Does prolactin influence the hypothalamo-pituitary GnRH-LH system in preovulatory-phase ewes?

The effects of prolonged infusions of prolactin (PRL) into the third ventricle of the brain of cycling ewes on the secretory activity of hypothalamic GnRH neurons and pituitary LH cells in the pars distalis during the proestrous day were studied. Mature Blackhead ewes were infused with vehicle (control, n=5) or with prolactin (200 mug/day, n=5) during 4 consecutive days prior to the next spontaneous ovulation. The dose of PRL was infused each day in 4 series of 50 mug/100 mul/h at 30-min. intervals, from 8.30 to 14.00 h. The animals were slaughtered on the 16th (proestrous) day of the estrous cycle immediately after the last infusion and their brains were fixed in situ. Plasma samples were collected for 6 h at 10 min. intervals, on days 12 (before the infusions) and 16 of the cycle. The distribution pattern, number and morphology of GnRH neurons in vehicle- and PRL-infused ewes were found to be similar and typical for the proestrous phase of the cycle. The immunoreactive (ir) GnRH stores in the median eminence were high and similar in both groups. There were no differences between control and PRL-treated ewes in the number or features of irLH cells. The area fraction and optical density for irLH cells and mRNA LHbeta-expressing cells did not differ between control and experimental groups. Irrespective of the kind of infusion, changes in LH secretion during the estrous cycle were similar in control and PRL-infused ewes. Mean plasma LH concentrations were higher (p<0.001) on day 16 compared to day 12 of the cycle. There were no differences in plasma LH concentrations or in the parameters of pulsatile LH secretion between groups. In conclusion, repeated, several-hour-long infusions of PRL into the CNS prior to the next spontaneous ovulation in ewes has no direct effect on the secretory activity of GnRH neurons, and/or the synthesis, accumulation, or tonic release of LH from the pituitary gonadotrophs.     

Developmental-stage-specific assembly of ParB complexes in Streptomyces coelicolor hyphae

In Streptomyces coelicolor ParB is required for accurate chromosome partitioning during sporulation. Using a functional ParB-enhanced green fluorescent protein fusion, we observed bright tip-associated foci and other weaker, irregular foci in S. coelicolor vegetative hyphae. In contrast, in aerial hyphae regularly spaced bright foci accompanied sporulation-associated chromosome condensation and septation.     

The Correlation between the Chromosome Variation in Callus and Genotype of Explants of Arabidopsis thaliana

Twelve callus lines of Arabidopsis thaliana were derived from four types of explants excised from diploid plants of two ecotypes (Columbia and Wilna) and autotetraploid plants of the Wilna ecotype. Cytogenetic analysis of the chromosome variation in particular callus lines was carried out for primary culture and callus during 5 months of culture. Ploidy levels of interphase nuclei were estimated by counting the number and size of chromocentres and nuclei of interphase cells. The first polyploid cells in all callus lines were observed during callogenesis. In primary culture the ploidy level ranged between 2 and 15x (10-75 chromosomes). The frequency of polyploid cells was higher in the 5-month old callus culture, but the ploidy level was the same. In the callus lines derived from autotetraploid plants, cells with reduced chromosome number appeared quite frequently along with diploid and polyploid cells.     

Hydrogen bonded supramolecular structures of cationic and anionic module assemblies containing square-planar oximate complex anions

The formation of the molecular assemblies consisting of anionic and cationic molecules may lead to their interesting structural properties. In this work, we present three new structures based on the tetradentate oxime and amide open-chain ligand CH₃-C(NOH)-C(O)-NH-(CH₂)₃-NH-C(O)-C(NOH)-CH₃ (PAP). All the structures contain the complex cations, complex anions [M(PAP-3H)]⁻ and solvating water molecules. These are H-bonded complex anions: [Ni(1,3-pn)₂(H₂O)₂][Ni(PAP-3H)]₂ · 4 H₂O (1), [Ni(Im)₄(H₂O)₂][Ni(PAP-3H)]₂ (2) and [Cu(Im)₄(H₂O)₂][Cu(PAP-3H)]₂ · 2H₂O (3) (Im=imidazole; 1-3-pn=1,3-diaminopropane). All compounds were synthesised by co-crystallisation of octahedral amine-containing cationic complex with oxime-containing anionic complexes in methanol solution. They were compared with some earlier reported assemblies based on the same ligand (PAP). The comparison of the structures reveals one distinct difference in the separation between the cis-situated oximate oxygen atoms O(1)?O(4) in the copper complexes. The consequence of this effect is the lengthening of the Cu-N distances. In the nickel complexes containing [Ni(PAP-3H)]⁻ anion this effect is much less pronounced.     

