Scorpion venom increases acetylcholine release by prolonging the duration of somatic nerve action potentials

Scorpionism is frequently accompanied by a massive release of catecholamines and acetylcholine from peripheral nerves caused by neurotoxic peptides present in these venoms, which have high specificity and affinity for ion channels. Tityus bahiensis is the second most medically important scorpion species in Brazil but, despite this, its venom remains scarcely studied, especially with regard to its pharmacology on the peripheral (somatic and autonomic) nervous system. Here, we evaluated the activity of T. bahiensis venom on somatic neurotransmission using myographic (chick and mouse neuromuscular preparations), electrophysiological (MEPP, EPP, resting membrane potentials, perineural waveforms, compound action potentials) and calcium imaging (on DRG neurons and muscle fibres) techniques. Our results show that the major toxic effects of T. bahiensis venom on neuromuscular function are presynaptically driven by the increase in evoked and spontaneous neurotransmitter release. Low venom concentrations prolong the axonal action potential, leading to a longer depolarization of the nerve terminals that enhances neurotransmitter release and facilitates nerve-evoked muscle contraction. The venom also stimulates the spontaneous release of neurotransmitters, probably through partial neuronal depolarization that allows calcium influx. Higher venom concentrations block the generation of action potentials and resulting muscle twitches. These effects of the venom were reversed by low concentrations of TTX, indicating voltage-gated sodium channels as the primary target of the venom toxins. These results suggest that the major neuromuscular toxicity of T. bahiensis venom is probably mediated mainly by α- and β-toxins interacting with presynaptic TTX-sensitive ion channels on both axons and nerve terminals

Neurotoxicity of Tityus bahiensis (brown scorpion) venom in sympathetic vas deferens preparations and neuronal cells

Systemic scorpion envenomation is characterized by massive neurotransmitter release from peripheral nerves mediated primarily by scorpion venoms neurotoxins. Tityus bahiensis is one of the medically most important species in Brazil, but its venom pharmacology, especially regarding to peripheral nervous system, is poorly understood. Here, we evaluated the T. bahiensis venom activity on autonomic (sympathetic) neurotransmission by using a variety of approaches, including vas deferens twitch-tension recordings, electrophysiological measurements (resting membrane potentials, spontaneous excitatory junctional potentials and whole-cell patch-clamp), calcium imaging and histomorphological analysis. Low concentrations of venom (≤ 3 μg/mL) facilitated the electrically stimulated vas deferens contractions without affecting postsynaptic receptors or damaging the smooth muscle cells; transient TTX-sensitive sustained contractions and resting membrane depolarization were mediated mainly by massive spontaneous ATP release. High venom concentrations (≥ 10 μg/mL) blocked the muscle contractions and induced membrane depolarization. In neuronal cells (ND7-23wt), the venom increased the peak sodium current, modified the current-voltage relationship by left-shifting the Nav channel activation curve, thereby facilitating the opening of these channels. The venom also caused a time-dependent increase in neuronal calcium influx. These results indicate that the sympathetic hyperstimulation observed in systemic envenomation is presynaptically driven, probably through the interaction of α- and β-toxins with neuronal sodium channels

