Takashi Akata

Effects of volatile anesthetics on acetylcholine-induced relaxation in the rabbit mesenteric resistance artery

Background Vascular endothelium plays an important role in the regulation of vascular tone. Volatile anesthetics have been shown to attenuate endothelium‐mediated relaxation in conductance arteries, such as aorta. However, significant differences in volatile anesthetic pharmacology between these large vessels and the small vessels that regulate systemic vascular resistance and blood flow have been documented, yet little is known about volatile anesthetic action on endothelial function in resistance arteries. Furthermore, endothelium‐dependent relaxation mediated by factors other than endothelium‐derived relaxing factor (EDRF) has recently been recognized, and there is no information available regarding volatile anesthetic action on non‐EDRF‐mediated endothelium‐dependent relaxation. Methods Employing isometric tension recording and microelectrode methods, the authors first characterized the endothelium‐dependent relaxing and hyperpolarizing actions of acetylcholine (ACh) in rabbit small mesenteric arteries, and tested the sensitivities of these actions to EDRF pathway inhibitors and Potassium sup + channel blockers. They then examined the effects of the volatile anesthetics isoflurane, enflurane, and sevoflurane on ACh‐induced endothelium‐dependent relaxation that was sensitive to EDRF inhibitors and that which was resistant to the EDRF inhibitors but sensitive to blockers of ACh‐induced hyperpolarization. The effects of the volatile anesthetics on endothelium‐independent sodium nitroprusside (SNP)‐induced relaxation were also studied. Results Acetylcholine concentration‐dependently caused both endothelium‐dependent relaxation and hyperpolarization of vascular smooth muscle. The relaxation elicited by low concentrations of ACh (less or equal to 0.1 micro Meter) was almost completely abolished by the EDRF inhibitors NG ‐nitro L‐arginine (LNNA), oxyhemoglobin (HbO sub 2), and methylene blue (MB). The relaxation elicited by higher concentrations of ACh (greater or equal to 0.3 micro Meter) was only attenuated by the EDRF inhibitors. The remaining relaxation, as well as the ACh‐induced hyperpolarization that was also resistant to EDRF inhibitors, were both specifically blocked by tetraethylammonium (TEA greater or equal to 10 mM). Sodium nitroprusside, a NO donor, produced dose‐dependent relaxation, but not hyperpolarization, in the endothelium‐denuded (E[‐]) strips, and the relaxation was inhibited by MB and HbO2, but not TEA (greater or equal to 10 mM). One MAC isoflurane, enflurane, and sevoflurane inhibited both ACh relaxation that was sensitive to the EDRF inhibitors and the ACh relaxation resistant to the EDRF inhibitors and sensitive to TEA, but not SNP relaxation (in the E[‐] strips). An additional finding was that the anesthetics all significantly inhibited norepinephrine (NE) contractions in the presence and absence of the endothelium or after exposure to the EDRF inhibitors. Conclusions The results confirm that ACh has a hyperpolarizing action in rabbit small mesenteric resistance arteries that is independent of EDRF inhibitors but blocked by the Potassium sup + channel blocker TEA. The ACh relaxation in these resistance arteries thus appears to consist of distinct EDRF‐mediated and hyperpolarization‐mediated components. Isoflurane, enflurane, and sevoflurane inhibited both components of the ACh‐induced relaxation in these small arteries, indicating a more global depression of endothelial function or ACh signaling in endothelial cells, rather than a specific effect on the EDRF pathway. All these anesthetics exerted vasodilating action in the presence of NE, the primary neurotransmitter of the sympathetic nervous system, which plays a major role in maintaining vasomotor tone in vivo. This strongly indicates that the vasodilating action of these anesthetics probably dominates over their inhibitory action on the EDRF pathway and, presumably, contributes to their known hypotensive effects in vivo. Finally, the vasodilating action of these anesthetics is, at least in part, independent from endothelium.

General anesthetics and vascular smooth muscle: direct actions of general anesthetics on cellular mechanisms regulating vascular tone.

