fMRI of the sensorimotor cortex in patients with traumatic brain injury after intensive rehabilitation.

For evaluating the patterns of brain activation in sensorimotor areas following motor rehabilitation, seven male patients diagnosed with TBI underwent an fMRI study before and after being subjected to motor rehabilitation. Six patients showed a reduction in the BOLD signal of their motor cortical areas during the second fMRI evaluation. A decrease in cerebellum activation was also observed in two patients. Newly activated areas, were observed in four patients after treatment. In addition, an increase in the activation of the supplementary motor area (SMA) following rehabilitation was observed in only one test subject. The findings show that motor rehabilitation in TBI patients produces a decrease in the BOLD signal for the sensorimotor areas that were activated prior to treatment. In addition, we observed the recruitment of different brain areas to compensate for functional loss due to TBI in line with the cortical reorganisation mechanism.

Bothrops jararacussu venom-induced neuromuscular blockade inhibited by Casearia gossypiosperma briquet hydroalcoholic extract

The hydroalcoholic extract of Casearia gossypiosperma Briquet (Flacourtiaceae) was standardized for the first time through quality control procedures including pharmacognostic methods, fingerprint chromatograms, defined amounts of marker substances and physicochemical characteristics. The pharmacological activity of C. gossypiosperma (Cg) hydroalcoholic extract was assayed by a traditional in vitro test, which involved irreversible neuromuscular blockade induced by Bothrops jararacussu (Bjssu) venom (60 µg/mL) in mouse phrenic nerve-diaphragm preparations. Bjssu venom blocked muscle activity for 26 (± 2.0) minutes (n = 6). Cg extract (0.1 mg/mL) induced changes on the baseline muscle activity without impairing the muscle function and inhibited 87.6 percent (± 1.8) (n = 6) of the Bjssu venom-induced blockade. Both flavonoids (0.624 g percent) and polyphenols (4.63 g percent) from the extract were spectrophotometrically quantified. Therefore, the present study confirms the antibothropic activity of Cg extract, supporting the ethnomedical use of Casearia sp. in the treatment of snakebite victims.

Low-level laser therapy decreases local effects induced by myotoxins isolated from bothrops jararacussu snake venom

The prominent myotoxic effects induced by Bothrops jararacussu crude venom are due, in part, to its polycationic myotoxins, BthTX-I and BthTX-II. Both myotoxins have a phospholipase A2 structure: BthTX-II is an active enzyme Asp-49 PLA2, while BthTX-I is a Lys-49 PLA2 devoid of enzymatic activity. In this study, the effect of low-level laser therapy (LLLT), 685 nm laser at a dose of 4.2 J/cm2 on edema formation, leukocyte influx and myonecrosis caused by BthTX-I and BthTX-II, isolated from Bothrops jararacussu snake venom, was analyzed. BthTX-I and BthTX-II caused a significant edema formation, a prominent leukocyte infiltrate composed predominantly by neutrophils and myonecrosis in envenomed gastrocnemius muscle. LLLT significantly reduced the edema formation, neutrophil accumulation and myonecrosis induced by both myotoxins 24 hours after the injection. LLLT reduced the myonecrosis caused by BthTX-I and BthTX-II, respectively, by 60 and 43 percent; the edema formation, by 41 and 60.7 percent; and the leukocyte influx, by 57.5 and 51.6 percent. In conclusion, LLLT significantly reduced the effect of these snake toxins on the inflammatory response and myonecrosis. These results suggest that LLLT should be considered a potential therapeutic approach for treatment of local effects of Bothrops species venom.