Tityus bahiensis scorpion venom activities on somatic and autonomic preparations

Orientadores: Edson Antunes, Edward Gerard RowanTese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciências MédicasResumo: O escorpionismo é caracterizado principalmente por manifestações locais mas que podem evoluir à sistêmicas, sendo acompanhadas por uma massiva liberação de catecolaminas e acetilcolina por nervos periféricos. Essa atividade é causada por peptídeos neurotóxicos presentes nestes venenos, que tem alta especificidade e afinidade por canais iônicos. Tityus bahiensis (T. bahiensis) é uma das espécies de importância médica no Brasil sendo responsável pela maioria dos acidentes escorpiônicos no estado de São Paulo (segundo maior causa de escopionismo no país), entretanto, apesar de sua relevância epidemiológica, seu veneno permanece pobremente estudado, especialmente em relação a sua farmacologia no sistema nervoso periférico. Este trabalho estudou a atividade do veneno de T. bahiensis na neurotransmissão somática motora e simpática por meio de abordagens miográficas (preparações neuromusculares de camundongo e ave, e canal deferente isolado de ratos), eletrofisiológicas (MEPP, EPP, SEJP, potenciais de membrana em repouso, formas de ondas perineurais, potenciais de ação compostos e whole-cell patch-clamp), e de imagem de cálcio (neurônios do DRG e neuroblastoma, e fibras musculares). Nossos resultados mostram que os maiores efeitos tóxicos promovidos pelo veneno de T. bahiensis na função neuromuscular e neuroefetora são de origem pré-sináptica. Baixas concentrações de veneno prolongam o potencial de ação axonal levando a uma despolarização prolongada do terminal nervoso que consequentemente promove a liberação de neurotransmissores e a facilitação da contração muscular. O veneno também estimulou a liberação espontânea de neurotransmissores provavelmente através da despolarização parcial do terminal nervoso. Altas concentrações de veneno bloqueiam a geração de potencial de ação e a contração muscular neurogênica. A farmacologia do veneno pode ser revertida por baixas concentrações de TTX, indicando que os canais de sódio dependentes de voltagem neuronais são um dos principais alvos dessas toxinas. O resultado deste trabalho sugere que a maioria da neurotoxicidade promovida pelo veneno de T. bahiensis são causados provavelmente por toxinas 'alfa' e 'beta' interagindo com canais iônicos pré-sinápticos sensíveis a TTX em ambos axônios e terminais nervososAbstract: Scorpionism is frequently accompanied by a massive release of catecholamines and acetylcholine from peripheral nerves caused by neurotoxic peptides present in theses venoms, which have high specificity and affinity for ion channels. Tityus bahiensis (T. bahiensis) is the second most medically important scorpion specie in Brazil (responsible for the major scorpion accidents in Sao Paulo State) but, despite its epidemiological relevance, its venom remains scarcely studied, especially with regarding to its pharmacology on peripheral nervous system. Here, we evaluated the activity of T. bahiensis venom on motor somatic and autonomic neurotransmission using myographic (chick and mouse neuromuscular preparations, and isolated rat vas deferens), electrophysiological (MEPP, EPP, SEJP, resting membrane potentials, perineural waveforms, compound action potentials) and calcium imaging (on DRG neurons and muscle fibres) techniques. Our results show that the major toxic effects of T. bahiensis venom on neuromuscular function are presynaptically driven. Low concentrations of venom prolong the axonal action potential leading to a longer depolarization of the nerve terminals that enhances neurotransmitter release and facilitates nerve-evoked muscle contraction. The venom also stimulates the spontaneous release of neurotransmitters, probably through partial neuronal depolarization. Higher concentration of venom blocks the generation of the action potential and resulting muscle twitches. The venom pharmacology was reversed by low concentrations of TTX, indicating voltage-gated sodium channels as one of the primary target of the venom toxins. These results suggest that major neurotoxicity promoted by T. bahiensis venom are probably caused mainly by 'alpha'- and 'beta'-toxins interacting with presynaptic TTX-sensitive ion channels on both axons and nerve terminalsDoutoradoFarmacologiaDoutora em Farmacologia2016/11319-6142460/2014-1FAPESPCNP

Bothrops fonsecai snake venom biological effects and the use of commercial bothropic antivenom as pharmacological tool

Orientador: Léa Rodrigues SimioniDissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências MédicasResumo: Venenos de serpentes botrópicas são caracterizados pela indução de extenso dano local associado a feitos sistêmicos. Bothrops fonsecai é uma serpente de distribuição geográfica restrita das regiões montanhosas de Mata Atlântica cujo veneno é pobremente estudado. Recente estudo proteômico identificou que esse veneno possui principalmente metaloproteases, fosfolipases A2, Lectinas tipo C e serinoproteases. Ensaios experimentais utilizando antiveneno botrópico comercial para neutralizar as atividades biológicas de venenos botrópicos possibilitam não somente respostas de cunho clínico (tratamento de acidentes), mas também se constitui em uma valiosa ferramenta farmacológica para avaliar a existência de diferenças na composição dos diversos venenos através da reatividade cruzada entre veneno e antiveneno. Levando em consideração esses fatos, este trabalho teve como objetivo estudar a toxicidade do veneno de B. fonsecai através da atividade bloqueadora sobre a junção neuromuscular, atividade miotóxica, hemorrágica, edematogênica e coagulante assim como a sua caracterização bioquímica. A neutralização dessas atividades por antiveneno botrópico comercial e a comparação das ligações antígeno:anticorpo (veneno:antiveneno) entre B. fonsecai e os antivenenos botrópico comercial e específico usando SE-HPLC e immunoblotting também foram avaliadas. O veneno de B. fonsecai apresentou atividade fosfolipásica, sendo essa provavelmente a maior responsável pelo bloqueio neuromuscular e miotoxicidade, além de ser uma das mediadoras da atividade edematogênica; a inibição parcial dessa atividade pelo antiveneno comercial corrobora com a neutralização parcial dos efeitos na junção neuromuscular e redução da atividade edematogênica, já a neutralização total da atividade miotóxica demonstra que as PLA2 catalíticas podem não ser as principais responsáveis pela miotoxicidade. Embora o veneno apresente metaloproteinases P-I e P-III, a atividade proteolítica não demonstrou-se marcante in vitro, diferente do ensaio de atividade hemorrágica (induzida principalmente por essas enzimas e que foi completamente inibida pelo antiveneno comercial) o que leva a crer que o ensaio enzimático esteja subestimando essa atividade. O veneno exibiu atividades esterásica e coagulante que são normalmente causadas por serinoproteases e ambas foram inibidas pelo antiveneno botrópico comercial, em menor grau na proporção 5:1 (recomendada pelo fabricante) que a obtida em 5:2. O antiveneno botrópico comercial apresentou reatividade-cruzada em todas as atividades testadas, porém sua afinidade com o veneno de B. fonsecai demonstrou-se baixa ressaltando que o veneno de B. fonsecai possui composição diferente do veneno das serpentes utilizadas na produção do antiveneno comercial, em especial as fosfolipases catalíticas, metaloproteinases P-I e serinoproteasesAbstract: Bothropic snake venoms are characterized for a marked local damage associated to systemic effects. Bothrops fonsecai is a Brazilian pit viper of restrict distribution (Atlantic Forest at high altitude regions) and its venom has been poorly studied but is known it has phospholipases A2, metalloproteases (P-I and P-III) and serine proteases. Experimental assays using commercial bothropic antivenom to neutralize the bothropic venom activities allows clinical responses (snakebites treatment) and also is a pharmacological tool to evaluate different venoms composition through venom:antivenom cross-reactivity. Thinking in all this, the aim of this work were the B. fonsecai venom toxicity analysis through the in vitro neuromuscular junction action, the myotoxic, hemorrhagic, edema-forming and coagulant activities, as well its biochemical characterization. The neutralization of those activities by the commercial bothropic antivenom and the comparison of venom:antivenom bindings (Bothrops fonsecai venom : Commercial and specific antivenoms) by SE-HPLC and immunoblotting were also evaluated. B. fonsecai venom had a high phospholipasic activity and this probably is the major responsible for neuromuscular blockade and miotoxicity caused by this venom. This activity is also one of edema-forming activity mediator (with metalloproteases P-I). The partial inhibition of this activity with both proportions (5:1 and 5:2) corroborates the partial neuromuscular blockade and edema neutralization; in contrast total the inhibition of miotoxicity allows us to suppose that the catalytic PLA2 are not the major responsible for the miotoxicity. The venom also showed a low metalloprotease activity (caused principally by P-I once is more abundant on this venom) and was not significantly inhibited by commercial bothropic antivenom at both proportions; these enzymes (especially P-III) also induced hemorrhage totally inhibited by commercial bothropic antivenom. The esterase and coagulant activities of this venom are caused mainly by serine-proteases; the commercial antivenom neutralization was more potent at 5:2 than 5:1 proportion. The commercial antivenom presented cross-reactivity in all tested activities but its low affinity with B. fonsecai venom indicates that the venom composition is a quite different than the composition of snake venoms used to produce the commercial antivenom, specially catalytic phospholipases, metalloproteases P-I and serine proteasesMestradoFarmacologiaMestra em Farmacologi

