William Freas

Attenuation of endothelium-mediated vasodilation by halothane.

To determine whether halothane alters endothelium-mediated vasodilation of vascular smooth muscle, isolated ring preparations of rabbit aorta and canine femoral and carotid arteries were suspended for isometric tension recordings in Krebs-Ringer bicarbonate solution at 37°C. Acetylcholine and bradykinin have been shown to relax these norepinephrine contracted arteries via an endothelium-dependent process. In this study, these relaxations were reversibly and significantly attenuated by 2% halothane. However, halothane did not affect relaxations caused by nitroglycerin, which, in these vessels, acts by an endothelium independent mechanism. These results suggest that halothane is not interfering with cyclic guanylate-monophosphate mediated relaxation of vascular smooth muscle, but may interfere with the synthesis, release, or transport of the endothelium-derived relaxing factor. In addition, during contractions evoked by norepinephrine, halothane caused significant decreases in tension in both the canine carotid and rabbit aortic preparations, but increased tension in the femoral artery rings. These effects were not altered by mechanical removal of the endothelium. These results suggest a direct action of halothane on the vascular smooth muscle, which can result in either an increase or decrease in tension, depending on the specific vessel. In addition to its direct vascular effect, this study suggests a new action of halothane; it interferes with endothelium-derived relaxing factor-mediated relaxation of vascular smooth muscle. This action may contribute in part to the vascular alterations seen clinically during administration of halothane.

Adrenergic nerve function and contractile activity of the caudal artery of the streptozotocin diabetic rat

The adrenergic nerve function and contractile responses of the densely innervated caudal artery of the 8-week streptozotocin (SZ) (65 mg/kg i.p.) diabetic rat were investigated. Segments of this artery were removed from diabetic and control rats, placed in Krebs-filled tissue baths (37 degrees C) and isometric tension recorded. Contractile responses to sympathetic nerve activation by electrical stimulation and to cumulative concentrations of norepinephrine (NE) and tyramine were recorded. In order to determine NE content, the NE was extracted from the caudal artery, isolated by adsorption chromatography, and quantified by HPLC with electrochemical detection. NE accumulation and release were also studied by quantifying the amount of tritiated NE [( 3H]NE) and its metabolites in extracts of the tissue or incubation medium. The responses of the caudal artery of SZ diabetic rats to electrical stimulation (4-16 Hz) and to tyramine (1 X 10(-5)-1 X 10(-4) M) were significantly less than those of arteries from control rats and the NE content reduced by 41%, while sensitivity to NE was unchanged. Diabetic arteries also accumulated and released more [3H]NE than did arteries from control rats. These results establish that neurovascular function of the isolated caudal artery of the 8-week SZ diabetic rat is abnormal and suggest that problems in the ability of adrenergic nerves to store and release NE may contribute to this dysfunction. Such changes may play a role in the cardiovascular disturbances associated with diabetic autonomic neuropathy.

Plasma histamine and hemodynamic responses following administration of nalbuphine and morphine.

A comparative study of plasma histamine levels following administration or morphine and nalbuphine in pentobarbital anesthetized dogs was performed. Two concentrations, 3 mg/kg and 0.3 mg/kg of these drugs were investigated. High dose morphine caused an immediate marked increase in plasma histamine from 5.0±0.4 to 340±72 ng/ml. Simultaneous with this increase in plasma histamine was a marked decrease in mean arterial blood pressure within the first minute. In contrast significant alterations in plasma histamine levels were not observed with high or low doses of nalbuphine. A low dose of morphine (0.3 mg/kg) did not increase plasma histamine levels. Heart rate was not changed by any drug treatment. The use of compound 48/80 a specific mast cell degranulating agent allowed for the identification of a specific pool of mast cells capable of responding to morphine.In vitro exposure of purified dog leukocytes to high doses of morphine did not result in histamine release. These results indicate that nalbuphine does not increase plasma histamine, while morphine does, and that the source of the increase in plasma histamine is from tissue mast cells.

The effects of morphine, nalbuphine, and butorphanol on adrenergic function in canine saphenous veins.

