat neocortex.

Results: Volatile anesthetics depressed spontaneous action potential firing of neocortical neurons in a concentration-dependent manner. The estimated median effective concentration (EC50) values were about one half the EC50 values for general anesthesia. In the presence of the GABA (A) antagonist bicuculline (20 [micro sign]M), the effectiveness of halothane, isoflurane, and enflurane in reducing the discharge rates were diminished by 48 - 65%, indicating that these drugs act via the GABAA receptor.     

Ketamine Blockade of Voltage-gated Sodium Channels: Evidence for a Shared Receptor Site with Local Anesthetics

Background : The general anesthetic ketamine is known to be an N-methyl-d-aspartate receptor blocker. Although ketamine also blocks voltage-gated sodium channels in a local anesthetic-like fashion, little information exists on the molecular pharmacology of this interaction. We measured the effects of ketamine on sodium channels.

Methods : Wild-type and mutant (F1579A) recombinant rat skeletal muscle sodium channels were expressed in Xenopus oocytes. The F1579A amino acid substitution site is part of the intrapore local anesthetic receptor. The effect of ketamine was measured in oocytes expressing wild-type or mutant sodium channels using two-electrode voltage clamp.

Results : Ketamine blocked sodium channels in a local anesthetic-like fashion, exhibiting tonic blockade (concentration for half-maximal inhibition [IC50] = 0.8 mm), phasic blockade (IC50 = 2.3 mm), and leftward shift of the steady-state inactivation; the parameters of these actions were strongly modified by alteration of the intrapore local anesthetic binding site (IC50 = 2.1 mm and IC50 = 10.3 mm for tonic and phasic blockade, respectively). Compared with lidocaine, ketamine showed greater tonic inhibition but less phasic blockade.     

Volatile General Anesthetics Produce Hyperpolarization of Aplysia Neurons by Activation of a Discrete Population of Baseline Potassium Channels

Background: The mechanism by which volatile anesthetics act on neuronal tissue to produce reversible depression is unknown. Previous studies have identified a potassium current in invertebrate neurons that is activated by volatile anesthetics. The molecular components generating this current are characterized here in greater detail.

Methods: The cellular and biophysical effects of halothane and isoflurane on neurons of Aplysia californica were studied. Isolated abdominal ganglia were perfused with anesthetic-containing solutions while membrane voltage changes were recorded. These effects were also studied at the single-channel level by patch clamping cultured neurons from the abdominal and pleural ganglia.

Results: Clinically relevant concentrations of halothane and isoflurane produced a slow hyperpolarization in abdominal ganglion neurons that was sufficient to block spontaneous spike firings. Single-channel studies revealed specific activation by volatile anesthetics of a previously described potassium channel. In pleural sensory neurons, halothane and isoflurane increased the open probability of the outwardly rectifying serotonin-sensitive channel (S channel). Halothane also inhibited a smaller noninactivating channel with a linear slope conductance of approximately 40 pS. S channels were activated by halothane with a median effective concentration of approximately 500 micro Meter (0.013 atm), which increased channel activity about four times. The mechanism of channel activation involved shortening the closed-time durations between bursts and apparent recruitment of previously silent channels.     

Interaction of Intravenous Anesthetics with Human Neuronal Potassium Currents in Relation to Clinical Concentrations

Background: Neuronal voltage-dependent potassium (K) currents are crucial for various cellular functions, such as the integration of temporal information in the central nervous system. Data for the effects of intravenous anesthetics on human neuronal K currents are limited. It was the authors' aim to evaluate the concentration-related effects of three opioids (fentanyl, alfentanil, sufentanil) and seven nonopioids (thiopental, pentobarbital, methohexital, propofol, ketamine, midazolam, droperidol) used in clinical anesthesia on neuronal voltage-dependent K currents of human origin.

Method: K currents were measured in SH-SY5Y cells using the whole cell patch-clamp technique. Currents were elicited by step depolarization from a holding potential of -80 to -50 mV through +90 mV, and their steady state amplitudes were determined.

Results: All drugs inhibited the K currents in a concentration-dependent and reversible manner. Because time dependence of inhibition differed among the drugs, effects were measured after 54-64 ms of the test pulse. The IC50 values (concentration of half-maximal inhibition) for current suppression ranged from 7 [mu]M for sufentanil to 2 mM for pentobarbital. Suppression of the K currents by the opioids occurred at 10-fold lower IC50 values (concentration of half-maximal inhibition) than that by the barbiturates. As estimated from the concentration-response curves, K-current suppression at clinical concentrations would be less than 0.1% for the opioids and approximately 3% for the other drugs.     

General Anesthetics Modify the Kinetics of Nicotinic Acetylcholine Receptor Desensitization at Clinically Relevant Concentrations

Background: General anesthetics are thought to induce anesthesia through their actions on ligand-gated ion channels. One such channel, the nicotinic acetylcholine receptor (nAcChoR), can be found in different subtypes in the central nervous system and at the periphery in the neuromuscular junction. The latter subtype of the nAcChoR is a useful model for examining interactions between general anesthetics and ligand-gated ion channels, because it can be isolated and purified in sufficient quantities to allow for biophysical and biochemical studies. This study examines the actions of general anesthetics on agonist-induced conversion of the nAcChoR to inactive desensitized conformational states.

Methods: Nicotinic acetylcholine receptor membranes were purified from the electric organ of Torpedo nobiliana. Agonist-induced desensitization was characterized from the time-dependent increase in fluorescence intensity that results from the binding of the fluorescent acetylcholine analog, Dns-C6 -Cho, to the nAcChoR.

Results: Mixing Dns-C6 -Cho with nAcChoR-rich membranes results in an increase in fluorescence that is characterized by four rate processes. Concentrations of isoflurane and butanol, which range from subclinical to toxic increase the rates of the third and fourth components of fluorescence, corresponding to fast and slow desensitization, respectively. At concentrations that are twice their EC sub 50 s for anesthesia, isoflurane, butanol, chloroform, methanol, and cyclopentane-methanol increase the apparent rates of fast and slow desensitization by an average of 92 plus/minus 22% and 108 plus/minus 22%, respectively.     

The Renin-Angiotensin-Aldosterone System During Haemodialysis With Acetate or Bicarbonate at Different Dialysate Sodium Concentrations

The hormones of the renin–angiotensin–aldosteronesystem were measured during regular haemodialysis with acetateor bicarbonate at dialysate sodium concentrations of 135, 140,145, and 150 mmol/l. Plasma renin activity and aldosterone concentrationwere higher during acetate haemodialysis than during bicarbonatehaemodialysis. At lower dialysate sodium concentrations, plasmarenin activity (acetate dialysis and bicarbonate dialysis) andaldosterone concentration (only acetate dialysis) were higherthan they were at higher dialysate sodium concentrations. Plasmarenin activity increased during acetate dialysis, but did notchange during bicarbonate dialysis. Aldosterone and potassiumconcentrations were positively correlated. Aldosterone decreasedduring haemodialysis (increase to predialysis values at theend of haemodialysis (4 h) at lower dialysate sodium concentrations).It is concluded that the renin-angiotensin-aldosterone systemis activated more during acetate dialysis than during bicarbonatedialysis. Aldosterone concentrations seem to be related moreclosely to serum potassium than to renin–angiotensin–aldosteronesystem and to serum sodium intradialytically.