Bothrops jararacussu venom-induced neuromuscular blockade inhibited by Casearia gossypiosperma Briquet hydroalcoholic extract

The hydroalcoholic extract of Casearia gossypiosperma Briquet (Flacourtiaceae) was standardized for the first time through quality control procedures including pharmacognostic methods, fingerprint chromatograms, defined amounts of marker substances and physicochemical characteristics. The pharmacological activity of C. gossypiosperma (Cg) hydroalcoholic extract was assayed by a traditional in vitro test, which involved irreversible neuromuscular blockade induced by Bothrops jararacussu (Bjssu) venom (60 ìg/mL) in mouse phrenic nerve-diaphragm preparations. Bjssu venom blocked muscle activity for 26 (± 2.0) minutes (n = 6). Cg extract (0.1 mg/mL) induced changes on the baseline muscle activity without impairing the muscle function and inhibited 87.6% (± 1.8) (n = 6) of the Bjssu venom-induced blockade. Both flavonoids (0.624 g%) and polyphenols (4.63 g%) from the extract were spectrophotometrically quantified. Therefore, the present study confirms the antibothropic activity of Cg extract, supporting the ethnomedical use of Casearia sp. in the treatment of snakebite victims.

Acute hepatotoxicity of Crotalus durissus terrificus (South American rattlesnake) venom in rats

Venom of the South American rattlesnake, Crotalus durissus terrificus (Cdt), presents myotoxic and neurotoxic outcomes, but reports on its effects on the liver are scarce. This study examined the hepatotoxicity resulting from Cdt venom administration (100, 200 and 300 miug/kg) in male Wistar rats. Animais were studies at 3, 9 and 12 hours after venom injection. The hepatotoxicity was assessed through serum levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (AP), gamma glutamyl transferase (GGT), bilirrubin and also by histopathological evaluation. All the different concentrations of Cdt venom resulted in increased levels of hepatic enzymes, when compared with the control group, except for the 100 miug/kg dose, which presented normal levels at 9 and 12 hours after venom administration. Bilirrubin levels remained unchanged by Cdt venom. Histological analysis revealed endothelial damage, inflammatory cell infiltration, as well as sinusoidal and portal congestion. Based on these observations, we may conclude that Cdt venom causes dose- and time-dependent hepatic damage in rats, characterized by elevated hepatic enzyme levels and histological alterations

Anti-inflammatory activity of Blutaparon portulacoides ethanolic extract against the inflammatory reaction induced by Bothrops jararacussu venom and isolated myotoxins BthTX-I and II

This article reports the anti-inflammatory effect of Blutaparon portulacoides (B. portulacoides), specifically the ethanolic extract of its aerial parts, on the edema formation and leukocyte influx caused by Bothrops jararacussu (B. jararacussu) snake venom and Bothropstoxin-I and II (BthTX-I and II) isolated from this venom as an alternative treatment for Bothrops snakebites. The anti-inflammatory effect of B. portulacoides ethanolic extract was compared with an animal group pretreated with dexamethasone. B. portulacoides ethanolic extract significantly inhibited paw edema induced by B. jararacussu venom and by BthTX-I and II. Also, results demonstrated that the extract caused a reduction of the leukocyte influx induced by BthTX-I. However, the extract was not capable of inhibiting the leukocyte influx induced by the venom and by BthTX-II. In conclusion, these results suggest that the ethanolic extract of this plant possess components able to inhibit or inactivate toxins present in B. jararacussu venom, including its myotoxins, responsible for the edema formation. However, the leukocyte migration caused by the venom and BthTX-II was not inhibited by the plant, probably due to the different mechanisms involved in the edema formation and leukocyte influx. This is the first report of B. portulacoides extract as anti-inflammatory against snake venoms and isolated toxins.(AU)

Cytoskeleton, endoplasmic reticulum and nucleus alterations in CHO-K1 cell line after Crotalus durissus terrificus (South American rattlesnake) venom treatment

Snake venoms are toxic to a variety of cell types. However, the intracellular damages and the cell death fate induced by venom are unclear. In the present work, the action of the South American rattlesnake Crotalus durissus terrificus venom on CHO-K1 cell line was analyzed. The cells CHO-K1 were incubated with C. d. terrificus venom (10, 50 and 100g/ml) for 1 and 24 hours, and structural alterations of actin filaments, endoplasmic reticulum and nucleus were assessed using specific fluorescent probes and agarose gel electrophoresis for DNA fragmentation. Significant structural changes were observed in all analyzed structures. DNA fragmentation was detected suggesting that, at the concentrations used, the venom induced apoptosis.

