Influence of temperature upon paralyzing and myotoxic effects of bothropstoxin-I on mouse neuromuscular preparations

Bothropstoxin-I (BthTX-I), from B. jararacussu venom, is a phospholipase A(2) (PLA(2)) homologue devoid of enzymatic activity. Besides inducing severe myonecrosis, BthTX-I promotes paralysis of both directly and indirectly evoked contractions in isolated neuromuscular preparations. We applied an experimental paradigm in order to characterize the steps involved in the toxic effects of BthTX-I on mouse neuromuscular junction. Myotoxicity was assessed by microscopic analysis of extensor digitorum longus muscles; paralyzing activity was evaluated through the recording of isolated contractions indirectly evoked in phrenic-diaphragm preparations. After 90 min at 35 degreesC, BthTX-I induced complete and irreversible paralysis, and damaged 30.3 +/- 2.7% of muscle fibers. In contrast, no effect was observed when tissues were incubated with BthTX-I at 10degreesC for 60 min and subsequently washed with toxin-free solution and maintained at 35 degreesC. These results indicate that the binding of BthTX-I to the cellular tissue surface is very weak at low temperature and that an additional factor is necessary. However, when tissues were submitted to BthTX-I (10degreesC for 60 min), and the temperature was elevated to 35 degreesC, omitting the washing step, it was observed muscle paralysis and damage in 39.04 +/- 4.2% of muscle fibers. These results indicate that a temperature-dependent step is necessary for BthTX-I to promote both its myotoxic and paralyzing activities. (C) 2004 Elsevier B.V.. All rights reserved.

Microscopic studies on the transition between the sigmoid sinus, the superior bulb of the jugular vein and the first portion of the internal jugular vein

The author studied the structure of the tissue components of the tunicae of the terminal segment of the sigmoid sinus, particularly at the level of the transition between the sigmoid sinus, the superior bulb of the jugular vein and the first portion of the human internal jugular vein; it was established that the transition between the sigmoid sinus and the first portion of the internal jugular vein occupies the whole extension of the superior bulb of the jugular vein up to the inferior third of the first portion of this vessel. These vascular walls exhibit a structure similar to that of the dura, i.e. the tunica adventitia is formed by fascicles of collagenic fibers which describe discontinuous spirals, more open proximal to the beginning of the first portion of the internal jugular vein. Approximately in the inferior third of the first portion of the internal jugular vein, there appear fascicles of smooth muscle fibers which are arranged similarly to those of the venous walls. The tunica intima of these vascular segments exhibits an endothelium resting on a network of elastic fibers which may play the role of an internal elastic lamina. From the bony border of the jugular foramen there originates a connective system whose fascicles of collagenic and elastic fibers incorporate to the wall of the internal jugular vein after describing a stretch in spiral around the vascular lumen.

Tensile strength and histological changes of abdominal aorta of malnourished rats

The resistance of the abdominal aorta of rats after 6, 7 and 8 weeks of malnutrition, compared with control animals, was evaluated by longitudinal tensiometry. Weakness of this vessel in malnourished rats was demonstrated; microscopic examination of the aorta stained by Masson, Calleja and hematoxylin-eosin methods showed a decrease in amorphous ground substance and an increase in the width of elastic laminae. There was no visible alteration either in the endothelial lining layer or in the smooth muscle fibers. Such alterations of the aorta are, to the authors' knowledge, the first reported modifications in the peripheral vasculature after malnutrition.

Features of myoneural junctions in the extraocular muscles of the opossum (Didelphis albiventris)

The myoneural junctions present in the opossum (Didelphis albiventris) extraocular muscles were studied through histochemical and ultrastructural techniques. Three different types of junction were shown: two single types, 'en grappe' and 'en plaque', present in the middle third of the muscle fibers; and one multiple type, more rare and observed in the distal third of the muscle.

Estudo dos efeitos da restrição alimentar nas características das fibras musculares de bovinos jovens confinados

The influence of restricting feed intake of young bulls in feedlots was evaluated in terms of structure of muscular fibers and respective areas, sampled by biopsy on the semitendinoso muscle. Sixty six crossbred Simental-Nelore bulls, 8 months old, averaging 220±34.03 kg were submited for 84 days in phase 1 (growing period), to three treatments: ad libitum (AL), restriction + whole soybean (RWS) and restriction + toasted whole soybean (RTS). The level of restriction of feed intake was 23%. Phase 2 was performed by splitting the animals in each treatment in phase 1 in two groups, feeding one with a diet containing soybeans and the other with poultry litter. The results showed that the animals AL presented more white fibers (FG), compared to the RWS and RST and a larger area of these fibers. A greater frequency of red fibers was observed in treatments RWS and RST. The conclusion was that the restriction of feed intake and consequent compensatory growth contributed for modulation of the muscular fibers increasing the frequency of the fast oxidative glycolitic (FOG) in 10.88% and decreasing of the slow oxidative (SO) and FG in 4.81 and 6.90%, respectively, with possible alteration on meat quality.