General anesthetics threaten cardiovascular stability by causing changes in cardiac function, vascular reactivity, and cardiovascular reflexes and significantly alter distribution of cardiac output to various organs. Their overall impact is often systemic hypotension, which is attributable to myocardial depression, peripheral vasodilation, and attenuated sympathetic nervous system activity. However, one could be more causative than the others, depending on anesthetic agents and cardiovascular factors inherent in patients (e.g., coexisting heart disease). It is generally believed that most general anesthetics attenuate sympathetic nervous system outflow from the central nervous system, thereby decreasing vascular resistance in peripheral circulations. Indeed, in previous in vivo studies, during administration of various general anesthetics, vascular resistance was decreased in most peripheral circulations; however, it was unaffected or increased in some peripheral circulations. General anesthetics may act directly on vascular smooth muscle and/or endothelial cells in various vascular beds, influencing total peripheral and/or regional vascular resistance, and hence organ blood flow. This article reviews previously reported direct (i.e., nonneural) vascular actions of general anesthetics and discusses their underlying mechanisms, their in vivo relevance, and the future of research for general anesthetic vascular pharmacology.

Mechanisms of Direct Inhibitory Action of Isoflurane on Vascular Smooth Muscle of Mesenteric Resistance Arteries

Background Isoflurane has been shown to directly inhibit vascular reactivity. However, less information is available regarding its underlying mechanisms in systemic resistance arteries. Methods Endothelium-denuded smooth muscle strips were prepared from rat mesenteric resistance arteries. Isometric force and intracellular Ca2+ concentration ([Ca2+]i) were measured simultaneously in the fura-2-loaded strips, whereas only the force was measured in the &bgr;-escin membrane-permeabilized strips. Results Isoflurane (3–5%) inhibited the increases in both [Ca2+]i and force induced by either norepinephrine (0.5 &mgr;m) or KCl (40 mm). These inhibitions were similarly observed after depletion of intracellular Ca2+ stores by ryanodine. Regardless of the presence of ryanodine, after washout of isoflurane, its inhibition of the norepinephrine response (both [Ca2+]i and force) was significantly prolonged, whereas that of the KCl response was quickly restored. In the ryanodine-treated strips, the norepinephrine- and KCl-induced increases in [Ca2+]i were both eliminated by nifedipine, a voltage-gated Ca2+ channel blocker, whereas only the former was inhibited by niflumic acid, a Ca2+-activated Cl− channel blocker. Isoflurane caused a rightward shift of the Ca2+-force relation only in the fura-2-loaded strips but not in the &bgr;-escin-permeabilized strips. Conclusions In mesenteric resistance arteries, isoflurane depresses vascular smooth muscle reactivity by directly inhibiting both Ca2+ mobilization and myofilament Ca2+ sensitivity. Isoflurane inhibits both norepinephrine- and KCl-induced voltage-gated Ca2+ influx. During stimulation with norepinephrine, isoflurane may prevent activation of Ca2+-activated Cl− channels and thereby inhibit voltage-gated Ca2+ influx in a prolonged manner. The presence of the plasma membrane appears essential for its inhibition of the myofilament Ca2+ sensitivity.

Volatile anesthetic actions on contractile proteins in membrane-permeabilized small mesenteric arteries

