Isoflurane but not halothane minimum alveolar concentration-sparing response of dexmedetomidine is enhanced in rats chronically treated with selective α2-adrenoceptor agonist

Halothane minimum alveolar concentration (MAC)-sparing response is preserved in rats rendered tolerant to the action of dexmedetomidine. It has been shown that halothane and isoflurane act at different sites to produce immobility. The authors studied whether there was any difference between halothane and isoflurane MAC-sparing effects of dexmedetomidine in rats after chronic administration of a low dose of this drug. Twenty-four female Wistar rats were randomly allocated into four groups of six animals: two groups received 10 μg/kg intraperitoneal dexmedetomidine for five days (treated groups) and the other two groups received intraperitoneal saline solution for five days (naive groups) prior to halothane or isoflurane MAC determination (one treated and one naive group of halothane and one treated and one naive group of isoflurane). Halothane or isoflurane MAC determination was performed before (basal) and 30 min after an intraperitoneal dose of 30 μg/kg of dexmedetomidine (post-dex) from alveolar gas samples at the time of tail clamp. Administration of an acute dose of dexmedetomidine to animals that had chronically received dexmedetomidine resulted in a MAC-sparing effect that was similar to that seen in naive animals for halothane; however, the same treatment increased the MAC-sparing response of dexmedetomidine for isoflurane. Isoflurane but not halothane MAC-sparing response of acutely administered dexmedetomidine is enhanced in rats chronically treated with this drug.     

The inhibitory effect of halothane on the emetic response in the ferret

Emesis and nausea are often associated with anaesthesia and continue to be a common clinical problem. Past clinical studies have demonstrated that halothane produces a higher incidence of vomiting compared with other anaesthetics, but some investigators have described an antiemetic effect. The purpose of this study was to investigate the effects of various doses of halothane on the emetic response in the decerebrate ferret. Following a control emetic response, a maximum of six increasing cumulative concentrations of halothane were delivered. At the end of each delivery period, the supradiaphragmatic vagal communicating branch, which has been shown to reproducibly elicit vomiting, was electrically stimulated and the emetic response was monitored. An increase in halothane concentration produced a marked depression of tongue, abdominal muscle, and diaphragm EMG activity as well as a decrease in central venous pressure. Licking, a prodromal response comparable to nausea in the human, appeared to be most sensitive. An increase in latency of the emetic response occurred as the concentration of halothane was increased. All phases of the response were observed at concentrations below 0.6 vol% halothane. At 0.6 vol% halothane, 75% of the animals vomited. At higher concentrations, the emetic response was completely abolished. One hour post-halothane, all latencies had returned to near control values. The methods utilized in this study provided a model that was not complicated by a large number of variables usually present in clinical studies. These data demonstrate that halothane exerts an inhibitory, concentration-dependent, and reversible effect on the emetic response in the ferret and provide further support that halothane alone does not possess emetic properties at clinical properties at clinical concentrations.     

Inhibition of Immune Complex-Induced Inflammation by A small Molecular Weight Selectin Antagonist

The anti-inflammatory effect of a small molecular weight antagonist of P- and E-selectin-dependent cell adhesion was examined. The glycolipid sulphatide was shown to block the adherence of thrombin-activated rat platelets to HL-60 cells. This interaction is known to be dependent on P-selectin. The rat dermal reverse passive Arthus reaction was used to assess the effect of sulphatide on a neutrophil dependent inflammatory response. Sulphatide dosedependently blocked both the vascular permeability increase and cell infiltration after intraperitoneal administration. These results show that a small molecular weight compound which blocks P- and E-selectin dependent adhesion in vitro can effectively block the inflammation due to immune complex deposition. A compound with this type of profile may have therapeutic potential in the treatment of immune complex mediated diseases.     

