L. Freire-Maia

Approaches to the treatment of scorpion envenoming

A total of 3866 patients stung by Tityus serrulatus scorpion was admitted to Hospital João XXIII, in Belo Horizonte, Brazil, over a 16-year period (an average of 241 cases per year). Of these, 73% were adults and 27% were children aged less than 14 years. The moderate or benign cases were treated with symptomatic measures and/or i.v. antivenom, whereas 168 severely envenomed children were treated in the Intensive Care Unit. Lung oedema was unilateral in several cases, with the presence of air bronchograms and a peripheral distribution, suggesting that a noncardiogenic factor is also involved in the genesis of lung oedema. The treatment consisted of symptomatic measures, support of vital functions and i.v. antivenom. The mortality was 1% among children and 0.28% for the total number of patients.

Acute pulmonary edema after Tityus serrulatus scorpion sting in children

Acute pulmonary edema due to scorpion toxins has been attributed to acute left ventricular failure resulting from massive catecholamine release or myocardial damage induced by the venom.1–3 It has been proposed that pulmonary edema could also result from increased pulmonary vascular permeability produced by vasoactive substances that may be released by the venom.4 This report presents radiologic, electrocardiographic, enzymatic, echocardiographic and histopathologic evidence that support both cardiogenic and noncardiogenic factors in the pathogenesis of acute pulmonary edema after Tityus serrulatus scorpion sting in children.

Evidence for a direct action of Tityus serrulatus scorpion venom on the cardiac muscle

The ability of toxins to activate the cardiovascular system plays an important role in the morbidity and lethality of the Tityus serrulatus scorpion envenoming. Most of the actions of the scorpion toxins are indirect and due to the release of adrenergic and cholinergic neurotransmitters. Accordingly, treatment following envenoming is targeted towards inhibition of adrenergic and cholinergic receptors. Here, we have sought evidence for a direct action of T. serrulatus venom on the isolated rat heart (Langendorffs method). We show that the bradycardia induced by T. serrulatus venom was completely blocked by atropine, a muscarinic receptor antagonist. Similarly, the increase in heart rate that follows the venom-induced bradycardia was totally inhibited by a beta(1)-adrenoceptor antagonist or by chemical sympathetic denervation with 6-hydroxydopamine. In contrast to these findings, the venom-induced increase in contractile force was not modified by beta(1)-adrenoceptor blockade or by chemical sympathetic denervation. The results clearly demonstrate that the chronotropic effects of T. serrulatus are dependent on neurotransmitter release, but the inotropic effects are not. The neurotransmitter-independent increase in contractility seems to be a direct action of the venom on cardiomyocytes. We suggest that this direct effect on cardiac fibers may play a role in the development of cardiac arrhythmias and contractility defects following envenoming with T. serrulatus scorpion.

Heparin or a PAF antagonist (BN-52021) prevents the acute pulmonary edema induced by Tityus serrulatus scorpion venom in the rat.

Experiments were performed in pentobarbital-anesthetized rats, to study the mechanism of the acute pulmonary edema induced by Tityus serrulatus scorpion venom. In control rats injection of venom (50 micrograms/100 g, i.v.) induced arterial hypertension and lung edema (lung/body index or LBI equal to 1.01 +/- 0.09). In rats pretreated with heparin (100 IU/100 g 30 min previously) the venom induced similar hypertensive effects, but no edema was detected (LBI = 0.63 +/- 0.06, P > 0.05). Similarly, in rats pretreated with the PAF antagonist BN-52021 (0.5 mg/100 g, i.v., 30 min previously), the venom-induced hypertension was not modified but the acute pulmonary edema was prevented (LBI = 0.67 +/- 0.08, P > 0.05). It is concluded that PAF plays an important role on the genesis of pulmonary edema induced by scorpion venom in the rat. It is suggested that the inhibitory action of heparin could be related to a decrease in the vascular permeability in the lungs.