PROPRIEDADE ANTIOFÍDICA DO EXTRATO METANÓLICO DE MIKANIA LAEVIGATA SOBRE AS AÇÕES BIOLÓGICAS INDUZIDAS PELO VENENO DE PHILODRYAS ALFERSII NA JUNÇÃO NEUROMUSCULAR

Embora negligenciadas pela maioria dos médicos, os acidentes com serpentes opistóglifas podem ser graves e até fatais. Envenenamentos por Philodryas olfersii demonstram efeitos local e sistêmico semelhantes aos envenenamentos por serpentes botrópicas. Para complementar a soroterapia, muitas plantas da medicina popular estão sendo estudadas por sua grande quantidade de compostos químicos e atividades farmacológicas, entre elas, Mikania laevigata, que demonstra ter os seguintes efeitos: antiinflamatório, antiedematogênico, antiulcerogênico, hipoglicêmico, antidiarréico, antimutagênico, antiasmático, broncodilatador, etc. Neste estudo, ensaios miográficos demonstraram que o extrato metanólico de M. laevigata (200 μL) inibiu 100% o bloqueio neuromuscular induzido pelo veneno bruto de P. olfersii (50 μg/mL). Conclui-se que o extrato metanólico da M. laevigata possui atividade antiofídica sob o parâmetro da junção neuromuscular

Hospitalization and mortality rates of malignant prostatic neoplasms in Brazil: A cross-sectional study from 2008 to 2018

Abstract Prostate cancer (PCa) is a highly prevalent condition among men worldwide, resulting in reduced quality of life and increased costs to health systems due to hospitalization and death. This study aimed to explore and understand the evolution of PCa in Brazil from 2008 to 2018. Data were obtained from the National Health System Department of Informatics (DATASUS) using code C61 for malignant prostatic neoplasms. We presented the hospitalization and mortality rates in a temporal-, regional- and age-dependent manner. From 2008 to 2018, a year-dependent increase in hospital admissions due to PCa was reported in Brazil, in which the Southeast region showed the highest prevalence. Men aged ≥80 and those 70-79 years old had similar hospitalization rates, followed by men aged 60-69, 50-59, 40-49 and 30-39 years old. Similarly, an increase in deaths due to PCa was reported during this period, with the highest rates seen in the Southeast. Men aged ≥80 years had higher mortality rates, followed by those aged 70-79, 60-69, 50-59, 40-49 and 30-39 years old. The results obtained indicate an age- and region-dependent increase in PCa morbidity and mortality in Brazil overtime and may contribute to the ongoing discussion on the role and future perspective of the health care system in Brazil