Extreme weather and air pollution effects on cardiovascular and respiratory hospital admissions in Cyprus.

In many regions of the world, climatic change is associated with increased extreme temperatures, which can have severe effects on mortality and morbidity. In this study, we examine the effect of extreme weather on hospital admissions in Cyprus, for inland and coastal areas, through the use of synoptic weather classifications (air mass types). In addition, the effect of particulate air pollution (PM10) on morbidity is examined. Our results show that two air mass types, namely (a) warm, rainy days with increased levels of water vapour in the atmosphere and (b) cold, cloudy days with increased levels of precipitation, were associated with increased morbidity in the form of hospital admissions. This was true both for cardiovascular and respiratory conditions, for all age groups, but particularly for the elderly, aged over 65. Particulate air pollution was also associated with increased morbidity in Cyprus, where the effect was more pronounced for cardiovascular diseases.

Opposite roles of GABA and excitatory amino acids on the control of GAD expression in cultured retina cells.

The mechanism of control of GAD expression by GABA and excitatory amino acids (EAAs) was studied in chick and rat retina cultures using immunohistochemical and PAGE-immunoblot detection of the enzyme, as well as by measuring enzyme activity. Aggregate cultures were prepared with retina cells obtained from chick embryos at embryonic days 8-9 (E8-E9). Organotypical cultures were also prepared with retinas from E14 chick embryos, post-hatched chicken and P21 rats. GABA (1-20 mM) fully prevented GAD expression in aggregate and organotypical cultures from chick embryo retinas. A substantial, but not complete, reduction of GAD was also observed in organotypical cultures of post-hatched chicken and P21 rats, in which both forms of the enzyme (GAD65 and 67) were affected. The GABA effect was not mimicked by THIP (100 microM), baclofen (100 microM) or CACA (300 microM), agonists of GABAa, b and c receptors, respectively. NNC-711, a potent inhibitor of GABA transporters, reduced by 50% the inhibition of GAD activity promoted by GABA. Aggregates exposed to GABA and treated with glutamate (5 mM) or kainate (100 microM) displayed an intense GAD-like immunoreactivity in many cell bodies, but not in neurite regions. Immunoblot analysis revealed that the increase in GAD-like immunoreactivity by EAA corresponded to a 67-kDa protein. However, GAD activity was not detected. Treatment of aggregates or retina homogenates with SNAP, a NO producing agent (but not its oxidized form), reduced GAD activity by more than 60% indicating that the lack of enzyme activity in GAD-like immunoreactive cells, could be due to NO production by EAA stimulation.

GABAergic system in the developing mammalian retina: dual sources of GABA at early stages of postnatal development.

In the present work, we have characterized the maturation of the GABAergic system in mammalian retina. Immunoreactivity for GABA, GAD (glutamic acid decarboxylase, EC 4.1.1.15) -65 and -67 in the adult rat retina was localized in cells in the inner nuclear and ganglion cell layers. This pattern was established around postnatal day 8 and included transient GABA and GAD-67 expression in horizontal cells. GAD activity was very low at P1 and P4, increasing after P8, reaching maximal activity by P21 and decreasing to attain adult values by P30. GABA content was approximately constant from P1 to P13, increasing thereafter to reach adult levels. GAD protein content increased progressively with postnatal development and the two isoforms could be distinguished at P8. The disparity between retinal GABA content vs. presence and activity of the synthesizing enzyme, led us to investigate the alternative pathway for GABA synthesis that utilizes putrescine as a substrate. Highest levels of ornithine decarboxylase activity (the limiting step for putrescine synthesis) were found between P1 and P4, decreasing to very low levels after P13. The same pattern was observed for putrescine content in the retina. Highest amounts were found at P1, that decreased and remained constant after P13. Additionally, approximately 40% of tritiated putrescine incorporated by P1, P4 and adult retinas was converted into GABA. Our results suggest the existence of two different sources of GABA in mammalian retina, one that uses glutamate as a precursor and predominates in the mature nervous system and another that utilizes putrescine and is present transiently at early developmental stages.

Developmental study of Müller cells in the rat retina using a new monoclonal antibody, RT10F7.