Background Volatile anesthetics have been shown to have vasodilating or vasoconstricting actions in vitro that may contribute to their cardiovascular effects in vivo. However, the precise mechanisms of these actions in vitro have not been fully elucidated. Moreover, there are no data regarding the mechanisms of volatile anesthetic action on small resistance arteries, which play a critical role in the regulation of blood pressure and blood flow. Methods With the use of isometric tension recording methods, volatile anesthetic actions were studied in intact and beta‐escin‐membrane‐permeabilized smooth muscle strips from rat small mesenteric arteries. In experiments with intact muscle, the effects of halothane (0.25–5.0%), isoflurane (0.25–5.0%), and enflurane (0.25–5.0%) were investigated on high Potassium sup + ‐induced contractions at 22 degrees Celsius and 35 degrees Celsius. All experiments were performed on endothelium‐denuded strips in the presence of 3 micro Meter guanethidine and 0.3 micro Meter tetrodotoxin to minimize the influence of nerve terminal activities. In experiments with membrane‐permeabilized muscle, the effects of halothane (0.5–4.0%), isoflurane (0.5–4.0%), and enflurane (0.5–4.0%) on the half‐maximal and maximal Calcium2+ ‐activated contractions were examined at 22 degrees Celsius in the presence of 0.3 micro Meter ionomycin to eliminate intracellular Calcium sup 2+ stores. Results In the high Potassium sup + ‐stimulated intact muscle, all three anesthetics generated transient contractions, which were followed by sustained vasorelaxation. The IC50 values for this vasorelaxing action of halothane, isoflurane, and enflurane were 0.47 vol% (0.27 mM), 0.66 vol% (0.32 mM), and 0.53 vol% (0.27 mM), respectively, at 22 degrees Celsius and were 3.36 vol% (0.99 mM), 3.07 vol% (0.69 mM), and 3.19 vol% (0.95 mM), respectively, at 35 degrees Celsius. Ryanodine (10 micro Meter) eliminated the anesthetic‐induced contractions but had no significant effect on the anesthetic‐induced vasorelaxation in the presence of high Potassium sup +. In addition, no significant differences were observed in the dose dependence of the direct vasodilating action among these anesthetics with or without ryanodine at either the low or the high temperature. However, significant differences were observed in the vasoconstricting actions among the anesthetics, and the order of potency was halothane > enflurane > isoflurane. The Calcium sup 2+ ‐tension relation in the membrane‐permeabilized muscle yielded a half‐maximal effective Calcium2+ concentration (EC50) of 2.02 micro Meter. Halothane modestly but significantly inhibited 3 micro Meter (approximately the EC50) and 30 micro Meter (maximal) Calcium sup 2+ ‐induced contractions. Enflurane slightly but significantly inhibited 3 micro Meter but not 30 micro Meter Calcium2+ contractions. Isoflurane did not significantly inhibit either 3 micro Meter or 30 micro Meter Calcium2+ contractions. Conclusions Halothane, isoflurane, and enflurane have both vasoconstricting and vasodilating actions on isolated small splanchnic resistance arteries. The direct vasoconstricting action appears to result from Calcium2+ release from the ryanodine‐sensitive intracellular Calcium2+ store. The vasodilating action of isoflurane in the presence of high Potassium sup + appears to be attributable mainly to a decrease in intracellular Calcium2+ concentration, possibly resulting from inhibition of voltage‐gated Calcium2+ channels. In contrast, the vasodilating actions of halothane and enflurane in the presence of high Potassium sup + appears to involve inhibition of Calcium2+ activation of contractile proteins as well as a decrease in intracellular Calcium2+ concentration in smooth muscle.

The Action of Sevoflurane on Vascular Smooth Muscle of Isolated Mesenteric Resistance Arteries (Part 1)Role of Endothelium

Background The direct action of sevoflurane on systemic resistance arteries is not fully understood. Methods Isometric force was recorded in isolated rat small mesenteric arteries. Results Sevoflurane (2–5%) enhanced contractile response to norepinephrine only in the presence of endothelium, but inhibited it in its absence. Sevoflurane still enhanced the norepinephrine response after inhibitions of the nitric oxide, endothelium-derived hyperpolarizing factor, cyclooxygenase and lipoxygenase pathways, or after blockade of either endothelin-1 ET-1), angiotensin-II, or sevotonin receptors. Sevoflurane (3–5%) inhibited contractile response to potassium chloride only in the absence of endothelium but did not influence it in its presence. In the endothelium-intact strips, inhibition of the norepinephrine response, which was enhanced during application of sevoflurane, was observed after washout of sevoflurane and persisted for approximately 15 min. In the endothelium-denuded strips, the inhibition of norepinephrine response was similarly prolonged after washout of sevoflurane. However, no significant inhibitions of potassium chloride response were observed after washout of sevoflurane in both the endothelium-intact and the endothelium-denuded strips. Conclusions The action of sevoflurane on norepinephrine contractile response consists of endothelium-dependent vasoconstricting and endothelium-independent vasodilating components. In the presence of endothelium, the former predominates over the latter, enhancing the norepinephrine response. The endothelium-independent component persisted after washout of sevoflurane, leading to prolonged inhibition of the norepinephrine response. The mechanisms behind the sevoflurane-induced inhibition of norepinephrine response are at least in part different from those behind its inhibition of potassium chloride response. Nitric oxide, endothelium-derived hyperpolarizing factor, cyclooxygenase products, lipoxygenase products, endothelin-1, angiotensin-II, and serotonin are not involved in the vasoconstricting action.

