The effect of heterologous insertions on gene conversion in mitotically dividing cells in Drosophila melanogaster

We examined the influence that heterologous sequences of different sizes have on the frequency of double-strand-break repair by gene conversion in Drosophila melanogaster. We induced a double-strand break on one X chromosome in female flies by P-element excision. These flies contained heterologous insertions of various sizes located 238 bp from the break site in cis or in trans to the break, or both. We observed a significant decrease in double-strand-break repair with large heterologous insertions located either in cis or in trans to the break. Reestablishing the homology by including the same heterologous sequence in cis and in trans to the double-strand break restored the frequency of gene conversion to wild-type levels. In one instance, an allelic nonhomologous insertion completely abolished repair by homologous recombination. The results show that the repair of a double-strand break by gene conversion requires chromosome pairing in the local region of the double-strand break.     

Electron microscope study of the myofibril partial disintegration and recovery in the mitotically dividing cardiac muscle cells

Summary The ultrastructure of the myocyte at all phases of mitosis as well as of early postmitotic cells has been studied in the myocardia of 14- and 18-day rat embryos and 5- and 7-day old rats. The myofibrils remain unchanged up to the late prophase. In prometaphase the majority of Z-disks in embryo myocyte myofibrils and considerable part of these disks in myofibrils of suckling rats are drastically disintegrated.This is followed by a progressive isolation and scattering of the myofilament bundles and of the whole sarcomeres during the subsequent phases of mitosis. Thick myofilaments seem to be unchanged but thin ones become frequently poorly outlined (mainly in embryos). The sarcoplasmic reticulum, including its typically differentiated subsarcolemmal cisternae, exhibits relatively few changes during mitosis. In the early postmitotic period there is a gradual restoration of contrast-rich Z-bands, interconnecting the previously isolated sarcomeres. Patterns of this process have much in common with early stages of myofibrillogenesis (appearance of subsarcolemmal Z-bodies, formation of skeins of thin filaments etc.). The cleavage furrow formation is either absent or considerably retarded up to the postmitotic period.Behaviour of some other organelles during myocyte mitosis has been described. Possible mechanisms and significance of the observed phenomena are discussed.The author is greatly indebted to the late Professor L. N. Zhinkin for his interest in this work. The valuable technical collaboration of N. V. Seina as well as the assistance of V. M. Semenov in operating the electron microscope are gratefully acknowledged.     

Neurotensin interacts with dopaminergic neurons in rat brain

Neurotensin (NT) injected intracerebroventricularly in rat increases dopamine (DA) turnover in the corpus striatum and nucleus accumbens. Significant increases in 3,4-dihydroxyphenylacetic acid (DOPAC) levels occurred within 15 minutes after injection with peak levels at 60 minutes. The effect on NT on DOPAC and homovanillic acid (HVA) accumulation was dose-dependent at 3–100 μg. NT, like haloperidol, stimulated 3,4-dihydroxyphenylalanine (DOPA) accumulation in striatal neurons, in the presence of DOPA decarboxylase inhibitor, after injection of gamma-butyrolactone (GBL). NT had a similar stimulatory effect on DOPA levels in the accumbens while haloperidol (0.25 mg·kg^?1) had no significant effect in this brain region. NT did not block the inhibitory effect of apomorphine on DOPA accumulation in both the striatum and accumbens, while haloperidol inhibited apomorphine effect in both regions. NT also failed to displace ³H-spiperone from DA receptors and the presence of NT in the binding assay did not alter the ability of DA to displace ³H-spiperone in either brain region. These experiments demonstrate that NT increases DA turnover in both the nigrostriatal and mesolimbic pathways.     

