Intensive Care Medicine

, Volume 31, Issue 3, pp 441–446 | Cite as

Direct vs. mediated effects of scorpion venom: an experimental study of the effects of a second challenge with scorpion venom

  • Lamia Ouanes-Besbes
  • Souhail El Atrous
  • Semir Nouira
  • Nicolas Aubrey
  • Alain Carayon
  • Mohamed El Ayeb
  • Fekri Abroug
Experimental

Abstract

Objective

To assess the respective roles of venom and of catecholamines following scorpion envenomation and to verify whether a second challenge with scorpion venom induces the same consequences than a first one.

Design and setting

Controlled animal study in a university research laboratory.

Subjects

Anesthetized and ventilated dogs.

Interventions

Fifteen dogs received intravenously a sublethal dose of scorpion venom (0.05 mg/kg). In the reenvenomated group (n=5) a second venom challenge with one-half sublethal venom dose was performed 30 min after the first one. The control group (n=10) received saline. Five additional animals served as sham.

Measurements and results

Plasma toxin and catecholamine levels and a set of usual hemodynamic measurements were repeatedly measured in the first hour following envenomation. In the reenvenomated group another set of measurements was performed 5 min after the second challenge. Changes in toxin, catecholamines, and the main hemodynamic parameters were compared between the study groups. Initial peak toxin levels were similar in the two groups. They induced a striking increase in circulating catecholamines, a fall in heart rate, and an increase in mean arterial and pulmonary artery occluded pressures and in systemic vascular resistance. In the reenvenomated group the second challenge with scorpion venom achieved a toxin blood level similar to the first peak. However, it was not associated with a significant effect either on catecholamines release or on hemodynamics. Subsequent trends in hemodynamic changes were similar to those observed in the control group.

Conclusions

These data emphasize the limited role of direct effects of scorpion venom on the cardiovascular system and the key role of catecholamines.

Keywords

Scorpion envenomation Hemodynamics Dog 

Introduction

Scorpion envenomation has a high attendant morbidity and mortality due mostly to acute heart failure presenting either as pulmonary edema or cardiogenic shock [1, 2]. Shortly following scorpion envenomation a catecholamine outpouring seems to account for its main cardiovascular features (increase in vascular resistance, hypertension, cardiac dysfunction, and pulmonary edema) [1, 2, 3, 4]. In addition to the role of catecholamines, a direct role of scorpion toxins on the cardiovascular system has been repeatedly reported, although this issue has recently been seriously questioned [5, 6, 7, 8, 9, 10]. However, this represents one of the most challenging issues in scorpion envenomation since it has major therapeutic implications. Indeed, the demonstration of a direct role of scorpion toxins on the cardiovascular system would provide substantial grounds to the use of scorpion antivenom to neutralize circulating levels of venom at any moment in the course of scorpion envenomation. Moreover, despite its short elimination half-life scorpion venom has been detected in blood several hours after envenomation in some circumstances [11]. On the other hand, there is compelling evidence suggesting that scorpion venom does no more than the ignition of a myriad of circulating mediators with potent cardiovascular action, which in turn elicit the main cardiorespiratory consequences of severe scorpion envenomation. In addition to catecholamines, many other mediators have been shown to be involved, including neuropeptide Y, endothelin, and cytokines [3, 8, 12, 13].

To further delineate the respective role of direct and mediated effects of scorpion venom we conducted an experimental study to determine whether a second challenge with scorpion venom administered 30 min after the first one induces similar cardiovascular effects. We hypothesized that since the physiological response to prolonged or repeated sympathetic stimulation is reduced because of exhaustion of the catecholamines stores, a second challenge with scorpion venom would not be associated with a substantial release of catecholamines, and that any further alteration in cardiovascular status would therefore be due to the direct effects of scorpion toxins.

Materials and methods

All conditions of animal anesthesia, catheterization, and killing conformed with the international guidelines and were approved by the local institutional review board on clinical and animal research.

Experimental preparation

Twenty mongrel dogs weighing 13.8±3 kg were perfused with saline throughout the study period. Anesthesia was performed with pentobarbital (10 mg/kg for the induction, renewed as required thereafter). Animals were intubated and mechanically ventilated (tidal volume=10–15 ml/kg, frequency=20 cycles/min, FIO2=50% and adjusted as mandated by blood gazes measurements). Right heart catheterization by a Swan-Ganz 5F-catheter was then carried out by denuding the femoral vein, and the pulmonary artery catheter was positioned through the check of vessels pressures. An arterial catheter was introduced in the femoral artery.