Nitric oxide and platelet energy metabolism

This study was undertaken to determine whether nitric oxide (NO) can affect platelet responses through the inhibition of energy production. It was found that NO donors: S-nitroso-N-acetylpenicyllamine, SNAP, (5-50 microM) and sodium nitroprusside, SNP, (5-100 microM) inhibited collagen- and ADP-induced aggregation of porcine platelets. The corresponding IC50 values for SNAP and SNP varied from 5 to 30 microM and from 9 to 75 microM, respectively. Collagen- and thrombin-induced platelet secretion was inhibited by SNAP (IC50 = 50 microM) and by SNP (IC50 = 100 microM). SNAP (20-100 microM), SNP (10-200 microM) and collagen (20 microg/ml) stimulated glycolysis in intact platelets. The degree of glycolysis stimulation exerted by NO donors was similar to that produced by respiratory chain inhibitors (cyanide and antimycin A) or uncouplers (2,4-dinitrophenol). Neither the NO donors nor the respiratory chain blockers affected glycolysis in platelet homogenate. SNAP (20-100 microM) and SNP (50-200 microM) inhibited oxygen consumption by platelets. The effect of SNP and SNAP on glycolysis and respiration was not reduced by 1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1-one, a selective inhibitor of NO-stimulated guanylate cyclase. SNAP (5-100 microM) and SNP (10-300 microM) inhibited the activity of platelet cytochrome oxidase and had no effect on NADH:ubiquinone oxidoreductase and succinate dehydrogenase. Blocking of the mitochondrial energy production by antimycin A slightly affected collagen-evoked aggregation and strongly inhibited platelet secretion. The results indicate that: 1) in porcine platelets NO is able to diminish mitochondrial energy production through the inhibition of cytochrome oxidase, 2) the inhibitory effect of NO on platelet secretion (but not aggregation) can be attributed to the reduction of mitochondrial energy production.     

Involvement of glial cells in rhythmic size changes in neurons of the housefly's visual system

In the housefly's first optic neuropile, or lamina, the axons of two classes of monopolar cell interneurons, L1 and L2, exhibit a daily rhythm of size changes: swelling during the day, and shrinking by night. At least for the L2 cells this rhythm is circadian. Moreover, epithelial glial cells that enwrap each lamina cartridge, its monopolar cell axons, and their surrounding crown of input photoreceptor terminals also change size, but in the opposite direction to the changes in L1 and L2-swelling by night and shrinking by day. The rhythmic changes in glia indicate the possible involvement of these cells in the lamina's circadian system. To examine their role in regulating the rhythmic changes of L1 and L2's axon sizes we have injected three chemicals into the haemolymph of the fly's head: fluorocitrate (FL) and iodoacetate (IAA), which affect the metabolism of glial cells, and octanol (OC), which closes gap junction channels. All chemicals exerted an effect on L1 and L2, which depended on the time of injection, the drug concentration, and the postinjection times at which we examined the fly's brains. Moreover, day/night changes in the axon sizes of L1 and L2 were increased in FL- and IAA-treated flies, indicating that glial cells may normally inhibit these changes by regulating the sizes of L1 and L2's axons during the day and night. In turn, lack of a day/night rhythm in L1 and L2 after OC injections shows that the rhythm's persistence depends on communication between the lamina cells through gap junction channels.     

Choice of isolation procedure for endotoxins from the outer membrane of the bacteria Desulfovibrio desulfuricans

The chemical structure determining properties and biological functions of endotoxins derived from Desulfovibrio desulfuricans species has not been recognized, which considerably hinders the choice of an effective extraction procedure of these lipopolysaccharides (LPS) from the bacterial outer cell membrane. We aimed at selecting the most effective method of LPS isolation from D. desulfuricans in terms of the most efficient extraction solution, the appropriate conditions of isolation and adequate purification technique. For this purpose we tested a few literature-based procedures utilizing various extraction mixtures (phenol-water, phenol-chloroform-petroleum ether and Tri-Reagent, i.e. aqueous solution of guanidinum thiocyanate and phenol). The best yield and purity of the isolated LPS were provided by the application of the extraction with phenol-water according to the modified by Shnyra et.al. (2000) procedure of Westphal et. al. (1952). A satisfactory method of isolation in micro scale appeared to be that based on Tri-Reagent and propagated by Yi and Hackett in 2000. The extraction of LPS from D. desulfuricans with phenol-chloroform-petroleum ether should not be recommended due to its low efficiency.     

Effect of Rho-associated kinase inhibition on actin cytoskeleton structure and calcium response in glioma C6 cells

The role of actin cytoskeleton functional state in glioma C6 cell morphology and calcium signaling was investigated through modification of myosin II activity by blocking Rho-associated kinase with the specific inhibitor Y-27632. Treatment of glioma C6 cells with ROCK inhibitor resulted in actin cytoskeleton reorganization and also in the changed shape and distribution of mitochondria. Changes in the distribution of ER, the main calcium store in glioma C6 cells, were not visible. The inhibition of myosin II activity influences the first phase of calcium signaling evoked by agonist, and both phases of thapsigargin-evoked calcium response. We suggest that the observed increase in Ca2+ release from intracellular stores induced by IP3 formation as well as inhibition of SERCA ATPase is at least in part related to severely affected mitochondria. Enhancement of capacitative calcium entry evoked by thapsigargin is probably associated with the reorganization of the acto-myosin II system. ATP-induced calcium response presents no changes in the second phase. We observed that ATP stimulation of Y-27632 pretreated cells leads to immediate morphological rearrangement of glioma C6 cells. It is a consequence of actin cytoskeleton reorganization: formation of stress fibers and relocation of phosphorylated myosin II to actin filaments. It seems that the agonist-evoked strong calcium signal may be sufficient for myosin II activation and the stress fiber organization. This is the first work showing the dependence between the functional state of the acto-myosin II system and calcium signaling stressing the reversible character of this relationship.