Paleobotanical studies inFagaceae of the European Tertiary

A systematic reassessment of megafossil records ofFagaceae in Central Europe has been undertaken on the basis of leaf cuticular characters. The oldest representatives date back to the Eocene:Quercus subhercynica spec. nova,Dryophyllum furcinerve (Rossm.)Schmalh.,Trigonobalanopsis rhamnoides (Rossm.) gen. & comb. nov. In the Oligocene other members of extant genera appear:Quercus rhenana (Weyl. & Kilpp.)Knobloch & Kvaek,Fagus attenuata Goepp.,Lithocarpus saxonicus spec. nova. In the Neogene these ancient taxa (except inFagus lineage), are gradually replaced by deciduous species ofQuercus andCastanea. Trigonobalanus andCastanopsis are recorded by fruits (or wood) only.     

Computerized ordering of experimental animals and test authorization

The authorization procedure required by law in Switzerland and the internal set-up at Roche for acquiring experimental animals has made a computerized system for monitoring authorizations and animal deliveries essential. The INQUIRE software program, which can be run on the central computer, was used to set-up databases with information on all personnel who place orders and perform experiments (PERI), authorization matters (BEWI), orders (ORDR), deliveries (SPED), animal species (SPEC), animal strains (STRE), populations (POPU) and the management of various data (BARA). The authorizations database (BEWI) permits sequential searches on specific questions. The animals ordered in the ORDR database are constantly updated in BEWI, thus ensuring that the authorized animal quotas are not exceeded. Expiry of an authorization or an unregistered experimenter will come to light in the course of the plausibility study. Through ORDR the experimenter has a good overview of the animals that he has ordered or have been ordered for him, and he can select the most appropriate strain or population for his studies in STRE or POPU, which contain data on the genetic and physiological characteristics as well as the breeding and keeping of all sublines and stocks. Realization of the IFIS project has made it a simple matter to keep a check on the legal requirements pertaining to animal experimentation and to update the information and evaluate the entire stock of data at any time.     

Xp-duplications with and without sex reversal

Duplications in Xp including the DSS (dosage sensitive sex reversal) region cause male to female sex reversal. We investigated two patients from families with Xp duplications. The first case was one of two sisters with karyotype 46,XY, der(22), t(X;22)(p11.3;p11)mat and unambiguous female genitalia. The living sister was developmentally retarded, and showed multiple dysmorphic features and an acrocallosal syndrome. The second case was a boy with a maternally inherited direct duplication of Xp21.3-pter with the breakpoint close to the DSS locus. He had multiple abnormalities and micropenis, but otherwise unambiguous male genitalia. We performed quantitative Southern blot analysis with probes from Xp22.13 to p21.2 to define the duplicated region. Clinical, cytogenetic, and molecular data from both patients were compared with those of previously reported related cases. A comparison of the extragenital symptoms revealed no differences between patients with or without sex reversal. In both cases, the symptoms were non-specific. Among 22 patients with a duplication in Xp, nine had unambiguous female genitalia and a well-documented duplication of the DSS region. Two patients with duplication of DSS showed ambiguous external genitalia. From these data, we conclude that induction of testicular tissue may start in these patients, but that the type of genitalia depends on the degree of subsequent degeneration by a gene in DSS.     

Excess amino acid polymorphism in mitochondrial DNA: contrasts among genes from Drosophila, mice, and humans

Recent studies of mitochondrial DNA (mtDNA) variation in mammals and Drosophila have shown an excess of amino acid variation within species (replacement polymorphism) relative to the number of silent and replacement differences fixed between species. To examine further this pattern of nonneutral mtDNA evolution, we present sequence data for the ND3 and ND5 genes from 59 lines of Drosophila melanogaster and 29 lines of D. simulans. Of interest are the frequency spectra of silent and replacement polymorphisms, and potential variation among genes and taxa in the departures from neutral expectations. The Drosophila ND3 and ND5 data show no significant excess of replacement polymorphism using the McDonald-Kreitman test. These data are in contrast to significant departures from neutrality for the ND3 gene in mammals and other genes in Drosophila mtDNA (cytochrome b and ATPase 6). Pooled across genes, however, both Drosophila and human mtDNA show very significant excesses of amino acid polymorphism. Silent polymorphisms at ND5 show a significantly higher variance in frequency than replacement polymorphisms, and the latter show a significant skew toward low frequencies (Tajima's D = -1.954). These patterns are interpreted in light of the nearly neutral theory where mildly deleterious amino acid haplotypes are observed as ephemeral variants within species but do not contribute to divergence. The patterns of polymorphism and divergence at charge-altering amino acid sites are presented for the Drosophila ND5 gene to examine the evolution of functionally distinct mutations. Excess charge-altering polymorphism is observed at the carboxyl terminal and excess charge-altering divergence is detected at the amino terminal. While the mildly deleterious model fits as a net effect in the evolution of nonrecombining mitochondrial genomes, these data suggest that opposing evolutionary pressures may act on different regions of mitochondrial genes and genomes.      