We produced the monoclonal antibody RT10F7, characterized its antigenic specificity and expression in the adult and developing retina, in cultured retinal cells and in other parts of the central nervous system. In metabolically-labelled retinal cultures RT10F7 immunoprecipitated a protein of approximately 36,000 mol. wt. In the adult, RT10F7 stained endfeet of Müller cells in the ganglion cell layer, four horizontal bands in the inner plexiform layer, and radial fibres in the outer plexiform layer which terminated at the outer limiting membrane. In the inner nuclear layer, most somata were underlined by Müller processes that wrapped around them, but some cell bodies were immunoreactive for RT10F7 in the cytoplasm. During development, postnatal day 21 was the first age at which the adult pattern of immunoreactivity was present, although a fourth band in the inner plexiform layer was less clear than for the adult. By 14 and eight days after birth, the pattern of RT10F7 immunoreactivity approximated that of the adult; however, only three bands and one band were present, respectively, in the inner plexiform layer. At earlier ages, postnatal days 4, 1 and embryonic ages 19 and 15, the monoclonal antibody stained Müller cell endfeet and radial fibres, from the inner plexiform layer through the neuroblastic layer to the outer limiting membrane. At these ages, the immunoreactivity was more prominent at the level of Müller cell endfeet. The monoclonal antibody stained glia in preparations of dissociated retinal cells maintained in culture but not astrocytes or oligodendrocytes from optic nerve cultures. In brain sections, tanycytes exhibited RT10F7 immunoreactivity. The monoclonal antibody RT10F7 recognized a specific cell type in the retina, the Müller cell. In the adult and developing retina, RT10F7 recognized an antigen that is present primarily in Müller cell processes. This feature allowed us to follow the maturation of the Müller cell and correlate it with developmental events in the retina. RT10F7 is a specific marker for Müller cells in vivo and in vitro and may be useful for studies of function of Müller cells after ablation or after injuries that are known to activate Müller cells.

Retinal ganglion cell depletion alters the phenotypic expression of GABA and GAD in the rat retina.

We have looked at the phenotypic expression of gamma-aminobutyric acid (GABA) and the two isoforms of its synthetic enzyme [glutamic acid decarboxylase (GAD)-65 and -67] in adult rat retinas that had the superior colliculus, pretectum and optic tract lesioned unilaterally at birth. It has been shown previously that this type of manipulation induces retrograde degeneration of retinal ganglion cells presumably without affecting other intraretinal neurons. We present evidence that GABAergic amacrine cells are affected by such manipulation. The number of cells immunoreactive for GABA, GAD-65 and GAD-67 decreased in the inner nuclear layer. In the retinal ganglion cell layer, however, the number of GABA- and GAD-65-labelled cells increased, while the number of GAD-67-labelled cells did not change. Biochemical assay showed that overall GAD activity was not altered in retinas of lesioned animals. Our results support the notion that, while neonatal lesion reorganizes the expression of GABA and GAD in the retina, enzyme activity is maintained within normal levels.

A correction to the calculation of the Gibbs free energy of adsorption for biomolecules in ion-exchange systems.

We wish to propose a correction to the methodology introduced by Gerstner et al. [J.A. Gerstner, J.A. Bell, S.M. Cramer, Biophys. Chem. 52 (1994) 97-106] for the calculation of Gibbs free energies of adsorption of biomolecules to ion-exchange systems. Our approach is based on the requirement that the mobile phase and stationary phase concentrations be expressed in exactly the same units and the equilibrium constant be strictly dimensionless. The Gibbs free energies of ion-exchange calculated based on this correction appear to be more negative than those originally calculated by Gertner et al.

Transient retinal ganglion cells in the developing rat are characterized by specific morphological properties.

To determine whether dendritic development of mammalian retinal ganglion cells (RGCs) is affected by axonal target specificity, the morphology of three populations of maturing RGCs was examined. These included RGCs that exhibited either a transient, topographically incorrect, projection to the caudal superior colliculus (SC), or a transient projection to the caudal inferior colliculus (IC), in addition to a control group that exhibited a topographically correct projection to the caudal SC. Projection populations were identified by retrograde transport of rhodamine labeled latex microspheres injected into target nuclei. Labeled RGCs were then injected in vitro with Lucifer yellow to reveal the details of their dendritic morphology. Retinal ganglion cells making target errors, most of which ultimately die, were found to undergo a remarkable degree of morphological differentiation and could be categorized according to the adult type I, II, or III criteria. However, the relative proportions of these cell types were different among RGCs making transient connections versus those whose projections were preserved. Approximately half of the RGCs making topographically incorrect projections to the SC belonged to type III, in contrast to 6% that made a topographically correct projection. In addition, the population of cells sending axons to caudal IC did not include type III RGCs, but consisted of small type II neurons. The development of the basic dendritic form of each RGC type was only modestly influenced by its projection pattern; dendritic trees of cells making transient projections were essentially normal with only a slight, but statistically significant, reduction in dimensions. Moreover, dendritic remodeling was evident during maturation of neurons making either transient or normal projections. Together, these findings indicate that target specificity plays a relatively minor role on dendritic development of retinal ganglion cells.

