Assessment of neuropharmacological potential of low molecular weight components extracted from Rhinella schneideri toad poison

Abstract

Background: Studies on toad poison are relevant since they are considered a good source of toxins that act on different biological systems. Among the molecules found in the toad poison, it can be highlighted the cardiotonic heterosides, which have a known mechanism that inhibit Na + /K +-ATPase enzyme. However, these poisons have many other molecules that may have important biological actions. Therefore, this work evaluated the action of the low molecular weight components from Rhinella schneideri toad poison on Na + /K +-ATPase and their anticonvulsive and / or neurotoxic effects, in order to detect molecules with actions of biotechnological interest. Methods: Rhinella schneideri toad (male and female) poison was collected by pressuring their parotoid glands and immediately dried and stored at-20 °C. The poison was dialysed and the water containing the low molecular mass molecules (< 8 kDa) that permeate the dialysis membrane was collected, frozen and lyophilized, resulting in the sample used in the assays, named low molecular weight fraction (LMWF). Na + /K + ATPase was isolated from rabbit kidneys and enzyme activity assays performed by the quantification of phosphate released due to enzyme activity in the presence of LMWF (1.0; 10; 50 and 100 µg/mL) from Rhinella schneideri poison. Evaluation of the L-Glutamate (L-Glu) excitatory amino acid uptake in brain-cortical synaptosomes of Wistar rats was performed using [3H]L-glutamate and different concentration of LMWF (10-5 to 10 µg/µL). Anticonvulsant assays were performed using pentylenetetrazole (PTZ) and N-methyl-D-aspartate (NMDA) to induce seizures in Wistar rats (n= 6), which were cannulated in the lateral ventricle and treated with different concentration of LMWF (0.25; 0.5; 1.0; 2.0; 3.0 and 4.0 µg/µL) 15 min prior to the injection of the seizure agent.

Toad venom bufadienolides and bufotoxins: An updated review

“…Among the toad venom active compounds, low‐molecular fractions mainly consisting of bufadienolides are expected to be responsible for Na + /K + ‐ATPase activity. Studying R. schneideri venom enabled such a conclusion (Baldo et al, 2019). A R. marina venom faction containing telocinobufagin (2.48%), marinobufagenin (65.50%), bufalin (8.88%), and resibufogenin (10.27%) demonstrated lower effect and thus higher IC 50 values in HBL (0.09 µg/mL), MM074 (0.1 µg/mL), and MM161 (0.35 µg/mL) melanoma cells compared with their resistant counterparts: HBL‐R (0.07 µg/mL), MM074‐R (0.05 µg/mL), and MM161‐R (0.06 µg/mL).…”

Toad venom bufadienolides and bufotoxins: An updated review

“…Among the toad venom active compounds, low‐molecular fractions mainly consisting of bufadienolides are expected to be responsible for Na + /K + ‐ATPase activity. Studying R. schneideri venom enabled such a conclusion (Baldo et al, 2019). A R. marina venom faction containing telocinobufagin (2.48%), marinobufagenin (65.50%), bufalin (8.88%), and resibufogenin (10.27%) demonstrated lower effect and thus higher IC 50 values in HBL (0.09 µg/mL), MM074 (0.1 µg/mL), and MM161 (0.35 µg/mL) melanoma cells compared with their resistant counterparts: HBL‐R (0.07 µg/mL), MM074‐R (0.05 µg/mL), and MM161‐R (0.06 µg/mL).…”

The Venom Revolution in Biomedicine: Unlocking Nature’s Toxin Toolkit for Therapeutic Innovation

“…Batrachotoxin produced by Phyllobates frogs targets voltage-gated sodium channels, causing sustained activation that leads to muscle contraction, convulsions, and respiratory paralysis [70] . Similarly, toad venom from Rhinella schneideri contains cardiotonic heterosides that inhibit Na⁺/K⁺-ATPase, resulting in cardiotoxic and neurotoxic effects [71] . Poison dart frogs exemplify toxin-based defence, secreting potent skin alkaloids that rapidly penetrate oral and mucosal tissues of predators, causing swelling, nausea, paralysis, or death [72] .…”

Action of heparin and acetylcholine modulators on the neurotoxicity of the toad Rhinella schneideri (Anura: Bufonidae) in Brazil

“…In recent years, several biological properties have been characterized from Rhinella schneideri 'Schneider's toad' poison (Rostelato-Ferreira et al, 2011;Rostelato-Ferreira et al, 2014;Abdelfatah, Lu, Schmeda-Hirschmann, & Efferth, 2019;Leal et al, 2020), a species widely distributed in Brazil (Wake, 2020), including the chemical identification and structural characterization of bufadienolides, e.g., marinobufagenin, bufalin, telocinobufagin, hellebrigenin, 20S21R-epoxymarinobufagin and β-sitosterol, which exhibit cytotoxic activity, modulatory action on the complement cascade, anti-inflammatory activity, inhibitory action of serine protease, presynaptic neuromuscular blocking activity and anticonvulsant action (Cunha-Filho et al, 2010;Anjolette et al, 2015;Sousa-Filho et al, 2016;Shibao, Anjolette, Lopes, & Arantes, 2015;Freitas et al, 2017;Rostelato-Ferreira et al, 2018;Baldo et al, 2019;Zheng et al, 2020).…”

