Carl Lynch

Volatile Anesthetics Depress Calcium sup 2+ Transients and Glutamate Release in Isolated Cerebral Synaptosomes

Background The current study was performed to determine whether volatile anesthetics may include as part of their action in the central nervous system the depression of presynaptic transmitter release by alteration in intrasynaptic [Calcium2+] ([Ca2+]i).

Isoflurane, but not halothane, induces protection of human myocardium via adenosine A1 receptors and adenosine triphosphate-sensitive potassium channels.

Background Volatile anesthetics produce differing degrees of myocardial protection in animal models of ischemia. The purpose of the current investigation was to determine the influence of isoflurane and halothane on myocardial protection in a human model of simulated ischemia and the role of adenosine A1 receptors and adenosine triphosphate–sensitive potassium (KATP) channels in the anesthetic pathway. Methods Human atrial trabecular muscles were superfused with oxygenated Krebs-Henseleit buffer and stimulated at 1 Hz, with recording of maximum contractile force. Fifteen minutes before a 30-min anoxic insult, muscles were pretreated for 5 min with either anoxia, the A1 agonist N6-cyclohexyladenosine, 1% halothane or 1.2% isoflurane. These treatments were also performed in the presence of either the KATP channel antagonist glibenclamide or the A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX). Anesthetic effects were also determined on KATP currents in isolated whole cell voltage-clamped human atrial myocytes. Results Recovery of force (recorded 60 min after anoxia) in isoflurane-pretreated muscles was reduced from 76.6 ± 7.5% of baseline to 43.7 ± 7.1% by pretreatment with glibenclamide, and to 52.5 ± 6.2% by pretreatment with DPCPX. Halothane treatment provided no cardioprotection and seemed to inhibit protection by anoxic preconditioning. Halothane decreased whole cell KATP currents in atrial myocytes, whereas isoflurane had no effects. Conclusions This study demonstrates the cardioprotective effects of isoflurane in contrast to the effects of halothane. Furthermore, A1 receptors and KATP channels seem to mediate the beneficial effects of anoxia and isoflurane in human myocardium.

Depression of myocardial contractility in vitro by bupivacaine, etidocaine, and lidocaine

The effects of local anesthetics in depressing myocardial contractility were studied in isolated guinea pig right ventric ular papillary muscles. Bupivacaine and etidocaine, 4 and 10 μM, showed reverse frequency-dependent depression of contractility, that is, less significant depression of contractility at higher stimulation frequencies (2–3 Hz) than at lesser frequencies (<1 Hz). Lidocaine, 40 μM, demon strated a similar trend. In contrast, the normal action potential maximum rate of depolarization (&OV0312;max), a measure of sodium channel conductance, was significantly more de pressed at 2–3 Hz by bupivacaine and etidocaine than by lidocaine. Consequently, contractile depression could be overcome only at higher stimulation frequencies, at which conduction was depressed. To explore the mechanism of the contractile depression, local anesthetic effects were studied on slow (calcium channel-mediated) action potentials in partially depolarized papillary muscles. Etidocaine and bupivacaine, 4 and 10 μM, and lidocaine, 40 and 100 μM, caused a marked depression of the late-peaking contractile responses, attributed to Ca2+ release from the sarcoplasmic reticulum. In contrast, only 10 μM bupivacaine caused any significant depression of the slow action potential rate of depolarization (to 89% of control), consistent with a possible small depression of Ca2 + entry.

