The neurotoxic effects of intrathecal midazolam and neostigmine in rabbits.

In parallel with improvements in understanding pain neurophysiology, many chemicals have recently been investigated for spinal anaesthesia and analgesia. However, studies discussing the effects of these drugs on neural tissue indicate that knowledge about some aspects of neurotoxicity is limited. Forty-nine New Zealand albino rabbits, weighing 2.2 +/- 0.2 kg, were randomly assigned to seven groups of seven animals each. Single dose groups received intrathecally through the atlantooccipital membrane 0.9% saline 1.5 ml; midazolam 100 microg/kg (low dose midazolam group) or 500 microg/kg (high dose midazolam group); neostigmine 10 microg/kg (low dose neostigmine group) or 50 microg/kg (high dose neostigmine group). Two groups had seven days of repeated dosing with either midazolam 100 microg/kg/day (repeat midazolam group) or 10 microg/kg/day neostigmine (repeat neostigmine group). The animals were sacrificed on day 8, and two spinal cord sections from the fourth cervical level and fourth lumbar level were removed and prepared for histopathological study. Transmission electron microscopic evaluations were performed on transverse spinal cord sections by a neuropathologist blinded to the group allocation. Twenty myelinated axons and neurones in the cervical and lumbar sections were investigated for the histopathological study. This study indicates that midazolam and neostigmine have different neurotoxic effects that depend on the dose and the repetition of dosing when these drugs are administered intrathecally.

Effect of left atrial size and function on P-wave dispersion: a study in patients with paroxysmal atrial fibrillation.

BACKGROUND: Paroxysmal atrial fibrillation (PAF), a common arrhythmia, is caused by the fractionated and nonhomogeneous propagation of sinus impulse. HYPOTHESIS: This study was undertaken to examine the effect of left atrial (LA) dimension and function on P-wave dispersion (deltaP) in unselected patients with PAF and health controls. METHOD: In this study, 62 consecutive patients with PAF (32 men, 30 women, mean age 55+/-11 years) and 62 age- and gender-matched healthy controls (33 men, 29 women, mean age 52+/-13 years) were studied to compare the effect of LA size, volume, and function on deltaP (difference between maximum and minimum P-wave duration on 12-lead electrocardiogram). RESULTS: P-wave dispersion in patients with PAF and normal LA diastolic diameter (LAD) was longer than that in controls with normal LA size (53+/-8 vs. 34+/-8 ms, p < 0.001). P-wave dispersion increased in patients with PAF (62+/-12 vs. 53+/-8 ms, p = 0.003) and controls (40+/-7 vs. 34+/-8 ms, p = 0.005) with increased LAD. Presence or absence of PAF did not interact with LAD for their effect on deltaP (2 x 2 analysis of variance test p = 0.20). In the PAF group, deltaP correlated with LAD (r = 0.43, p = 0.002), LA diastolic volume (r = 0.6, p < 0.001), and LA ejection fraction (AEF) (r = - 0.33, p = 0.05). The AEF was preserved when LAD increased in the patients without PAF (0.52+/-0.07 vs. 0.57+/-0.10, p = NS), however was significantly decreased in the PAF group (0.37+/-0.12 vs. 0.49+/-0.10, p = 0.01). On multivariate logistic regression analysis, only deltaP retained significance on development of PAF. CONCLUSION: It was concluded that deltaP increased in patients with PAF and normal LA size. In controls with increased LA size, deltaP increased but did not reach the levels attained in patients with PAF. The AEF was decreased in patients with PAF but was preserved in those without PAF. These findings can be explained by the changes in LA microarchitecture which concurrently decreased atrial myocardial contraction, increased deltaP, and predisposed to PAF.

Diffusion-weighted MR imaging after angioplasty or angioplasty plus stenting of arteries supplying the brain.

