Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) Stimulates Adenylyl Cyclase and Phospholipase C Activity in Rat Cerebellar Neuroblasts

Abstract: The presence of receptors for the novel neuropeptide pituitary adenylate cyclase-activating polypeptide (PACAP) has been recently demonstrated in the external granule cell layer of the cerebellum, a germinative matrix that generates the majority of cerebellar interneurons. In the present study, we have taken advantage of the possibility of obtaining a culture preparation that is greatly enriched in immature cerebellar granule cells to investigate the effect of PACAP on the adenylyl cyclase and phospholipase C transduction pathways. The two molecular forms of PACAP, i.e., 27-(PACAP27) and 38-(PACAP38) amino-acid forms of PACAP, induced a dose-dependent stimulation of cyclic AMP production in granule cells. The potencies of PACAP27 and PACAP38 were similar (ED₅₀ = 0.12 ± 0.01 and 0.23 ± 0.07 n M , respectively), whereas vasoactive intestinal polypeptide (VIP) was ∼100 times less potent. PACAP27 and PACAP38 also induced a dose-dependent stimulation of polyphosphoinositide breakdown (ED₅₀ = 19.1 ± 6.3 and 13.4 ± 6.0 n M , respectively), whereas VIP had no effect on polyphosphoinositide metabolism. The effect of PACAP38 on inositol phosphate formation was significantly reduced by U-73122 and by pertussis toxin, indicating that activation of PACAP receptors causes stimulation of a phospholipase C through a pertussis toxin-sensitive G protein. In contrast, forskolin and dibutyryl cyclic AMP did not affect PACAP-induced stimulation of inositol phosphates. Taken together, the present results demonstrate that PACAP stimulates independently the adenylyl cyclase and the phospholipase C transduction pathways in immature cerebellar granule cells. These data favor the concept that PACAP may play important roles in the control of proliferation and/or differentiation of cerebellar neuroblasts.     

Performances of a full-scale novel multiplate anaerobic reactor treating cheese whey effluent

A 450-m(3) multiplate anaerobic reactor (MPAR) has been started-up in April 1992 for treating wastewater (whey permeate and domestic wastewater) at the Nutrinor (Lactel) cheese factory in Chambord (Québec, Canada). The MPAR consists of four superimposed sections. The liquid flows upwards from one section to the next, while the gas is collected below each plate and evacuated through side-outlets. The wastewater is concurrently distributed at the bottom of the first, second, and third sections, as 50%, 33%, and 17% of the total influent stream, respectively. Granular anaerobic sludge at an initial concentration of 30 kg of volatile suspended solids (VSS) per cubic meter of reactor liquid volume was used to inoculate the reactor. Under normal operation of the factory, the chemical oxygen demand (COD) concentration of the influent ranged from 20 to 37 kg COD m(-3). The reactor organic loading rate (OLR) fluctuated between 9 and 14.7 kg COD m(-3) d(-1) for hydraulic retention times (HRT) maintained between 55 and 68 h. At the highest OLR, the MPAR showed an efficiency of 98% and 92% for soluble and total COD removal, respectively, and a methane production rate averaging around 4 m(3) m(-3) d(-1).Biomass-specific activities ranged between 7 and 51, 1.3 and 8.5, 5.3 and 12.2, 60 and 119, and 119 and 211 mmol g(-1) VSS d(-1) for glucose, propionate, acetate, formate, and hydrogen, respectively. Average equivalent-diameter of the granules was around 0.65 mm. The MPAR reactor generally showed a large capacity for solid retention with a biomass content between 32 and 37 kg VSS m(-3). (c) 1995 John Wiley & Sons, Inc.     

