Influence of imidazole on behavioral effects induced by dopaminergic agonists in rats

Imidazole (IMI) (from 18.7 to 300 mg/Kg) i.p. injected in adult rats induced shaking, which was antagonized by both morphine (MOR) and haloperidol (HALO) but not by methysergide (MET). I.p. IMI pretreatment inhibited the penile erections (PE) and stretching and yawning (SY) typically elicited by N-n-propylnorapomorphine (NPA), a well-known CNS dopamine (DA) receptor stimulant, injected either i.p. or i.c.v., whereas it enhanced stereotyped behavior (SB). IMI had similar effects on the same parameters considered when injected before lisuride, an ergot derivative also active as a central DA receptor agonist. In this case not only SB but also and above all aggressiveness were markedly potentiated, both the signs appearing at doses of lisuride which were "per se" ineffective. Aggressiveness, like SB, was not sex linked and was antagonized by HALO and MOR, but not by MET. IMI alone potentiated the fighting induced by electrical shock, an effect which was abolished by HALO pretreatment. Considering the results obtained as a whole it is submitted that IMI antagonizes PE and SY through a selective blockade of a class of DA receptors, presumably DA presynaptic autoinhibitors, thus potentiating SB and aggressiveness, which involve stimulation of DA postsynaptic receptors.     

Identification of Tyr115 labeled by S-(4-bromo-2,3-dioxobutyl)glutathione in the hydrophobic substrate binding site of glutathione S-transferase, isoenzyme 3-3.

Incubation of S-(4-bromo-2,3-dioxobutyl)glutathione (S-BDB-G), a reactive analogue of glutathione, with the 3-3 isoenzyme of rat liver glutathione S-transferase at pH 6.5 and 25 degrees C results in a time-dependent inactivation of the enzyme. The kobs exhibits a nonlinear dependence on S-BDB-G concentration from 50 to 900 microM, with a kmax of 0.073 min-1 and KI = 120 microM. The addition of 5 mM S-hexylglutathione, a competitive inhibitor with respect to glutathione, completely protects against inactivation by S-BDB-G. About 2.0 mol of [3H]S-BDB-G/mol of enzyme subunit is incorporated concomitant with 100% inactivation, whereas only 0.96 mol of reagent/mol subunit is incorporated in the presence of S-hexylglutathione when activity is fully retained. Modified enzyme, prepared by incubating glutathione S-transferase with [3H]S-BDB-G in the absence or in the presence of S-hexylglutathione, was reduced with NaBH4, reacted with N-ethylmaleimide, and digested with trypsin. Analysis of the tryptic digests, fractionated by reverse-phase high-performance liquid chromatography, revealed Tyr115 as the amino acid whose reaction with S-BDB-G correlates with inactivation. Examination of the stability of S-(4-bromo-2,3-dioxobutyl)glutathione and modified enzyme in the absence and presence of dithiothreitol and under acidic conditions suggests that for stable linkage to peptides, the carbonyl moieties of the reagent should be reduced immediately after modification of a protein. Comparison of results from the 4-4 and 3-3 isoenzymes of rat liver glutathione S-transferase (both of the mu gene class) indicates: the 4-4 isoenzyme exhibits a greater affinity for S-BDB-G; Cys86 is labeled by [3H]S-BDB-G in both isoenzymes but is nonessential for activity; in the 3-3 isoenzyme, Cys86 is more accessible to S-BDB-G; and Tyr115 is an important residue in the hydrophobic binding site of both enzymes.     

Screening for mutations in the neurofibromatosis type 2 (NF2) gene in sporadic meningiomas

Meningiomas are benign tumors of the central nervous system. They are usually sporadic but can also occur associated with the neurofibromatosis type 2 (NF2) syndrome. The gene responsible for NF2, recently isolated from chromosome 22, encodes a membrane-organizing protein that shows high sequence homology to a protein family thought to link the cytoskeleton with membrane proteins. Mutations of the NF2 gene have been described in sporadic meningiomas, exclusively in tumors that show loss of heterozygosity (LOH) of 22q. These preliminary results indicate that the NF2 gene is involved in the pathogenesis of at least a subset of meningiomas, where it does indeed behave as a tumor suppressor gene. In order to characterize better the role of the NF2 gene in the genesis of meningiomas we have examined the entire coding sequence of the gene in 125 meningiomas by single-strand conformational polymorphism analysis; furthermore, LOH analysis for markers of 22q has been carried out. Inactivating mutations were identified in 30% of our samples, all of which also showed LOH of 22q. The majority of mutations identified were frameshifts and nonsense mutations, which are predicted to produce a truncated or non-functional protein. We also found two missense and three in-frame deletions that may pinpoint specific regions of the protein critical to its function. Furthermore, the distribution of mutations throughout the gene, suggested that exons 2, 3, 5, 11 and 13 are more frequently involved. Our results reconfirm the importance of the NF2 gene in the pathogenesis of meningiomas and also suggest that there may be a nonrandom clustering of mutations throughout the gene.     