Saphenous vein rings mounted in organ chambers containing Krebs–Ringer solution were used to determine if the venodilator effects of morphine, nalbuphine, and butorphanol are the result of interference with adrenergic neurotransmission or are caused by direct depressant actions on venous smooth muscle cells. Morphine (5 × 10–5 M and 2 × 10–4 M) caused a dose-dependent depression of the contractile response to transmural electrical stimulation. H1-and H2- histamine antagonists did not attenuate the inhibitory effect of morphine. Concentrations of morphine and nalbuphine lower than 5 × 10–5 M had no effect, whereas 5 × 10−6 M butorphanol significantly depressed the evoked tension response to electrical stimulation. The contractile responses of the veins to exogenous norepinephrine (NE) were not altered by morphine, indicating a presynaptic site of action rather than a direct action on the venous smooth muscle. Transmural electrical stimulation was used to evoke release of endogenous NE. Morphine (5 × 10−5 M and 2 × 10−4 M), nalbuphine (2 × 10–4 M), and butorphanol (4 × 10–6 M) significantly decreased release of NE. Naloxone did not alter NE release and did not attenuate the inhibition of NE release observed with the opiates, indicating that the effect of morphine on this neuroeffector junction is not mediated by a naloxone-sensitive opiate receptor. Blockade of presynaptic alpha receptors by phenoxybenzamine or phentolamine augments NE release caused by transmural electrical stimulation; morphine inhibited this augmentation. The results of these experiments indicate that high concentrations of morphine may decrease NE release, an effect that may contribute to the venodilation and hypotension observed following administration of high doses of morphine in humans. In the usual analgesic doses, the venodilatory effects of morphine cannot be explained by local action on either NE release or venous smooth muscle contractility.

Effects of bufalin on norepinephrine turnover in canine saphenous vein.

Abundant experimental data suggest that an endogenous digitalislike factor is responsible for some essential hypertension. Some forms of hypertension have also been associated with increased levels of catecholamines. We therefore designed experiments to investigate the role of digitalislike factors in the regulation of norepinephrine turnover in the neurovascular junction. We chose bufalin, an amphibian-derived compound that shares many of the physiological properties postulated as characteristic of digitalislike compounds, as a model of the mammalian compound. In vitro experiments in canine saphenous veins showed that, in addition to inhibiting norepinephrine uptake, bufalin increased norepinephrine overflow by an amount larger than could be explained solely by uptake inhibition. The effect of bufalin on norepinephrine overflow is inhibited by tetrodotoxin, which suggests a dependence of this response on Na+ influx through the neuronal membranes. We propose that Na+,K+-ATPase inhibition resulting in neuronal depolarization is responsible for the augmented norepinephrine turnover caused by bufalin and that these indirect effects of norepinephrine on the cardiovascular system may play a role in the etiology of hypertension.

Plasma histamine and catecholamine levels during hypotension induced by morphine and compound 48/80.

Histamine receptors are present in adrenergic terminals, and histamine is reported to inhibit release of the neurotransmitter norepinephrine (NE) at certain neuroeffector junctions. However, a physiological role for histamine in modifying adrenergic neurotransmission has not been established. To examine the interaction of elevated plasma histamine and catecholamine release, two compounds that release histamine, morphine (3 mg/kg), and compound 48/80 (0.5 mg/kg), were administered intravenously (i.v.). Plasma norepinephrine (NE) levels were used to monitor sympathetic nervous system activity, and plasma epinephrine (Epi) levels were used to monitor adrenal activity. Both morphine and compound 48/80 caused an immediate and marked increase in plasma histamine. Simultaneous with this increase, a marked decrease in mean arterial pressure occurred. Plasma NE levels increased in animals administered compound 48/80, but in morphine-treated animals, plasma NE levels did not change from pretreatment values. Plasma Epi levels increased in both groups, but the magnitude and duration of the responses differed. The results indicate that elevated plasma catecholamines can increase in response to histamine-induced hypotension but this effect can be suppressed by the central actions of morphine.