BACKGROUND AND PURPOSE: There has been concern regarding the safety of revascularization procedures of vessels supplying the brain vessels because of the risk of cerebral embolization during the procedure. We have observed a high incidence of hyperintense lesions on diffusion-weighted MR images of the brain after stenting at the carotid bifurcation. The hypothesis of this study is that diffusion-weighted MR imaging of the brain can reveal new diffusion abnormalities after angioplasty or angioplasty plus stenting of arteries supplying the brain, other than at the carotid bifurcation. Therefore, we prospectively obtained diffusion-weighted MR images of the brain before and after such revascularization procedures. METHODS: Thirty-seven revascularization procedures were performed in 32 patients. Eleven interventions were performed at the distal internal carotid artery, two at the external carotid artery, two at the common carotid artery, five at the innominate artery, five at the vertebral artery, and 12 at the proximal subclavian artery. Diffusion-weighted MR imaging of the brain was performed before and 24 hours after the procedures. RESULTS: After eight (22%) of 37 procedures, new hyperintensities were visible on the diffusion-weighted MR images. With six of these eight procedures, the hyperintensities occurred in the vascular territory supplied by the treated vessel. In total, 35 new cerebral lesions could be seen, 33 (94%) of which occurred in the vascular territory supplied by the treated vessel. None of the patients in whom new diffusion abnormalities were found had new neurologic symptoms or deficits. No new lesions could be seen after procedures at the subclavian artery. CONCLUSION: Revascularization procedures of arteries supplying the brain were associated with new lesions on the diffusion-weighted MR images of the brain after 22% of the procedures, provided that MR imaging could be performed, indicating the occurrence of cerebral microemboli during such procedures. Diffusion-weighted MR imaging of the brain can be used as a tool to assess the impact of modifications of procedural technique and/or the use of cerebral protection devices on the occurrence of such lesions.

[Cerebral protection with balloon occlusion during carotid artery stent implantation--first experiences]. / Zerebrale Protektion mit Ballonokklusion bei der Stentimplantation der A. carotis--Erste Erfahrungen.

PURPOSE: To assess the technical feasibility and the results of cerebral protection with the GuardWire Plus Temporary Occlusion & Aspiration System during carotid artery stenting for high-grade stenosis. PATIENTS AND METHODS: In 20 patients 20 carotid artery stenoses were treated with stent placement under cerebral protection. A contralateral carotid occlusion was an exclusion criteria for the use of the protection device. In all cases only aspiration, but no flushing was used before deflation of the occlusion balloon. In 17 of 20 patients diffusion-weighted (DW-)MRT imaging of the brain was performed before and 24 hours after the procedure. RESULTS: The stent implantation was successfully performed in all patients. In 3 patients neurologic symptoms occurred during the occlusion time. In these 3 patients the symptoms immediately disappeared after deflation of the balloon. In one case there was dilatation of the internal carotid artery at the site of the balloon inflation. In 3 of the 17 DW-MR images new ipsilateral cerebral lesions, in one case a new contralateral lesion occurred after the procedure. CONCLUSIONS: The cerebral protection procedure is technically feasible. The occlusion of the internal carotid artery was not tolerated by all patients. The DW-MR imaging demonstrated cerebral lesions indicating the occurrence of cerebral microemboli during the procedure. Further investigations are necessary to determine if the use of the cerebral protection device will improve the results of the carotid artery stenting for high-grade stenoses.

Clinical results of cerebral protection with a filter device during stent implantation of the carotid artery.

PURPOSE: To assess the technical feasibility and the clinical results of cerebral protection with the Angioguard emboli-capture guidewire system during carotid artery stenting (CAS) for high-grade stenoses. METHODS: In 20 patients 20 stenoses of the internal carotid artery were treated with stent implantation. In all patients diffusion-weighted (DW) magnetic resonance imaging (MRI) of the brain was performed before and 24 hr after the procedure. RESULTS: The filter device was effectively employed during all steps of the procedure in 16 of 20 patients, in two only postdilation could be protected, and in two no protection was possible. After three of the 20 procedures new ipsilateral cerebral lesions were visualized by DW-MRI. No new permanent neurologic deficits occurred. CONCLUSION: Cerebral protection with the filter device is technically feasible in most cases. DW-MRI demonstrated new cerebral lesions indicating the occurrence of cerebral microemboli during the protected procedures. Further investigations are necessary to determine whether the use of the cerebral protection device will improve the results of CAS.