Halothane depression of myocardial slow action potentials

Effects of halothane on myocardial electrophysiologic and contractile properties were studied by simultaneous measurement of action potentials (APs) and contractions in guinea pig papillary muscle. Muscles were stimulated by field electrodes and normal responses measured before, during, and after recovery from halothane application. Halothane was administered in 0.5 per cent to 4 per cent concentrations in 5 per cent CO2–95 per cent O2 bubbled through standard Tyrode perfusing solutions. Slow action potentials were then induced with 10−7 isoproterenol in partially depolarized muscles (typically −40 mV in 26 mM K+ media). AP characteristics and accompanying contractions were again measured before, during, and after halothane application. The maximum rate of rise (+&OV0312;max) of the normal (fast) AP was not depressed in any concentration of halothane, although amplitude and duration were decreased in 3 per cent halothane. In contrast, halothane depressed +&OV0312;max of the slow AP to 61 per cent, 28 per cent, and 14 per cent of control, in concentrations of 1, 2 and 3%, respectively. Decreased duration and decreased amplitude (85% of control of the slow AP), or loss of excitability (4 of 7 muscles) occurred in 3 per cent halothane. Initially, halothane application caused a 5 per cent enhancement of tension with both fast and slow APs. In 0.5 per cent halothane, contractions subsequently declined to steady-state levels of 66 per cent (fast AP) and 76 per cent (slow AP) of control. Contractions were depressed linearly with log dose to 18 per cent (fast AP) and 5 per cent (slow AP) of control in 3 per cent halothane. Halothane concentrations of 1 per cent and greater inhibit slow (Na+ – Ca++) channels which mediate the slow action potentials. The negative inotropic effect of halothane may be due in part to decreased Ca++ influx through the slow channel. The negative inotropic effect of 0.5 per cent halothane, in which the slow AP is unaffected, suggests that additional mechanisms, not involving the slow channel, also participate in the negative inotropic action of halothane.

Differential depression of myocardial contractility by halothane and isoflurane in vitro.

Depressant effects of halothane and isoflurane on isolated right ventricular guinea pig papillary muscle bathed in Tyrodes solution at 37°C were examined. Contractions were elicited by stimulation through external field electrodes while tension was recorded continuously and the intracellular cardiac action potential (AP) was monitored simultaneously by microelectrodes. The time differential of tension (dT/dt) and of membrane potential (V) was determined electronically and recorded also. Contractions after rest and at stimulation rates of 0.1, 0.25, 0.5, 1, 2, and 3 Hz were studied. With normal APs, isoflurane (1.3 and 2.5%) depressed peak tension significantly less at high frequencies than did equivalent doses of halothane (0.75 or 1.5%). Isoflurane depressed dT/dt max less than halothane at all frequencies. At 0.3 Hz stimulation, isoflurane (1–4%) significantly increased the normal AP duration by 7–11%. Slow calcium-dependent APs and accompanying contractions were studied in partially depolarized muscles (−40 to −45 mV resting potential in 26mM K+Tyrodes solution) stimulated with 0.1 M isoproterenol. Following rest and at 0.1, 0.25, 0.5, 1, 2, and 3 Hz, both isoflurane (1.3% or 2.5%) and enflurane (1.7% or 3.5%) markedly depressed the late-peaking slow AP contraction observed with low-frequency stimulation. Halothane (0.75% or 1.5%) caused a similar contractile depression (40–60%) at all frequencies. In contrast, isoflurane depressed early peaking tension and the dT/dt max at frequencies greater than 1 Hz significantly less than did halothane or enflurane. At 0.3 Hz, 2% and 4% isoflurane caused 9% and 17% depression of slow AP maximum rate of depolarization (Vmax), but significantly prolonged the AP duration. Isoflurane altered the pattern of tension development in a different manner than halothane, suggesting differing mechanisms of myocardial depression by these anesthetics.