Toad venom bufadienolides and bufotoxins: An updated review

“…Among the toad venom active compounds, low‐molecular fractions mainly consisting of bufadienolides are expected to be responsible for Na + /K + ‐ATPase activity. Studying R. schneideri venom enabled such a conclusion (Baldo et al, 2019). A R. marina venom faction containing telocinobufagin (2.48%), marinobufagenin (65.50%), bufalin (8.88%), and resibufogenin (10.27%) demonstrated lower effect and thus higher IC 50 values in HBL (0.09 µg/mL), MM074 (0.1 µg/mL), and MM161 (0.35 µg/mL) melanoma cells compared with their resistant counterparts: HBL‐R (0.07 µg/mL), MM074‐R (0.05 µg/mL), and MM161‐R (0.06 µg/mL).…”

The Venom Revolution in Biomedicine: Unlocking Nature’s Toxin Toolkit for Therapeutic Innovation

“…Batrachotoxin produced by Phyllobates frogs targets voltage-gated sodium channels, causing sustained activation that leads to muscle contraction, convulsions, and respiratory paralysis [70] . Similarly, toad venom from Rhinella schneideri contains cardiotonic heterosides that inhibit Na⁺/K⁺-ATPase, resulting in cardiotoxic and neurotoxic effects [71] . Poison dart frogs exemplify toxin-based defence, secreting potent skin alkaloids that rapidly penetrate oral and mucosal tissues of predators, causing swelling, nausea, paralysis, or death [72] .…”

Action of heparin and acetylcholine modulators on the neurotoxicity of the toad Rhinella schneideri (Anura: Bufonidae) in Brazil

“…In recent years, several biological properties have been characterized from Rhinella schneideri 'Schneider's toad' poison (Rostelato-Ferreira et al, 2011;Rostelato-Ferreira et al, 2014;Abdelfatah, Lu, Schmeda-Hirschmann, & Efferth, 2019;Leal et al, 2020), a species widely distributed in Brazil (Wake, 2020), including the chemical identification and structural characterization of bufadienolides, e.g., marinobufagenin, bufalin, telocinobufagin, hellebrigenin, 20S21R-epoxymarinobufagin and β-sitosterol, which exhibit cytotoxic activity, modulatory action on the complement cascade, anti-inflammatory activity, inhibitory action of serine protease, presynaptic neuromuscular blocking activity and anticonvulsant action (Cunha-Filho et al, 2010;Anjolette et al, 2015;Sousa-Filho et al, 2016;Shibao, Anjolette, Lopes, & Arantes, 2015;Freitas et al, 2017;Rostelato-Ferreira et al, 2018;Baldo et al, 2019;Zheng et al, 2020).…”

Toad venom bufadienolides and bufotoxins: An updated review

“…Among the toad venom active compounds, low‐molecular fractions mainly consisting of bufadienolides are expected to be responsible for Na + /K + ‐ATPase activity. Studying R. schneideri venom enabled such a conclusion (Baldo et al, 2019). A R. marina venom faction containing telocinobufagin (2.48%), marinobufagenin (65.50%), bufalin (8.88%), and resibufogenin (10.27%) demonstrated lower effect and thus higher IC 50 values in HBL (0.09 µg/mL), MM074 (0.1 µg/mL), and MM161 (0.35 µg/mL) melanoma cells compared with their resistant counterparts: HBL‐R (0.07 µg/mL), MM074‐R (0.05 µg/mL), and MM161‐R (0.06 µg/mL).…”

The Venom Revolution in Biomedicine: Unlocking Nature’s Toxin Toolkit for Therapeutic Innovation

“…Batrachotoxin produced by Phyllobates frogs targets voltage-gated sodium channels, causing sustained activation that leads to muscle contraction, convulsions, and respiratory paralysis [70] . Similarly, toad venom from Rhinella schneideri contains cardiotonic heterosides that inhibit Na⁺/K⁺-ATPase, resulting in cardiotoxic and neurotoxic effects [71] . Poison dart frogs exemplify toxin-based defence, secreting potent skin alkaloids that rapidly penetrate oral and mucosal tissues of predators, causing swelling, nausea, paralysis, or death [72] .…”

Action of heparin and acetylcholine modulators on the neurotoxicity of the toad Rhinella schneideri (Anura: Bufonidae) in Brazil

“…In recent years, several biological properties have been characterized from Rhinella schneideri 'Schneider's toad' poison (Rostelato-Ferreira et al, 2011;Rostelato-Ferreira et al, 2014;Abdelfatah, Lu, Schmeda-Hirschmann, & Efferth, 2019;Leal et al, 2020), a species widely distributed in Brazil (Wake, 2020), including the chemical identification and structural characterization of bufadienolides, e.g., marinobufagenin, bufalin, telocinobufagin, hellebrigenin, 20S21R-epoxymarinobufagin and β-sitosterol, which exhibit cytotoxic activity, modulatory action on the complement cascade, anti-inflammatory activity, inhibitory action of serine protease, presynaptic neuromuscular blocking activity and anticonvulsant action (Cunha-Filho et al, 2010;Anjolette et al, 2015;Sousa-Filho et al, 2016;Shibao, Anjolette, Lopes, & Arantes, 2015;Freitas et al, 2017;Rostelato-Ferreira et al, 2018;Baldo et al, 2019;Zheng et al, 2020).…”