The early protective effects of L-arginine and Ng-nitro-L-arginine methyl ester after experimental acute spinal cord injury. A light and electron microscopic study.

The purpose of this study was to investigate the early protective effects of L-arginine and Ng-nitro-L-arginine methyl ester (L-NAME) after acute spinal cord injury. Acute spinal cord injury was performed by epidural application of an aneurysm clip at thoracic (T) 7 - 11 level. L-arginine at a dose of 750 microg/kg/min was administered 10 min before acute spinal cord injury and continued for 30 min to 10 animals (Group II). L-NAME at a dose of 250 microg/kg/min was administered 10 min before acute spinal cord injury and continued for 30 min to 10 animals (Group III). No drug was administered to 10 animals after acute spinal cord injury (Group I). Light and electron microscopic analysis were performed in all of the groups. Oedema of perineural, axoplasm or white matter in the L-arginine-treated group was less than in Group I and Group III. Thickening in the walls of the arterioles and venules in the L-arginine-treated group was much milder than in Group I and Group III. Degeneration of myelinated axons in the L-arginine-treated group was milder than in the control group. But there was no different between Group II and Group III.

Ca2+-induced inhibition of adenylyl cyclase in turkey erythrocyte membranes.

We investigated the effects of calcium ions (Ca2+) on the adenylyl cyclase activity in purified turkey erythrocyte membranes. Results showed the following: (i) Ca2+ inhibits cAMP accumulation stimulated by isoproterenol (1 micromol/l), NaF + AlCl3 (10 mmol/l + 20 micromol/l) or forskolin (10 micromol/l) in EGTA-washed turkey erythrocyte membranes. IC50 of free [Ca2+] is approximately 0.1 mmol/l in the presence of Mg2+ (2.5 mmol/l) and isobutylmethylxanthine (1 mmol/l). (ii) The potency of Ca2+ to inhibit cAMP accumulation is independent of the type of stimulus used to activate the adenylyl cyclase. We also evaluated the calcium sensitivity of the basal cAMP accumulation in the presence of GTP (10 micromol/l) and Mg2+ (2.5 mmol/l) which was also inhibited by Ca2+ with the same potency. (iii) The inhibition pattern of cAMP accumulation is not affected by the presence of added calmodulin (100 nmol/l). (iv) Ca2+ is ineffective on the binding of isoproterenol to the beta-adrenoceptors. (v) Increasing the concentration of Ca2+ does not induce an observable activation of cyclic nucleotide phosphodiesterase in the present experimental conditions. Thus, we concluded that the inhibition of cAMP accumulation is due to an inhibition of the adenylyl cyclase rather than the activation of phosphodiesterase(s). The presence of a yet unidentified isoform of adenylyl cyclase that can be directly inhibited by Ca2+ or a Gi protein that can be activated by Ca2+ seems to explain these results. In either case, these results provide an additional mode of cross-talk that can take place between the Ca2+- and cAMP-signaling systems.

Tissue and concentration-dependent effects of sodium selenite on muscle contraction.