Modulation of IL-1, tumor necrosis factor, and C5a-mediated murine neutrophil migration by alpha-melanocyte-stimulating hormone

This study was designed to examine the effects of i.p.-injected alpha-melanocyte stimulating hormone (MSH) on murine neutrophil migration into subcutaneously implanted sponges in response to IL-1-alpha, TNF-alpha, and C5a. The results show that as little as 0.1 ml of 5 x 10(-7) M MSH injected i.p. significantly blocked the accumulation of neutrophils in sponges in response to IL-1. This action of MSH was dose dependent, reversible, and was maximally effective if MSH was given at the same time as the injection of IL-1. This effect of MSH was not restricted to IL-1-induced neutrophil emigration, because MSH also antagonized the accumulation of neutrophils in response to both TNF and C5a. The proopiomelanocortin-derived peptide ACTH which contains the MSH sequence also significantly reduced neutrophil accumulation in response to IL-1, although less effectively than MSH. Similar studies with beta-endorphin showed that it had no effect on neutrophil accumulation in this system. The direct injection of MSH, beta-endorphin and ACTH into sponges or i.p. did not stimulate a neutrophil emigration and eliminated the possibility that MSH or ACTH suppressed the neutrophil influx in response to IL-1, TNF, or C5a by competing for circulating neutrophils. The action of MSH on IL-1, TNF, and C5a-induced neutrophil emigration suggests that this peptide may be an important regulator of the inflammatory response.     

Fatty acids markedly lower the threshold for halothane-induced calcium release from the terminal cisternae in human and porcine normal and malignant hyperthermia susceptible skeletal muscle

Malignant hyperthermia is caused by an abnormal increase in Ca2+ levels in skeletal muscle in response to anesthetics, including halothane. Since fatty acid production is elevated in skeletal muscle from individuals with malignant hyperthermia, the effects of fatty acids on the threshold of halothane-induced Ca2+ release were examined. In the absence of fatty acids halothane caused Ca2+ release from porcine and human heavy sarcoplasmic reticulum fractions, but only at concentrations above the clinically relevant range. Oleic acid (20 microM), an unsaturated fatty acid, reduced the threshold at which halothane induced Ca2+ release to concentrations used for anesthesia. Stearic acid, a saturated fatty acid had considerably less effect on the threshold of halothane action. The greater sensitivity of malignant hyperthermia muscle to halothane can be explained by elevated fatty acid production.     

Effects of urethane, alphaxolone/alphadolone, or halothane with or without neuromuscular blockade on survival during repeated episodes of global cerebral ischaemia in the rat.

The effect of 4 anaesthetic regimens on blood pressure and survival was investigated during repeated episodes of cerebral ischaemia in the rat induced by diathermy of the vertebral arteries and reversible occlusion of the carotid arteries. The best results were obtained with inspired halothane with neuromuscular blockade and artificial ventilation, followed in order by halothane, intravenous alphaxolone/alphadolone, and intraperitoneal urethane with spontaneous ventilation.     

In vitro responses of cat skeletal muscle to halothane and caffeine

Strips of soleus (100% type I) and gracilis (90% type II) muscle were obtained from anesthetized cats and mounted in organ baths filled with aerated Krebs-Ringer solution (37 degrees C). The contractile patterns in response to electrical stimulation (0.1 Hz, 25 V, 5 ms), caffeine, halothane, and caffeine in the presence of halothane were examined in the two fiber types. The ability of 25 microM dantrolene to alter the contractile patterns was also evaluated. In vitro contractile properties in response to electrical stimulation were similar to properties observed in situ, except that twitch tension in soleus muscle was significantly less in vitro than in situ. In the presence of halothane, type I soleus muscle developed a rapid contracture. The contracture was blocked by pretreatment with dantrolene and was reversed by addition of dantrolene at the peak of the response. Halothane-induced contractures were not observed at any time in type II gracilis. Type I soleus was also significantly more sensitive both to caffeine alone and to caffeine in the presence of halothane than was type II gracilis. In both fiber types, halothane increased the sensitivity of the muscles to caffeine. Dantrolene attenuated caffeine-induced contractures in both fiber types, but the attenuating effect was less in the presence of halothane. The findings of a halothane-induced contracture in the cat soleus and differential sensitivities of the two muscle fiber types to caffeine indicate that further studies in these two muscles may be useful for delineating the mechanisms inducing contracture in muscle from individuals susceptible to malignant hyperthermia.     