Collected parameters

The pressures in the systemic artery, pulmonary artery, occluded pulmonary artery, and the right atrium were recorded. Cardiac output measurements were performed by the thermodilution technique. The usually derived parameters were calculated: cardiac output adjusted to animal weight, stroke volume (ml/kg), and systemic and pulmonary vascular resistance. Blood samples were also withdrawn for the dose of catecholamines and of scorpion toxin Aah-1.

Experimental protocol

Three groups of animals were studied in this experiment. The first group (n=5) served as sham preparation. The second group (n=10; control) had one challenge with scorpion venom allowing a characterization of the main hemodynamic and neurohormonal consequences of scorpion envenomation. In the third group (n=5; reenvenomation) two injections with scorpion venom were made, the first after baseline values has been obtained and a second challenge after 30 min. In envenomated animals a first dose of 0.05 mg/kg body weight of the purified venom toxic fraction (G50 fraction of the scorpion Androctonus australis hector) was injected by the forearm vein after verification of the preparation stability (systemic artery pressure and heart rate variation of less than 10% for at least 10 min). The above hemodynamic parameters were measured at baseline (prior to the venom injection) and at 5, 30, and 60 min following venom injection. In the reenvenomation group (n=5) a second injection of a one-half dose venom (0.025 mg/kg) was injected after 30 min. This period was chosen because catecholamine stores were thought to be exhausted following the first venom challenge. In addition to the above points of time, another hemodynamic set of measurements was made after 35 min in the reenvenomation group (5 min after the second venom challenge). Blood samples (for the dose of catecholamines and scorpion toxin Aah-1) were withdrawn at each of these points of time and at 15 min.

Biological measurement

All blood samples were collected and centrifuged at 4°C. Plasma was stored at −20°C until analysis of plasma catecholamines were quantified by radioenzymatic assay (CAT-A-KITTM assay system; TRK895, Amersham; detection limit 20 pg/ml). The scorpion toxin Aah-1 levels were measured by a recombinant scFv/streptaridum-binding peptides fusion, protein method (detection limit: 0.3 ng/ml) [14]. Collected data in the control group were previously published in a study evaluating the effects of scorpion antivenom administered at various doses and at different times of experimental evenomation [3].

Statistics

Data are presented as medians with interquartile range. Intragroup comparisons were made with Friedman’s test while between groups comparisons for each point of time used the Kruskall-Wallis tests. Comparison between the hemodynamic effects of the two venom challenges in the reenvenomated group was made with the Wilcoxon test. Differences at the level of p<0.05 were considered statistically significant.

Results

Scorpion toxin levels in blood

Figure1 shows the blood levels of the toxin Aah-1 in both control and reenvenomation groups. In the control group peak toxin level was measured at 5min. A sustained decrease in toxin levels was recorded thereafter. In the reevenomation group a biphasic course was observed with a first blood peak measured by 5 min achieving a level similar to that recorded in the control group (NS). Five minutes after the second injection of scorpion venom (35 min) a second peak was measured with an equivalent level to the first one (NS).
Fig. 1

Change in toxin Aah-1 levels in the control group (filled bars) and the reenvenomation group (open bars)

Hemodynamic and neurohormonal changes following scorpion venom administration

Hemodynamic variables at baseline were similar in control and reenvenomation groups (Table 1). Similar variations in hemodynamic parameters were observed after scorpion envenomation in the two groups. Scorpion venom induced a fall in heart rate and an increase in mean arterial pressure as a consequence of a sharp increase in systemic resistance (p<0.05 by Friedman’s test, for each of these parameters). The increase in left ventricular afterload was associated with a sharp increase in pressure of occluded pulmonary artery while stroke volume which was initially maintained close to the preenvenomation level eventually decreased. Regarding the neurohormonal effects of scorpion venom a striking release of both epinephrine and norepinephrine was recorded by 5 minute (almost a 40-fold increase in both catecholamines circulating levels) in both groups (Fig. 2).
Table 1

change in hemodynamic parameters after the first and the second venom challenge. No statistical difference was observed in comparisons between control and reenvenomation groups (HR heart rate, MAP mean arterial pressure, SV stroke volume, PAOP pressure of occluded pulmonary artery)

Baseline

5 min

30 min

60 min

HR (bpm)

  Sham

201±43

189±66

192±67

196±64

  Control

204±39

150±68

146±69

166±89*

  Reenvenomation

216±63

152±89

136±55

101±34*

MAP (mmHg)