Effects of Propofol and Thiopental in Isolated Rat Aorta and Pulmonary Artery

This study was performed to determine if direct arterial dilating actions of propofol contribute to the drugs hypotensive actions. The effects of propofol were compared with those of thiopental on isolated vascular ring preparations from rat thoracic aorta and pulmonary artery. Thoracic aortic ring responses were evaluated in the presence and absence of endothelium, indomethacin, and N omega-nitro-L-arginine methyl ester (LNAME; a specific inhibitor of endothelium-derived relaxing factor-nitric oxide [EDRF/NO] synthase). Pulmonary artery responses were investigated with intact endothelium. After the induction of active isometric force by a predetermined EC50 dose of phenylephrine for each ring, effects of propofol (30, 100, 300 microM) and thiopental (10, 30, 100 microM) were examined. Propofol caused significant vasodilation in endothelium-intact, endothelium-denuded, and LNAME-treated aortic rings. In the endothelium-intact aortic and pulmonary artery rings, the initial vasodilation due to 30 and 100 microM propofol showed gradual and partial recovery over 15 min; 300 microM propofol caused sustained vasodilation. Endothelium-denuded rings and LNAME-pretreated endothelium-intact rings showed constant and sustained vasodilation with all propofol concentrations. Propofol also caused marked vasodilation in pulmonary arteries. In contrast, thiopental had no vasodilating effect in aortic or pulmonary artery preparations. In control experiments, propofol vehicle (Intralipid) also had no effect on vascular rings. Indomethacin pretreatment induced a dose-dependent vasoconstriction by thiopental in endothelium-intact rings and decreased the vasodilation due to propofol.(ABSTRACT TRUNCATED AT 250 WORDS)

Propofol and Thiopental Depression of Myocardial Contractility A Comparative Study of Mechanical and Electrophysiologic Effects in Isolated Guinea Pig Ventricular Muscle

The purpose of the study was to compare the actions of propofol and thiopental on myocardial contractility and cellular electrophysiologic behavior. Isometric tension of isolated guinea pig right ventricular papillary muscle was studied in normal and 26 mM potassium Tyrodes solutions at various stimulation rates (after rest up to 3 Hz). Normal and slow action potentials were also recorded by conventional microelectrodes. Propofol (30,100, and 300 μM) applied in the commercial 10% Intralipid emulsion caused dose-dependent depression of contractions at all stimulation rates, whereas Intralipid alone had no effect. Thiopental (10, 30, and 100 μM) caused depression similar to the threefold greater concentrations of propofol. Although neither drug altered the normal action potential (AP) amplitude or dV/dt max, thiopental (30 μM) increased AP duration. In the partially depolarized (26 mM potassium) β-adrenergically stimulated myocardium, propofol and thiopental caused dose-dependent contractile depression similar to that in normal Tyrodes solution. Whereas propofol did not alter slow AP characteristics, 30–100 μM thiopental increased slow AP duration (consistent with decreased potassium conductance), and 100 μM thiopental depressed dV/dt max (consistent with decreased calcium channel ionic influx). Comparing the clinical plasma concentration ranges required for an equivalent anesthetic effect, propofol depresses myocardial contractility less than thiopental.

Convergent and reciprocal modulation of a leak K+ current and Ih by an inhalational anaesthetic and neurotransmitters in rat brainstem motoneurones

Neurotransmitters and volatile anaesthetics have opposing effects on motoneuronal excitability which appear to reflect contrasting modulation of two types of subthreshold currents. Neurotransmitters increase motoneuronal excitability by inhibiting TWIK‐related acid‐sensitive K+ channels (TASK) and shifting activation of a hyperpolarization‐activated cationic current (Ih) to more depolarized potentials; on the other hand, anaesthetics decrease excitability by activating a TASK‐like current and inducing a hyperpolarizing shift in Ih activation. Here, we used whole‐cell recording from motoneurones in brainstem slices to test if neurotransmitters (serotonin (5‐HT) and noradrenaline (NA)) and an anaesthetic (halothane) indeed compete for modulation of the same ion channels ‐ and we determined which prevails. When applied together under current clamp conditions, 5‐HT reversed anaesthetic‐induced membrane hyperpolarization and increased motoneuronal excitability. Under voltage clamp conditions, 5‐HT and NA overcame most, but not all, of the halothane‐induced current. When Ih was blocked with ZD 7288, the neurotransmitters completely inhibited the K+ current activated by halothane; the halothane‐sensitive neurotransmitter current reversed at the equilibrium potential for potassium (EK) and displayed properties expected of acid‐sensitive, open‐rectifier TASK channels. To characterize modulation of Ih in relative isolation, effects of 5‐HT and halothane were examined in acidified bath solutions that blocked TASK channels. Under these conditions, 5‐HT and halothane each caused their characteristic shift in voltage‐dependent gating of Ih. When tested concurrently, however, halothane decreased the neurotransmitter‐induced depolarizing shift in Ih activation. Thus, halothane and neurotransmitters converge on TASK and Ih channels with opposite effects; transmitter action prevailed over anaesthetic effects on TASK channels, but not over effects on Ih. These data suggest that anaesthetic actions resulting from effects on either TASK or hyperpolarization‐activated cyclic nucleotide‐gated (HCN) channels in motoneurones, and perhaps at other CNS sites, can be modulated by prevailing neurotransmitter tone.

