Unusual A1/A2-visual pigment conversion during light/dark adaptation in marine fish.

Changes in visual pigments were studied in two marine fish species, the masked greenling Hexagrammos octogrammus and the prickleback Pholidapus dybowskii. A microspectrophotometric (MSP) analysis showed that the rods and cones of the fish collected from the natural marine environment in summer or kept in a tank at a high illumination level predominantly contained porphyropsins based on chromophore A2. As a result, λmax of the double cones significantly shifted to longer wavelengths, reaching 625 and 609 nm, respectively. After several weeks of dark adaptation, the spectra of all the photoreceptor types shifted to shorter wavelengths, as the A1:A2 ratio switched to A1. The MSP data from the fish kept under controlled light conditions were confirmed by chromatography (HPLC), which showed that the changes in the chromophore ratio were reversible and independent of the water temperature. After the preliminary deep dark adaptation, the first noticeable shift in the pigment ratio from A1 to A2 occurred within two weeks of exposure to bright light. A novel finding in this study was a reverse polarity of A1/A2 changes, unlike the case in most other fish species, where A2 chromophore predominated after the dark exposure. This demonstration of the unusual phenomenon of visual pigment transformation suggests a modification or a new way for the activation of specific biochemical mechanisms of A1:A2 conversion at both high and low illumination levels.

Metalloproteinase inhibitor GM6001 delays regeneration in holothurians.

The effect of the GM6001 metalloproteinase inhibitor on the regeneration of ambulacral structures in Eupentacta fraudatrix has been investigated. Inhibition of proteinase activity exerts a marked effect on regeneration, being dependent on the time when GM6001 is injected. When administration of the inhibitor begins on day 3 post-injury, regeneration is completely abolished, and the animals die. This means that early activation of proteinases is crucial for triggering the regenerative process in holothurians. When GM6001 in first injected on day 7 post-injury, the regeneration rate decreases. However, this effect has proven to be reversible: when inhibition ceases, the regeneration resumes. The effect of the inhibitor is manifested as a retarded degradation of the extracellular matrix, the lack of cell dedifferentiation, and, probably, a slower cell migration. The gelatinase activity is detected in all the regenerating organs of E. fraudatrix. In the holothurian Cucumaria japonica, which is not capable of healing skin wounds and ambulacrum reparation, no gelatinase activity was observed at the site of damage. A suggestion is made that proteinases play an important role in regeneration in holothurians. The most probable morphogenesis regulators are matrix metalloproteinases with gelatinase activity.

Specific features of analgesic effect of docosahexaenoic acid in rats with neuropathic pain syndrome.

Experiments on white rats showed that sciatic nerve ligation induced the development of neuropathic pain syndrome: thermal pain threshold decreased, significant reduction in weight bearing of the injured limb, and degenerative changes in the foot tissues. Administration of docosahexaenoic acid reduces activity and duration neuropathic pain syndrome, promoted reversion of weight bearing asymmetry, and prevented the development of degenerative changes in the foot tissues.

Histo- and immunocytochemical detection of inducible NOS and TNF-alpha in the locus coeruleus of human opiate addicts.

In order to determine the role of inducible nitric oxide synthase (iNOS) and tumor necrosis factor-alpha (TNF-alpha) in the pathogenesis of opiate addiction in humans, the expression of both was analyzed in the locus coeruleus (LC) of patients who died from heroin overdose. In control subjects, NADPH-diaphorase (NADPHd) histochemistry was mainly observed in non-noradrenerdic neurons, some glial and endothelial cells. However, in the brain of opiate addicts, NADPHd and iNOS expression was detected in noradrenergic LC cells, correlating with an increase in iNOS and TNF-alpha expression in glial cells as revealed by immunohistochemical and Western blot analyses. These findings indicate that sustained overproduction of cytokine and NO via iNOS expression may be responsible, at least in part, for some neurochemical changes in the locus coeruleus caused by chronic opiate usage in humans.

Effects of NO-modulating agents on the development of acute painful reaction in rats.

Painful reaction of rats to intraperitoneal injections of L-arginine, Nw-nitro-L-arginine, and agmatine was studied on the model of formalin-induced inflammation. All drugs exhibited a dubious effect on the patterns of nociceptive behavior depending on the phase of painful reaction. The dynamics of nitrate/nitrite content in animal blood and serum indicated the presence of NO-dependent and NO-independent components in the mechanisms of pharmacological effects of these drugs.