Immunofluorescence evidence for the absence of histone H1 in a mitotically dividing, genetically inactive nucleus

Antibodies directed against whole histone and purified lysine-rich histone H1 extracted from isolated macronuclei of the ciliate Tetrahymena were obtained and conjugated to fluorescein isothiocyanate. The fluorescein-antibody conjugates were used to directly label Tetrahymena cells. Both macro- and micronuclei were visibly fluorescent in cells stained with anti-whole histone conjugate. However, the anti-H1 conjugate only labeled macronuclei. This in situ demonstration of the lack of positive immunofluorescent staining of micronuclei with anti-H1 conjugate provide further evidence for the absence of H1 in the genetically inactive, mitotically dividing Tetrahymena micronucleus.     

Localization of GRF-like immunoreactive neurons in the rat brain

The localization of human GRF1-44-immunoreactive neurons was studied in the rat brain. A dense accumulation of GRF-containing fibers was noted in the external layer of the median eminence. Cell bodies were observed in colchicine-treated rats. The most intensely fluorescent cluster of cells was contained in the arcuate nucleus. Other cells were seen on the base of the hypothalamus, within the median forebrain bundle, dorsal and ventral aspects of the ventromedial nucleus, zona incerta and dorsal part of the dorsomedial nucleus. These cells may influence the pulsatile release of pituitary growth hormone.     

Preparation of neurons and glial cells from rat brain by zonal centrifugation

A method for routine preparation of cell fractions from rat brain is described. A suspension of undamaged cells was obtained by means of a combined enzymatic and mechanical procedure. The cell yield was about 25% of brain DNA.     

Ultrastructure of the surface of the epithelial cells in the rat retina

Summary The epithelial cells in the retina of rats were examined by scanning and transmission electron microscopy.The apical surface of the epithelial cells is covered by a large number of microvilli, which are embedded in an extracellular material. Distinct holes remaining after detached photoreceptor cell segments are numerous. The cells are polyhedronal, usually hexagonal and holes are scattered in their plasma membrane. Many of the epithelial cells are binucleated. The borders between adjacent cells are always close to each other. Numerous cytoplasmic folds or foot processes occur at the base of the cells as well as an extracellular space.The characteristic surface structures are discussed in relation to the function of the retinal epithelial cells.Supported by grants from the Swedish Medical Research Council (B70-12X-2543-02) and Riksföreningen mot Cancer (69171, 265-B69-01X).     

Ultrastructure of distal nephron cells in rat renal cortex

Distal nephron segments in the rat renal cortex contain distal convoluted tubule cells (DCT cells), connecting tubule cells (CNT cells), intercalated cells (I cells), and principal cells (P cells). The present study was carried out to expand present knowledge on the ultrastructure of these cells. The cells were sampled from superficial cortex and analyzed by electron microscopy. Several morphometric parameters were determined and statistical comparison between cell types was performed. Significant structural differences between the cell types were demonstrated. DCT cells showed the highest volume density of mitochondria whereas the amplification of basolateral membranes was higher in CNT cells than in I and P cells. The surface density of the membrane that bounds intermediate vesicles in the apical cytoplasm was twofold higher in I cells than in the other cell types. The morphological differentiation found in the present study adds to available evidence indicating a functional differentiation between the cell types and provides a reference for structure-function correlations in these cells.     

The renin-angiotensin system in the rat brain. Immunocytochemical localization of angiotensinogen in glial cells and neurons

The distribution of angiotensinogen containing cells was determined in the brain of rats using immunocytochemistry. Specific angiotensinogen immunoreactivity is demonstrated both in glial cells and neurons throughout the brain, except the neocortical and cerebellar territories. Positive neurons are easily and invariably detected in female brains, and haphazardly in male brain (sex hormone dependent). Angiotensinogen immunoreactivity in male brain neurons can be induced by water deprivation or binephrectomy in some areas and particularly in paraventricular nuclei. Finally, the highest concentrations of positive neurons are found in the anterior and lateral hypothalamus, preoptic area, amygdala and some well known nuclei of the mesencephalon and the brainstem. Our results confirm the wide distribution of angiotensinogen mRNA in the brain reported recently by Lynch et al. (1987). Thus the demonstration of angiotensinogen in neurons and glial cells allows a greater understanding of the biochemical and physiological data in accordance with multiple brain renin angiotensin systems.     