Polypeptide amino acid composition and isoelectric point. II. Comparison between experiment and theory.

Experimental isoelectric points and amino acid compositional data for 58 proteins were compiled and organized. The experimental isoelectric points correlated well with the acidic to basic amino acid molar ratio. This agreement proved the usefulness of a recently presented analytical expression correlating explicitly protein isoelectric point to acid-base composition. Regressed acidic and basic dissociation constants were determined to be pK(alpha) = 4.9 and pK(beta) = 10.0, in fair agreement with the expected values of pK(alpha) = 4.2 and pK(beta) = 11.2. Theoretical isoelectric points determined by a more complete computational procedure were on the average in as good an agreement with the experimental values as those calculated via the theoretical approximation using the regressed dissociation constants. Thus, the analytical approximation is a powerful tool for the convenient and accurate calculation of protein isoelectric point from the amino acid composition.

Dendritic remodelling of retinal ganglion cells during development of the rat.

Investigation of the morphology of ganglion cells in the cat retina has shown that a remarkable reduction in the number of dendritic spines and branches occurs during development of the alpha and beta cell classes. To learn whether dendritic remodelling represents a generalized mechanism of mammalian retinal ganglion cell development, we have examined the morphology of ganglion cells in the retina of the developing rat. The present study has concentrated on type II cells, which retain a great number of dendritic spines and branches in the adult and comprise a large proportion of the population of rat retinal ganglion cells. To reveal fine dendritic and axonal processes, Lucifer yellow was injected intracellularly in living retinae maintained in vitro. Size and complexity of the dendritic trees were found to increase rapidly during an initial stage of development lasting from late fetal life until approximately postnatal day 12 (P12). Dendrites and axons of immature ganglion cells expressed several transient morphological features comprising an excessive number of dendritic branches and spine-like processes, and short, delicate axonal sidebranches. The following developmental stage was characterized by a remarkable decrease in the morphological complexity of retinal ganglion cells and a slowed growth of their dendritic fields. The number of dendritic branches and spines of types I and II retinal ganglion cells declined after P12 to reach a mature level by the end of the first postnatal month. Thus, even cells that retain a highly complex dendritic tree into the adult state undergo extensive remodelling. These results suggest that regressive modifications at the level of the dendritic field constitute a generalized mechanism of maturation in mammalian retinal ganglion cells.

Continuous and discrete T-maze alternation in rats: effects of intertrial and interrun intervals.

1. Recent concepts concerning animal memory have emphasized the kind of information processed in memory. Reference memory provides information relevant over several trials, i.e., it codes expectancy-based information. Working memory provides information critical for only one trial, i.e., it codes data-based information. Some investigators consider that a continuous alternation task in a T-maze depends on the reference memory of a series of left-right responses, whereas a discrete alternation task is thought to depend on working memory. 2. In the present report, we tested rats in a continuous alternation task with different intertrial intervals (ITI's). Rats were first subjected to 10 or 12 sessions at each of the following ITI's: 0, 55, 100, 200 and 600 s, and then tested at varying ITI's within each session for 12 sessions in the following sequence: 0, 55, 100, 200, 600 and 0 s. Next, the same rats were trained to perform discrete alternation with ITI's and interrun intervals (IRI's) varying across sessions but only IRI's changing within sessions, i.e., IRI = 0 or 55 with ITI = 0, 55, 100, 200 or 600 s across sessions, and IRI = 0, 55, 100, 200 and 600 s with ITI = 0 within sessions. 3. Rats performed both alternation tasks at high levels when ITI's and IRI's changed across sessions. However, when intervals changed within sessions, rats showed a better performance in the continuous task at intervals of 55, 100 and 200 s compared to their performance at these same intervals in the discrete task. In addition, for the discrete task with IRI's changing within sessions, errors probably due to proactive interference occurred more frequently with progressively increasing IRI's. 4. The hypothesis that performance of continuous alternation depends on reference memory and performance of discrete alternation depends on working memory is supported by these data.