Intracellular pH in individual pituitary cells: measurement with a dual emission pH indicator

Intracellular pH (pHi) can now be measured at the single cell level using dual emission wavelength microspectrofluorimetry with the fluorescent pH indicator SNARF 1 and its membrane permeant acetoxymethyl ester (SNARF 1/AM). We measured pHi of individual pituitary cells under both basal and stimulated conditions. The emitted fluorescence of SNARF 1 probe was calibrated following experimental manipulations of pHi in two types of rat pituitary cells. The calibration curves obtained in the two cell types were identical. We observed a Gaussian distribution of individual pHi with a wide dispersion (6.95 to 8) in the two cell populations. TRH (10(-7) M) and ionomycin (5 microM) induced a transient acidification followed by a sustained alkalinization, whereas K+ (50 mM) depolarization only exerted a transient acidification. These results show that the dual emission pH indicator SNARF 1 can be used to reliably investigate changes in pHi in individual endocrine cells.     

Interactions of ubiquinones with membrane models

The effect of the insertion of coenzyme Q10 and some of its shorter chain homologues in membrane models (Reverse Micelles, Small Unilamellar Vesicles and Liposomes) has been studied by NMR and IR spectroscopies. By 1H-NMR we have found that the stretched conformation of the isoprenoid side-chain observed in solution is maintained in membrane models. Interaction between the quinonoid moiety of the Q's and the phosphatidic groups of the phospholipids has been evidenced by 31P-NMR. A large effect of this interaction on the water microdynamics and distribution around the charged groups of the phospholipids has been observed by measurements of 1H and 2H relaxation times and by infrared spectra. The 13C-NMR spectra of the backbone of the acyl chains of phospholipids does not seem to be influenced to a noticeable extent by the presence of the Q's.     

Methylthioadenosine phosphorylase from the archaeon Pyrococcus furiosus. Mechanism of the reaction and assignment of disulfide bonds.

The extremely heat-stable 5'-methylthioadenosine phosphorylase from the hyperthermophilic archaeon Pyrococcus furiosus was cloned, expressed to high levels in Escherichia coli, and purified to homogeneity by heat precipitation and affinity chromatography. The recombinant enzyme was subjected to a kinetic analysis including initial velocity and product inhibition studies. The reaction follows an ordered Bi-Bi mechanism and phosphate binding precedes nucleoside binding in the phosphorolytic direction. 5'-Methylthioadenosine phosphorylase from Pyrococcus furiosus is a hexameric protein with five cysteine residues per subunit. Analysis of the fragments obtained after digestion of the protein alkylated without previous reduction identified two intrasubunit disulfide bridges. The enzyme is very resistant to chemical denaturation and the transition midpoint for guanidinium chloride-induced unfolding was determined to be 3.0 M after 22 h incubation. This value decreases to 2.0 M in the presence of 30 mM dithiothreitol, furnishing evidence that disulfide bonds are needed for protein stability. The guanidinium chloride-induced unfolding is completely reversible as demonstrated by the analysis of the refolding process by activity assays, fluorescence measurements and SDS/PAGE. The finding of multiple disulfide bridges in 5'-methylthioadenosine phosphorylase from Pyrococcus furiosus argues strongly that disulfide bond formation may be a significant molecular strategy for stabilizing intracellular hyperthermophilic proteins.     

Cohort study of multiple brain lesions in sport divers: role of a patent foramen ovale.

OBJECTIVE: To investigate the role of a patient foramen ovale in the pathogenesis of multiple brain lesions acquired by sport divers in the absence of reported decompression symptoms. DESIGN: Prospective double blind cohort study. SETTING: Diving clubs around Heidelberg and departments of neuroradiology and neurology. SUBJECTS: 87 sport divers with a minimum of 160 scuba dives (dives with self contained underwater breathing apparatus). MAIN OUTCOME MEASURES: Presence of multiple brain lesions visualised by cranial magnetic resonance imaging and presence and size of patent foramen ovale as documented by echocontrast transcranial Doppler ultrasonography. RESULTS: 25 subjects were found to have a right-to-left shunt, 13 with a patent foramen ovale of high haemodynamic relevance. A total of 41 brain lesions were detected in 11 divers. There were seven brain lesions in seven divers without a right-to-left shunt and 34 lesions in four divers with a right-to-left shunt. Multiple brain lesions occurred exclusively in three divers with a large patent foramen ovale (P = 0.004). CONCLUSIONS: Multiple brain lesions in sport divers were associated with presence of a large patent foramen ovale. This association suggests paradoxical gas embolism as the pathological mechanism. A patent foramen ovale of high haemodynamic relevance seems to be an important risk factor for developing multiple brain lesions in sport divers.     

Global asymptotic stability for a vector disease model with spatial spread

Summary We analyze the global behaviour of a vector disease model which involves spatial spread and hereditary effects. This model can be applied to investigate growth and spread of malaria. No immunization is considered. We prove that, if the recovery rate is less than or equal to a threshold value, the disease dies out, otherwise the infectious people density tends to a homogeneous distribution. Our results follow using contracting convexes techniques and agree with the results given by K. L. Cooke for the model without diffusion.Work supported by C.N.R., Grant No. 79.00696.01.     

