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PubMed2001

Estimating the size of the capillary-to-fiber interface in skeletal muscle: a comparison of methods.

Current evidence suggests that the size of the capillary-to-fiber (C/F) interface is a major determinant of O2 flux into muscle fibers, and methods have been developed for estimating the size of this region via the C/F perimeter ratio in perfusion-fixed material (Mathieu-Costello O, Ellis CG, Potter RF, MacDonald IC, and Groom AC. Am J Physiol Heart Circ Physiol 261: H1617-H1625, 1991) and the quotient of the individual, fiber-based C/F number ratio and fiber perimeter (C/F perimeter exchange index) in muscle biopsies (Hepple RT. Can J Appl Physiol 22: 11-22, 1997). The purpose of this study was to compare the two methods and examine how differences in muscle tissue preparation between perfusion fixation and frozen biopsy can influence the estimate of the size of the C/F interface. The left medial gastrocnemius muscle of nine purpose-bred dogs was perfusion fixed in situ, and a sample from the midportion of the midbelly was processed for microscopy. A corresponding sample from the right gastrocnemius muscle obtained by open biopsy in six of the nine animals was frozen for histochemistry. A significant correlation was found between the two estimates of the size of the C/F interface in the same sections of perfusion-fixed material (r = 0.75, P < 0.05). However, estimates of the size of the C/F interface were smaller in biopsies than perfusion-fixed material, and there was no significant relationship between the estimates in the two preparations. This was due to differences in fiber size (33% larger fiber cross-sectional area in biopsy material after normalization for sarcomere length; P < 0.05) and muscle sampling between the two tissue preparations.

Animals↗
PubMed1999

Ultrastructural analysis of ectopic synaptic boutons arising from peripherally regenerated primary afferent fibers.

The central axons of peripherally regenerated Abeta primary sensory neurons were impaled in the dorsal columns of alpha-chloralose-anesthetized cats 9-12 mo after axotomy. The adequate peripheral stimulus was determined, and the afferent fibers intracellularly stimulated while simultaneously recording the resulting cord dorsum potentials (CDPs). Fibers that successfully had reinnervated the skin responded to light tactile stimulation, and evoked CDPs that suggested dorsally located boutons were stained intracellularly with horseradish peroxidase (HRP). Two HRP-stained regenerated Abeta afferent fibers were recovered that supported large numbers of axon collaterals and swellings in laminae I, IIo, and IIi. Sections containing the ectopic collateral fibers and terminals in the superficial dorsal horn were embedded in plastic. Analyses of serial ultrathin sections revealed that ectopic projections from both regenerated fibers supported numerous synaptic boutons filled with clear round vesicles, a few large dense core vesicles (LDCVs) and several mitochondria (>3). All profiles examined in serial sections (19) formed one to three asymmetric axo-dendritic contacts. Unmyelinated portions of ectopic fibers giving rise to en passant and terminal boutons often contained numerous clear round vesicles. Several boutons (47%) received asymmetric contacts from axon terminals containing pleomorphic vesicles. These results strongly suggest that regenerated Abeta fibers activated by light tactile stimuli support functional connections in the superficial dorsal horn that have distinct ultrastructural features. In addition, the appearance of LDCVs suggests that primary sensory neurons are capable of changing their neurochemical phenotype.

Animals↗
PubMed1987

Morphometry and ultrastructure of the squirrel monkey (Saimiri sciureus) vestibular nerve.

Vestibular nerves of squirrel monkeys (Saimiri sciureus) embedded in plastics and epoxies were examined with light microscopy (LM) and transmission electron microscopy (TEM), and computerized measures were obtained and analyzed statistically. An average of 12,412 perikarya and 12,005 myelinated nerve fibers was obtained. Approximately 0.7% of the perikarya appeared unmyelinated under LM. About 500 unmyelinated fibers were counted. The cross-sectional area of 1,864 perikarya was 200-650 micron 2. The cross-sectional area of 1,346 nerve fibers was 3-11 micron 2 for the axoplasm and 11-12 micron 2 for the myelin sheath of the same fiber. Myelin thickness was directly proportional to the axoplasm cross-sectional area of the nerve fibers. The cross-sectional area of central axons and peripheral dendrites differed significantly (p less than 0.001). The initial segments of peripheral dendrites were usually smaller, but longer than the initial segments of the central axons. Both initial segments increased in diameter after the first node of Ranvier. Schmidt-Lantermann incisures were more abundant in thick and heavily myelinated fibers than in thin and lightly myelinated fibers. Larger perikarya usually had larger fibers and vice versa, within the first 100-200 micron from the first node of Ranvier. No major ultrastructural differences were found between myelinated and unmyelinated perikarya, except at the hillock region. The Nissl substance was preferentially located in the peripheral cytoplasm.

