Mitochondrial DNA (mtDNA) mutations have been found associated with several neuromuscular diseases. In order to address further mechanisms of pathogenesis of the specific mtDNA mutations in responsible for these diseases, the ρ° human cell culture system has been established and has been widely used in recent years (4)'(5). This system is based upon the repopulating of human cells, which are completely depleted of mtDNA (termed ρ° cells), with exogenous mitochondria containing the mtDNA of interest. Molecular genetic and biochemical analysis showed that transformants can be obtained with various mitochondrial donors and the respiratory competence can be restored. In this study, the ultrastructural and immunocytochemical variations of mitochondria in human cell lines cultured in this system observed by transmission electron microscopy are reported.

Cell cultures for wild-type 143B, ρ206, and transmitochondrial cell lines have been described previously. Cells were trypsinized and collected from individual culture plates, and then were fixed with 3% paraformaldehyde and 0.04% glutaradehyde in 0.1 M phosphate-buffered saline (PBS) with 4% sucrose (pH 7.6) at RT.

The gregarine Pterospora is a parasite found in the body cavities of bamboo worms (Polychaeta: Maldanidae). The gamont stage of Pterospora has a bizarre structure with a main cell body and multiple posterior cytoplasmic extensions (Fig. 1). The cells are found in pairs within the coelom of the host and move by cytoplasmic streaming as they fill and empty their posterior extensions. Reports of this parasite in the literature are few (see references 1 & 2 for a review) and no ultrastructural details have been published regarding the genus. This study examines the fine structure of the gamont stage with particular emphasis on the structure of the pellicle.

The maldanid worm Axiothella mucosa was collected by shovel in St. Andrew Bay, Florida and returned to the Troy State University campus. Pterospora spp. gamonts were pipetted from minced setigers of the worms and fixed in 3% glutaraldehyde buffered with 0.05M sodium cacodylate, pH 7.5, for 1-6 hours.

Aureobasidins are cyclic depsipeptide natural products that have been shown to be fungicidal against a wide variety of fungi. We have studied the effect of an aureobasidin-like compound, SCH 56301 on the morphology and cytology of yeast cells. For both, Candida albicans and Saccharomyces cerevisiae, the normal cell cycle is disrupted by sub-lethal concentrations of the compound. In Saccharomyces, cells accumulate at the G2/M border of the cell cycle, DNA replication is essentially complete and a short nuclear spindle is visible; however, nuclear migration and spindle elongation do not occur and consequently a small round daughter cell remains attached to the mother cell. Changes in cell wall morphology are visible at old bud scars remaining on mother cells. Under the conditions of preparation of the specimens, there may have been insufficient cross-linking during treatment with SCH 56301 so that intracellular pressure appears to be pushing on a weakened cell wall at these points.

Monilinia vaccinii-corymbosi is the cause of mummy berry disease of commercial blueberries (Vaccinium spp.) grown in the United States and Canada (1). Ovaries of blueberry flowers are infected by the conidia (asexual spores) of this pathogen. However, despite their importance in disease initiation, little is known of the ultrastructural features of these conidia. The only ultrastructural data on these spores has come from the work of Batra (2) who used scanning electron microscopy to describe the morphology of the so-called “disjunctors” which connect adjacent conidia of M. vaccinii-corymbosi. The presence or absence of disjunctors is an important taxonomic character in the genus Monilinia (1) but little is known of their development and exact nature. The objective of this study was, therefore, to elucidate details of conidium and disjunctor development in M. vaccinii-corymbosi.

The Foraminifera are an enormously successful group of predominantly marine protists. They first appeared in the rock record >500 million years ago and currently are found in most planktonic and benthic marine and marginal marine environments. Though complete life cycles are known for only ˜30 of the >10,000 described extant species, these taxa illustrate reproductive patterns that are among the most diverse of all protists. The foraminiferal life cycle typically is characterized by an alternation of sexual and asexual generations during which meiosis occurs in the early phases of schizogony in the agamont. Sexual reproduction involves the production of biflagellated, triflagellated, or amoeboid gametes, and fertilization may be gametogamous, gamontogamous, or autogamous. Of these options, most Foraminifera for which sexual reproduction has been documented (44 species) produce many thousands of small biflagellated gametes that are shed directly into the surrounding seawater where gametogamous fertilization occurs.

