Neurotoxic effects evoked by cyanobacterial extracts suggest multiple receptors involved in electrophysiological responses of molluscan (CNS, heart) models

The responses of the snail central neurons (Helix pomatia, Lymnaea stagnalis) and the isolated Helix heart were characterized evoked by cyanobacterial extracts (Cylindrospermopsis raciborskii ACT strains) isolated from Lake Balaton (Hungary). The nicotinergic acetylcholine (ACh) receptors in the CNS (both excitatory and inhibitory) were blocked by the extracts of ACT 9502 and ACT 9505 strains and the anatoxin- a (homoanatoxin-a) producing reference strain of Oscillatoria sp. (PCC 6506), similar to the inhibitory effects of the pure anatoxin-a. The enhancement of the ACh responses by the ACT 9504 extract suggests additional, probably acetylcholine esterase inhibitory mechanisms. On the isolated Helix heart the crude ACT 9505 and PCC 6506 extracts evoked frequency increase and transient twitch contraction, opposite to the ACh evoked heart relaxation. Anatoxin-a similarly contracted the heart but did not increase its contration frequency. These data suggest the involvement of some non-cholinergic mechanisms, acting very likely by direct modulation of the electrical or contractile system of the isolated heart. Diversity of the effects evoked by the cyanobacterial extracts in the CNS and heart suggest pharmacologically different neuroactive components among the secondary metabolites of the cyanobacteria acting on both (anatoxin-a like) cholinergic and (unidentified) non-cholinergic receptors.     

Phe-met-arg-phe (FMRF)-amide is a substrate source of NO synthase in the gastropod nervous system

The possible involvement of the L-arginine-containing Phe-met-arg-phe (FMRF)-amide (FMRFa) in neuronal nitric oxide (NO) biosynthesis was studied in a gastropod species. We found NADPH-diaphorase-positive neurons and FMRFa-containing fibers in close proximity in the enteric nervous system. Administration of L-arginine and FMRFa induced quantitatively similar nitrite production in both intact intestinal tissues and tissue homogenates. These changes could be prevented by the presence of NOARG (an NO synthase inhibitor). Neither chemically modified FMRFa (D-arginine instead of L-arginine) nor amino acid constituents of FMRFa (methionine, phenylalanine) affected basal nitrite production. FMRFa-induced alterations were reduced in the presence of Na⁺ channel blockers (tetrodotoxin, amiloride, lidocaine), the Na⁺/K⁺ATPase inhibitor ouabain, or protease inhibitors (leupeptine, pepstatine-a). FMRFa and its amino acid constituents were analyzed by paper chromatography. When FMRFa was added to tissue homogenates, the peptide was eliminated within 1–2 min, whereas methionine, phenylalanine, arginine, and citrulline levels were elevated simultaneously. We tested the effects of FMRFa, L-arginine, and NOARG on intestinal contractile activity. FMRFa relaxed the intestine for 1–2 min and then induced contractions for 20–40 min. In the presence of NOARG, no relaxant effect of FMRFa was recorded. As administration of L-arginine strongly inhibits the mechanical activity of the intestinal muscle, NO production presumably plays a substantial role in the action of FMRFa, at least in the initial phase. Our biochemical data indicate a direct involvement of FMRFa in NO biosynthesis. FMRFa might be hydrolyzed by extracellular peptidases and then the locally released arginine might be transported into the cells and broken-down to produce NO. Depolarization-induced NO production attributable to the activation of amiloride-sensitive Na⁺ channels might also be involved.     

Electromyographic analysis during pull,forward punch,elevation and overhead throw after conservative treatment or capsular shift at patient with multidirectional shoulder joint instability

PurposeThe aim of this study was to compare the muscle activity of patients with multidirectional instability treated in a conservative or complex manner (capsular shift with postoperative rehabilitation) and the muscle activity of stable shoulder joints before and after treatment during pull, push, and elevation of upper extremities and during overhead throw.ScopeThe study was carried out on 34 patients with multidirectional shoulder instability treated non-operatively, on 31 patients with multidirectional shoulder instability treated operatively, and on 50 healthy subjects. Signals were recorded by surface EMG from eight different muscles. The mean and standard deviation of the maximum amplitude of normalized voluntary electrical activity for the different movement types and time broadness values during overhead throw were determined for each muscle in all groups and compared with each other.ConclusionThe centralization of the glenohumeral joint and the reduction of instability is attempted to be ensured by the organism through increasing the role of rotator cuff muscles (p = 0.009) and decreasing the role of the deltoid, biceps brachii, and pectoralis maior muscles (p = 0.007). At patients after short-term and long-term conservative treatment, the maximum amplitude of normalized voluntary electrical activity of stabilizer muscles is significantly higher (p = 0.006), and that of accelerator muscles is significantly lower (p = 0.005) and the time broadness is significantly longer (p = 0.01) than that of the control group. At patients after complex treatment (open capsular shift with postoperative conservative rehabilitation) the characteristic of the muscle pattern is similar (p = 0.19) to the control group.The complex treatment resolves the labral ligamentous abnormalities by operative treatment and restores the impaired muscular control by postoperative rehabilitation, whereas the conservative treatment restores only the muscular control.     

