Gyroxin increases blood-brain barrier permeability to Evans blue dye in mice

Abstract

Gyroxin is a serine protease enzyme component of the South American rattlesnake (Crotalus durissus terrificus) venom. This toxin displays several activities, including the induction of blood coagulation (fibrinogenolytic activity), vasodilation and neurotoxicity, resulting in an effect called barrel rotation. The mechanisms involved in this neurotoxic activity are not well known. Because gyroxin is a member of a potentially therapeutic family of enzymes, including thrombin, ancrod, batroxobin, trypsin and kallicrein, the identification of the mechanism of gyroxin's action is extremely important. In this study, gyroxin was isolated from crude venom by affinity and molecular exclusion chromatography. Analysis of the isolated gyroxin via sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed a single protein band with a molecular weight of approximately 28 kDa, confirming the identity of the molecule. Furthermore, intravenous administration of purified gyroxin (0.25 μg/g of body weight) to mice resulted in symptoms compatible with barrel rotation syndrome, confirming the neurotoxic activity of the toxin. Mice treated with gyroxin showed an increase in the concentration of albumin-Evans blue in brain extracts, indicating an increase in the blood-brain barrier (BBB) permeability. This gyroxin-induced increase in BBB permeability was time-dependent, reaching a peak within 15 min after exposure, similar to the time span in which the neurotoxic syndrome (barrel rotation) occurs. This work provides the first evidence of gyroxin's capacity to temporarily alter the permeability of the BBB.