  Sham

136±33

139±36

140±23

141±33

  Control

125±38

212±34

161±44

127±35*

  Reenvenomation

136±34

213±37

141±80

90±60*

SV (ml/kg)

  Sham

19±8

17±9

16±9

17±9

  Control

19±10

20±10

16±10

12±9*

  Reenvenomation

16±10

18±12

13±13

12±14*

SVR (dyne s−1 m−5)

  Sham

2820±1712

2875±1698

2613±1735

2912±1678

  Control

2450±1873

5775±3853

6787±5675

6515±3815*

  Reenvenomation

3650±1445

4693±2474

7096±7999

5835±4015*

PAOP (mmHg)

  Sham

3±2

2±3

1±2

3±2

  Control

2±2

23±8

20±14

8±5.5*

  Reenvenomation

4±1

24±13

5±19

6±9*

Fig. 2

Change in norepinephrine (top) and epinephrine (bottom) levels in control (filled bars) and reenvenomation (open bars) groups

Effects of the second challenge with scorpion venom

The second administration of venom did not elicit significant changes in hemodynamic and neurohormonal effects of scorpion envenomation. In comparison to the consequences of the first injection of scorpion venom in the reenvenomated group, and despite similar blood levels of Aah-1 toxin, there was almost no variation in pressure of occluded pulmonary artery, heart rate, or stroke volume while mean arterial pressure tended to decrease instead of the recorded increase following the first venom administration (Table 2). Sustained catecholamine decrease was recorded despite reenvenomation. In fact the natural trend over time of neurohormonal and hemodynamic parameters following envenomation was not altered by the second challenge with scorpion venom. Table 1 shows the lack of any statistically significant difference between 30 and 60 min in control and reenvenomation groups with regard to the variations in the hemodynamic parameters of interest.
Table 2

Comparison in the reenvenomated group of absolute variations following initial and second venom challenges (HR heart rate, MAP mean arterial pressure, SV stroke volume, PAOP pressure of occluded pulmonary artery)

Variation 5 min after initial envenomation

Variation 5 min after reenvenomation

p

HR (bpm)

−88±88

−43±35

0.043

SV (ml/kg)

6.4±21

1±5

0.34

MAP (mmHg)

52±48

−21±35

0.003

PAOP (mmHg)

21±13

0±6

0.005

Epinephrine (%)

8,952±1,100

−1,862±4,522

0.035

Norepinephrine (%)

16,982±5,630

−735±4,092

0.034

Discussion

Despite similar toxin levels achieved in blood by a first and a second challenge with scorpion venom we did not observe similar pathophysiological effects. Indeed, the catecholamine storm and the major hemodynamic changes usually associated with experimental scorpion envenomation (increases in vascular resistance in left ventricular filling pressure and in systemic pressure and a decrease in heart rate) were not observed following the second venom challenge. Obviously our findings cannot firmly rule out a direct toxic effect on cardiovascular and neurohormonal effects of envenomation; however, they do emphasize the major role of catecholamine blood levels.

Two possible explanations may explain our findings. The first is that scorpion toxin does not account by itself for the observed cardiovascular impairment following envenomation, acting rather through the release of mediators. The second is that following scorpion envenomation there is a period during which no further impairment in hemodynamic status is induced (refractory period). We hypothesize that this refractory period concerns either the stimulation of the sympathetic system (saturation of venom receptors) or that of adrenergic receptors (a phenomenon of sympathetic receptors’ saturation or downregulation). The lack of catecholamine increase following the second venom challenge lessens the putative role of the sympathetic receptors’ downregulation and emphasizes the major role of the lack of catecholamine release underlying the lack of a protracted hemodynamic impairment. The lack of a secondary increase in circulating catecholamines observed in our study may be due to saturation of scorpion venom receptors achieving a persistent maximum effect. It may also be related to the fact that the physiological response to prolonged or repeated sympathetic stimulation is reduced because of an exhaustion of the catecholamines stores [15]. Hence all assumptions converge toward the key role of catecholamines in the occurrence of hemodynamic consequences of severe scorpion envenomation. Although speculative, opinions differ regarding the mechanisms underlying catecholamine release following scorpion envenomation. Peripheral sympathetic stimulation, central spinal and sympathetic preganglionic stimulation, hypothalamic stimulation, and adrenal medullary secretory effect, alone or in combination, have been advocated [1].