Effects of shell status and social context on the agonistic behavior of the tropical hermit crab,Clibanarius signatus

In the hermit crabClibanarius signatus, the influence of both shell status and social context on agonistic behavior was examined. The experimental hermits were of the same size and sex, but differed in the status of the inhabited shell, which was either optimal, sub-optimal or damaged. The evidence from this experiment suggests that the agonistic behavior ofC. signatus is influenced to a great extent by asymmetries in the resource value; attacks are mainly performed by crabs which inhabit shells in a relatively poor condition and are mostly directed towards opponents with better-fitting shells. However, this effect differs according to the social context, being more pronounced when the potential gain is greater. Shell fights are decidedly more complex than most animal combats; more work is certainly needed to understand the mechanisms underlying resource assessment and decisions made during hermit crabs' interactions.     

Acquisition of desiccation tolerance by isolated maize embryos exposed to different conditions: the questionable role of endogenous abscisic acid

The effect of exogenous ABA on acquisition of desiccation tolerance has been well documented for the embryos of several species. including maize ( Zea mays L.). It has also been suggested that endogenous ABA plays a role in regulating the same phenomena. To test this hypothesis, endogenous ABA was quantified by radioimmunoassay. Our results show that: (1) during embryogenesis in maize, endogenous ABA increase-concomitantly with the acquisition of desiccation tolerance: (2) ABA deficient embryos of the vp 5 mutant are desiccation intolerant, but tolerance can he induced by exogenous ABA: and (3) desiccation tolerance is acquired if desiccation sensitive embryos undergo a slow drying treatment, during which ABA increases. However, when embryos were preincubated in fluridone to prevent ABA accumulation during slow drying, desiccation tolerance was induced in spite of the low level of endogenous ABA in the embryo. Our results cast doubts on an exclusive role of ABA in the acquisition of desiccation tolerance in maize embryo.     

The nitrite reductase gene of Pseudomonas aeruginosa: Effect of growth conditions on the expression and construction of a mutant by gene disruption

Abstract The expression of nitrite reductase has been tested in a wild-type strain of Pseudomonas aeruginosa (Pao1) as a function of nitrate concentration under anaerobic and aerobic conditions. Very low levels of basal expression are shown under non-denitrifying conditions (i.e. absence of nitrate, in both aerobic and anaerobic conditions); anaerobiosis is not required for high levels of enzyme production in the presence of nitrate. A Pseudomonas aeruginosa strain, mutated in the nitrite reductase gene, has been obtained by gene replacement. This mutant, the first of this species described up to now, is unable to grow under anaerobic conditions in the presence of nitrate. The anaerobic growth can be restored by complementation with the wild-type gene.     

Cumulus expansion is impaired with advanced reproductive age due to loss of matrix integrity and reduced hyaluronan

Reproductive aging is associated with ovulatory defects. Age-related ovarian fibrosis partially contributes to this phenotype as short-term treatment with anti-fibrotic compounds improves ovulation in reproductively old mice. However, age-dependent changes that are intrinsic to the follicle may also be relevant. In this study, we used a mouse model to demonstrate that reproductive aging is associated with impaired cumulus expansion which is accompanied by altered morphokinetic behavior of cumulus cells as assessed by time-lapse microscopy. The extracellular matrix integrity of expanded cumulus–oocyte complexes is compromised with advanced age as evidenced by increased penetration of fluorescent nanoparticles in a particle exclusion assay and larger open spaces on scanning electron microscopy. Reduced hyaluronan (HA) levels, decreased expression of genes encoding HA-associated proteins (e.g., Ptx3 and Tnfaip6), and increased expression of inflammatory genes and matrix metalloproteinases underlie this loss of matrix integrity. Importantly, HA levels are decreased with age in follicular fluid of women, indicative of conserved reproductive aging mechanisms. These findings provide novel mechanistic insights into how defects in cumulus expansion contribute to age-related infertility and may serve as a target to extend reproductive longevity.     

PYK2 is overexpressed in chronic lymphocytic leukaemia: A potential new therapeutic target

Chronic Lymphocytic Leukaemia (CLL) is the most common adult B-cell leukaemia and despite improvement in patients' outcome, following the use of targeted therapies, it remains incurable. CLL supportive microenvironment plays a key role in both CLL progression and drug resistance through signals that can be sensed by the main components of the focal adhesion complex, such as FAK and PYK2 kinases. Dysregulations of both kinases have been observed in several metastatic cancers, but their role in haematological malignancies is still poorly defined. We characterized FAK and PYK2 expression and observed that PYK2 expression is higher in leukaemic B cells and its overexpression significantly correlates with their malignant transformation. When targeting both FAK and PYK2 with the specific inhibitor defactinib, we observed a dose–response effect on CLL cells viability and survival. In vivo treatment of a CLL mouse model showed a decrease of the leukaemic clone in all the lymphoid organs along with a significant reduction of macrophages and of the spleen weight and size. Our results first define a possible prognostic value for PYK2 in CLL, and show that both FAK and PYK2 might become putative targets for both CLL and its microenvironment (e.g. macrophages), thus paving the way to an innovative therapeutic strategy.