Ultrastructure of the interstitial cells of the renal papilla in the white rat

Interstitial cells of the renal papilla have the phenotypic appearance, being more of a fibroblast than of a histiocyte character. Interstitial cells differ from the "typical fibroblasts" in the number of lipid droplets, in the development of cisternae of the Golgi complex and rough endoplasmic reticulum, and in the number of lysosome-like bodies and mitochondria.     

DNA topoisomerase I from rat brain neurons

The DNA topoisomerase I has been isolated from neurons of rat cerebral cortex. The most homogeneous fraction purified contains only one polypeptide of Mr approx. 100 000. The enzyme relaxes supercoiled DNA in the absence of ATP or Mg2+. The optimum monovalent cation concentration for the relaxation of superhelical DNA under conditions of DNA excess is found to be 175-200 mM. The neuron enzyme is similar to other mammalian type I DNA topoisomerases in that it links to the 3' ends of the broken DNA strands. Like calf thymus DNA topoisomerase I, the neuron topoisomerase can be selectively inhibited by poly(dG) but not by other homopolymerical deoxyribonucleotides.     

Transport of thymidine during the cell cycle in mitotically synchronized CHO cells

The kinetics of total uptake of thymidine into the cell were determined for cells which had been mitotically synchronized, plated into scintillation vials and pulsed with five concentrations of [³H]-thymidine at various times during the cell cycle. From Lineweaver-Burk plots of these rates, V_max and K_m values were determined for the transport of thymidine. The V_max values ranged from a low of 2.0 pmoles/ min/10⁶ cells in mid-G1 to a high of 99.7 in mid-S before a decline in late S and G2. K_m values displayed only a 5-fold range in values.     

Ultrastructure of venules in the cat brain

Summary Intracerebral venules of the cat were examined to establish criteria for a distinct separation between the venous and arterial system, and to characterize, in greater detail, the mural construction of individual venules. The intracerebral venules were compared with those of other organs. Venules do not have a vascular wall composed clearly of endothelium, media, and adventitia, as is characteristic of arteries and arterioles. The venous endothelium has a similar structure to that of capillaries. The periendothelial cells of the venule differ in shape depending on the vascular diameter. The number of periendothelial cell processes in postcapillary venules increases progressively. Segments in which the basal lamina of the endothelium merges with that of the glia cover a smaller portion of the circumference than in venous capillary loops. In collecting venules, the endothelium is almost completely enveloped by periendothelial cells which have a larger number of filaments. There are no typical smooth muscle cells in the intracerebral venules. The perivascular space becomes wider in collecting venules, contains adventitial cells, phagocytes and a great number of collagen fibers.     

Ultrastructure of arterioles in the cat brain

Summary A total of 110 arterioles were examined in the brains of cats; different sites were studied including the cortex, putamen, pons and crus cerebri. No internal elastic laminae were seen in the subendothelial space, although occasional fragments of elastic material were present in the larger arterioles. The media was composed of one, two or three layers of smooth muscle cells which interlocked in such a way that the vessel wall thickness was constant. Numerous tight junctions were seen between adjacent smooth muscle cells and between the endothelium and smooth muscle cells. Apart from the usual cell organelles, the smooth muscle cells of arterioles had numerous dense patches on the cell surface. The structure of the adventitia varied according to the diameter of the vessel and the site in the brain; it contained adventitial cells, bundles of collagen fibres and nerve fibres. Innervation of arterioles was more constant in the brain stem than in the cortex. Metarterioles had less specialised, atypical smooth muscle cells, a discontinuous media and numerous, extensive myoendothelial tight junctions; they were not innervated by nerve fibres. The diameter of metarterioles was less than 10 m whereas that of arterioles was 10–45 m. The possible functional aspects of arteriolar innervation are discussed.