In experimental models scorpion venom has been shown to achieve significant levels of concentration in many tissues of which the heart, lungs, and kidneys, suggesting a direct action of scorpion venom on these tissues [6]. Scorpion venom is indeed a potent activator of the sodium, potassium, and calcium channels at the cellular level and therefore has the potential for direct action on many tissues. Nevertheless, the fact that the sympathetic system is usually stimulated following scorpion envenomation represents a confounding factor in any attempt to separate direct venom effects on the cardiovascular system from those mediated by catecholamine release. Few experimental studies have addressed the issue of respective effects of direct and mediated cardiovascular consequences of scorpion envenomation. Almeida et al. [9] and Couto et al. [10] studied the direct effects of scorpion venom on the isolated pig heart and atria. They recorded a dose-dependent inotropic effect of scorpion venom on isolated pig heart. At the atrium level the addition of toxin purified from Tityus serrulatus scorpion venom induced a transient increase in contractile force that was prevented by a β-blocker (metoprolol) reflecting the adrenergic origin of this inotropic effect, emphasizing again the difficulty even in the experimental setting to separate the direct effects of venom from those mediated by the catecholaminergic release. Our study provides additional evidence in favor of the mediator theory although its experimental design does not allow ruling out direct toxic effects since catecholamines-induced hemodynamic disturbances are prominent.

The issue addressed by the current study is both pathophysiologically and clinically relevant. Apart from its contribution to the understanding of the pathophysiology of scorpion envenomation, our study helps to answer one of the recurrent questions surrounding the treatment of severe scorpion envenomation: should we systematically antagonize scorpion toxin that persists in the blood of envenomated patients [11, 16, 17, 18, 19, 20, 21, 22, 23, 24]? The answer to this question leads to two questions. Firstly, what is the relevance of measured toxin levels in comparison to their toxic levels in scorpion envenomation. Secondly, can pathophysiological features of scorpion envenomation be reproduced without any limit of time each time scorpion toxin is administered? Our study provides a piece of answer to the second issue. It suggests that during the first 30 min following scorpion envenomation circulating scorpion toxin (even at levels similar to that associated with severe envenomation’s features) is no longer able to induce harm. Determining the duration of this refractory period was beyond the scope of the current study, but this parameter is probably important to delineate. Whether serum antivenom should be administered beyond the 30-min limit remains an open question. Nevertheless, our findings suggest that antagonizing catecholamine effects on receptors would be more promising than neutralizing the venom.