Vascular Interactions of Calcium and Reactive Oxygen Intermediates Produced Following Photoradiation

This study was designed to exarmine vascular smooth muscle contractile properties following cnhanced proudction of reactive oxygen intermediates (ROIs), which were produced by pretreating rat caudal arteries and aortas with a photosensitizer, hematoporphyrin deravative. and then illuminating them with red laser light. This treatment produced a long-lasting contraction that was dependent on the presence of extracellular calcium. Reduction in extracellular calcium relaxed the smooth muscle and replacement of calcium 30 min later increased the tension. Oxygen radical scavengers did not block the contractile effect postillumination when calcium was returned to the hathing solution: however, verapamil (5.5 μM) and nifedipine (10 μM) attenuated this contraction. The contractions were dependent on oxygen in the aerating gas mixture. Production of ROIs by isolated blood vessels was supported biochemically by a significant increase in both bath and tissue levels of oxidation products. reactive with thiobarbituric acid. and by a reduction in the tissue stain, nitroblue tetrazolium. These ROIinduced contractions were observed in vitro on large conduit arteries and also in vivo on small ear arteries. The vascular response following this acute production of ROIs may be similar to vascular abnormalities in certain pathologicl conditions where ROI production is reported to be elevated. Therefore. these results could contribute to a further understanding of mechanisms involved in these ROI-dependent vascular changes.

Effects of enflurane on adrenergic function in canine mesenteric artery and vein

We examined the pre-and postsynaptic effects of enflurane on the superior mesenteric artery and vein. We measured the release of norepinephrine (NE) from sympathetic nerve terminals caused by electrical stimulation (ES) or potassium and changes in vascular smooth muscle tension. The effect of enflurane was also examined in the presence of alpha1- and alpha2-adrenoceptor antagonists. Enflurane (1.4 to 1.6 minimum alveolar anesthetic concentration [MAC] in the dog) did not alter basal tension, but it significantly inhibited tension development caused by 40 mM KCl and ES in the canine mesenteric artery and vein. Enflurane inhibited the KCl- and ES-induced release of NE from sympathetic nerve terminals in the canine mesenteric vein. The alpha1- and alpha2-adrenoceptor antagonists, corynanthine or yohimbine, did not modify the effect of enflurane on the ES-induced frequency response curve of the mesenteric artery and vein. These data suggest that enflurane has at least two sites of action in mesenteric vascular beds. One site is presynaptic where it inhibits NE release, and the other is a postsynaptic site distal to the adrenergic receptor where it inhibits the smooth muscle contraction process. (Anesth Analg 1995;81:265-71)

Interactions of Volatile Anesthetics and Reactive Oxygen Intermediates on Vascular Smooth Muscle

Reactive oxygen intermediates (ROIs, also referred to as oxygen-derived free radicals or reactive oxygen metabolites) have been implicated in several physiological and pathological processes.1–3 In particular, they have been reported to be associated with several vascular abnormalities, including those that occur during hypertension,4 reperfusion injury,5 transplant rejection,3 inflammation, premature aging,6 radiation injury, diabetes7 and endotoxic shock.8 An increase in PaO2, especially after hypoxia, is a major stimulus for production of ROIs.9

Further isolation of endogenous factors in canine and human plasma that inhibit [3H]norepinephrine accumulation

We have previously shown that both homologous canine plasma and a crude extract of this plasma contain substances that inhibit accumulation of [(3)H]norepinephrine ([(3)H]NE) by the canine saphenous vein. The purpose of this study was to further purify these substances and to determine if similar factors were present in human plasma. Crude extracts of plasma were purified with a Folch extraction in which most of the biological activity was recovered in the bottom or organic phase. This phase significantly inhibited [(3)H]NE uptake by the canine saphenous vein (23.5 +/- 7.6% by concentrate from 9.1 ml of original plasma/ml incubate solution) and increased development of tension following transmural electrical stimulation by 91.5 +/- 23.3% (extract from 1 ml of plasma/ml bath solution). The Folch extracts obtained from 100ml of plasma were purified by column and thin layer (TLC) chromatography. Samples were applied to a silicic acid column and eluted with chloroform, acetone, and methanol. The [(3)H]NE uptake inhibitory activity was primarily recovered in the methanol fraction. TLC of the methanol fraction of canine plasma on silica gel G plates (with pre-absorbent) resulted in five zones which were then assayed for their ability to inhibit [(3)H]NE accumulation by the saphenous vein. In the first zone (concentrate from 27.5 ml plasma/ml bath solution) there was significantly greater inhibitory activity (55.4 +/- 8.3%), than in the corresponding zone obtained from solvent blanks (20.7 +/- 4.1%). These results indicate that there is a factor or possibly factors in canine and human plasma that have thin layer chromatographic properties of a polar lipid, which inhibit [(3)H]NE accumulation and enhance the contractile response of vascular smooth muscle to transmural electrical stimulation.