Physiotherapy vs. capsular shift and physiotherapy in multidirectional shoulder joint instability

PurposeThe aim of the study was to compare the kinematic parameters and the on–off pattern of the muscles of patients with multidirectional instability (MDI) treated by physiotherapy or by capsular shift and postoperative physiotherapy before and after treatment during elevation in the scapular plane.ScopeThe study was carried out on 32 patients with MDI of the shoulder treated with physiotherapy, 19 patients with MDI of the shoulder treated by capsular shift and postoperative physiotherapy, and 25 healthy subjects. The motion of skeletal elements was modeled by the range of humeral elevation, scapulothoracic angle and glenohumeral angle, scapulothoracic (ST) and glenohumeral (GH) rhythms, and relative displacement between the rotation centers of the humerus and scapula. The muscle pattern was modeled by the on–off pattern of muscles around the shoulder, which summarizes the activity duration of the investigated muscles.ResultsThe different ST and GH rhythms and the increased relative displacement between the rotation centers of the scapula and the humerus were observed in MDI patients. The physiotherapy strengthened the rotator cuff, biceps brachii, triceps brachii, deltoid muscles, and increase the neuromuscular control of the shoulder joints. Capsular shift and physiotherapy enabled bilinear ST and GH rhythms and the normal relative displacement between the rotation centers of the scapula and humerus to be restored. After surgery and physiotherapy, the duration of muscular activity was almost normal.ConclusionThe significant alteration in shoulder kinematics observed in MDI patients cannot be restored by physiotherapy only. After the capsular shift and postoperative physiotherapy angulation at 60° of ST and GH rhythms, the relative displacement between the rotation centers of the scapula and humerus and the duration of muscular activity were restored.     

Synthesis and binding characteristics of a novel enkephalin analogue, [3H]Tyr-D-Ala-Gly-Phe-D-Nle-Arg-Phe

The endogenous opioid heptapeptide (Tyr-Gly-Gly-Phe-Met-Arg-Phe; MERF) has been shown to interact with multiple opioid as well as non-opioid sites in mammalian brain membranes. To increase the stability and bioavailability of MERF, new synthetic derivatives with D-amino acid substitutions were prepared and studied. One of the new compounds in this series, Tyr-D-Ala-Gly-Phe-D-Nle-Arg-Phe (DADN), had only moderate affinity in competing with [3H]MERF, whereas it displayed the highest potency in producing antinociception following intrathecal administration. DADN was radiolabeled with 41Ci/mmol specific activity. Specific binding of [3H]DADN was saturable, stereoselective and of high affinity. Chemical stability, increased micro-receptor selectivity, and hydrophobicity of the peptide all contribute to the effectiveness observed in biochemical and pharmacological studies.     

Interaction of a synthetic polyanion with rat liver mitochondria

The effect of a polyanion (a copolymer of methacrylate, malaete and styrene in a 1:2:3 proportion with an average molecular weight of 10 000) on respiration, ATPase activity and ADP/ATP exchange activity of rat liver mitochondria and submitochondrial particles has been studied.The polyanion (at 17–150 μg/ml concentration, 100 μg polyanion corresponding to 0.83 μequiv. of carboxylic groups) inhibits the oxidation of succinate and NAD-linked substrates in state 3 in a concentration-dependent manner. The extent of this inhibition can be decreased by elevating the concentration of ADP. State 4 respiration is not affected by the polyanion. It has also a slight inhibitory effect on the oxidation of the above mentioned substrates in the uncoupled state (a maximum inhibition of 37% at 166 μg/ml polyanion concentration), which is unaffected by ADP. The strong inhibition of state 3 respiration can be relieved by 2,4-dinitrophenol to the low level observed in the uncoupled state. Ascorbate+TMPD oxidation is slightly inhibited in state 3, while it is not inhibited at all in the uncoupled state.The polyanion, depending on its concentration, strongly inhibits also the DNP-activated ATPase activity of mitochondria (50% inhibition at 40 μg/ml polyanion concentration).The ATPase activity of sonic submitochondrial particles is also inhibited. However, this inhibition is incomplete (reaching a maximum of 65%) and higher concentrations of the polyanion are required than to inhibit the ATPase activity of intact mitochondria.The polyanion inhibits the ADP/ATP translocator activity of mitochondria, measured by the “back exchange” of [2-³H]ADP. After a short preincubation of the mitochondria with the polyanion, the concentration dependence of the inhibition by the polyanion corresponds to that of the DNP-activated ATPase activity of intact mitochondria.It is concluded that, in intact mitochondria, the polyanion has at least a dual effect, i.e. it partially inhibits the respiratory chain between cytochrome b and cytochrome c, and strongly oxidative phosphorylation by blocking the ADP/ATP translocator.     