Dual Actions of Halothane on Intracellular Calcium Stores of Vascular Smooth Muscle

Background Halothane has been reported to affect the integrity of intracellular Calcium2+ stores in a number of tissues including vascular smooth muscle. However, the actions of halothane on intracellular Calcium2+ stores are not yet fully understood. Methods Employing the isometric tension recording method, the action of halothane in isolated endothelium‐denuded rat mesenteric arteries under either intact or beta‐escin‐membrane‐permeabilized conditions was investigated. Results Halothane (0.125–5%) produced concentration‐dependent contractions in Calcium2+ free solution in both intact and membrane‐permeabilized muscle strips. Ryanodine treatment or repetitive application of phenylephrine eliminated both caffeine‐ and halothane‐induced contractions in the Calcium2+ free solution. When either halothane and caffeine, caffeine and halothane, phenylephrine and halothane, or inositol 1,4,5‐triphosphate and halothane were applied consecutively in the Calcium2+ free solution in either intact or membrane‐permeabilized muscle strips, the contraction induced by application of the second agent of the pair was inhibited compared to application of that agent alone. However, when procaine was applied before and during application of the first agent, the contraction induced by the first agent was inhibited and the contraction induced by the second agent was restored. Heparin inhibited the inositol 1,4,5‐triphosphate‐mediated contraction, but not contractions induced by halothane or caffeine. Halothane (0.125–5%), applied during Calcium2+ loading, produced concentration‐dependent inhibition of the caffeine contraction (used to estimate the amount of Calcium2+ in the store) in both intact and membrane‐permeabilized muscle strips. In contrast, halothane applied with procaine during Calcium2+ loading produced concentration‐dependent enhancement of the caffeine contraction. This enhancement was observed only in the intact but not in the membrane‐permeabilized condition. Conclusions Halothane has two distinct actions on the intracellular Calcium2+ stores of vascular smooth muscle, a Calcium2+ releasing action and a stimulating action on Calcium2+ uptake. Halothane releases Calcium2+ from the stores that are sensitive to both caffeine/ryanodine and phenylephrine/inositol 1,4,5‐triphosphate through a procaine‐sensitive mechanism. The observed inhibitory effect on Calcium2+ uptake is probably caused by the Calcium2+ releasing action, whereas the stimulating action on Calcium2+ uptake after blockade of Calcium2+ release may be membrane‐mediated.

Role of Endothelium in the Action of Isoflurane on Vascular Smooth Muscle of Isolated Mesenteric Resistance Arteries

BackgroundIt is believed that isoflurane decreases blood pressure predominantly by decreasing systemic vascular resistance with modest myocardial depression. Nevertheless, little information is available regarding the direct action of isoflurane on systemic resistance arteries. MethodsWith use of the isometric force recording method, the action of isoflurane on contractile response to norepinephrine, a neurotransmitter that plays a central role in sympathetic maintenance of vascular tone in vivo, was investigated in isolated rat small mesenteric arteries. ResultsIn the endothelium-intact strips, the norepinephrine response was initially enhanced after application of isoflurane (2–5%), but it was subsequently almost normalized to the control level during exposure to isoflurane. However, the norepinephrine response was notably inhibited after washout of isoflurane. In the endothelium-denuded strips, the norepinephrine response was gradually inhibited during exposure to isoflurane (≥ 3%), and the inhibition was prolonged after washout of isoflurane. The isoflurane-induced enhancement of norepinephrine response was still observed after inhibitions of the nitric oxide, endothelium-derived hyperpolarizing factor, cyclooxygenase and lipoxygenase pathways, or after blockade of endothelin-1, angiotensin-II, and serotonin receptors; however, it was prevented by superoxide dismutase. ConclusionsIn isolated mesenteric resistance artery, the action of isoflurane on contractile response to norepinephrine consists of two distinct components: an endothelium-dependent enhancing component and an endothelium-independent inhibitory component. During exposure to isoflurane, the former counteracted the latter, preventing the norepinephrine response from being strongly inhibited. However, only the endothelium-independent component persists after washout of isoflurane, causing prolonged inhibition of the norepinephrine response. Superoxide anions may be involved in the enhanced response to norepinephrine.

Heparin prevents the vasodilating actions of protamine on human small mesenteric arteries

Despite the wide clinical use of protamine, the precise mechanisms of its hypotensive effects during reversal of heparin anticoagulation have not been elucidated fully. We, therefore, investigated the effects of protamine on isolated human small mesenteric arteries, both in the absence and presence of heparin, employing the isometric tension recording method. Protamine exerted vasodilating actions in the absence of heparin: 1) protamine (> or = 50 or 150 micrograms/mL) inhibited (P < 0.05) both norepinephrine (1 microM)- and high K+ (40 mM)-induced contractions in the presence of extracellular Ca2+ both in endothelium-intact and -denuded tissues; and 2) protamine inhibited (P < 0.05) norepinephrine (1 microM)-induced, but not caffeine (10 mM)-induced, contractions in the absence of extracellular Ca2+. Such vasodilating actions were blocked almost completely in the presence of heparin. We conclude that only protamine, but not a heparin-protamine complex, has a vasodilating action on the human arteries.