Neuromuscular activity of Bothrops neuwiedi pauloensis snake venom in mouse nerve-muscle preparations

The pharmacological effects of Bothrops neuwiedi pauloensis venom on mouse phrenic nerve-diaphragm (PND) preparations were studied. Venom (20 mug/ml) irreversibly inhibited indirectly evoked twitches in PND preparations (60 ± 10 percent inhibition, mean ± SEM; p<0.05; n=6). At 50 mug/ml, the venom blocked indirectly and directly (curarized preparations) evoked twitches in mouse hemidiaphragms. In the absence of Ca2+, venom (50 mug/ml), produced partial blockade only after an 80 min incubation, which reached 40.3 ± 7.8 percent (p<0.05; n=3) after 120 min. Venom (20 mug/ml) increased (25 ± 2 percent, p< 0.05) the frequency of giant miniature end-plate potentials in 9 of 10 end-plates after 30 min and the number of miniature end-plate potentials which was maximum (562 ± 3 percent, p<0.05) after 120 min. During the same period, the resting membrane potential decreased from - 81 ± 1.4 mV to - 41.3 ± 3.6 mV 24 fibers; p<0.01; n=4) in the end-plate region and from - 77.4 ± 1.4 to -44.6 ± 3.9 mV (24 fibers; p<0.01; n=4) in regions distant from the end-plate. These results indicate that B. n. pauloensis venom acts primarily at presynaptic sites. They also suggest that enzymatic activity may be involved in this pharmacological action.(AU)

Effects of different protocol doses of low power gallium-aluminum-arsenate (Ga-Al-As) laser radiation (650 nm) on carrageenan induced rat paw ooedema.

The purpose of the present study was to investigate the effect of the low power laser therapy on the acute inflammatory process. Male Wistar rats were used. The rat paw oedema was induced by sub-plantar injection of carrageenan, the paw volume was measured before and 1, 2, 3 and 4 h after the injection using a hydroplethysmometer. To investigate the mechanism action of the Ga-Al-As laser on inflammatory oedema, parallel studies were performed using adrenallectomized rats or rats treated with sodium diclofenac. Different laser irradiation protocols were employed for specific energy densities (EDs), exposure times and repetition rates. The rats were irradiated with the Ga-Al-As laser during 80 s each hour. The ED that produced an anti-inflammatory effect were 1 and 2.5 J/cm(2), reducing the oedema by 27% (P<0.05) and 45.4% (P<0.01), respectively. The ED of 2.5 J/cm(2) produced anti-inflammatory effects similar to those produced by the cyclooxigenase inhibitor sodium diclofenac at a dose of 1 mg/kg. In adrenalectomized animals, the laser irradiation failed to inhibit the oedema. Our results suggest that low power laser irradiation possibly exerts its anti-inflammatory effects by stimulating the release of adrenal corticosteroid hormones.

Histopathological changes in avian kidney caused by Bothrops insularis (jararaca ilh a) venom and a phospholipase A2-containing fraction.

The histopathological changes induced in avian kidney by the intramuscular injection of Bothrops insularis (jararaca ilh a) venom and its phospholipase A2 (PLA2)-containing fraction were examined. Acute experiments (3 h and 24 h) with B. insularis crude venom (20 microg and 80 microg) or its PLA2-contaning fraction (10 microg and 40 microg) resulted in significant structural damage to the kidneys of 5-12-day-old chicks. Histopathological analysis indicated that the venom and its fraction acted on the renal tubules and glomeruli. The morphological changes, although widespread, varied in intensity from cell to cell, and from tubule to tubule in venom-injected chicks. The tubular and glomerular changes produced by the venom and its PLA2-containing fraction may be the result of a direct cytotoxic effect potentiated by ischemia-related disturbances in the regional hemodynamics. The venom and its fraction affected more segments along reptilian-type nephrons than along mammalian ones. This divergent sensitivity to the venom and its fraction may reflect the species-specific characteristics of B. insularis snake, an example of geographical isolation influencing its diet which is almost exclusively avian.