Energy transduction in photosynthetic bacteria. VIII. Activation of the energy-transducing ATPase by inorganic phosphate

ATPase activity and ATP-induced energization of photosynthetic membranes from Rhodopseudomonas capsulata are stimulated by phosphate; the maximum stimulatory effect occurs at a concentration between 1 and 2 mM.The sensitivity of the ATPase to oligomycin increases in the presence of phosphate since all the P_i-stimulated activity is inhibited by this antibiotic. Aurovertin, which has no effect on ATPase in the absence of phosphate, inhibits completely the activity elicited by this anion.The addition of P_i induces a substantial increase in the V of ATPase activity without changing the affinity of the enzyme for ATP or ADP.Arsenate, at the same concentrations, produces effects very similar to those of phosphate. The stimulation by arsenate of the transfer of energy from ATP to the membrane suggests a non-hydrolytic role of this anion as a modifier of the ATPase activity.     

Biological Aspects of Medorippe lanata (Linnaeus, 1767) (Brachyura: Dorippidae) from the Eastern Ligurian Sea (Western Mediterranean)

The aim of the present study is to investigate the demographic structure and to identify some aspects of the biology of an exploited population of Medorippe lanata (Brachyura: Dorippidae) in the eastern Ligurian Sea, western Mediterranean. 1364 specimens (639 males and 725 females) of M. lanata were collected on a monthly basis from January to December 2001, in a wide area of the eastern Ligurian Sea usually exploited by the Viareggio ‘rapido’ trawl fleet. M. lanata represented an important fraction of the discard, both in weight and in number of individuals. Maximum abundance of this species occurred in late summer-early autumn (up to 3369 ind. km⁻² and 50.6 kg km⁻² in August). The overall females:males sex-ratio was 1.13:1, while the monthly sex-ratio did not differ statistically from 1:1 in all months, except in September and October, when females significantly outnumbered males. The sampled population was composed of two cohorts from November to April. Sizes ranged from 10 to 29 mm carapace length (CL) for females and from 9 to 29 mm CL for males. The von Bertalanffy growth curve, computed for both sexes, gave a higher growth rate in males than in females. Recently moulted males and females were observed throughout the year, except in summer, when the highest number of ovigerous females was present. Females with external eggs were collected from March to November, with peaks in August and September. The monthly evolution of the ovarian maturity stages showed no clear temporal trend. At 21 mm CL, 50% of females were ovigerous or showed macroscopically mature ovaries. According to the dimorphism in chelae size, the presence of adult males (post-puberty stage) was observed all year round, from 18 to 29 mm CL, without evident temporal trends.     

The poor health of deep-water species in the context of fishing activity and a warming climate: will populations of Molva species rebuild or collapse?

Many deep-water fish populations, being K-selected species, have little resilience to overexploitation and may be at serious risk of depletion as a consequence. Sea warming represents an additional threat. In this study, the condition, or health, of several populations of common ling (Molva molva), blue ling (Molva dypterygia) and Mediterranean or Spanish ling (Molva macrophthalma) inhabiting different areas in the North Atlantic and the Mediterranean was evaluated, to shed light on the challenges these deep-water species are facing in the context of fishing activity and a warming climate. The data on the condition of Molva populations which are analysed here have been complemented with data on abundance and, for the southernmost species (Mediterranean ling), with two other health indicators (parasitism and hepato-somatic index). Despite some exceptions (e.g., common ling in Icelandic waters), this study shows that the condition of many populations of Molva species in the northeastern Atlantic and the Mediterranean Sea has worsened, a trend which, in recent decades, has usually been found to be accompanied by a decline in their abundance. In addition, the poor health status of most populations of common ling, blue ling and Mediterranean ling considered in this analysis points to a lower sustainability of these populations in the future. Overall, the health status and abundance of Molva populations in the northeastern Atlantic and the Mediterranean suggest that only some populations located in the North Atlantic may be able to rebuild, whereas the populations in southern North Atlantic and the Mediterranean, which are probably most at risk from sea warming, are facing serious difficulties in doing so. In the context of fisheries and global warming, this study's results strongly indicate that management bodies need to consider the health status of many of the populations of Molva species, particularly in southern European waters, before implementing their decisions.     

Characterization of the first adult de novo case of 46,X,der(Y)t(X;Y)(p22.3;q11.2)

Herein, we describe a case of an infertile man detected in postnatal diagnosis with FISH characterization and array-CGH used for genome-wide screening which allowed the identification of a complex rearrangement involving sex chromosomes, apparently without severe phenotypic consequences. The deletion detected in our patient has been compared with previously reported cases leading us to propose a hypothetical diagnostic algorithm that would be useful in similar clinical situations, with imperative multi disciplinary approach integrated with genetic counseling. Our patient, uniquely of reproductive age, is one of six reported cases of duplication of Xp22.3 (~ 8.4 Mb) segment and contemporary deletion of Yq (~ 42.9 Mb) with final karyotype as follows:

46,X,der(Y),t(X;Y)(Ypter → Yq11.221::Xp22.33 → Xpter).ish der(Y) (Yptel+,Ycen+,RP11-529I21+,RP11-506M9-Yqtel −,Xptel +). arrXp22.33p22.31(702–8,395,963, 8,408,289x1), Yq11.221q12 (14,569,317x1, 14,587,321–57,440,839x0)