In this study, we demonstrated that sodium selenite with high doses (> or = 10(-3) M) were potent in inducing a contracture type effect on heart and smooth muscles. Selenite (Se), at a concentration of 10(-3) M, caused a contracture effect in heart preparations. Also, low Se concentrations did not have any significant effect. Although low concentrations of Se (> or = 10(-5) M) had a biphasic effects on acetylcholine (ACh) induced and spontaneous ileum contractions, 10(-3) M selenite enhanced once more a contracture effect similar to that of the heart preparations. Replacing Ca2+ concentration of the bathing solution by twofold Ca2+ or Ca2+-free did not change the effects of selenite (10(-5) M) on contractility of ileum preparations. In vascular smooth muscle, low concentration of selenite (< 10(-4)) had no significant effects on KCl, and phenylephrine-induced contractions and acetylcholine-induced endothelium-dependent relaxations of isolated rabbit aorta. However, the contractions induced by phenylephrine and the relaxations induced by acetylcholine in rabbit aorta were depressed significantly by high concentration of selenite (10(-3) M). The results obtained by selenite exposure from these three different types of tissue preparations first suggest that the high concentration of selenite exposure induces some alterations in the functions of muscles and endothelium in a tissue- and dose-dependent manner. Second, this observed irreversible type of dysfunction of tissues induced by 10(-3) M selenite is not directly dependent on the Ca2+ entrance into the cytosol, but might be induced by the increase of intracellular Ca2+ with the disturbance of Ca2+ regulation.

TEA inhibits ACh-induced EDRF release: endothelial Ca(2+)-dependent K+ channels contribute to vascular tone.

The effects of K(+)-channel blockers on the acetylcholine (ACh)-induced relaxation of vascular smooth muscle, intracellular free Ca2+ concentration ([Ca2+]i) elevation, and ACh-evoked outward K+ current of endothelial cells of rabbit aorta were studied using bioassay, spectrofluorimetry, and patch-clamp techniques, respectively. In bioassay experiments, ACh caused relaxation of endothelium-denuded aortic rings in a concentration-dependent manner when perfused through an endothelium-intact donor segment of aorta but not when perfused directly onto the recipient aortic ring. ACh-induced relaxation was inhibited by perfusion of tetraethylammonium ions (TEA; 5 mM) through the donor but not by perfusion directly onto the recipient segment. Glibenclamide had no effect on ACh-induced relaxation of the bioassay ring in either situation. ACh increased [Ca2+]i at the endothelial surface of aortic strips but not at the adventitial surface. TEA inhibited ACh-induced [Ca2+]i elevation, whereas glibenclamide had no effect. In patch-clamp experiments with freshly isolated endothelial cells, ACh evoked a biphasic outward current which was completely abolished by TEA (3 mM). It is concluded that Ca(2+)-dependent K+ channels are important for increasing [Ca2+]i during agonist stimulation and consequently for the synthesis/release of endothelium-derived relaxing factors (EDRFs). Furthermore, endothelial ATP-sensitive K+ channels do not contribute to ACh-induced relaxation or evoke an increase in endothelial [Ca2+]i of rabbit thoracic aorta.

The passive calcium leak in cultured porcine aortic endothelial cells.

Unidirectional 45Ca fluxes were measured to characterize the calcium-leak pathway in cultured endothelial cells of pig aorta. Lanthanum, but not other calcium channel blockers such as verapamil and diltiazem, inhibited the passive 45Ca uptake. Passive uptake of 45Ca across the plasma membrane increased progressively as extracellular pH was raised from 5.2 to 9.2. SK&F 96365, a putative inhibitor of receptor-mediated Ca2+ entry, selectively blocked ATP-induced 45Ca uptake. The Ca2+ leak pathway may play an important role in endothelial cell Ca2+ homeostasis which controls the basal and stimulated release of endothelium-derived relaxing factors (i.e., EDRF, prostacyclin) in physiological or pathological situations.

Inhibition of iloprost of the contractile effect of noradrenaline in mesenteric artery rings: evidence for a possible calcium-dependent mechanism.