Polyamines in Human Brain: Regional Distribution and Influence of Aging

Abstract: Depolarization of adult rat forebrain slices with veratrine induced the release of excitatory amino acids (glutamate and aspartate), the synthesis of nitric oxide (NO), and increases in cyclic GMP (cGMP). The NO synthase inhibitors N ^ω-monomethyl- l -arginine and N ^ω-nitro- l -arginine methyl ester decreased the release of NO and the levels of cGMP without affecting the release of excitatory amino acids. In contrast, the antiepileptic drug lamotrigine inhibited the release of excitatory amino acids and of NO, and decreased the levels of cGMP without causing a significant direct inhibition of the NO synthase. Furthermore, the synthesis of NO and the increases in cGMP induced by veratrine were partially blocked by the N -methyl- d -aspartate (NMDA) receptor antagonist MK-801 but not by 6-nitro-7-sulphamobenzo( f )quinoxaline-2,3-dione, a non-NMDA receptor antagonist. Neither of these compounds inhibited directly the NO synthase or the release of excitatory amino acids. Thus, these three types of compound act as an inhibitor of voltage-sensitive sodium channels (lamotrigine), as a receptor antagonist (MK-801), or as direct inhibitors of the NO synthase, to block the pathway leading to increased cGMP after veratrine depolarization. It is likely that some of the pharmacological and therapeutic actions shared by these three types of compound are, at least in part, a consequence of inhibition of the synthesis of NO.     

Physicochemical characterization and in vitro interaction with brain capillary endothelial cells of artificially monoacylated ribonucleases A

Acylated proteins play a crucial role in cellphysiology because of their increased interaction withmembranes. Their isolation is difficult as aconsequence of their low cellular concentration andtheir chemical preparation is problematic due tosolubility problems. Through the use of reversedmicelles, we produced tens of milligrams of acylatedribonucleases A, chosen as a model, purified them bysemi-preparative high performance liquidchromatography (HPLC) and characterized them by analyticalHPLC, capillary electrophoresis, mass spectrometry, peptide mapping, Edman degradation and enzyme activity. We nextscrutinized the interaction with an in vitro blood–brainbarrier model and demonstrated that palmitoylated andstearoylated ribonucleases A are transported from onecompartment to the other across the cellular monolayer,in contrast to the native enzyme.     

Propagation of prion strains through specific conformers of the prion protein.

Two prion strains with identical incubation periods in mice exhibited distinct incubation periods and different neuropathological profiles upon serial transmission to transgenic mice expressing chimeric Syrian hamster/mouse (MH2M) prion protein (PrP) genes [Tg(MH2M) mice] and subsequent transmission to Syrian hamsters. After transmission to Syrian hamsters, the Me7 strain was indistinguishable from the previously established Syrian hamster strain Sc237, despite having been derived from an independent ancestral source. This apparent convergence suggests that prion diversity may be limited. The Me7 mouse strain could also be transmitted directly to Syrian hamsters, but when derived in this way, its properties were distinct from those of Me7 passaged through Tg(MH2M) mice. The Me7 strain did not appear permanently altered in either case, since the original incubation period could be restored by effectively reversing the series of passages. Prion diversity enciphered in the conformation of the scrapie isoform of PrP (PrP(Sc)) (G. C. Telling et al., Science 274:2079-2082, 1996) seems to be limited by the sequence of the PrP substrates serially converted into PrP(Sc), while prions are propagated through interactions between the cellular and scrapie isoforms of PrP.     

Modelling of fluid-phase endocytosis kinetics in the amoebae of the cellular slime mouldDictyostelium discoideum. A multicompartmental approach

Fluid-phase endocytosis (pinocytosis) kinetics were studied inDictyostelium discoideum amoebae from the axenic strain Ax-2 that exhibits high rates of fluid-phase endocytosis when cultured in liquid nutrient media. Fluorescein-labelled dextran (FITC-dextran) was used as a marker in continuous uptake- and in pulse-chase exocytosis experiments. In the latter case, efflux of the marker was monitored on cells loaded for short periods of time and resuspended in marker-free medium. A multicompartmental model was developed which describes satisfactorily fluid-phase endocytosis kinetics. In particular, it accounts correctly for the extended latency period before exocytosis in pulse-chase experiments and it suggests the existence of some sorts of maturation stages in the pathway.