Integrated palaeoecological and historical data in the service of fine-resolution land use and ecological change assessment during the last 1000 years in Rõuge, southern Estonia

Aims Our aim is to reconstruct decadal scale development of historical landscapes during the last 1000 years by means of fossil pollen analysis of annually laminated lake sediments, and detailed historical maps and documents. Location Lake Rõuge Tõugjärv (Estonia), a small lake with annually laminated lake sediments situated in a dense prehistoric setting. Methods The chronology of the palaeodata is based on the annual laminations supported by AMS ¹⁴C and ²¹⁰Pb dating and ¹³⁷Cs, ²⁴¹Am, and spheroidal carbonaceous particle marker horizons. The time‐scale and resolution allows fine sampling (the pollen samples generally comprise 3.5 years) and vegetation change reconstruction. Relevant source area of pollen (RSAP) of the lake was estimated, and the statistical zonation, rate of change, palynological richness, and DCA and PCA ordinations were generated on the basis of the pollen data. The historical calibration data set (maps, numerical information on population, domestic stock, farmland division, etc.) is based on archival material preserved in the Estonian Historical Archives. Results The topmost part (0–180 cm) of the sediment column of Lake Rõuge Tõugjärv, covering the last 1000 years, is visibly laminated carbonaceous gyttja. The varve chronology extends from ad 2000 to ad 1339, with a cumulative ± 9‐year error estimate. Beyond this the chronology is extrapolated using the ¹⁴C date and varve age–depth estimations. The simulation of the RSAP of Lake Tõugjärv shows that the major portion of the pollen loading originating from local vegetation is derived from plants growing within 2000 m of the sampling site. The pollen record divides into five statistically significant subgroups, which fall on the PCA plot into three clusters reflecting the general openness–closedness of the landscape. During the period between ad 1000 and 1200 (RT 1) the Rõuge area was generally wooded with birch, spruce and pine forests. The advancement of extensive farming gradually opened up the landscape between ad 1200 and 1650 (RT 2 and RT 3). The maximum openness of the landscape was reached between ad 1650 and 1875 (RT 4), with the most open period in the late eighteenth century. Historical maps from 1684 and 1870–99 and available quantitative data on population, domestic stock, farmland division, etc. show the same trend. The pollen data covering the last 125 years, and maps from 1935 and 1995, show the reduction of arable land in RSAP of the lake under investigation and the reduction of open land to an extent comparable with the end of the seventeenth century. Main conclusions The formation and development of the cultural landscape at Rõuge over the last 1000 years is characterized by rapid changes in floristic richness and rates of vegetation change attributed to certain historic processes in the RSAP. Five phases of landscape and social development are clearly distinguished during the last 1000 years. The decadal scale vegetation response to human‐induced forcing agrees with historical maps and documents and could be used for past landscapes prior to the period with solid historical data.     

The effect of administration of estradiol and testosterone on body growth of young male rats.

The influence of estradiol and testosterone on body growth of young male Wistar rats was investigated. In the first experiment, estradiol was given to intact ad libitum fed male rats at 32, 37 and 42 days of age. Moreover, two untreated groups of animals were used: one was fed restrictedly according to the food intake of animals receiving estradiol and another was fed ad libitum. The animals were sacrificed at 47 days of age. Both untreated groups of animals achieved significantly higher body weight and length of tibia than estradiol treated animals. Also the growth of the tail of untreated animals was more intensive than that of estradiol treated animals. In the second experiment, estradiol was given to intact ad libitum fed male rats at 30, 35 and 45 days of age. Moreover, testosterone was given to a half of these animals at 45, 50 and 55 days of age. The animals were sacrificed at 60 days of age. Administration of testosterone significantly increased the growth of the tail and tibia in comparison to the animals which did not receive testosterone after estradiol administration. The results of the present study show that the inhibitory effect of estradiol on body growth of young male rats is not only the result of decreased food intake and that testosterone can improve the skeletal growth of male rats altered by previously given estradiol.     