Iloprost caused a concentration-dependent decrease in the response to noradrenaline in the rabbit isolated endothelium denuded rings from superior mesenteric artery but not thoracic aorta. Similar inhibition was obtained by verapamil using identical concentrations. In Ca(2+)-free EGTA containing medium noradrenaline both at lower and higher concentrations elicited a reduced contractile response and further addition of Ca2+ (2.5 mM) to the medium produced a second contraction in both mesenteric artery and aortic rings which was significantly and equally inhibited by iloprost and verapamil using identical concentrations in mesenteric artery but not in aortic rings. Prior addition of iloprost to the medium did not protect the inhibitory effect of phenoxybenzamine against noradrenaline-induced contraction. These results were taken as an evidence for the possible Ca2+ entry reducing effect of iloprost in mesenteric artery but not thoracic aorta. These results were also taken as an indirect evidence supporting the hypothesis that increased synthesis of prostacyclin by noradrenaline in the vascular wall may inhibit the contractile effect of the agonist by a (-) feedback mechanism mediated by Ca2+ entry into the vascular smooth muscle.

Endothelium modulates the effects of alpha-adrenoceptor agonists in vascular smooth muscle.

1. The contraction induced by clonidine and guanfacine but not phenylephrine was enhanced in endothelium-denuded and methylene blue pretreated rabbit aortic, pulmonary artery rings and isolated perfused whole superior mesenteric, carotid and femoral arteries from the same species. 2. The responses to acetylcholine in guanfacine preconstricted superior mesenteric and carotid arteries were also augmented when compared with phenylephrine and clonidine preconstricted segments. No difference was observed to the relaxing activity of acetylcholine in the aortic, pulmonary artery rings and whole perfused femoral artery contracted by phenylephrine, clonidine and guanfacine. 3. Acetylcholine also produced a biphasic effects in the pulmonary artery rings precontracted with the used alpha-adrenoceptor agonists. The contractile effect was observed with the concentration up to 10(-6) M of acetylcholine and was higher for guanfacine than phenylephrine and clonidine precontracted rings. 4. These results were taken as an evidence for the specificity of alpha 2-adrenoceptor agonists on the production and release of EDRF from vascular endothelium and in this respect guanfacine seems to have higher potency than clonidine. 5. These results also indicate that production or release of EDRF from vascular endothelium may vary depending on the regional vascular bed.

Possible calcium channel modulating activity of iloprost in rabbit isolated vascular segments.

1. Iloprost produced a concentration-dependent decrease in the contractile responses to K+ and NA in CoeA and SMA strips without altering the responses in RA, MPA, CaA and ThA. 2. Nifedipine and verapamil also inhibited the contractile responses to NA and K+ in CoeA and SMA rings in a concentration dependent manner. 3. Removal of endothelium did not alter the inhibitory effects of iloprost and Ca2+ channel blockers in the investigated arterial segments. 4. In Ca2+ free and high K+ medium both iloprost and Ca2+ channel blockers potentially inhibited the contractile effect of external Ca2+. 5. Iloprost at lowest concentration which is not influence the contractile response of K+ can potentiate the inhibitory effect of nifedipine in CoeA and SMA rings without altering its inhibitory effect on NA responses. 6. These results suggest that iloprost may modulate Ca2+ entry into the cell probably by acting through the potential-operated Ca2+ channels in CoeA and SMA. These results also indicate that Ca2+ channels are heterogenously functioned in various vascular segments of rabbit.

Ergotamine enhances acetylcholine-induced relaxation in various vascular segments of rabbits.

The vasodilator response to acetylcholine and-betanechol in segments of rabbit carotid, superior mesenteric and femoral arteries preconstricted with phenylephrine and with intact endothelium was enhanced when the vessels were preincubated with ergotamine but not with ergometrine for 60 min or longer. The vasodilator response to acetylcholine and its enhancement by ergotamine were completely prevented after methylene blue pretreatment or endothelium removal. These results were taken as evidence that the potentiation by ergotamine of the vasodilator response to acetylcholine is probably due to an increased production of EDRF from the vascular endothelium.

Effect of endothelium on phenylephrine-induced contraction in the rat isolated aortic strips.