Sevoflurane and bradykinin-induced calcium mobilization in pulmonary arterial valvular endothelial cells in situ: sevoflurane stimulates plasmalemmal calcium influx into endothelial cells.

Kinins locally synthesized in the cardiovascular tissue are believed to contribute to the regulation of cardiovascular homeostasis by stimulating the endothelial cells to release nitric oxide, prostacyclin, or a hyperpolarizing factor via autocrine-paracrine mechanisms. This study was designed to investigate the action of sevoflurane on bradykinin-induced Ca2+ mobilization in endothelial cells in situ. Utilizing fura-2-loaded rat pulmonary arterial valve leaflets, the effects of sevoflurane were examined on bradykinin-induced increases in intracellular Ca2+ concentration ([Ca2+]i) in endothelial cells in situ. In the presence of extracellular Ca2+ (1.5 m M), bradykinin (3–30 &mgr;M) produced an initial phasic and a subsequent tonic increase in [Ca2+]i in a concentration-dependent manner. However, it produced only the phasic increase in [Ca2+]i in the absence of extracellular Ca2+. Sevoflurane (5%, 0.67 m M) inhibited both the phasic and tonic responses to bradykinin. In these experiments, sevoflurane (3–5%) generated sustained increases (approximately 20–40% of the bradykinin-induced maximal increase in [Ca2+]i) in the resting [Ca2+]i level. Sevoflurane still increased [Ca2+]i after depletion of the intracellular Ca2+ stores with ionomycin (0.1 &mgr;M). However, the sevoflurane-induced increase in [Ca2+]i was eliminated by removal of the extracellular Ca2+ and attenuated by NiCl2 (1–3 m M). In conclusion, in the pulmonary arterial valvular endothelial cells, sevoflurane inhibits both bradykinin-induced Ca2+ release from the intracellular stores and bradykinin-induced plasmalemmal Ca2+ influx. In addition, sevoflurane appears to stimulate the plasmalemmal Ca2+ influx and thereby increase the endothelial [Ca2+]i level. Sevoflurane might influence the pulmonary vascular tone through its direct action on the pulmonary arterial valvular endothelial cells.

Changes in Body Temperature Following Deflation of Limb Pneumatic Tourniquet

STUDY OBJECTIVES To investigate changes in both core and peripheral skin-surface temperatures during and after application of a unilateral leg pneumatic tourniquet in adult patients. DESIGN Prospective, observational clinical study. SETTING University hospital. PATIENTS 21 ASA physical status I and II adult patients scheduled for elective leg orthopedic surgery with lumbar epidural anesthesia. INTERVENTIONS Rectal and fingertip skin-surface temperatures were recorded every minute after steady-state lumbar epidural anesthesia was established. MEASUREMENTS AND MAIN RESULTS Significant (p < 0.05) increases in both rectal and fingertip temperatures were observed during tourniquet application for 91 +/- 6 minutes from 36.5 +/- 0.14 degrees C to 37.0 +/- 0.17 degrees C and from 32.6 +/- 0.79 degrees C to 35.5 +/- 0.44 degrees C, respectively. In contrast, both rectal and fingertip temperatures progressively decreased following tourniquet release; significant (p < 0.05) decreases in the rectal and fingertip temperatures were observed 6 and 5 minutes after tourniquet release, respectively. Decreases (approximately maximum) in the rectal and fingertip temperatures 15 minutes after tourniquet release were 0.25 +/- 0.05 degrees C and 1.26 +/- 0.26 degrees C, respectively. In each case, changes in fingertip temperature were approximately six times greater than those in the rectal temperature. CONCLUSIONS Limb tourniquets appear to cause thermal perturbations during epidural anesthesia. The progressive increases in core temperature during tourniquet application presumably resulted from constraint of metabolic heat to the core thermal compartment, and the greater increases in the skin-surface temperature during tourniquet application appear to represent vasodilation in response to the core hyperthermia. On the other hand, redistribution of body heat and the efflux of hypothermic venous blood from the tourniqueted area into systemic circulation following tourniquet deflation probably decreased the core temperature, which might switch off the thermoregulatory vasodilation, leading to the decreases in skin-surface temperature. Recognition of these thermal perturbations are useful in diagnosing intraoperative thermal perturbations.