An unusual presynaptic action of Bothrops insularis snake venom mediated by phospholipase A2 fraction.

A phospholipase A2-containing fraction was isolated from the venom of Bothrops insularis by a combination of gel filtration on Sephadex G-150 and ion exchange chromatography on DEAE-Sephadex. Peak IV of the latter chromatography containing all of the phospholipase A2 (PLA2) activity, was assayed on isolated neuromuscular preparations. In the mouse phrenic nerve-diaphragm incubated in Tyrode at 37 degrees C, the PLA2 fraction produced an initial increase in the twitch tension and in the frequency of the mepps, followed by a dose-dependent, irreversible blockade. The replacement of 1.8 mM Ca2+ by 4 mM Sr2 inhibited the neuromuscular blocking effect of the fraction. In the chick hiventer cervicis preparation incubated with Krebs solution at 37 degrees C, the PLA2 fraction induced blockade but did not affect the response to acetylcholine and K+, excluding the involvement of post-synaptic and direct muscular effects. A low temperature (18-22 degrees C) incubation prevented the neuromuscular effect from developing. These results suggest that the PLA2-containing fraction acts predominantly at presynaptic sites at the neuromuscular junction. This fraction also accounts for most of the pharmacological effects of the crude venom.

Characterization of a myotoxin from the Duvernoy's gland secretion of the xenodontine colubrid Philodryas olfersii (green snake): effects on striated muscle and the neuromuscular junction.

A myotoxin has been isolated from the Duvernoy's gland (DG) secretion of the xenodontine colubrid Philodrvas olfersii (green snake) by gel filtration on Sephadex G-100 SF. Under non-reducing and reducing conditions in SDS-PAGE, the myotoxin migrates as a single band with a mol. wt. of 20000. The toxin has 182 amino acid residues (approximately 20% acidic), a pI of 4.8 and a blocked N-terminal. In the chick biventer cervicis preparation, P. olfersii myotoxin partially blocks potassium-evoked contractures without affecting either the twitch-tension resulting from indirect stimulation or the contractures evoked by acetylcholine. Both the DG secretion and the myotoxin increase the serum creatine kinase (CK) levels of mice and stimulate the release of CK from the biventer cervicis preparation in a dose- and time-dependent manner. The varying degrees of muscle cell lysis and extensive widening of the intercellular spaces caused by the DG secretion are reproduced by the myotoxin, with the exception that in the latter the partial or total loss of transverse muscle striations is restricted to the muscle periphery. This myotoxin is the first such protein to be characterized from a DG secretion.

The effects of Duvernoy's gland secretion from the xenodontine colubrid Philodryas olfersii on striated muscle and the neuromuscular junction: partial characterization of a neuromuscular fraction.