Enzyme immobilization via monoclonal antibodies I. Preparation of a highly active immobilized carboxypeptidase A

A novel method for the preparation of highly active immobilized enzymes is described. It is based on the binding of enzymes to suitable carriers via monoclonal antibodies, which bind to the enzyme with high affinity without affecting its catalytic activity. The applicability of the method forwarded has been illustrated by the preparation of two samples of highly active immobilized carboxypeptidase A (CPA) preparations as follows: A mouse monoclonal antibody (mAb 100)to CPA that binds to the enzyme with a high-affinity constant without affecting its catalytic activity was prepared, purified, and characterized. Covalent binding of this monoclonal antibody to Eupergit C (EC) or noncovalent binding to Sepharose-protein A (SPA)yielded the conjugated carriers EC-mAb and SPA.mAb, respectively, which reacted specifically with CPA to give the immobilized enzyme preparations EC-mAb.CPA and SPA.mAb.CPA displaying full catalytic activity and improved stability. At pH 7.5 and a temperature range of 4-37 degrees C an apparent binding constant of approximately 10(8)M(-1) characterizing the interaction of CPA with EC-mAb and SPA.mAb, was obtained. To compare the properties of EC-mAb.CPA and SPA.mAb.CPA with those of immobilized CPA preparations obtained by some representative techniques of covalent binding of the enzyme with a corresponding carrier, the following immobilized CPA preparations were obtained and their properties investigated: EC-CPA (I), a preparation obtained by direct binding of EC with CPA; EC-NH-GA-CPA (II), a derivative obtained by covalent binding of CPA to aminated EC via glutaraldehyde; EC-NH-Su-CPA (III), a CPA derivative obtained by binding the enzyme to aminated EC via a succinyl residue; and EC-HMD-GA-CPA (IV), obtained by binding the enzyme via glutaraldehyde to a hexamethylene diamine derivative of EC. Full enzymic activity for all of the bound enzyme, such as that recorded for the immobilized CPA preparations EC-mAb.CPA and SPA.mAb.CPA, was not detected in any of the insoluble covalently bound enzyme preparations.     

Isolation and characterization of a large,neurite-associated glycoconjugate from neuroblastoma cells

A high molecular weight glycoconjugate has been isolated from neurite-producing neuronal tumor cells in culture and has been designated as I(0) based on its elution characteristics in gel filtration chromatography. This molecule cannot be found in a variety of nonneuronal cells. I(0) is found in the substratum-attached material or cell fraction of neurite-producing neuroblastoma cells, depending upon culture conditions. It is found in the substratum-bound fraction of B104 rat neuroblastoma cells during serum starvation and in the EGTA-detached cell fraction of B104 cells grown in chemically defined N2 medium. It occurs only in the cell fraction of the human neuroblastoma line Platt. Examination of behavioral variants of the B104 rat line further strengthens the association of I(0) with neurite production; the constitutive neurite-producing E(R)B9 variant contains I(0) while the non-neurite-producing E(R)A11 variant does not. I(0) is large, eluting in the void volume of sepharose-CL2B columns. Radioiodination of intact cells with lactoperoxidase shows I(0) to be a cell surface component. Metabolic radiolabeling studies show that it contains a high proportion of polysaccharide to protein, does not contain mannose, and is unsulfated. Alkaline borohydride reduction release two size classes of large polysaccharide chain. The alkaline reduction results, along with the mannose incorporation studies, show the presence of O-glycosidic linkages and few, if any, N-linkages. Resistance to nitrous acid deamination, insensitivity to glycosaminoglycan lyases, and the absence of sulfation, indicate that I(0) does not contain the glycosaminoglycans hyaluronic acid, chondroitin-, dermatan-, or heparin- sulfates. Affinity column chromatography reveals high binding affinity of I(0) to polyornithine and no binding to gelatin (collagen) or the glycosaminoglycans hyaluronate and heparin. These studies describe a unique high molecular weight glycoconjugate on the surface of neurite-producing neuroblastoma cell lines from two species.     