Genetic effects on an anesthetic-sensitive pathway in the brain of Drosophila

General anesthetics are known to inhibit the electrically induced escape response of the fruitfly through action within the brain. We examined this response and its sensitivity to anesthetics in several mutants that cause significant disruption of the mushroom body and other structures of the central brain in adult flies. Because we show here that anesthesia sensitivity is influenced by genetic background, we have used a set of congenic mutant lines. Sensitivity to halothane is normal in most of these lines, indicating that the anesthetic target is unaffected by the gross status of the central brain. Thus, for the escape response, anesthetic sensitivity is not a global feature but reflects action at a localized target. Only the mushroom body defect (mud) line showed an increased sensitivity of the escape response to halothane. Sensitivity to two other anesthetics is also perturbed in this line, albeit less dramatically so. The behavior of mud/+ heterozygotes and the comparison of brain anatomy among all the mutant lines imply that the effect of the mud mutation on anesthesia is not via gross alteration of central brain structures. The possibility that an adventitious mutation in the mud line is responsible for the effects on anesthesia is disfavored by the behavior of a heterozygote between two mud alleles. Although we do not yet know whether the mud gene encodes an anesthetic target or influences the functioning of an anesthetic-sensitive neuron in this pathway, our work indicates that this gene regulates the effects of halothane on a circumscribed pathway.     

Anesthesia alters pulmonary vasoregulation by angiotensin II and captopril.

We investigated the effects of an intravenous (pentobarbital sodium) and inhalational (halothane) general anesthetic on the pulmonary vascular responses to angiotensin II and angiotensin-converting enzyme inhibition (CEI). Multipoint pulmonary vascular pressure-flow (P/Q) plots were generated in conscious pentobarbital- (30 mg/kg iv) and halothane-anesthetized (approximately 1.2% end-tidal) dogs in the intact (no drug) condition, during angiotensin II administration (60 ng.kg-1.min-1 iv), and during CEI (captopril 1 mg/kg plus 1 mg.kg-1.h-1 iv). In conscious dogs, angiotensin II increased (P less than 0.001) the pulmonary vascular pressure gradient [pulmonary arterial pressure--pulmonary arterial wedge pressure (PAP-PAWP)] over the empirically measured range of Q; i.e., angiotensin II caused pulmonary vasoconstriction. Pulmonary vasoconstriction (P less than 0.01) in response to angiotensin II was also observed during pentobarbital sodium anesthesia. In contrast, angiotensin II had no effect on the P/Q relationship during halothane anesthesia. In conscious dogs, CEI decreased (P less than 0.001) PAP-PAWP over the empirically measured range of Q; i.e., CEI caused pulmonary vasodilation. However, CEI caused pulmonary vasoconstriction (P less than 0.02) during pentobarbital sodium and had no effect on the P/Q relationship during halothane. Thus, compared with the conscious state, the pulmonary vasoconstrictor response to angiotensin II is unchanged or abolished, and the pulmonary vasodilator response to CEI is reversed to vasoconstriction or abolished during pentobarbital sodium and halothane anesthesia, respectively.     

The neutrophil migration induced by tumour necrosis factor alpha in mice is unaffected by glucocorticoids

Macrophages harvested from the peritoneal cavities of rats release a neutrophil chemotactic factor (MNCF) in response to stimulation with Gram-negative bacterial lipopolysaccharide (LPS). MNCF has been shown to be active in rats treated with dexamethasone, a glucocorticoid that usually inhibits the neutrophil migration induced in this species by interleukin (IL)-1, tumour necrosis factor alpha (TNFalpha), IL-8, C5a and leukotriene B(4) (LTB(4)). Here we report that macrophages harvested from peritoneal cavities of mice, and stimulated in vitro with LPS, also release a factor that induces neutrophil migration in dexamethasone-treated animals. This chemotactic activity was neutralized by the incubation of the LPS-stimulated macrophage supernatants with a purified polyclonal IgG anti-mouse TNFalpha. In addition, significant amounts of TNF were detected in the supernatants. The neutrophil migration induced by intraperitoneal administration of recombinant murine TNFalpha was also unaffected by pretreatment of the mice with dexamethasone. Moreover, neutrophil migration induced by intraperitoneal injection of LPS was completely blocked by pretreatment of the mice with a monoclonal antibody against murine TNFalpha. In conclusion, our results support the hypothesis that, in contrast to the role of TNF in rats (where it indirectly induces neutrophil migration), in mice, it may be an important mediator in the recruitment of neutrophils to inflammatory sites.     

