Primary cultures as a model for studying ependymal functions: glycogen metabolism in ependymal cells

Ependymal cells form a single-layered, ciliated epithelium at the interface between the cerebrospinal fluid and the brain parenchyma. Although their morphology has been studied in detail, ependymal functions remain largely speculative. We have established and characterized a previously described cell culture model to investigate ependymal glycogen metabolism. During growth in minimal medium lacking many non-essential amino acids including L-glutamate, but containing glucose at physiological concentration, the cells contained negligible amounts of glycogen (7 +/- 3 nmol glucosyl residues/mg protein) despite the presence of insulin. However, during a period of 24 h, the cells accumulated glycogen to very high levels after transferal to a medium containing insulin, glucose at a 5-fold higher concentration. and all proteinogenic amino acids except L-asparagine and L-serine (990 +/- 112 nmol glucosyl residues/mg protein). omission of insulin resulted in a 50%, reduction in glycogen accumulation. Upon glucose deprivation, glycogen was degraded with a half-life of 21 min. The ependymal primary cultures contained 80 +/- 5 mU glycogen phosphorylase (Pho)/mg protein and stained positively with antibodies raised against this enzyme. Astroglial cultures built up less glycogen and had less Pho activity under identical conditions. Ependymal glycogen was mobilized by noradrenaline and serotonin. Our results indicate that ependymal cells maintain glycogen as a functional energy store, subject to rapid turnover dependent on the availability of energy substrates and the presence of appropriate signal molecules. Thus ependymocytes appear to be active players in the multitude of processes resulting in normal brain function, and ependymal primary cultures are suggested as a suitable model for studying the role of ependymal cells in these processes. (C) 2001 Elsevier Science B.V. All rights reserved

Angiotensin receptor type 1 blockade in astrocytes decreases hypoxia-induced cytotoxicity and inflammation

The present study investigated the role of angiotensin receptors (AT-R) in the survival and inflam- matory response of astroglia upon hypoxic injury. Expo- sure of rat astroglial primary cultures (APC) to hypoxic conditions (HC) led to decreased viability of the cells and to a 3.5-fold increase in TNF-alpha release. AT-R type1 (AT1-R) antagonist losartan and its metabolite EXP3174 decrease the LDH release (by 36 ± 9%; 45 ± 6%) from APC under HC. Losartan diminished TNF-alpha release (by 40 ± 15%) and the number of TUNEL-cells by 204 ± 38% under HC, alone and together with angiotensin II (ATII), while EXP3174 was dependent on ATII for its effect on TNF-alpha. The AT2-R antagonist, PD123.319, did not influence the release of LDH and TNF-alpha under normoxic (NC) and HC. These data suggest that AT1-R may decrease the susceptibility of astrocytes to hypoxic injury and their propensity to release TNF-alpha. AT1-R antagonists may therefore be of therapeutic value during hypoxia-associated neurodegeneration

The prognostic fingerprint of quality of life in older inpatients Relationship to geriatric syndromes' and resources' profile

Background The Comprehensive Geriatric Assessment (CGA) provides essential information about older hospitalized patients but is either not systematically adopted or not adopted at all in clinical routine. As a consequence, critical factors influencing patients' trajectories, like personal resources (geriatric resources, GR), geriatric syndromes (GS), health-related quality of life (HRQoL) and multidimensional prognosis often escape routine diagnostics. Objective To investigate the association between HRQoL and GR/GS as well as its prognostic signature. Material and methods In this study 165 inpatients older than 65 years admitted to an internal medicine department of a German large metropolitan hospital were assessed by a CGA-based calculation of the multidimensional prognostic index (MPI). Ten different GR and 17 GS, as well as HRQoL were collected. After 3, 6 and 12 months the patients were followed-up by telephone. Results The HRQoL was associated with MPI (p < 0.001), number of GS (p < 0.001) and survival days after discharge (p = 0.008). Additionally, significant associations were found between HRQoL and number of GR (p < 0.001). GS displaying risk for physical dependence like instability (p < 0.001) and chronic pain (p = 0.007) and single GR/GS that influence patient's confidence like isolation (p < 0.001), depression (p < 0.001) and emotional resources (p = 0.002) were also associated with HRQoL. Conclusion The HRQoL is significantly associated to specific risk and protective factor profiles of GR and GS. To improve quality of life, targeted, patient-centered diagnostics and treatment of GS as well as stabilization of GR should be encouraged in the management of older, multimorbid patients outside geriatric settings

Biosynthesis of Wdrl6, a marker protein for kinocilia-bearing cells, starts at the time of kinocilia formation in rat, and wdr16 gene knockdown causes hydrocephalus in zebrafish

The rat ortholog of the WD40 repeat protein Wdr16 is abundantly expressed in testis and cultured ependymal cells. Low levels are found in lung and brain, respectively, while it is absent from kinocilia-free tissues. In testis and ependymal primary cultures, Wdr16 messenger RNA appears concomitantly with the messages for sperm-associated antigen 6, a kinocilia marker, and for hydin, a protein linked to ciliary function and hydrocephalus. In testis, ependyma and respiratory epithelium, the Wdr16 protein is up-regulated together with kinocilia formation. The wdr16 gene is restricted to genera in possession of kinocilia, and it is strongly conserved during evolution. The human and zebrafish proteins are identical in 62% of their aligned amino acids. On the message level, the zebrafish Wdr16 ortholog was found exclusively in kinocilia-bearing tissues by in situ hybridisation. Gene knockdown in zebrafish embryos by antisense morpholino injection resulted in severe hydrocephalus formation with unaltered ependymal morphology or ciliary beat. Wdr16 can be considered a differentiation marker of kinocilia-bearing cells. In the brain, it appears to be functionally related to water homeostasis or osmoregulation

Comparison of lactate transport in astroglial cells and monocarboxylate transporter 1 (MCT 1) expressing Xenopus laevis oocytes. Expression of two different monocarboxylate transporters in astroglial cells and neurons

The transport of lactate is an essential part of the concept of metabolic coupling between neurons and glia. Lactate transport in primary cultures of astroglial cells was shown to be mediated by a single saturable transport system with a Km value for lactate of 7.7 mM and a Vmax value of 250 nmol/(min x mg of protein). Transport was inhibited by a variety of monocarboxylates and by compounds known to inhibit monocarboxylate transport in other cell types, such as alpha-cyano-4-hydroxycinnamate and p-chloromercurbenzenesulfonate. Using reverse transcriptase-polymerase chain reaction and Northern blotting, the presence of mRNA coding for the monocarboxylate transporter 1 (MCT1) was demonstrated in primary cultures of astroglial cells. In contrast, neuron-rich primary cultures were found to contain the mRNA coding for the monocarboxylate transporter 2 (MCT2). MCT1 was cloned and expressed in Xenopus laevis oocytes. Comparison of lactate transport in MCT1 expressing oocytes with lactate transport in glial cells revealed that MCT1 can account for all characteristics of lactate transport in glial cells. These data provide further molecular support for the existence of a lactate shuttle between astrocytes and neurons