Animals↗
PubMed1996

The cricothyroid space: topography and clinical implications.

The inferior border of the thyroid cartilage and the superior rim of the cricoid arch are connected by the cricothyroid membrane and partly covered by the cricothyroid muscles ventrally. This region has been termed the cricothyroid space (CS) and is important with regard to surgical procedures, the spread of laryngeal cancer and traumatic lesions of the larynx. The precise topographic relations of the CS were investigated in plastinated serial sections of 21 normal adult human specimens. The CS consists of a caudal and a cranial portion, which are different with regard to their cartilaginous framework and their relationship to intralaryngeal structures. The caudal portion is bordered by the cricoid cartilage and reveals a complete separation of extra- and intralaryngeal regions by the cricothyroid membrane. It provides an easy access to the subglottic airways. The cranial portion of the CS is mainly bordered by the thyroid cartilage. It is characterized by a gap lateral to the median cricothyroid ligament. This allows a connection of extra- and intralaryngeal adipose tissue. There, extralaryngeal extension of laryngeal cancer preferentially occurs. Traumatic subglottic ruptures of the larynx involve both the caudal and cranial portions of the CS, which often results in extensive scar tissue formation.

Aged↗
PubMed1997

Plastination in the neurosciences. Keynote lecture.

Plastination is a unique technique of tissue preservation developed by Dr. Gunther von Hagens in Heidelberg, Germany, in 1978. In this process, water and lipids in biological tissues are replaced by curable polymers which are subsequently hardened, resulting in dry, odorless and durable specimens. In neuroanatomy silicone and polyester resins are used. Silicone rubber is used for whole brains and brain dissections resulting in natural looking specimens. Polyester resin is used for brain slices resulting in an excellent distinction between gray and white matter. The silicone S 10 standard technique and the polyester P 35 technique are described in detail. The advantages of plastinated specimens in neuroanatomy teaching and research are discussed.

Brain↗
PubMed1997

Setting up and features of a database of images of plastinated materials.

A database of plastinated materials was set up and distributed with a CD-ROM on the Internet. A total of 197 specimens, prepared at the Medical Museum of the University of Tokyo between 1991 and 1995, were catalogued in the database on a personal computer with commercial and original software. The database was originally designed to catalogue anatomical specimens and drawings for curators and researchers, but it also services as a worldwide digital museum.

Anatomy↗
PubMed1997

The potentially best stereographic presentation of the plastinated anatomical specimen.

Volumegraph, which is an improved version of the parallax panoramagram, has been studied as a method of stereophotography of a plastinated human body specimen. The volumegraph provides a steric view of the specimen in a natural color appearance. The potential advantages in using the volumegraph for the presentation of anatomical specimens are discussed taking account of its technical details.

Anatomy↗
PubMed1997

Reclamation of acetone in plastination laboratories: a simple and inexpensive method.

Since the introduction of the plastination process by von Hagens [Anat Rec 194/2: 247-256, 1979], the cost of acetone used for the dehydration step has been considered an important factor in the cost of plastination. We have developed a three-step method that permits the reuse of acetone. The first step simply consists of storing the contaminated acetone in the freezer and separating the congealed fat by filtration. The second step is vacuum distillation of the acetone and can be conducted with the freezer and vacuum pump (found in any plastination laboratory) with just a few additions. It produces 95-97% pure acetone. The last step uses a desiccant to take away the residual water from the distilled acetone and brings the purity to 99.5%. With this method, we have reduced the amount of acetone to be purchased to a minimum and completely eliminated the cost of discarding used acetone. In addition vaporized acetone released during the impregnation step of plastination is recaptured.

