In-Vitro Neutralization of the Neurotoxicity of Coastal Taipan Venom by Australian Polyvalent Antivenom: The Window of Opportunity

Abstract

Coastal taipan (Oxyuranus scutellatus) envenoming causes life-threatening neuromuscular paralysis in humans. We studied the time period during which antivenom remains effective in preventing and arresting in vitro neuromuscular block caused by taipan venom and taipoxin. Venom showed predominant pre-synaptic neurotoxicity at 3 µg/mL and post-synaptic neurotoxicity at 10 µg/mL. Pre-synaptic neurotoxicity was prevented by addition of Australian polyvalent antivenom before the venom and taipoxin and, reversed when antivenom was added 5 min after venom and taipoxin. Antivenom only partially reversed the neurotoxicity when added 15 min after venom and had no significant effect when added 30 min after venom. In contrast, post-synaptic activity was fully reversed when antivenom was added 30 min after venom. The effect of antivenom on pre-synaptic neuromuscular block was reproduced by washing the bath at similar time intervals for 3 µg/mL, but not for 10 µg/mL. We found an approximate 10-15 min time window in which antivenom can prevent pre-synaptic neuromuscular block. This time window is likely to be longer in envenomed patients due to the delay in venom absorption. Similar effectiveness of antivenom and washing with 3 µg/mL venom suggests that antivenom most likely acts by neutralizing pre-synaptic toxins before they interfere with neurotransmission inside the motor nerve terminals.

Keywords: antivenom; paralysis; post-synaptic; pre-synaptic; taipan; venom.

Figure 1

In vitro neurotoxicity of O. scutellatus venom: (a) concentration-dependent inhibition of indirect twitches in chick biventer nerve-muscle preparation by venom. (* indicates significantly different from control at the corresponding time, p < 0.05, one-way ANOVA followed by Tukey’s multiple comparison test, n = 3–5); (b) effect of venom on responses to exogenous agonists acetylcholine (ACh), carbachol (CCh) and KCl (* indicates significantly different from control, p < 0.05, one-way ANOVA followed by Tukey’s multiple comparison test, n = 4–5).

Figure 2

Effect of the Australian polyvalent antivenom on O. scutellatus venom pre-synaptic neurotoxicity: (a) effect of the antivenom (AV) on preventing/arresting the inhibition of indirect twitches caused by 3 µg/mL venom (* indicates significantly different from control at 180 min, ** indicates significantly different from both control and venom at 180 min, p < 0.05, one-way ANOVA followed by Tukey’s multiple comparison test, n = 4–5); (b) the effect of antivenom, added at different time points, on the venom-mediated response to exogenous agonists acetylcholine (ACh), carbachol (CCh) and KCl.

Figure 3

Effect of Australian polyvalent antivenom (AV) on taipoxin-mediated pre-synaptic neurotoxicity: (a) reversibility of the inhibition of indirect twitches caused by taipoxin (* indicates significantly different from control at 180 min, p < 0.05, one-way ANOVA followed by Tukey’s multiple comparison test, n = 3–5); (b) effect of taipoxin on responses to exogenous agonists acetylcholine (ACh), carbachol (CCh) and KCl.

Figure 4

Effect of washing the preparation on O. scutellatus venom pre-synaptic neurotoxicity: (a) effect of washing on the inhibition of indirect twitches caused by venom (3 µg/mL). (* indicates significantly different from control at 180 min, ** indicates significantly different from both control and venom at 180 min, p < 0.05, one-way ANOVA followed by Tukey’s multiple comparison test, n = 4-5); (b) the effect of washing at different time points on the venom response to exogenous agonists acetylcholine (ACh), carbachol (CCh) and KCl.

Figure 5

Effect of washing and Australian polyvalent antivenom (AV) on the neuromuscular block caused by 10 µg/mL O. scutellatus venom, when applied after different time intervals following the application of venom: the effect of washing and antivenom on the venom-mediated inhibition of indirect twitches and the response of the neuromuscular preparation to exogenous agonists acetylcholine (ACh), carbachol (CCh) and KCl when added or commenced after 30 min (a,b), 15 min (c,d) and 5 min (e,f) (# indicates the twitch height of the AV added preparations is significantly different to preparations washed at the corresponding time point, p < 0.05, Mann Whitney test, n = 3–5; * indicates significantly different from control, p < 0.05, one-way ANOVA followed by Tukey’s multiple comparison test, n = 3–5).