The effect of Philodryas olfersii Duvernoy's secretion was studied in vivo in mice and chicks as well as in the mouse phrenic nerve-diaphragm and the chick biventer cervicis preparations. The whole secretion (20-40 micrograms/ml) increased the creatine kinase (CK) levels in mice but had no effect on the mouse phrenic nerve-diaphragm preparation. In the chick, the secretion caused head drop and paresia as well as irreversible blockade of the twitch-tension evoked by indirect stimulation in the chick biventer cervicis preparation (50% paralysis in 34.5 +/- 2.7 min, n = 4). The secretion also caused muscle contracture (30% of the maximal twitch-tension generated) after a latency of nearly 9 min. Following fractionation on a Superose 12 FPLC column, the neuromuscular activity was recovered in the high mol. wt fraction (Peak I). At a concentration of 10 micrograms/ml in the chick biventer cervicis preparation, Peak I caused 50% paralysis within 18.5 +/- 3.0 min (n = 4), and evoked a strong contracture (70% of the maximal twitch-tension generated). The contractile responses of the chick preparation to ACh and KCL were partially blocked (90%) by the whole secretion and totally blocked by Peak I. CK release was increased by the whole secretion but not by Peak I. The whole secretion also produced various degrees of muscle cell lysis and extensive widening of the intercellular spaces. The latter showed a loosely arranged membranous network. In general, Peak I caused only minor morphological alterations compared with the whole secretion, although these were still significantly different from those observed in the control preparations. The changes principally involved hypercontraction of the muscle fibers. Based on the above results, we conclude that Peak I contains the factor(s) responsible for the in vitro effects on neuromuscular transmission, whereas the direct myotoxic effect is apparently caused by at least one other component of the Duvernoy's secretion.

The ability of specific antivenom and low temperature to inhibit the myotoxicity and neuromuscular block induced by Micrurus nigrocinctus venom.

In the isolated mouse diaphragm preparation, Micrurus nigrocinctus venom produced a dose-dependent contracture and blockade of the contractile response to direct and indirect electrical stimulation of the muscle. This effect could not be completely reversed by repeated washing of the preparation nor by the addition of neostigmine or 3, 4-diaminopyridine. The observation that the direct blockade had to be preceded by indirect blockade together with the capacity for venom to prevent the ACh- but not the KCl-induced contractures in biventer cervicis and chronically denervated preparations strongly suggests a curarimimetic action for the venom. The temperature at which the experiment was performed greatly influenced the neuromuscular blocking and myotoxic actions of the venom and suggests that the venom component responsible for these effects is thermolabile. Both the neuromuscular blocking action and the myotoxicity of the venom could be prevented by a specific M. nigrocinctus antivenom regardless of whether this was added together with or after the venom. The muscle morphological changes induced by the venom were accompanied by a corresponding increase in the release of creatine kinase (CK) into the incubation medium. This release was, however, submaximal (35%) when compared to that induced by the detergent Triton X-100. In contrast to what has been demonstrated for other Micrurus venoms (M. frontalis, M. corallinus, M. lemniscatus and M. spixii), our results show that the myotoxic effect induced by M. nigrocinctus venom is important for the development of blockade of the muscle contractile response.

Effect of Bothrops insularis venom on the mouse and chick nerve-muscle preparation.

The effects of Bothrops insularis venom were examined in vivo in mice and chicks and in vitro using the mouse phrenic nerve diaphragm and chick biventer cervicis muscle preparations. Incubation of the indirectly or directly stimulated mouse preparation with B. insularis venom (20-80 micrograms/ml) produced an initial increase in twitch tension followed by irreversible blockade. With direct stimulation in the presence of D-tubocurarine, no increase in twitch tension was observed prior to the onset of blockade. A venom-induced effect on presynaptic activity was suggested by the marked increase in the frequency of the mepps recorded in vitro 5-15 min after venom addition. A direct muscular effect was shown by the dose- and time-dependent reduction in the resting membrane potential of the diaphragm. Chick preparations were more sensitive than those of the mouse. In the isolated chick biventer cervicis muscle preparation, B. insularis venom induced a contracture and a dose-dependent block of responses to indirect stimulation. At low venom concentrations (1-5 micrograms/ml), no significant release of creatine kinase (CK) was observed from this preparation. However, a dose-dependent release of CK was detected at higher doses (10-80 micrograms/ml). For morphological studies, B. insularis venom was injected into the chick left pectoralis muscle. At low doses (0.4 microgram), only an inflammatory reaction was present, while at high doses (20-80 micrograms) increasing numbers of necrotic fibers were observed as well as occlusive thrombosis and hemorrhage. The muscular effect, also observed on the incubated muscle, points to a direct myolytic action of the whole venom.