Ependymal cells in tissue culture

Summary Ciliated ependymal cells from various locations of the ventricular system of mammals showed outgrowth either in the form of epithelial sheets or in the form of pools and elongated double rows, dependant on the site from which the ependyma was taken and on the original orientation of the cells in respect to the cover slip.The ependymal cilia in such cultures were shown to be moving at a rate of 51/2 to 6 times per second. The movement of the cilia of adjacent cells was apparently well coordinated causing the surrounding fluid to flow in a particular direction.The effect of physiological saline, morphine sulfate, ethyl and methyl alcohol on the ciliary motility has been studied in living cells with phase contrast cinematography.Grateful acknowledgement is made to Messers G. Lefeber, E. Pitsinger and J. Carnes for indispensible aid with photographic work. Mrs. J. Finerty and Mrs. W. Hild contributed generously in the management of the tissue cultures and in executing staining procedures.This investigation was supported by a research grant [PHS B-364 (C3)] from the National Institute of Neurological Diseases and Blindness, of the National Institutes of Health, Public Health Service, administered by C. M. Pomerat.     

Evidence for "twisted plane" undulations in golden hamster sperm tails

Motile spermatozoa from the golden hamster have been arrested by rapid freezing and then fixed with glutaraldehyde at low temperature after substitution with ethylene glycol. As far as can be judged, the flagellar waveforms thus stabilized are similar to those seen in living sperm; in contrast, fixation in glutaraldehyde, without prior freezing, induces agonal changes in flagellar conformation. The characteristics waveform after freeze substitution contains three bends. Approx. half of these flagella are entirely planar. The rest are three dimensional, with the third bend displaced in a regular way from the plane containing the proximal two bends. From the geometry of these flagella, it is concluded that the plane of action of a given bending cycle undergoes a clockwise twist (from a forward viewpoint) as the cycle is succeeded by new bending cycles. This "twisted plane" undulation is quite different from helical movement. The twisting seems to occur abruptly, between cycles, as if each bending cycle has a preferred plane of action. The mechanism underlying the twisting is uncertain. However, on the basis of the angular displacements between the preferred planes, and the findings from electron microscopy, the following idea is presented as a working hypothesis: that, if the most proximal plane of bending is topographically determined by peripheral doublet 1, then successive distal planes of action are influenced predominantly by doublets 2, 3, etc., in clockwise sequence. The merits and weaknesses of this hypothesis are discussed.     

Proton transport by bacteriorhodopsin through an interface film

Summary Interface films of purple membrane and lipid containing spectroscopically intact and oriented bacteriorhodopsin have been used as a model system to study the function of this protein. Small positive charges in surface potential (<1 mV) are detected upon illumination of these films at the air-water interface. These photopotentials, are not affected by overlaying the interface film with a thin layer (0.3 mm) of decane. However, they are dramatically increased when lipid soluble proton carriers FCCP or DNP are added to the decane. The polarity of the photopotential indicates that, in the light, positive charges are transported through the interface from the aqueous to the organic phase. The action spectrum of the photopotential is identical to the absorption spectrum of bacteriorhodopsin. Since bacteriorhodopsin molecules are oriented with their intracellular surface towards the aqueous subphase, the characteristics of the photopotential indicate that in the light bacteriorhodopsin translocates protons from its intracellular to its extracellular surface. The kinetics of the photopotential reveal that the rate and extent of proton transport are proportional both to the fraction of bacteriorhodopsin molecules excited and to the concentration of proton acceptor. The photopotentials result from changes in the ionic distribution across the decane-water interface and can be cancelled by lipid soluble anions.     

Aspects of turnover and biogenesis of synaptic vesicles at locust neuromuscular junctions as revealed by zinc iodide-osmium tetroxide (ZIO) reacting with intravesicular SH-groups

Retractor unguis nerve muscle preparations from the locust were subjected to the zinc iodide-osmium tetroxide reaction (ZIO) after pre-fixation in glutaraldehyde. Applied for 18 h at 4 degrees C in the dark, ZIO reacts at pH 4.2--4.0 fairly selectively with the matrix of synaptic vesicles. Approximately 53% of the vesicles are completely and 4% partially stained. The percentage of ZIO-positive vesicles is increased to nearly 90% and reduced to 4% or less by pretreatment with SH-protecting (dithiothreitol) or SH-blocking (N-ethylmaleimide, p-chloromercuriphenyl sulfonic acid) and SH-oxidizing (azodicarboxylic acid-bis-dimethylamide) reagents, respectively. Stimulation of the motor nerve at 20 Hz for 7 min, partially fatiguing synaptic transmission, reduces the number of vesicles per square micrometer of terminal area by approximately 52%; 2 min of rest restores this number of its pre-stimulation level. These changes are chiefly accounted for by changes in the number of completely ZIO-positive vesicles. 2 min after the end of stimulation, partially ZIO-positive vesicles are three times more frequent than before. With all experimental conditions, the average volume of vesicles was as follows: ZIO-negative less than partially ZIO-positive less than completely ZIO-positive. The average volume of ZIO-positive vesicles is almost unaffected by stimulation; that of ZIO-negative vesicles is decreased by 25% immediately after stimulation, increasing with subsequent rest to the initial level after 1 h. It is suggested (a) that ZIO demonstrates intravesicular protein(s) containing SH-groups and (b) that the completely ZIO-positive vesicles represent the mature ones ready to be used for transmitter release. How the ZIO reaction differentiates between different developmental stages of vesicles which could arise from the smooth endoplasmic reticulum is discussed.