The average somatic nucleus is only 10 microns in diameter, yet contains 2 meters of DNA coiled within its volume. This compaction must be accomplished in such a way so that the DNA remains accessible to the machinery of transcription, replication and repair. Sperm nuclei also have compacted DNA; however, structurally and functionally the arrangement of DNA in sperm is vastly different from that of somatic DNA. Somatic DNA is initially compacted by coiling around histone octamers, while sperm DNA is bound to elongate, highly basic proteins called protamines. Protamines bind the major groove of the double helix and allow adjacent DNA strands to come in close contact by neutralizing the negative charge of the sugar-phosphate backbone. Figure 1 illustrates the two most widely accepted models of DNA compaction in both somatic and sperm nuclei.

The arrangement of DNA within sperm nuclei is difficult to study because it is so highly compressed.

Plant vascular system is the principal means to carry water and food throughout the plant body. It is composed of two types of conducting vascular tissues, xylem and phloem. Xylem carries water and minerals from the root to the shoot, whereas phloem transports photosynthates from leaves to other parts of the body. The evolution of vascular system which solved the problem of water and food transport was considered to be one of the key events for the successful emergence of vascular plants on the land from aquatic environments. Although vascular tissues in almost all vascular plants consist of xylem and phloem, diverse arrangements of vascular tissues within the bundles and of vascular bundles in the stele were evolved (1). The occurrence of diverse vascular patterns in vascular plants offers an excellent opportunity to study the evolutionary mechanisms controlling pattern formation. In this report, we present our studies on the vascular differentiation and pattern formation in the inflorescence stems of Arabidopsis thaliana.

The formation of abnormal mitosis associated with cancer has been intriguing for many decades. While microtubules had been the focus of previous studies, recent research has focused on centrosomes, microtubule organizing centers which organize the mitotic apparatus during cell division. During normal mitosis centrosomes form two poles but in cancer, centrosomes can form three, four, or more poles, and organize tripolar, quadripolar, and multipolar mitoses, respectively. This has severe consequences for genomic stability because chromosomes are separated unequally to three, four, or more poles. This can result in aneuploidy and gene amplifications with multiple defects in cellular regulation. It can result in malignancy that is accompanied by cell cycle imbalances and abnormal cell proliferation. While radiation and chemical agents are known to damage DNA and can lead to cell cycle abnormalities, the damage of centrosome structure leading to abnormal mitosis deserves also consideration.

Spironolactone bodies (SB) were first described in 1963 by Janigan. These laminated, whorled structures are seen in cells of the adrenal zona glomerulosa in patients treated with the drug spironolactone. Spironolactone is an aldosterone antagonist. Hyperaldosteronism may result from excess production by the adrenal cortex. By both light microscopy and transmission electron microscopy (TEM), SB have a distinctive, laminated appearance. Kovacs, et al. observed that SB are composed of cellular constituents. To our knowledge, SB have not been analyzed using scanning electron microscopy (SEM) and electron probe microanalysis technology.

An adrenal gland with a 1 cm cortical mass was removed from a 39 year old female and received in 10% buffered formalin. Histologic examination of the mass showed a monotonous population of cells recapitulating zona glomerulosa cells. Intracytoplasmic structures compatible with SB were identified. Portions of the adrenal gland were processed for TEM and SEM analysis.

Our goal was to determine the cellular localization of left-handed Z-RNA, within preeguatorial zone (PZ) epithelium of the normal adult dog ocular lens (1.5 yr) (Fig. 1), employing anti-Z-RNA IgG polyclonal antibodies. B-DNA has the ability to adopt the Z-DNA configuration in vitro(1). A-RNA can be transformed into Z-RNA under certain conditions (2). Z-RNA has been localized in cultured cells (3). Strong evidence supports the presence of Z-DNA in vivo (1). Elimination of DNA binding proteins by certain fixatives can initiate DNA supercoiling which stabilizes Z-DNA sequences (1). Z-DNA may play a role in regulating in vivo transcriptional enhancement (1).

Anti-Z-RNA antibody probes were produced in 3 rabbits immunized with injections of Z-RNA: Br-poly[ribosomal(G-C)]. Concerning light microscopy [immunohistochemistry (ABC method)], lens tissues were fixed in Carnoy's, embedded in paraffin and sectioned (3 μm) (Fig. 2). Image analysis was performed using a Leitz DM-RB microscope and Leica Quantiment 500 + image analyzer.