Cellular changes during the infusion of high dose intravenous immunoglobulin

With the ever widening group of autoimmune conditions that are beneficially affected by infusions of high dose immunoglobulin the possible mechanisms of action of such therapy appear increasingly complex. Fc mediated blockade of the mononuclear phagocyte system is an acknowledged early effect. This is, however, accompanied by a decrease of neutrophil counts which suggests that IgG binding to the neutrophil may be a mechanism of action. The decrease of neutrophil counts is transient but in immune thrombocytopenia is inversely proportional to the platelet response observed. In parallel to the effect on the neutrophil there are changes in the lymphocyte subsets with reversal of the T helper/suppressor ratio and alterations in the individual cellular constituents of each subset that correlate with the clinical response. The observed changes in B cell numbers and function suggest that T dependent and independent antibody production is effected by intravenous immunoglobulin. It is increasingly clear that in ITP at least the clinical response to IV IgG is a summation of several cellular events and their balance reflects the ultimate outcome. It may eventually be possible to use these observations to predict the likely outcome in the individual patient of this mode of therapy.     

Effects of anaesthetics on membrane mobility and locomotor responses of human neutrophils

Abstract The morphological response of neutrophils to chemotactic factors is characterized by an immediate change (in seconds) from a spherical to an irregular shape. Within two or three minutes, the cells assume the head-tail polarity typical of locomotor cells. In this study the effects of the anaesthetic drugs, propofol and thiopentone, on the time-sequence of the morphological response of human neutrophils to the chemotactic peptide fMet-Leu-Phe were examined. At concentrations seen in the plasma during anaesthesia, both drugs inhibited both the rate and degree of the neutrophil chemotactic response. The effect of propofol was not attributable to its lipid vehicle, as 10% intralipid alone had no effect on neutrophil polarization. Plasma membrane reorganization occurs during polarization of neutrophils, resulting in morphological and functional changes which prepare the cells for chemotaxis and phagocytosis. Fluorescence recovery after photobleaching (FRAP) was used to investigate effects of the anaesthetics on membrane lipid behaviour. With a lipid probe, the proportion of mobile lipid in neutrophils exposed to propofol or thiopentone was reduced. There was a less significant reduction with intralipid which also caused reduction in velocity of lateral diffusion of the probe. These findings suggest that the inhibitory effects of anaesthetics on neutrophil locomotion are related to reductions in fluid mobility of the plasma membranes of anaesthetic-treated cells.     

Genetic effects on an anesthetic‐sensitive pathway in the brain of drosophila

General anesthetics are known to inhibit the electrically induced escape response of the fruitfly through action within the brain. We examined this response and its sensitivity to anesthetics in several mutants that cause significant disruption of the mushroom body and other structures of the central brain in adult flies. Because we show here that anesthesia sensitivity is influenced by genetic background, we have used a set of congenic mutant lines. Sensitivity to halothane is normal in most of these lines, indicating that the anesthetic target is unaffected by the gross status of the central brain. Thus, for the escape response, anesthetic sensitivity is not a global feature but reflects action at a localized target. Only the mushroom body defect (mud) line showed an increased sensitivity of the escape response to halothane. Sensitivity to two other anesthetics is also perturbed in this line, albeit less dramatically so. The behavior of mud/+ heterozygotes and the comparison of brain anatomy among all the mutant lines imply that the effect of the mud mutation on anesthesia is not via gross alteration of central brain structures. The possibility that an adventitious mutation in the mud line is responsible for the effects on anesthesia is disfavored by the behavior of a heterozygote between two mud alleles. Although we do not yet know whether the mud gene encodes an anesthetic target or influences the functioning of an anesthetic‐sensitive neuron in this pathway, our work indicates that this gene regulates the effects of halothane on a circumscribed pathway. © 2000 John Wiley & Sons, Inc. J Neurobiol 42: 69–78, 2000     