Morphology and histochemistry of the hyolingual apparatus in chameleons

We reexamined the morphological and functional properties of the hyoid, the tongue pad, and hyolingual musculature in chameleons. Dissections and histological sections indicated the presence of five distinctly individualized pairs of intrinsic tongue muscles. An analysis of the histochemical properties of the system revealed only two fiber types in the hyolingual muscles: fast glycolytic and fast oxidative glycolytic fibers. In accordance with this observation, motor-endplate staining showed that all endplates are of the en-plaque type. All muscles show relatively short fibers and large numbers of motor endplates, indicating a large potential for fine muscular control. The connective tissue sheet surrounding the entoglossal process contains elastin fibers at its periphery, allowing for elastic recoil of the hyolingual system after prey capture. The connective tissue sheets surrounding the m. accelerator and m. hyoglossus were examined under polarized light. The collagen fibers in the accelerator epimysium are configured in a crossed helical array that will facilitate limited muscle elongation. The microstructure of the tongue pad as revealed by SEM showed decreased adhesive properties, indicating a change in the prey prehension mechanics in chameleons compared to agamid or iguanid lizards. These findings provide the basis for further experimental analysis of the hyolingual system. © 2001 Wiley-Liss, Inc.

Mouse extensor digitorum longus and soleus show distinctive ultrastructural changes induced by veratrine

We investigated whether veratrine (5 μl, 10 ng/kg) injected into the mouse extensor digitorum longus (EDL) (fast-twitch) and soleus (SOL) (slow-twitch) muscles provokes distinctive ultrastructural disturbances 15, 30 and 60 min later. The mitochondria in SOL were affected earlier (within 15 min) than in EDL. Swelling of the sarcoplasmic reticulum terminal cisternae was more marked in EDL than in SOL and caused distortion of sarcomeres so that fragmentation of myofilaments was more pronounced in EDL. Hypercontracted sarcomeres were seen mainly in SOL and veratrine caused infoldings of the sarcolemma only in this muscle. In both muscles, the T-tubules remained unaffected and by 60 min after veratrine most of the above alterations had reverted to normal. Pretreatment with tetrodotoxin prevented the alterations induced by veratrine. This suggests that most of the alterations resulted from the enhanced influx of Na+ into muscle fibers. These results emphasize the importance of considering the type of muscle when studying the action of myotoxic agents.

Histoenzymology and morphometry of the masticatory muscles of tufted capuchin monkey (Cebus apella Linnaeus, 1758)

Samples of the anterior and posterior regions of the masseter and temporal muscles and of the anterior belly of the digastric muscle of 4 adult male tufted capuchin monkeys (Cebus apella) were removed and stained with HE and submitted to the m-ATPase reaction (with alkaline and acid preincubation) and to the NADH-TR and SDH reactions. The results of the histoenzymologic reactions were similar, except for acid reversal which did not occur in fibers of the fast glycolytic (FG) type in the mandibular locomotor muscles. FG fibers had a larger area and were more frequent in all regions studied. No significant differences in frequency or area of each fiber type were detected, considering the anterior and posterior regions of the masseter and temporal muscles. The frequency of fibers of the fast oxidative glycolytic (FOG) and slow oxidative (SO) types and of FOG area differed significantly between the anterior belly of the digastric muscle and the mandibular locomotor muscle. The predominance of fast twitch (FG and FOG) fibers and the multipenniform and bipenniform internal architecture of the masseter and temporal muscles, respectively, are characteristics that permit the powerful bite typical of tufted capuchin monkeys.