Effects of tachykinin NK1 or PAF receptor blockade on the lung injury induced by scorpion venom in rats

In cases of severe human scorpion envenoming, lung injury is a common finding and frequently the cause of death. In the rat, two distinct mechanisms account for oedema following the intravenous injection of the venom -- acute left ventricular failure resulting from a massive release of catecholamines and an increase in pulmonary vascular permeability. In the present work, we investigated the effects of a tachykinin NK1 receptor antagonist (CP96,345, the dihydrochloride salt of (2S,3S)-cis-2-(diphenylmethyl)-N-((2-methoxyphenyl)methyl)-1-az abicycol[2.2.2]octan-3-amine) and its 2 R-3 R inactive enantiomer (CP96,344) on the acute lung injury induced by the i.v. injection of Tityus serrulatus venom in rats. Lung injury was assessed by evaluating the extravasation of Evans blue dye in the bronchoalveolar lavage fluid and in the lung of venom-treated and control animals. The effects of the platelet-activating factor (PAF) receptor antagonist WEB2170 (2-methyl-1-phenylimidazol[4,5c]pyridine) were evaluated for comparison. The i.v. injection of the venom induced the extravasation of Evans blue in the bronchoalveolar lavage fluid and into the left lung. Pretreament with the tachykinin NK1 receptor antagonist CP96,345, but not CP96,344, inhibited Evans blue dye extravasation in the bronchoalveolar lavage fluid and in the lung by 96% and 86%, respectively. The PAF receptor antagonist WEB2170 inhibited the increase in vascular permeability in the bronchoalveolar lavage fluid by 60% and had no effect on the extravasation to the lung parenchyma of venom-injected animals. In addition to abrogating lung injury, pretreatment of rats with CP96,345, but not CP96,344 or WEB2170, decreased by 70% the mortality induced by the venom. This is the first study to show the relevance of the tachykinin NK1 receptor in mediating lung injury and mortality in animals injected with the neurotoxic T. serrulatus venom. Blockade of the tachykinin NK1 receptor may represent an important strategy in the treatment of patients with signs of severe envenoming and clearly deserves further studies.

Mechanism of the pulmonary edema induced by intravenous injection of scorpion toxin in the rat

The intravenous injection of purified scorpion toxin (tityustoxin, TsTX) into unanesthetized rats induces a severe systemic hypertension followed by a hemorrhagic edema of the lungs. The edema is focal or diffuse, whereas the hemorrhage is always focal and less prominent than the edema. Anesthesia of the rats prevents the appearance of pulmonary edema. It seems likely that this protective action of the anesthesia is due, at least in part, to an interference with the hypertension induced by TsTX. The pulmonary edema is prevented by bilateral adrenalectomy, guanethidine or phenoxybenzamine. It is suggested that the edema depends on a sympathetic-adrenal discharge and that catecholamines released by TsTX act onalpha adrenergic receptors. The mean kininogen content of the rat plasma, 1 h after TsTX injection, is not significantly different from that found in the control animals. The possible role played by kinins and other mediators in the early phases of the pulmonary edema induced by TsTX is under investigation.

Immunotherapy for scorpion envenoming in Brazil

Using the ELISA we have shown that in rats subcutaneously injected with Tityus serrulatus scorpion venom there is a fast absorption rate, a fast and high distribution of venom to tissues, a great affinity of the venom for the tissues and a slow elimination half-life. Because of these experimental data, i.v. immunotherapy should be given to patients stung by scorpions as soon as possible after hospital admission. The severity of scorpion envenoming is related to plasma venom concentration (ELISA). The high levels of plasma scorpion venom antigens (ELISA) were cleared 1 h after the infusion of antivenom (5-30 ml of Fab2 fragment) and high concentrations of circulating antivenom persisted for at least 24 h, confirming the efficacy of immunotherapy to neutralise circulating venom. Some symptoms (e.g. local pain and vomiting) decreased 1 h after the starting of immunotherapy, whereas the other symptoms disappeared from 12-48 h later. Using our tripartite approach of treating scorpion envenoming (symptomatic measures, support of vital functions and serotherapy), the mortality rate was very low (0.28%).

Lung oedema induced by Tityus serrulatus scorpion venom in the rat.