Anesthetic stabilization of protein intermediates: myoglobin and halothane

Halothane, an inhaled anesthetic, destabilizes the folded structure of myoglobin. To determine whether this is due to preferential interactions with less stable folded conformers of myoglobin versus the completely unfolded state, we used photoaffinity labeling, hydrogen exchange, fluorescence spectroscopy, and circular dichroism spectroscopy. Apomyoglobin was used as a model of a less stable conformer of myoglobin. Halothane destabilizes myoglobin and binds with low affinity and stoichiometry but stabilizes and binds with higher affinity to apomyoglobin. The same halothane concentration has no effect on cytochrome c stability. The apomyoglobin/halothane complex is favored at pH 6.5 as compared to pH 4.5 or pH 2.5. Halothane photoincorporates into several sites in apomyoglobin, some allosteric to the heme pocket. Guanidinium unfolding of myoglobin, monitored by CD spectroscopy, shows destabilization at less than 1.3 M Gdm but stabilization at greater than 1.3 M Gdm, consistent with the hypothesis that less stable conformers of myoglobin bind halothane preferentially. We suggest the structural feature underlying preferential binding to less stable conformers is an enlarged cavity volume distribution, since myoglobin has several intermediate-sized cavities, while cytochrome c is more well packed and has no cavities detected by GRASP. Specific binding to less stable intermediates may underlie anesthetic potentiation of protein activity.     

Effect of halothane on the Ca2+-transport system of surface membranes isolated from normal and malignant hyperthermia pig skeletal muscle

Trapezius muscle from normal and malignant hyperthermia (MH) pigs was used to investigate the effects of halothane on contractile properties and on the calcium transport system of isolated surface membranes. We observed that (i) halothane, diluted in dimethyl sulfoxide, induced a higher isometric contracture response in MH muscle than in normal muscle, (ii) halothane had a more pronounced inhibitory effect on the sarcolemmal Ca2+-ATPase activity in MH membrane, and (iii) the actively accumulated calcium was released in higher amounts in MH muscle than in normal muscle. These results suggest that halothane might induce, in vivo, an important influx of extracellular calcium ions through the MH sarcolemmal membranes and this pool of intracellular calcium may constitute the trigger for the defective sarcoplasmic reticulum "calcium-induced calcium-release" system.     

Halothane, an inhalation anesthetic, activates protein kinase C and superoxide generation by neutrophils

The rate of superoxide generation of guinea pig intraperitoneal neutrophils by a chemotactic peptide or 12-O-tetradecanoylphorbol-13-acetate (TPA) was increased by 2-bromo-2-chloro-1,1,1,-trifluoroethane (halothane), an inhalation anesthetic. This increase was inhibited by 1-(5-isoquinolinesulfonyl)methylpiperazine dihydrochloride (H-7), a specific inhibitor of Ca2+- and phospholipid-dependent protein kinase C (PKC). Halothane was found to significantly activate partially purified PKC. The activation required phosphatidylserine (PS) and Ca2+. Dioleoylglycerol- or TPA-activated PKC activity was further increased by halothane. The cytoplasmic proteins of guinea pig neutrophils phosphorylated by halothane-activated PKC were similar to those phosphorylated by PMA-activated PKC. The phosphorylation of a 48 kDa protein, a phosphorylated protein required for NADPH oxidase activation, was also increased by halothane. These data suggest that the increase of superoxide production by halothane is correlated with its activation of PKC.     

Changes in cytoplasmic free calcium caused by halothane. Role of the plasma membrane and intracellular Ca2+ stores

Malignant hyperthermia is a muscle disease characterized by an abnormal response to anaesthetics, stress, and exercise. It is typified by muscle contracture and a dramatic elevation in body temperature. A defect in the regulation of the concentration of cytoplasmic free calcium, [Ca2]i, is thought to underlie this disease, but the actual [Ca2+]i was not measurable until recently. We have shown that the anaesthetic halothane increases [Ca2+]i in isolated lymphocytes from malignant hyperthermia-susceptible humans and pigs but not in the normal counterparts. In this report we extend these observations to a larger number of cases and analyze the molecular mechanisms responsible for the increase. The halothane-mediated rise in [Ca2+]i required external Ca2+ and was prevented by nifedipine, an inhibitor of the voltage-sensitive Ca2+ channels of the cell membrane. In addition, the effect of halothane on the releasable Ca2+ from intracellular stores was determined by measuring the size of the releasable pool before and after addition of the anaesthetic. After addition of halothane, about 73% of this Ca2+ pool was still available for release by the Ca2+ ionophore ionomycin in cells from normal humans and pigs. In contrast, only about 45% of the free Ca2+ in intracellular stores was left after treatment with halothane in cells from malignant hyperthermia-susceptible humans and swine. These results indicate that halothane acts both at the cell membrane and at intracellular organelles, and that this action results in a net increase in [Ca2+]i in malignant hyperthermia, but not in normal cells. The action at the cell membrane appears to be on the voltage-sensitive Ca2+ channels.(ABSTRACT TRUNCATED AT 250 WORDS)     