Phenylephrine tested on the isolated endothelium intact rat aortic strips elicited an enhanced response in the second assay when compared with the response obtained in the first trial. Removal of the endothelium and the pretreatment of the strips in the endothelium with methylene blue almost completely prevented the enhanced response to phenylephrine in the second assay. Pretreatment of the strips with lysine acetyl salicylate failed to abolish the potentiated response to phenylephrine in the second test. These data were taken as an evidence that the basal production or release of endothelium-derived relaxing factor(s) but not cyclooxygenase products of arachidonic acid from the endothelium of the rat aorta can influence the contractile effect of phenylephrine and the enhanced response to this alpha-adrenoceptor agonist in the second assay is probably due to the recuded production or release of these endogenously occurring endothelium-originated substance(s).

Iloprost preserves kidney function against anoxia.

Tissue protective activity of iloprost against anoxia was studied in the isolated perfused rabbit kidney. Addition of iloprost to the perfusion medium at concentrations between 10(-9)-10(-7) M attenuated the release of noradrenaline due to periarterial stimulation and decreased urine outflow. Iloprost also caused a concentration-dependent decrease in perfusion pressure. The potentiation by angiotensin II of the vasoconstriction due to periarterial stimulation and increase in urine volume were also decreased by further addition of iloprost into the medium. Iloprost at concentrations below 10(-7) M did not alter the vasoconstrictor effect of exogenously applied noradrenaline. UK 38 485, a powerful thromboxane A2 synthetase inhibitor, significantly suppressed the vascular but greatly potentiated the diuretic effects of angiotensin II. In kidneys exposed to anoxia for 24 hours in Krebs medium, the vascular and diuretic effects of angiotensin II and the release of noradrenaline due to periarterial stimulation were significantly diminished. In addition, interation between UK 38 485 and angiotensin II in both perfusion pressure and urine volume was also reduced after anoxia for 24 hours. On the other hand, no significant loss was observed in all investigated parameters measured in this study, in kidneys exposed to anoxia for 48 hours in the presence of iloprost. From these results it was concluded that iloprost preserves kidneys functionally against anoxia and possible mechanisms of this effect are discussed.

Iloprost maintains acetylcholine relaxations of isolated rabbit aortic strips submitted to hypoxia.

The protective activity of iloprost against hypoxia was studied in isolated rabbit aortic strips precontracted with angiotensin II on the relaxant effect of acetylcholine. Exposure of the strips to hypoxia in the medium of Krebs alone for 48 h caused a highly significant decrease in the relaxing effect of acetylcholine without altering that of papaverine. Iloprost (10(-9) mol/l), when added to the incubation medium before hypoxia, prevented the loss of the relaxing effect of acetylcholine in angiotensin II precontracted strips. These results were taken as evidence indicating possible protective activity of iloprost in vascular endothelium.

Modulation by endothelium of the vascular effects of angiotensin II.

Modulation by vascular endothelium of the effects of AII was studied in the isolated rabbit aortic and superior mesenteric artery strips. The contractile effect of AII was enhanced in rubbed aortic strips. Similar enhancement was obtained in hydroquinone pretreated unrubbed strips. The relaxing effect of acetylcholine in AII--induced precontracted aortic strips was abolished after rubbing and hydroquinone pretreatment. However, no significant changes were observed in the contractile response to AII on aspirin and nicotine pretreated strips. In the isolated mesenteric artery strips AII produced a biphasic responses. The contractile effect of AII was enhanced in rubbed strips. Similar potentiation was also obtained in hydroquinone, aspirin and nicotine pretreated unrubbed strips. The relaxation phase of AII response was completely abolished in rubbed strips but partially inhibited in hydroquinone, aspirin and nicotine pretreated unrubbed strips. From these results it was concluded that EDRF is the main endothelial humoral factor which modulates the effect of AII in the rabbit aorta while both EDRF and PGI2 are involved for the modulation of the effects of octapeptide in the mesenteric artery.