The multiple forms of brain acetylcholinesterase

Summary The pattern of the multiple forms of the acetylocholinesterase (AChE, E.C. 3.1.1.7) of the rat brain is investigated using polyacrylamide gradient micro-gel electrophoresis with regard to a possible functional importance of this individual forms. The patterns of the AChE-forms of selected regions of the CNS are compared and certain differences could be shown. After increased cholinergic input (into the hippocampus by electrical stimulation of the nc. septi medialis) an aggregation of AChE subunits is detectable. Subletal intoxication with an irreversible inhibitor of AChE is followed by a faster recovery of the smaller forms. A suggestion of a possible functional role of the multiple forms of AChE is discussed.The research reported in this paper was supported by the Ministerium für Wissenschaft und Technik der DDR     

Drought and biodiversity in Grasslands

Summary The local species richness of four different grassland fields fell an average of 37% during a 1988 drought that decreased above-ground living plant mass by an average of 47%. Despite the return to more normal plant mass and precipitation during the next two years, there was no significant recovery in species richness in the 46 permanent plots, suggesting that local species richness became recruitment limited. The drought led to the loss of annual species independent of their abundance. For perennial grasses, perennial forbs, legumes and woody species, the probability of a species being lost from a plot was significantly negatively correlated with its predrought abundance. These results demonstrate that environmentally extreme conditions can limit species richness by causing the local extinction of rare species. Because droughts of this intensity occur about every 50 years in the prairie, periodic drought may have limited prairie diversity. Moreover, if the accumulation of greenhouse gases leads to a more variable or extreme climate, it could cause increased rates of species extinctions.     

Preen oil composition of Pied Flycatchers is similar between partners but differs between sexes and breeding stages

Preen oil, the secretion of the uropygial gland, may be an important source of body odour in birds. By characterizing the chemical composition of preen oil, we can describe the olfactory phenotypes of birds and investigate whether odours could have a function in sexual signalling or other chemical communication. Here we analysed the preen oil of a wild passerine, the European Pied Flycatcher Ficedula hypoleuca, to find out whether it holds socially relevant information. We sampled both the female and male of breeding pairs during nestling rearing to test for sex differences and within-pair similarity. We additionally sampled the females during incubation to test for changes across breeding stages and for individual repeatability of chemical profiles. Pair mates had similar chemical profiles in comparison with other breeding adults. Furthermore, we found evidence for sex differences and for changes across breeding stages. Notably, the preen oil of females was more diverse and more volatile than that of males, and the preen oil secreted by females during incubation was more volatile than that secreted during nestling rearing. However, we found no evidence for individual repeatability of chemical profiles across breeding stages in females. Our results point towards a function of preen oil in sexual signalling, although other functions should not be excluded. Our study is a first step towards understanding the role of odours in the social life of an important avian model species used in the study of mate choice and sexual selection.     

Perforin and Fas act together in the induction of apoptosis, and both are critical in the clearance of lymphocytic choriomeningitis virus infection

In this report we questioned the current view that the two principal cytotoxic pathways, the exocytosis and the Fas ligand (FasL)/Fas-mediated pathway, have largely nonoverlapping biological roles. For this purpose we have analyzed the response of mice that lack Fas as well as granzyme A (gzmA) and gzmB (FasxgzmAxB(-/-)) to infection with lymphocytic choriomeningitis virus (LCMV). We show that FasxgzmAxB(-/-) mice, in contrast to B6, Fas(-/-), and gzmAxB(-/-) mice, do not recover from a primary infection with LCMV, in spite of the expression of comparable numbers of LCMV-immune and gamma interferon-producing cytotoxic T lymphocytes (CTL) in all mouse strains tested. Ex vivo-derived FasxgzmAxB(-/-) CTL lacked nucleolytic activity and expressed reduced cytolytic activity compared to B6 and Fas(-/-) CTL. Furthermore, virus-immune CTL with functional FasL and perforin (gzmAxB(-/-)) are more potent in causing target cell apoptosis in vitro than those expressing FasL alone (perfxgzmAxB(-/-)). This synergistic effect of perforin on Fas-mediated nucleolysis of target cells is indicated by the fact that, compared to perfxgzmAxB(-/-) CTL, gzmAxB(-/-) CTL induced (i) an accelerated decrease in mitochondrial transmembrane potential, (ii) increased generation of reactive oxygen species, and (iii) accelerated phosphatidylserine exposure on plasma membranes. We conclude that perforin does not mediate recovery from LCMV by itself but plays a vital role in both gzmA/B and FasL/Fas-mediated CTL activities, including apoptosis and control of viral infections.     