Proteolytic Extraction of Salmon Oil and PUFA Concentration by Lipases

Commercial proteases (Alcalase^®, Neutrase^®, and Flavourzyme^™) were tested for their ability to release the oil content of marine by-products (salmon heads). The amount of oil (17%) obtained after 2 hours was close to that obtained by the chemical extraction method (20%). Lipolysis of the oil was carried out with Novozym SP398 to obtain a mixture of free fatty acids and acylglycerols (24 hours 45% hydrolysis). The mixture was filtered on a hydrophobic membrane to discriminate between high melting saturated fatty acids and low melting acylglycerols. The sum of total polyunsaturated fatty acids increased from 41.6% in the crude oil to 46.5% in the permeate. The docosahexaenoic acid content increased from 9.9% to 11.6%, and the eicosapentaenoic acid changed from 3.6% to 5.6%. Data from differential scanning calorimetry DSC and from thin layer chromatography coupled with flame ionization detection (TLC-FID) differed significantly between permeate and retentate. A re-esterification of the free fatty acids in the permeate with Lipozyme IM was carried out to increase the amount of long chain acylglycerols.     

Light distribution in a novel photobioreactor – modelling for optimization

The paper reports a novel photobioreactor developed to achievehomogeneous and flexible illumination inside the reactor. This is toovercome the problem of studying kinetics in standard photobioreactors,which are characterized by strong light gradients and light fluxes that cannotbe controlled. The reactor is used for the study of microalgal kinetics formodelling purposes.The new reactor combines the advantages of a stirred reactor(homogeneity) and a plate reactor (short path length). The light inputsystem consists of an external light source, a fibre-optical ring-light and alight emitting tube. Light is generated in a light source arranged externallyand directed into the reactor using optical fibres. The fibres are spread ina ring-light to provide a uniform illumination in the concentrically arrangedcylinder. Any focusable light source can be applied; by using a shuttermodule, light fluctuations can be generated in a wide range of frequencies.In order to change the light quality, spectral filters are placed between thelamp and the optical fibre.A model based approach was used to optimize the illumination: lightdistribution was calculated employing a Monte-Carlo simulation. Lightemission characteristics, reflection, refraction, scattering in the suspensionand on rough surfaces were studied numerically. Propositions were derivedhow to optimize the reactor, e.g. now to achieve higher light intensities anda more uniform illumination. Finally, mean photon flux densities of 100± 15 mol m^-1 s^-1 were achieved at theilluminated surface.The simulation results revealed that the light distribution at constantbiomass concentration is mainly determined by the geometrical parametersof the lightening device mentioned above and that any simplifications leadto serious misinterpretations.     

Effects of capture and video‐recording on the behavior and breeding success of Great Tits in urban and forest habitats

Behavioral research often involves capturing and video‐recording birds, but these procedures may have undesired effects on the behavior of birds that have rarely been quantified. In addition, birds in urban and more natural areas may differ in their sensitivity to disturbance. We examined the possible effects of both capturing, weighing and measuring, and taking a blood sample, and the presence of video‐cameras on the behavior of male and female Great Tits (Parus major) breeding in urban and forest habitats. Using a 2 × 2 block design, we compared the behavior and breeding success of parents that either were or were not captured on their nests a few days before behavioral observations, and of parents that either were or were not habituated to the presence of a concealed video‐recorder mounted on nest boxes. We found no significant effects of habituation to the camera on bird behavior, but males captured in their nest boxes were more vigilant and hesitated longer before entering nest boxes, and also had slightly lower provisioning rates than males that had not been captured. Captured females also tended to be more vigilant than females that had not been captured, but their provisioning rates were not affected. Capturing males also influenced the behavior of their non‐captured mates, but capturing females had no effect on the behavior of their non‐captured mates. We found no difference in the effects of capture on Great Tits in urban and forest habitats, and our treatments also had no effect on the mass, size, and survival of nestlings until fledging. Our results suggest that, for Great Tits, being captured results in sex‐dependent behavioral effects that can last for at least several days. As such, we suggest that the possibility of similar effects in other species of birds should be considered in behavioral studies where birds must be captured, and recommend either that behavioral data be collected before capturing birds or that all birds in a study should be captured and handled in a standardized way.