The objective of this study was to evaluate the biological responses of three different mixtures of a bone substitute material placed in tooth extraction sites to determine which mixture best enhances bone formation. Under general anesthesia three adult mongrel canine had bilateral extractions of the second and fourth premolar bicuspids. Each site was modeled into 3-wall defect with box-like dimensions and having the window on the buccal (outer) side of the mandible. A synthetic hydroxyapatite bone cement (HAC) was prepared three different ways: 1) 2.5 gms HAC + 0.75 cc sterile water (W); 2) 2.5 gms HAC + 0.75 cc 2% Hydroxypropylmethylcellulose in sterile water (HPMC); 3) 2.5 gms HAC + 0.75 cc Sodium Phosphate [45mM/15mL Phosphate + 60 MEq/15mL Sodium] (SP). Each mixture was syringe injected into a different extraction site and the fourth site received no cement to serve as a control site, and after sites were filled, a periodontal flap closure was accomplished with a 4-0 VICRYL suture.

Aminoacyl-tRNA synthetases are the family of enzymes responsible for correctly linking tRNA molecules with amino acids so that the latter may be subsequently incorporated into growing polypeptide chains. A unique feature of aminoacyl-tRNA synthetases from higher eukaryotes is the existence of a high molecular mass multisynthetase complex. This approximately 1 × 106 Da particle measures approximately 27 nm in diameter and appears to be a cup or elongated “U“ shape as suggested by distinct orientations visible in negatively stained electron micrographs. A working model of the structure has been proposed which divides the particle into three domains. The composition of each domain as well as relative positions of individual polypeptides within each have been proposed, but to date there has been no precise localization of any specific component or site within the multisynthetase complex. This study describes the first localization of a particular polypeptide within the particle.

In principle, electron energy loss spectroscopy (EELS) in the scanning transmission electron microscope (STEM) should be able to detect single atoms of phosphorus in biological macromolecules. Such a capability would provide a direct method for measuring the degree of phosphorylation or nucleotide binding which are important in determining the functional states of protein assemblies. Although it has already been possible to map heavy elements, e.g., uranium, at near single atom sensitivity, this has not yet been achievable for lighter atoms like phosphorus. One difficulty in imaging light atoms in a biological specimen is that there is insufficient contrast to visualize the atoms directly so, in a stationary probe analysis, it is not known where to place the probe. Identifying single phosphorus atoms therefore necessarily involves compositional mapping in which EELS information is collected pixel by pixel, a technique known as spectrum-imaging. Recently, improvements have been made in the acquisition of such images, especially the ability to correct for specimen drift in a robust fashion.

Huntington’s Disease (HD) is a progressive neurodegenerative disease that is characterized by loss 19 of motor function and a decline in cognitive functions. HD is characterized morphologically by loss of neurons in the basal ganglia. During the progression of HD, striatal projection neurons die but cholinergic and somatostatin-containing intemeurons survive. The mutated gene for HD has been described and the protein transcribed by the disease (huntingtin or HT) described. Within the striatum, there is a lack of correlation between presence of HT and neuronal vulnerability. HT has been shown to be associated with intracellular vesicles in human tissues.

Female rats were transcardially perfused with 4% paraformaldehyde, 0.1 M lysine, 10 mM sodium periodate (PLP) in 0.1 M phosphate buffer. Following fixation, the brain was removed and the cortex and striatum removed from the brain as 5 mm3 pieces and transferred to fresh PLP containing 0.25% glutaraldehyde.

For the past ten years, research in our laboratory has focused on the synaptic circuitry of GABAergic and glutamatergic networks in the monkey basal ganglia, a group of subcortical brain structures involved in the control of motor and psychoaffective behaviors. Elucidating the distribution and synaptic localization of different subtypes of glutamate and GABA receptors and transporters is essential to understand the basic functioning of these brain structures and develop better therapies for neurodegenerative diseases such as Parkinson's and Huntington's diseases. Although immunoperoxidase labeling allows to localize receptors at the cellular level, this method is generally not suitable for quantification and exact synaptic location of receptors due to the diffuse nature of the peroxidase reaction product. Here, we describe a highly sensitive post-embedding immunogold technique on Lowicryl-embedded ultrathin sections to analyze the subsynaptic distribution of these receptors at the electron microscope level.