[A study of quantitative dynamics of F-actin during oocyte maturation in the starfish Asterias amurensis].

We studied the actin cytoskeleton state in Asterias amurensis oocytes within 30 min after the 1-methyladenine-induced maturation until the germinal vesicle breakdown. The total amount of actin remained unchanged during oocyte maturation. In immature oocytes, the major part of actin is not a part of filaments, but in the presence of 1-methyladenine massive actin polymerization began already within 20 min. Electron immunocytochemistry methods demonstrated joint localization of actin and alpha-protein in the cytoplasm. They were redistributed from the cortex to the cytoplasm in the presence of 1-methyladenine. A possible involvement of actin cytoskeleton in transmembrane transduction of the hormonal signal at the postreceptor stages is discussed.

[Effect of 1-methyladenine on localization of Galpha1-protein in starfish oocytes]. / Vliianie 1-metiladenina na lokalizatsiiu Galpha1-belka v ootsitakh morskoi zvezdy.

Localization and quantitative dynamics of alpha i subunit of G protein was studied by electron immunocytochemistry and immunoenzyme assay after hormonal induction of maturation in Asterias amurensis starfish oocytes. G alpha i protein was chiefly localized in the plasma membrane of immature oocytes; 1-methyladenine induced redistribution of the alpha i protein from the plasma membrane to intracellular structure up to the germinal physical breakdown.

[Identification and isolation of GTP-binding regulator protein from plasma membranes of oocytes from the starfish Asterias amurensis]. / Identifikatsiia i vydelenie GTF-sviazyvaiushchego regulatornogo belka iz plazmaticheskikh membran ootsitov morskoi zvezdy Asterias amurensis.

A method for isolating a GTP-binding regulatory protein from starfish oocytes is described. The protein consists of three subunits with molecular weights of 40, 37, and about 8 kDa. It is shown that the 40-kDa subunit has a high GTPase activity and is susceptible to ADP-ribosylation by pertussis toxin. The latter property of this subunit proved to decrease upon its incubation with nonhydrolyzable GTP analogues. These data provide evidence that the plasma membrane of starfish oocytes contains a 40-kDa GTP-binding protein with properties characteristic of the alpha subunit of the inhibitory Gi protein. The role of this protein in the transmembrane signal transmission from the 1-methyladenine receptor to intracellular effectors is discussed.

[Polyclonal antibodies, labeling radial nerve cells of the starfish Asterias amurensis]. / Poliklonal'nye antitela, markiruiushchie kletki radial'nogo nerva morskoi zvezdy Asterias amurensis.

The results of preliminary studies suggest that the cytoskeletal fraction of the radial nerve of the starfish Asterias amurensis contained a 32 kDa protein, which is tissue specific. This protein was isolated from the radial nerve by preparative electrophoresis and used as an antigen for raising polyclonal antibodies. When testing these antibodies on sections of the starfish tissues, it was shown that they interact only with the proteins present in the radial nerve cells. A conclusion was drawn that the raised antibodies may be used as a cell marker when studying regeneration of the nervous system in starfish.

Muscle regeneration in the holothurian Stichopus japonicus.

The regeneration of longitudinal muscle bands (LMBs) in the sea cucumber Stichopus japonicus was studied using light and electron microscopic and immunocytochemical methods. Previous investigations of holothurian organs showed the presence of some cytoskeletal proteins which were specific for LMBs only. One of them, the 98 KDa protein, was isolated by means of SDS-electrophoresis and used as an antigen to obtain polyclonal antibodies. When tested on paraffin sections of sea cucumber organs, the antibodies were shown to interact only with coelomic epithelial cells covering the LMBs. The antibodies were used to study LMB regeneration after transverse cutting. During regeneration no signs of myocyte dedifferentiation or mitotic division were observed. In the wound region, damaged myocytes degenerated and muscle bundles desintegrated. However, the coelomic epithelial cells dedifferentiated and began to invade the LMB. Just beneath the surface these cells formed clusters (muscle bundle rudiments). The number and size of the clusters gradually increased, the cells lengthened and developed contractile filaments. These observations suggest that new muscle bundles arise from coelomic epithelial cells covering the LMBs. The migration of coelomic epithelial cells into the damaged LMBs and their myogenic transformation are the basic mechanism of holothurian muscle regeneration.