The effects of drugs were investigated on the induction of acute lung oedema by scorpion Tityus serrulatus venom in male Wistar rats (200-230 g) anaesthetized with sodium pentobarbital (40 mg/kg, i.p.). Intravenous (i.v.) injection of scorpion venom (0.5 mg/kg) into 12 rats induced arterial hypertension and severe lung oedema, whereas i.v. injection of scorpion venom into 16 rats previously injected with commercial heparin induced arterial hypertension, but only a slight lung oedema. It is suggested that the inhibitory effect of commercial heparin on the genesis of lung oedema may be due to a decrease in vascular permeability in the lungs. Previous i.v. injection of aprotinin did not prevent the arterial hypertension and the lung oedema induced by scorpion venom. Previous injections of platelet-activating factor antagonists (BN-52021 and WEB-2170) or of an inhibitor of lipo- and cyclooxygenase (Nordihydroguaiaretic acid) did not prevent the arterial hypertension induced by scorpion venom, but decreased the magnitude of the lung oedema elicited by the venom. Previous injections of inhibitors of 5-lipoxygenase (MK-886) or cyclooxygenase (aspirin or indomethacin) significantly decreased the magnitude of the lung oedema induced by scorpion venom. It is concluded that the release of vascular permeability factors, such as platelet-activating factors, leukotrienes, and prostaglandins may play a role in the induction of acute lung oedema by scorpion venom in rats.

Evidence for a role of mast cells in the lung edema induced by Tityus serrulatus venom in rats

In the most severe cases of human poisoning by Tityus serrulatus, pulmonary edema is a frequent finding and can be the cause of death. Mast cells can release a range of mediators known to be involved in the development of lung edema following T. serrulatus venom injection. The present work was designed to investigate whether mast cells participated in the acute lung injury induced by T. serrulatus scorpion venom and could, thus, be an intermediate between neuropeptide release and activation of the inflammatory cascade. To this end, mast cells were depleted using compound 48/80. Pulmonary edema, as assessed by the levels of extravasation of Evans blue dye in the bronchoalveolar lavage and in the left lung, was completely inhibited in compound 48/80-treated animals. Moreover, the number of animals surviving 60min after injection of venom rose from 20 to 60%. Our results demonstrate an important role for mast cells in the development of lung injury and lethality following the intravenous administration of T. serrulatus venom.

Pharmacokinetics of Tityus serrulatus scorpion venom determined by enzyme-linked immunosorbent assay in the rat

Experiments were performed in two groups of anaesthetized rats to study the genesis of pulmonary oedema and to determine the pharmacokinetic parameters following a subcutaneous (s.c.) injection of Tityus serrulatus scorpion venom. In group I, the rats were anaesthetized with pentobarbital (4 mg/100 g, i.p.); the s.c. injection of scorpion venom at the dose of 50 micrograms/100 g did not induce arterial hypertension, but unilateral pulmonary oedema was observed in three of six rats. The injection of a higher dose of venom (200 micrograms/100 g, N = 6) induced arterial hypertension and bilateral (N = 3) or unilateral (N = 1) pulmonary oedema. These data indicate that it is possible to evoke unilateral pulmonary oedema without previous arterial hypertension induced by the venom. For the study of pharmacokinetic parameters a second group of six rats was anaesthetized with urethane (140 mg/ 100 g, i.p.) and the venom injected at a dose of 200 micrograms/100 g, s.c. The plasma concentrations of venom were determined by enzyme-linked immunosorbent assay, at times 0, 5, 30, 60, 180, 360, and 720 min after venom injection. A biphasic curve was obtained with an ascending phase followed by a descending phase. The maximum plasma scorpion venom concentration was reached at 60 min. The pharmacokinetic parameters showed a fast absorption rate (Ka = 0.058 min-1), a fast and high distribution of venom to tissues (t1/2 alpha = 31.50 min and Vdarea = 6800.47 ml.kg-1, respectively), a great affinity of the venom for the tissues (KCT = 0.056 min-1 and KTC = 0.002 min-1) and a slow elimination half-life (t1/2 beta = 173.25 min).