Effect of protease and protease inhibitors on the adhesion of Ehrlich ascites tumour cells to plastic

The adhesion of Ehrlich ascites tumour cells to plastic was inhibited by the addition to the incubation medium of either of two chemically unrelated trypsin inhibitors at non-toxic concentrations. Supplementing the medium with low concentrations of trypsin enhanced adhesion. These findings are compatible with the existence of intrinsic proteolytic activity at the surface of the tumour cell and support the view that such a protease would affect the adhesive characteristics of the cell.     

A single cDNA encodes two isoforms of stathmin, a developmentally regulated neuron-enriched phosphoprotein

Stathmin, a 19-kDa neuron-enriched soluble phosphoprotein, has been recently proposed as an ubiquitous intracellular relay for the diverse extracellular signals regulating cell proliferation, differentiation, and functions through various second messenger pathways (Sobel, A., Boutterin, M.C., Beretta, L., Chneiweiss, H., Doye, V., and peyro-Saint-Paul, H. (1989) J. Biol. Chem. 264, 3765-3772). Internal sequences of the protein from rat brain were determined after purification by two-dimensional polyacrylamide gel electrophoresis, electrotransfer onto Immobilon, and in situ proteolysis. Oligonucleotide mixtures based on these sequences were used to clone a cDNA for stathmin from a rat PC12 cell lambda gt 10 library. The deduced amino acid sequence reveals partial homologies with the coiled coil structural regions of several intracellular matrix phosphoproteins. Using this cDNA as a probe, we show that the expression of stathmin mRNA parallels that of the protein during brain ontogenesis, reaching a maximum at the neonatal stage. In vitro translation of the derived cRNA yielded all the known molecular forms of stathmin, namely its alpha and beta isoforms in their unphosphorylated and phosphorylated states. Thus, a single cDNA codes for both biologically relevant isoforms of the protein, indicating that they differ by co- or post-translational modifications.     

A telescopic method for photographing within 8×8 cm minirhizotrons

The volatile organic compounds produced during a sequence of soil incubations under controlled conditions, with either added NH₄ ⁺-N or NO₃ ^--N, were collected and identified. The nature and relative amounts of the volatile organic compounds produced by the microorganisms in the soils were remarkably reproducible and consistent.     

Relative apparent synapomorphy analysis (RASA). I: The statistical measurement of phylogenetic signal

We have developed a new approach to the measurement of phylogenetic signal in character state matrices called relative apparent synapomorphy analysis (RASA). RASA provides a deterministic, statistical measure of natural cladistic hierarchy (phylogenetic signal) in character state matrices. The method works by determining whether a measure of the rate of increase of cladistic similarity among pairs of taxa as a function of phenetic similarity is greater than a null equiprobable rate of increase. Our investigation of the utility and limitations of RASA using simulated and bacteriophage T7 data sets indicates that the method has numerous advantages over existing measures of signal. A first advantage is computational efficiency. A second advantage is that RASA employs known methods of statistical inference, providing measurable sensitivity and power. The performance of RASA is examined under various conditions of branching evolution as the number of characters, character states per character, and mutations per branch length are varied. RASA appears to provide an unbiased and reliable measure of phylogenetic signal, and the general approach promises to be useful in the development of new techniques that should increase the rigor and reliability of phylogenetic estimates.      

Molecular evolution of voltage-sensitive ion channel genes: on the origins of electrical excitability

We have analyzed nucleic acid and amino acid sequence alignments of a variety of voltage-sensitive ion channels, using several methods for phylogenetic tree reconstruction. Ancient duplications within this family gave rise to three distantly related groups, one consisting of the Na+ and Ca++ channels, another the K+ channels, and a third including the cyclic nucleotide-binding channels. A series of gene duplications produced at least seven mammalian homologues of the Drosophila Shaker K+ channel; clones of only three of these genes are available from all three mammalian species examined (mouse, rat, and human), pointing to specific genes that have yet to be recovered in one or another of these species. The Shaw-related K+ channels and the Na+ channel family have also undergone considerable expansion in mammals, relative to flies. These expansions presumably reflect the needs of the high degree of physiological and neuronal complexity of mammals. Analysis of the separate domains of the four-domain channels (Ca++ and Na+) supports their having evolved by two sequential gene duplications and implies the historical existence of a functional two-domain channel.