Agrobacterium phytochrome as an enzyme for the production of ZZE bilins

Photoconversion of phytochrome from the red-absorbing form Pr to the far-red-absorbing form Pfr is initiated by a Z to E isomerization around the ring C-ring D connecting double bond; the chromophore undergoes a ZZZ to ZZE isomerization. In vivo, phytochrome chromophores are covalently bound to the protein, but several examples of noncovalent in vitro adducts have been reported which also undergo Pr to Pfr photoconversion. We show that free biliverdin or phycocyanobilin, highly enriched in the ZZE isomer, can easily be obtained from chromophores bound in a noncovalent manner to Agrobacterium phytochrome Agp1, and used for spectral assays. Photoconversion of free biliverdin in a methanol/HCl solution from ZZE to ZZZ proceeded with a quantum yield of 1.8%, but was negligible in neutral methanol solution, indicating that this process is proton-dependent. The ZZE form of biliverdin and phycocyanobilin were tested for their ability to assemble with Agp1 and cyanobacterial phytochrome Cph1, respectively. In both cases, a Pfr-like adduct was formed but the chromophore was bound in a noncovalent manner to the protein. Agp1 Pfr undergoes dark reversion to Pr; the same feature was found for the noncovalent ZZE adduct. After dark reversion, the chromophore became covalently bound to the protein. In analogy, the PCB chromophore became covalently bound to Cph1 upon irradiation with strong far-red light which initiated ZZE to ZZZ isomerization. Agrobacterium Agp2 belongs to a yet small group of phytochromes which also assemble in the Pr form but convert from Pr to Pfr in darkness. When the Agp2 apoprotein was assembled with the ZZE form of biliverdin, the formation of the final adduct was accelerated compared to the formation of the ZZZ control, indicating that the ZZE chromophore fits directly into the chromophore pocket of Agp2.     

Microinjection of heme oxygenase genes rescues phytochrome-chromophore-deficient mutants of the moss Ceratodon purpureus

 In protonemal tip cells of the moss Ceratodon purpureus (Hedw.) Brid., phototropism and chlorophyll accumulation are regulated by the photoreceptor phytochrome. The mutant ptr116 lacks both responses as a result of a defect in the biosynthesis of phytochromobilin, the chromophore of phytochrome, at the point of biliverdin formation. The rescue of the phototropic response and of chlorophyll synthesis were tested by injecting different substances into tip cells of ptr116. Microinjection was first optimised with the use of fluorescent dyes and an expression plasmid containing a green fluorescent protein (GFP) gene. Injected phycocyanobilin, which substitutes for phytochromobilin, rescued both the phototropic response and light-induced chlorophyll accumulation in ptr116. The same results were obtained when expression plasmids with heme oxygenase genes of rat (HO-1) and Arabidopsis thaliana (L.) Heynh. (HY1) were injected. Heme oxygenase catalyses the conversion of heme into biliverdin. Whereas HY1 has a plastid target sequence and is presumably transferred to plastids, HO-1 is proposed to be cytosolic. The data show that ptr116 lacks heme oxygenase enzyme activity and indicate that heme oxygenases of various origin are active in Ceratodon bilin synthesis. In addition, it can be inferred from the data that the intracellular localisation of the expressed heme oxygenase is not important since the plastid enzyme can be replaced by a cytosolic one. Received: 8 March 1999 / Accepted: 30 July 1999     

Role for Akt3/protein kinase Bgamma in attainment of normal brain size

Studies of Drosophila and mammals have revealed the importance of insulin signaling through phosphatidylinositol 3-kinase and the serine/threonine kinase Akt/protein kinase B for the regulation of cell, organ, and organismal growth. In mammals, three highly conserved proteins, Akt1, Akt2, and Akt3, comprise the Akt family, of which the first two are required for normal growth and metabolism, respectively. Here we address the function of Akt3. Like Akt1, Akt3 is not required for the maintenance of normal carbohydrate metabolism but is essential for the attainment of normal organ size. However, in contrast to Akt1-/- mice, which display a proportional decrease in the sizes of all organs, Akt3-/- mice present a selective 20% decrease in brain size. Moreover, although Akt1- and Akt3-deficient brains are reduced in size to approximately the same degree, the absence of Akt1 leads to a reduction in cell number, whereas the lack of Akt3 results in smaller and fewer cells. Finally, mammalian target of rapamycin signaling is attenuated in the brains of Akt3-/- but not Akt1-/- mice, suggesting that differential regulation of this pathway contributes to an isoform-specific regulation of cell growth.     

Chronically active: activation of microglial proteolysis in ageing and neurodegeneration

One of the microglial cell functions is the removal of modified extracellular proteins in the brain. The connection between protein oxidation, proteolysis, and microglial activation is the topic of this review. The effect of various activation agents on microglial cells with regard to changes in substrate uptake, proteolytic capacity and degradation efficiency of different types of oxidized protein materials is reviewed. It is shown that different activation stimuli initiate substrate-specific modulation for uptake and proteolysis, influencing an array of factors including receptor expression, lysosomal pH, and proteasome subunit composition. Age-related alterations in activation and proteolytic capacity in microglial cells are also discussed. In ageing, proteolytic effectiveness is diminished, while microglial cells are chronically activated and lose the oxidative burst ability, possibly supporting a 'vicious circle' of macrophage-induced neurodegeneration.