The cerebellar cortex of albino mice, hamsters, teleost fishes, primates and human have been examined by correlative microscopy to study the Golgi cell soma, dendritic processes, axonal plexus and synaptic connections in the granular and molecular layers. For light microscopy (LM) toluidinc blue stained-plastic embedded scmithin sections and Golgi light microscopy preparations were used. For confocal laser scanning microscopy (CLSM) of hamster cerebellum the FM4-64 fluorescent stain was used as intracellular tracer (1). Conventional and high resolution scanning electron microscopy (SEM) of teleost fishes, primates and human were coated with gold-palladium and chromium (2). I Transmission electron microscopy (TEM). either by ullrathin sections or frccze-clching replicas, were examined to characterize synaptic connections in the granular and molecular layers. The Golgi cells appeared in the granular layer as polygonal, stellate, round or fusiform microncurons. 10-25 μm in maximal dimension, surrounded by the granule cell groups. Golgi light microscopy.

Borrelia burgdorferi is the causative agent of Lyme disease. In addition to the characteristic effects of acute infection, Lyme disease can cause chronic effects including cardiac and neurological complications [1]. Several laboratories have shown the ability of B. burgdorferi to attach and invade several cell types such as lymphocytes [2] and fibroblasts [3]. Cultured normal human astrocytes were used to investigate the possible interaction between a cell type of the central nervous system and the Lyme disease spirochete.

Initially scanning electron microscopy was used to monitor and characterize the surface interaction between astrocytes and spirochetes. Astrocyte cell cultures were co-incubated with low passage B31 cultures over a time course from 1 hr to 18 hr. The co-cultures were washed extensively to discourage non-specific binding, glutaraldehyde fixed, critical point dried and mounted for field emission scanning electron microscopy. A time dependent association between the cultured astrocytes and B. burgdorferi were found to range from approximately 20% after 1 hr to almost 100% after 18 hr.

The signaling pathways which lead to the wave of photoreceptor cell death in the rd mouse during the second week after birth have yet to be defined. The photoreceptors of these mice develop normally until they begin to elaborate phototransducing outer segments at around postnatal day (P)8. At this point, their differentiation slows as intracellular levels of cGMP rise due to the inherited mutation in the phosphodiesterase-β subunit.

We have used an immunocytochemical approach to investigate potential signaling pathways leading to photoreceptor cell apoptosis in this mutant. The earliest change detected in the photoreceptor layer was an induction of CREB immunoreactivity in photoreceptor nuclei, commencing at P8. Although CREB is generally thought to be constitutively expressed and its function controlled primarily by its phosphorylation, we find that immature photoreceptors of normal mice are immunonegative for CREB until the time of eye opening and the initiation of phototransduction at around P14.

Copepods are an abundant and diverse group of crustaceans. One order of free living copepods, the calanoids, are usually < 3 mm in length, planktonic, and possess very rapid escape responses. These animals dominate planktonic communities and their escape reactions contribute to their success. Although all calanoids respond to hydrodynamic disturbances with an escape jump, minimum reaction times vary among species, ranging from 1.5 to 6 msec. To help us understand the physiological basis underlying the phenomenally short reaction times we initiated a comparative study of the internal structures by using transmission electron microscopy (TEM).

Traditional chemical fixation was adequate for several species of copepods, and sensory structures were clearly shown. In these calanoids, minimum reaction times ranged from 3 msec to 6 msec and can be partially explained by giant axons, the most common way invertebrates increase the rate of conduction of nerve impulses. However, the internal structures of two copepod species, Euchaetarimana and Undinula vulgaris, were distorted (Fig. 1).

Chronic wasting disease (CWD) is a transmissible spongiform encephalopathy (TSE) of mule deer and elk. Affected animals are characterized clinically by a combination of abnormal behavior and gradual loss of body condition. The disease is invariably fatal. Diagnosis of CWD is made by histologic examination of the central nervous system and microscopic lesions of CWD are typical of the TSEs. Brains from CWD-suspect elk and mule deer can be examined by transmission electron microscopy (TEM) for the presence of scrapie-associated fibrils (SAF). Western blot methods identify abnormal prion protein (PrPres) which accumulates in the brains of animals with TSE, including CWD.

Conventional SAF purification procedures for TEM examination of CWD-suspect brain tissue require both high speed and ultracentrifugation steps followed by Proteinase K enzyme treatment. Modifications used in this experiment include early Proteinase K, or Proteinase K and Collagenase treatment prior to high speed centrifugation and the elimination of the ultracentrifugation step.