Erasmo G. Mendes

A tetrodotoxin-like substance found in the Brazilian frog Brachycephalus ephippium

A tetrodotoxin-like substance, denoted ephippiotoxin, was obtained from the tissue of Brachycephalus ephippium, a small pumpkin-coloured frog collected in the Atlantic Forest of the southeast region of Brazil. Ephippiotoxin is a dialyzable substance soluble in water, methanol and ethanol, but insoluble in organic solvents such as chloroform and other apolar solvents. After treatment with active charcoal (Norit-A) and purification with ion-exchange Amberlite IRC-50 resin (NH4 + form), a freeze-dried residue was obtained, with a toxicity of c. 117 micrograms/kg (mice, i.p.). Ephippiotoxin showed the same mobility as crystalline tetrodotoxin (Sankyo) when submitted to thin-layer chromatography (silica gel G) using seven different solvent systems. White mice (20 +/- 1 g) injected i.p. with either B. ephippium tissue extracts or semi-purified toxin showed partial paralysis of the hind limbs, lethargy, altered breathing rhythm and clonic convulsions. Death occurred within 1.5-30 min after injection, depending on the dose. Ephippiotoxin induced atrioventricular diastolic blockade in the toad heart. It also inhibited the response of toad striated muscle to direct and indirect electric stimulation and blocked the compound action potential of isolated frog sciatic nerve.

Indirect cardiotoxic activity of the caecilian Siphonops paulensis (Gymnophiona, Amphibia) skin secretion

The skin secretion of the caecilian S. paulensis has an indirect cardiotoxic effect. It induces a rapid blockage of electrical activity and subsequent diastolic arrest on in situ toad heart preparations. This action is not blocked by atropine and is therefore not mediated through a cholinergic mechanism. In addition, S. paulensis skin secretion is ineffective in inducing any response in isolated perfused toad heart assay, in the spontaneously beating isolated toad atria or in the electrically driven toad ventricle strip. These results, and the observation that S. paulensis skin secretion exhibits an hemolytic activity ([Schwartz, E. F., Schwartz, C. A., Sebben, A., Mendes E. G. (1997) Cardiotoxic and hemolytic activities on the caecilian Siphonops paulensis skin secretion. J. Venom. Anim. Toxins 3(1), 190]), suggest that the skin secretion cardiotoxicity could be mediated through an endogenous agent. The cardiotoxic action of S. paulensis skin secretion was investigated in isolated toad cardiac tissues in the presence of toad red blood cells (TRBC). In both atria and ventricle it evoked the same responses observed in the in situ heart. The potassium concentrations of the suspending media, as determined by atomic absorption spectroscopy were 9.5 and 11.7 times greater, respectively, in the presence of TRBC than in its absence. Given that increased levels of potassium also showed a negative inotropic effect in the isolated ventricle strip, the action of the skin secretion was attributed to the hemolytic activity, which causesan increase in K+ concentration of the medium.

Pharmacological studies on the invertebrate non-striated muscles, I. The response to drugs

Abstract 1. The lantern muscles of the sea urchins Echinometra lucunter and Echinus esculentus are adequate preparations for the assay of cholinergic drugs. They are easily dissected, exhibit no spontaneous activity, and relax fast enough after treatment. 2. Threshold doses of acetylcholine for the protractor muscle of E. lucunter and both protractor and retractor muscles of E. esculentus are respectively 0·1, 0·5, and 5·0 μg. (in a 10-ml. bath). Dose-effect studies show that the responses to acetylcholine follow a straight line with an insignificant deviation from linearity. 3. The responses of the protractor muscle of E. lucunter to acetylcholine, propionylcholine, and butyrylcholine indicate a decline in activity when the acetyl group is departed from. 4. The protractors of E. lucunter respond also to acetyl-beta-methylcholine, carbamylcholine, and nicotine; they are insensitive to bethanechol, pilocarpine, adrenaline, tyramine, tryptamine, serotonine, G.A.B.A., I-glutamate, and histamine. 5. The responses to acetylcholine can be blocked (at least, partially) by atropine, methantheline, propantheline, D-tubocurarine, Flaxedil (gallamine triethiodide), mytolon, and bebeerine, but not by succinylcholine. 6. The responses to drugs which are either more markedly muscarinic or nicotinic and the blockings obtained with both atropine and curares may suggest the presence in the muscles of either undifferentiated receptors for acetylcholine or the two kinds of receptors known in higher animals (muscarinic and nicotinic). 7. The action of acetylcholine is potentiated by eserine and prostigmine. 8. Muscle homogenates are active against both acetylcholine and acetyl-beta-methylcholine, suggesting the presence of true cholinesterase.

The respiratory responses of an amphibious snail Pomacea lineata (spix, 1827), to temperature and oxygen tension variations

Departamento de Fisiologia e Farmacologia Centro de Ciencias Biologicas Universidade Federal de Pernambuco, 50000 Recife, PEI

Effect of body size and temperature on oxygen uptake in the water snakes Helicops modestus and Liophis miliaris (colubridae)

Abstract 1. 1. Oxygen uptake during rest was measured in the water snakes Helicops modestus and Liophis miliaris and related to body size and temperatures between 10–35°C at 5°C intervals. 2. 2. The relationships between oxygen uptake and body size at 25°C is expressed by the relation v o 2 = 0.411 BW 0.560 for H. modestus and V o 2 = 0.339 BW0.647 f 3. 3. The significantly higher b value ( v o 2 = aBW b ) for L. miliaris may be related to the size attained by the fully grown specimens as well as to the larger lung surface area. 4. 4. There was no significant difference between slopes of males and females despite sexual dimorphism in size in both species. 5. 5. When the exponent b was estimated at various temperatures and compared with the value at 25°C the differences were not significant for any of the species. 6. 6. H. modestus and L. miliaris showed a metabolic plateau between 25 and 30°C, and within this temperature range there were significant differences in the b values between species.