Study of the venom glands in Ectatomma quadridens (Hymenoptera: Formicidae). Evolutionary hypothesis in subfamily ponerinae

The venom glands of worker ants of the species Ectatomma quadridens morphologically resemble an elongated sac or reservoir ending in a narrower portion that has the function of releasing the secretion to the exterior. Two external secretory filaments are individually inserted into the proximal portion of the gland and end inside the convoluted gland. The venom gland of workers of E. quadridens is, therefore, morphologically subdivided into four distinct portions: a) sac-shaped reservoir measuring approximately 1mm in length; b) excretory duct, proximal portion of the reservoir that joins the gland to the sting apparatus; c) convoluted gland, final portion of the external secretory filaments located inside the reservoir; and d) two secretory filaments measuring about 2 mm in length; their free extremities end blindly and are individually inserted into the reservoir wall at the proximal region of the venom gland. The histological data showed that the filaments and the convoluted gland are composed of cubic cells of secretory function. The reservoir consists of a simple cubical epithelium externally surrounded by muscle fibers. A thick cuticle internally coats the epithelium of the reservoir. The application of histochemical tests allowed us to establish that the final secretion of the venom gland of Ectatomma quadridens is of glycoproteic nature. This secretion undergoes several modifications at the secretory filaments, at the convoluted gland, and in the reservoir before reaching the excretory duct, the point at which the secretion is released in its final composition, namely the venom. Based on the differences among various Ponerinae species we propose a hypothesis suggesting a probable evolutionary process that the venom glands of members of this subfamily might have undergone.

Morphological and histochemical analysis of the human vestibular fold

A morphological and histochemical study of the human vestibular fold was carried out using routine histological techniques. Seven μm-thick histological sections stained with hematoxylin-eosin (HE) and Calleja showed the presence of elastic collagen fibers and seromucous glands in the vestibular fold. Muscle fibers forming the ventricular muscle were also identified. Ultrastructural analyses of the epithelial layer by scanning electron microscopy (SEM) revealed ciliated cells and gland ducts opening on the epithelial surface. Histochemical analyses were performed on ventricular muscles submitted to nicotinamide-adenine-dinucleotide tetrazolium reductase (NADH-TR), succinate dehydrogenase (SDH), and myofibrillar adenosine triphosphatase (mATPase) reactions. Based on these reactions, it was observed that the muscle is formed by three types of muscle fibers: slow-twitch oxidative (SO), fast-twitch oxydative glycolytic (FOG) and fast-twitch glycolytic (FG) fibers distributed in a mosaic pattern. The fiber frequency was 22.7%, 69.9% and 7.4%, respectively. The higher frequency of SO and FOG fibers characterized the muscle as having aerobic metabolism and resistance to fatigue. The ventricular muscle was considered fast. The study of the neuromuscular junctions performed after nonspecific esterase reaction showed that they are of the en-plaque type and have multiple occurrences in the ventricular muscle.

A comparative study of myocardial function and morphology during fasting/refeeding and food restriction in rats

Background: This study compared the influence of fasting/refeeding cycles and food restriction on rat myocardial performance and morphology. Methods: Sixty-day-old male Wistar rats were submitted to food ad libitum (C), 50% food restriction (R50), and fasting/refeeding cycles (RF) for 12 weeks. Myocardial function was evaluated under baseline conditions and after progressive increase in calcium and isoproterenol. Myocardium ultrastructure was examined in the papillary muscle. Results: Fasting/refeeding cycles maintained rat body weight and left ventricle weight between control and food-restricted rats. Under baseline conditions, the time to peak tension (TPT) was more prolonged in R50 than in RF and C rats. Furthermore, the maximum tension decline rate (-dT/dt) increased less in R50 than in RF with calcium elevation. While the R50 group showed focal changes in many muscle fibers, such as the disorganization or loss of myofilaments, polymorphic mitochondria with disrupted cristae, and irregular appearance or infolding of the plasma membrane, the RF rats displayed few alterations such as loss or disorganization of myofibrils. Conclusion: Food restriction promotes myocardial dysfunction, not observed in RF rats, and higher morphological damage than with fasting/refeeding. The increase in TPT may be attributed possibly to the disorganization and loss of myofibrils; however, the mechanisms responsible for the alteration in -dT/dt in R50 needs to be further clarified. © 2010 Elsevier Inc. All rights reserved.