L-phenylisopropyladenosine (L-PIA) diminishes halothane anesthetic requirements and decreases noradrenergic neurotransmission in rats

The effect of L-phenylisopropyladenosine (L-PIA), the A1 adenosine agonist, on the depth of anesthesia was investigated in halothane-anesthetized rats. L-PIA treatment reduced the minimum anesthetic concentration (MAC) of halothane that prevented 50% of animals from moving in response to a painful stimulus by 49%. MAC experiments performed with L-PIA given in conjunction with A1 adenosine receptor antagonists which either permeate the blood-brain barrier (8-phenyltheophylline [8-PT] or do not (8-sulphophenyltheophylline [8-So-PT]) indicate that central mechanisms are involved. Noradrenergic neurotransmission was diminished following L-PIA administration in halothane-anesthetized rats in all brain regions. These data suggest that acute L-PIA treatment decreases central noradrenergic neurotransmission and may represent the mechanism for the decrease in halothane dose to achieve an anesthetic endpoint anesthetic response to halothane.     

A 14 000-year record of paleoenvironmental change in the western basin of China's third largest lake,Lake Taihu

Effect of leaf extract of Ocimum sanctum on (i) the specific and non-specific immune responses and (ii) disease resistance against Aeromonas hydrophila was investigated in Oreochromis mossambicus. Sheep red blood cells (SRBC) and Heat aggregated bovine serum albumin (HA-BSA) were used as antigens for specific and non-specific immune response studies, respectively. Antigens were administered through intraperitoneal route. Anti-SRBC antibody titres were determined by direct haemagglutination and anti-bacterial antibodies were determined by bacterial agglutination. Nitroblue tetrazolium (NBT) assay was used to determine neutrophil activity. Disease resistance was measured in terms of relative percent survival (RPS). The immunostimulatory effect of the leaf extract of O. sanctum, when administered through intraperitoneal and oral routes was obvious. Leaf extract of O. sanctum, when administered intraperitoneally, stimulated both antibody response and neutrophil activity. Dietary intake of O. sanctum also enhanced the antibody response and disease resistance against A. hydrophila. Possibility of using O. sanctum as immunostimulant in the maintenance of finfish health in intensive freshwater aquaculture is suggested.     

Modification of susceptibility to Klebsiella pneumoniae during murine cytomegalovirus infection

The effect of murine cytomegalovirus (MCMV) infection on susceptibility to bacterial infection was studied in mice by a combination of intraperitoneal (ip) inoculation of a sublethal dose of MCMV with subsequent ip challenge of 2 X 10(3) cfu of a strain of Klebsiella pneumoniae (KP). When given alone, KP produced a mortality of 30-40%. Mortality was increased when KP was given 1 to 7 days after MCMV injection with the peak increase at the 4th to 5th day when 100% mortality occurred. Virus levels in various organs of mice infected with MCMV alone, or superinfected with KP did not differ. Bacterial counts on the other hand, showed that increased mortality in mixed MCMV and KP infected mice was due to an uncontrolled growth of bacteria at the site of primary lodgment, i.e., the peritoneum, and severe systemic infection. Neutrophil response to growth of KP during the first 3 days of bacterial infection was defective in MCMV infected mice. While the initial clearance of KP from the blood was more efficient in MCMV infected mice, probably due to activated reticuloendothelial function, it did not protect the mice against KP infection. Using the in vivo model, it was shown that poor neutrophil response and possibly other defective neutrophil functions were responsible for increased mortality to KP infection in MCMV infected mice.