Multiple ionic mechanisms mediate inhibition of rat motoneurones by inhalation anaesthetics

1 We studied the effects of inhalation anaesthetics on the membrane properties of hypoglossal motoneurones in a neonatal rat brainstem slice preparation. 2 In current clamp, halothane caused a membrane hyperpolarization that was invariably associated with decreased input resistance; in voltage clamp, halothane induced an outward current and increased input conductance. Qualitatively similar results were obtained with isoflurane and sevoflurane. 3 The halothane current reversed near the predicted K+ equilibrium potential (EK) and was reduced in elevated extracellular K+ and in the presence of Ba2+ (2 mm). Moreover, the Ba2+‐sensitive component of halothane current was linear and reversed near EK. The halothane current was not sensitive to glibenclamide or thyrotropin‐releasing hormone (TRH). Therefore, the halothane current was mediated, in part, by activation of a Ba2+‐sensitive K+ current distinct from the ATP‐ and neurotransmitter‐sensitive K+ currents in hypoglossal motoneurones. 4 Halothane also inhibited Ih, a hyperpolarization‐activated cationic current; this was primarily due to a decrease in the absolute amount of current, although halothane also caused a small, but statistically significant, shift in the voltage dependence of Ih activation. Extracellular Cs+ (3 mm) blocked Ih and a component of halothane‐sensitive current with properties reminiscent of Ih. 5 A small component of halothane current, resistant to Ba2+ and Cs+, was observed in TTX‐containing solutions at potentials depolarized to ∼−70 mV. Partial Na+ substitution by N‐methyl‐D‐glucamine completely abolished this residual current, indicating that halothane also inhibited a TTX‐resistant Na+ current active near rest potentials. 6 Thus, halothane activates a Ba2+‐sensitive, relatively voltage‐independent K+ current and inhibits both Ih and a TTX‐insensitive persistent Na+ current in hypoglossal motoneurones. These effects of halothane decrease motoneuronal excitability and may contribute to the immobilization that accompanies inhalation anaesthesia.

HCN Subunit-Specific and cAMP-Modulated Effects of Anesthetics on Neuronal Pacemaker Currents

General anesthetics have been a mainstay of surgical practice for more than 150 years, but the mechanisms by which they mediate their important clinical actions remain unclear. Ion channels represent important anesthetic targets, and, although GABAA receptors have emerged as major contributors to sedative, immobilizing, and hypnotic effects of intravenous anesthetics, a role for those receptors is less certain in the case of inhalational anesthetics. The neuronal hyperpolarization-activated pacemaker current (Ih) is essential for oscillatory and integrative properties in numerous cell types. Here, we show that clinically relevant concentrations of inhalational anesthetics modulate neuronal Ih and the corresponding HCN channels in a subunit-specific and cAMP-dependent manner. Anesthetic inhibition of Ih involves a hyperpolarizing shift in voltage dependence of activation and a decrease in maximal current amplitude; these effects can be ascribed to HCN1 and HCN2 subunits, respectively, and both actions are recapitulated in heteromeric HCN1-HCN2 channels. Mutagenesis and simulations suggest that apparently distinct actions of anesthetics on V1/2 and amplitude represent different manifestations of a single underlying mechanism (i.e., stabilization of channel closed state), with the predominant action determined by basal inhibition imposed by individual subunit C-terminal domains and relieved by cAMP. These data reveal a molecular basis for multiple actions of anesthetics on neuronal HCN channels, highlight the importance of proximal C terminus in modulation of HCN channel gating by diverse agents, and advance neuronal pacemaker channels as potentially relevant targets for clinical actions of inhaled anesthetics.