Evaluation of intracellular calcium in golden retriever muscular dystrophy

The present study had the objective of evaluating calcium accumulations in muscle fibers and their correlation with the canine muscular dystrophy. After the deaths of the animals (13 dystrophic and 3 non-dystrophic), samples of the skeletal muscles were collected. The material was stained with hematoxylin-eosin, Gomori's modified trichrome and alizarin red S technique (pH 4.3). The histopathological changes were analyzed and the proportions of calcium-positive (CPF) and negative muscle fibers were evaluated. Histopathological changes such as muscle fiber diameter changes, necrosis, hyalinization, presence of inflammatory infiltrate and fatty atrophy were identified in all the dystrophic muscles. Statistically significant differences in numbers of CPF between dystrophic muscles and non-dystrophics were observed for the masseter (6%), brachial biceps (5%) and triceps, sartorius and femoral biceps (4%) muscles. The identifying calcium is of interest as a parameter for helping in diagnostic screening.

Alterações eletromiográficas e histopatológicas da musculatura de cães naturalmente infectados por Leishmania infantum

The aim of this study was to evaluate the electromyographic and histopathological changes in skeletal muscles of dogs naturally infected by L. infantum. Twenty five mixed breed adult dogs with parasitological, molecular and serological diagnosis were selected. The evaluated muscles were: triceps brachial, extensor carpi radialis, biceps femoris and gastrocnemius. One dog had locomotor clinical signs with hind limbs paresis associated with severe muscle atrophy. Twenty-three (92%) had some type of muscular change, and in 22 (88%) such changes were directly identified by electromyography. Even without any clinical signs of the disease, 10 (40%) dogs had electromyographic and histopathological changes. Leishmania antigens were detected in muscles of four (16%) dogs. The electromyographic evaluation indicated the occurrence of chronic polymyositis in 13 (52%) dogs, the presence of both acute and chronic muscle inflammation four (16%), acute myopathy in two (8%) and absence of electromyographic abnormalities in three (12%) dogs. The most frequently observed histopathological changes were degeneration and necrosis of myofibers and inflammatory infiltration observed in 12 (48%) dogs. Other changes were decreased diameter of muscle fibers in 15 (60%) and peri or endomysial fibrosis in 14 (56%) animals. The changes observed in the present study showed that even in the absence of clinical signs, most dogs infected by Leishmania infantum have chronic polymyositis.

Bothropstoxin-I reduces evoked acetylcholine release from rat motor nerve terminals: Radiochemical and real-time video-microscopy studies

Understanding the biological activity profile of the snake venom components is fundamental for improving the treatment of snakebite envenomings and may also contribute for the development of new potential therapeutic agents. In this work, we tested the effects of BthTX-I, a Lys49 PLA2 homologue from the Bothrops jararacussu snake venom. While this toxin induces conspicuous myonecrosis by a catalytically independent mechanism, a series of in vitro studies support the hypothesis that BthTX-I might also exert a neuromuscular blocking activity due to its ability to alter the integrity of muscle cell membranes. To gain insight into the mechanisms of this inhibitory neuromuscular effect, for the first time, the influence of BthTX-I on nerve-evoked ACh release was directly quantified by radiochemical and real-time video-microscopy methods. Our results show that the neuromuscular blockade produced by in vitro exposure to BthTX-I (1 μM) results from the summation of both pre- and postsynaptic effects. Modifications affecting the presynaptic apparatus were revealed by the significant reduction of nerve-evoked [3H]-ACh release; real-time measurements of transmitter exocytosis using the FM4-64 fluorescent dye fully supported radiochemical data. The postsynaptic effect of BthTX-I was characterized by typical histological alterations in the architecture of skeletal muscle fibers, increase in the outflow of the intracellular lactate dehydrogenase enzyme and progressive depolarization of the muscle resting membrane potential. In conclusion, these findings suggest that the neuromuscular blockade produced by BthTX-I results from transient depolarization of skeletal muscle fibers, consequent to its general membrane-destabilizing effect, and subsequent decrease of evoked ACh release from motor nerve terminals. © 2012 Elsevier Ltd.

Undesirable effect of cosmetic lip augmentation with autologous fat tissue

Background: Facial cosmetic procedures are commonplace nowadays, especially techniques that aim to increase lip volume. Full lips provide a youthful, healthy, feminine and sensual appearance. There are many techniques and materials used to recover the loss of contour that occurs with age. Case presentation. An unusual case of fat tissue accumulation following cosmetic upper lip augmentation in a 61 year-old female was reported. Surgical treatment was performed for esthetic concerns. Microscopically, the tissue removed was composed of muscle fibers and mature adipocytes. Conclusion: Undesirable effects of esthetic treatment can occur and the clinician should be familiar with such complications to diagnose and manage them. © 2013 Kaminagakura et al.; licensee BioMed Central Ltd.