In both experiments, the animals were compared to control animals (n = 8), which received only saline by i.p. injection in the same volume. The survival rate of animals was observed 24 and 48 h after inoculation of venom or saline. At the end of the experiment, the surviving animals were euthanized with sodium pentobarbital (225 mg/kg). The Ts-MG and Ts-DF venoms were evaluated for their ability to induce behavioral and physiological changes in mice. All groups of mice that were part of the experiment for the determination of LD50 were observed during the first
three hours after venom or saline injection. Those effects observed in animals that received venom and were absent in control animals AZD2281 molecular weight (saline) were considered as behavioral
and physiological changes. A set of behavioral and physiological effects was previously defined (see Table 1). The ability of T. serrulatus venom from DF and MG to induce acute pulmonary edema in rats was evaluated as done by Matos et al., 1999 and Matos et al., 2001, with some modifications, as follows. Eighteen rats were divided into three experimental groups (n = 6): Ts-MG venom (0.5 mg/kg of T. serrulatus venom from MG), Ts-DF venom (0.5 mg/kg of T. serrulatus venom from DF) see more and control group (150 mM NaCl). Saline or venom was injected by i.v. route (200uL). The rats were firstly anesthetized with a mixture of xylazine hydrochloride (10 mg/kg) and ketamine (75 mg/kg) i.p. The animals were observed for a period of 1 h after the treatment. After that, animals were euthanized by an overdose of sodium pentobarbital (225 mg/kg) and their hearts and lung were rapidly removed; the lungs were weighted and both organs were prepared for histology. The lungs were immediately weighed and the presence of acute pulmonary edema (APE) was determined according to Magalhães et al.
(1998) using the formula: APE = Pulmonary Mass × 100/Body Mass. The presence of pulmonary edema activity was assessed by differences between the APE obtained from animals injected with venom and the APE obtained from the control group. Hearts and lungs were fixed in 10% buffered formalin and embedded in paraffin (Prophet et al., 1992). Histological sections (4 mm thick) were stained with haematoxylin eosin (HE) and analyzed under an optical microscope. After the morphological analyses, the tissues were classified according Mannose-binding protein-associated serine protease to Matos et al. (1997) with some adjustments, as described: 1) normal tissue: tissue without morphological changes when compared to the control group; 2) mild pulmonary edema: usually irregular, sub-pleural, with extravasations of plasma; 3) moderate pulmonary edema, multifocal, with large plasma leakage; 4) severe pulmonary edema: diffuse interstitial (intra-alveolar) in all lung lobes, sometimes associated with hemorrhagic foci. All animals used in the induction of pulmonary edema were also subjected to blood sampling right after being euthanized.