References

  1. 1.
    Ismail M (1995) The scorpion envenoming syndrome. Toxicon 33:825–858CrossRefGoogle Scholar
  2. 2.
    Gueron M, Ilia R, Sofer S (1992) The cardiovascular system after scorpion envenomation. A review. J Toxicol Clin Toxicol 30:245–258Google Scholar
  3. 3.
    Abroug F, Nouira S, El Atrous S, Besbes L, Boukef R, Boussarsar M, Marghli S, Eurin J, Barthelemy C, El Ayeb M, Dellagi K, Carayon A (2003) A canine study of immunotherapy in scorpion envenomation. Intensive Care Med 29:2266–2276CrossRefPubMedGoogle Scholar
  4. 4.
    Zeghal K, Sahnoun Z, Guinot M, Richer C, Giudicelli JF (2000) Characterization and mechanisms of the cardiovascular and haemodynamic alterations induced by scorpion venom in rats. Fundam Clin Pharmacol 14:351–361Google Scholar
  5. 5.
    Silveira NP, Moraes-Santos T, Azevedo AD, Freire-Maia L (1991) Effects of Tityus serrulatus scorpion venom and one of its purified toxins (toxin gamma) on the isolated guinea-pig heart. Comp Biochem Physiol C Pharmacol Toxicol 98:329–336CrossRefGoogle Scholar
  6. 6.
    Ismail M, Abd-Elsalam MA (1988) Are the toxicological effects of scorpion envenomation related to tissue venom concentration? Toxicon 26:233–256CrossRefGoogle Scholar
  7. 7.
    Drumond YA, Couto AS, Moraes-Santos T, Almeida AP, Freire-Maia L (1995) Effects of toxin Ts-gamma and tityustoxin purified from Tityus serrulatus scorpion venom on isolated rat atria. Comp Biochem Physiol C Pharmacol Toxicol Endocrinol 111:183–190CrossRefGoogle Scholar
  8. 8.
    Matos IM, Teixeira MM, Leite R, Freire-Maia L (1999) Pharmacological evidence that neuropeptides mediate part of the actions of scorpion venom on the guinea pig ileum. Eur J Pharmacol 368:231–236CrossRefGoogle Scholar
  9. 9.
    Almeida AP, Alpoim NC, Freire-Maia L (1982) Effects of a purified scorpion toxin (tityustoxin) on the isolated guinea pig heart. Toxicon 20:855–865CrossRefGoogle Scholar
  10. 10.
    Couto AS, Moraes-Santos T, Azevedo AD, Almeida AP, Freire-Maia L (1992) Effects of toxin Ts-gamma, purified from Tityus serrulatus scorpion venom, on the isolated rat atria. Toxicon 30:339–343CrossRefGoogle Scholar
  11. 11.
    De Rezende NA, Dias MB, Campolina D, Chavez-Olortegui C, Diniz CR, Amaral CF (1995) Efficacy of antivenom therapy for neutralizing circulating venom antigens in patients stung by Tityus serrulatus scorpions. Am J Trop Med Hyg 52:277–280Google Scholar
  12. 12.
    Freire-Maia L, de Matos IM (1993) Heparin or a PAF antagonist (BN-52021) prevents the acute pulmonary edema induced by Tityus serrulatus scorpion venom in the rat. Toxicon 31:1207–1210CrossRefGoogle Scholar
  13. 13.
    Fukuhara YD, Reis ML, Dellalibera-Joviliano R, Cunha FQ, Donadi EA (2003) Increased plasma levels of IL-1beta, IL-6, IL-8, IL-10 and TNF-alpha in patients moderately or severely envenomed by Tityus serrulatus scorpion sting. Toxicon 41:49–55CrossRefGoogle Scholar
  14. 14.
    Aubrey N, Devaux C, Billiald P (2002) [Rapid immunotitration of individual toxins from Androctonus australis venom]. Bull Soc Pathol Exot 95:194–196Google Scholar
  15. 15.
    Rona G (1985) Catecholamine cardiotoxicity. J Mol Cell Cardiol 17:291–298Google Scholar
  16. 16.
    Gueron M, Ilia R (1999) Is antivenom the most successful therapy in scorpion victims? Toxicon 37:1655–1657CrossRefGoogle Scholar
  17. 17.
    Abroug F, El Atrous S, Nouira S, Haguiga H, Touzi N, Bouchoucha S (1999) Serotherapy in scorpion envenomation: a randomised controlled trial. Lancet 354:906–909CrossRefGoogle Scholar
  18. 18.
    Bawaskar HS, Bawaskar PH (1991) Treatment of cardiovascular manifestations of human scorpion envenoming: is serotherapy essential? J Trop Med Hyg 94:156–158Google Scholar
  19. 19.
    Belghith M, Boussarsar M, Haguiga H, Besbes L, Elatrous S, Touzi N, Boujdaria R, Bchir A, Nouira S, Bouchoucha S, Abroug F (1999) Efficacy of serotherapy in scorpion sting: a matched-pair study. J Toxicol Clin Toxicol 37:51–57CrossRefGoogle Scholar
  20. 20.
    Possani LD (2000) Antivenom for scorpion sting. Lancet 355:67Google Scholar
  21. 21.
    Soulaymani-Bencheikh R, Faraj Z, Semlali I, Khattabi A, Skalli S, Benkirane R, Badri M (2002) Epidémiologie des piqres de scorpion au Maroc. Rev Epidemiol Sante Publique 50:341–347Google Scholar
  22. 22.
    Ghalim N, El-Hafny B, Sebti F, Heikel J, Lazar N, Moustanir R, Benslimane A (2000) Scorpion envenomation and serotherapy in Morocco. Am J Trop Med Hyg 62:277–283Google Scholar
  23. 23.
    Isbister GK, Graudins A, White J, Warrell D (2003) Antivenom treatment in arachnidism. J Toxicol Clin Toxicol 41:291–300CrossRefGoogle Scholar
  24. 24.
    Sofer S, Shahak E, Gueron M (1994) Scorpion envenomation and antivenom therapy. J Pediatr 124:973–978Google Scholar

Copyright information

© Springer-Verlag 2005

Authors and Affiliations

  • Lamia Ouanes-Besbes
    • 1
  • Souhail El Atrous
    • 2
  • Semir Nouira
    • 1
  • Nicolas Aubrey
    • 3
  • Alain Carayon
    • 4
  • Mohamed El Ayeb
    • 5
  • Fekri Abroug
    • 1
  1. 1.Intensive Care UnitCHU F BourguibaMonastirTunisia
  2. 2.Unité de Recherche (UR/06/02)CHUMonastirTunisia
  3. 3.Museum National d’Histoire NaturelleParisFrance
  4. 4.Laboratoire de BiochimieUpres 1390 CHU Pitié SalpétrièreParisFrance
  5. 5.Institut PasteurTunisTunisia

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