The influence of size, temperature and oxygen tension upon the respiratory metabolism of the terrestrial amphipod Talitrus (Talitroides) pacificus Hurley, 1955

1. 1. The oxygen consumption of Talitrus pacificus, measured at 20°C and in normoxic conditions, varied with body weight according to the equation O2 = 1.245 W0.462 for N = 45(1.4–34.1 mg) and r = 0.725 (P < 0.05). 2. 2. The weight (mg) specific oxygen consumption (“respiratory rate”) of T. pacificus ranged from 1.171 (1.4 mg animal) to 0.148 (34.1 mg animal) in μ 102mg/hr. 3. 3. The respiratory rates of T. pacificus acclimated to 20°C increased with rising temperatures, particularly in the range 10–15°C (Q10 = 3.65); a Q10 = 1.09 indicated a zone of respiratory homeostasis between 25 and 30°C. 4. 4. In declining oxygen tensions, T. pacificus regulated the rate of oxygen consumption down to 105 or even 70 mm Hg of O2. 5. 5. No animals out of groups of 10 survived after 40–45 min in an anoxic condition (100% nitrogen).

Oxygen consumption related to size and sex in the tropical millipede Pseudonannolene tricolor (Diplopoda, Spirostreptida)

Abstract 1. 1. The oxygen consumption of the tropical millipede, Pseudonannolene tricolor (Spirostreptida, Pseudonannolenidae) was studied in both male and female animals (body mass varying from 0.242 to 2.802 g) using a Warburg microrespirometer at 25°C. 2. 2. The allometric equation M = a W b was used in order to check the metabolic increases with increasing body mass. The b exponents were, respectively, 0.68 for males and 0.60 for females. 3. 3. Results are discussed in terms of the meaning of the b values in Diplopoda and animals in general. 4. 4. A relationship between volume and body mass in P. tricolor is also reported.

The respiratory metabolism of the lantern muscles of the sea urchin Echinometra lucunter L.-I. The respiratory intensity

Abstract 1. 1. The respiratory rates of the lantern muscles of the sea urchin Echinometra lucunter L. were measured at 25°C in different combinations of muscle integrity, oxygen tension and suspending medium. 2. 2. The results showed that, probably due to their low metabolism, it is irrelevant to respiration whether the isolated muscles, whole or scraped, in natural or artificial seawater, are exposed to air or to oxygen. 3. 3. The respiratory intensity of the lantern muscles is higher than that of the holothurian longitudinal muscles and this is explained by their being engaged in a more continuous activity.

Pharmacological studies on the invertebrate non-striated muscles. II. The tunicate siphon muscles

Abstract 1. 1. The siphon muscles of the tunicate, Ascidia nigra, respond only to acetylcholine, some other choline esters, and acetylcholine-like substances, such as nicotine and pilocarpine. Dose-effect studies show that the responses to acetylcholine follow a straight line with an insignificant deviation from linearity. 2. 2. On a molar basis the responses to acetylcholine and nicotine are about the same; there is a decline in activity on departing from the acyl group; the threshold is higher to mecholyl and pilocarpine and no response is obtained with benzoylcholine. 3. 3. The action of acetylcholine is potentiated by eserine. 4. 4. The responses to acetylcholine are depressed by previous treatment with atropine, methantheline, propantheline, and mytolon. 5. 5. Succinylcholine either enhances the response of the siphon to acetylcholine or is active per se on the preparation; the tendency with flaxedil is also to potentiate the action of acetylcholine rather than to block it; d-tubocurarine can act as an ACh blocker. 6. 6. On quiet preparations bursts of activity can be produced by the application of succinylcholine, flaxedil, d-tubocurarine, and mytolon, but not with atropine. 7. 7. On spontaneously active preparations acetylcholine greatly increases the tonus and atropine markedly depresses it. 8. 8. It is concluded that the siphon muscles are endowed with a myogenic activity, are acetylcholineceptive, and that their acetylcholine receptors are differentiated in a muscarinic rather than in a nicotinic direction.

Behavioral modifications of Liriope tetraphylla (Chamisso and Eysenhardt) (Cnidaria, Hydrozoa, Trachymedusae) induced by hyposmotic conditions

The osmoregulatory capability and swimming behavior of Liriope tetraphylla (Chamisso and Eysenhardt) were analyzed under hyposmotic conditions. These hydromedusae are shown to be osmoconforming animals that are not able to maintain constant osmotic and ionic concentrations of their body fluids when exposed to anisosmotic media. Swimming behavior (as measured by the pulsation rate and the swimming index) is transiently affected by osmotic shock, being restored 30 min after exposure. However, even when swimming activity was restored, two different swimming patterns were observed depending on the hypotonic period. Thirty minutes exposure led to an increase of the quiescent period with no changes on the swimming burst characteristics while five hours exposure to hypotonicity shortened both the quiescent period and burst durations. Such behavioral modifications were related to the changes in body fluid ionic composition (mainly to extracellular K+ levels) that occur during the osmotic adjustment process. The fact that swimming activity was impaired during hypoionic shocks and this condition caused morphological changes in the animals suggests that these early metazoans are endowed with mechanisms to compensate for ionic gradient changes induced by hyposmotic, but not for the ones caused by hypoionic condition.