Acetone↗
PubMed1997

Design of lifting gear for a plastination laboratory.

Conventional lifting gear based on steel components can be safely used in a laboratory where acetone is used provided that: (a) the circumstances under which significant amounts of acetone vapour will be present are known and can be controlled and (b) operating practices are imposed to prevent the use of the lifting gear at times when acetone vapour may be present in explosive concentrations.

Acetone↗
PubMed1997

Sheet plastination as a clinically based teaching aid at the University of Auckland.

As a concerted move toward closer integration of the clinical and preclinical aspects of the undergraduate medical curriculum at the University of Auckland, the Department of Anatomy with Radiology has implemented a number of clinical procedures, pathological observations and diagnostic methods into the course resulting in a structured program of clinically based teaching of gross anatomy to second- and third-year medical students. The recently developed method of E12 epoxy resin sheet plastination of sectioned cadaver specimens has expanded the learning process twofold, firstly as a developmental microscopy teaching aid enabling us to link histological slides directly to gross anatomical specimens and secondly as a radiographic training tool in the correlation of clinical imaging techniques of magnetic resonance (MRI) and computed tomography (CT) with the teaching of gross anatomy. The direct comparison between E12 serial sectioned cadaver specimens and the equivalent MRI and CT images provides the student with a much clearer understanding of anatomical structures in relation to clinical diagnosis.

Aged↗
PubMed1997

Learning relational anatomy by correlating thin plastinated sections and magnetic resonance images: preparation of specimens.

Plastination is a process whereby the tissue water and part of the tissue fat of anatomical specimens is replaced with a curable polymer. Several variations of this technique are available, depending on both the type of specimen and polymer being used. In this study, the efficiency of BIODUR PEM 11-prepared cross sections as a teaching aid for radiographic anatomy is investigated. A human cadaveric head was scanned on a magnetic resonance (MR) imager, recording images in a coronal plane of approximately 6 mm in thickness. Corresponding slices were then taken of this head and plastinated by the BIODUR PEM 11 method. In general, a good correlation existed between the plastinated sections and the MR scans. The plastinated slices displayed superior differentiation between musculature compared to the scans. In conclusion, the plastinated sections, used in conjunction with the MR scans, are a valuable tool in learning radiographic anatomy. Additionally, BIODUR PEM 11 is a viable alternative to plastination by other methods.

Anatomy↗
PubMed1997

Correlating sheet plastinated slices, computed tomography images and magnetic resonance images of the pelvic girdle: a teaching tool.

Sheet plastination is currently used to produce anatomical slices of different body structures, allowing one to study and teach their topography in an anatomically correct state. Correlation with computed tomography (CT) and magnetic resonance imaging (MRI) techniques gives more insight into their anatomy. Using two female cadaver pelvises CT and MRI were performed. One pelvis was used to prepare 2-mm-thick coronal plastinated slices according to the technique described by von Hagens. We found a good overall correlation between plastinated slices, CT and MRI images. This combined approach provides a unique anatomical insight and is a valuable addition to other teaching tools used by medical students, radiologists and anatomists.

Anatomy, Cross-Sectional↗
PubMed1997

Plastinated canine gastrointestinal tracts used to facilitate teaching of endoscopic technique and anatomy.

Plastinated specimens, when prepared with a design for endoscopic use, can serve as a practical model for teaching. Intact alimentary canals were excised from fresh canine cadayers. Cannulas in excess of the intended endoscope size (9.6 mm diameter) were placed in restrictive openings [cardiac ostium (ostium cardiacum), pyloric ostium (ostium pyloricum) and cecocolic orifice (ostium cecocolicum)]. These cannulas allowed ingesta to be removed and maintained adequate diameters for endoscoping. After flushing out the gastrointestinal contents, specimens were formaldehyde-fixed overnight in a dilated anatomical conformation. Prior to S10/S3 impregnation, fixative was flushed from the specimens and they were dehydrated in acetone. After impregnation, slow cure (elongation of S3 molecules at room temperature) was allowed to proceed for approximately 1 week. The gastrointestinal tracts were maintained in a dilated conformation by a positive pressure air flow. When polymer seepage was minimal, they were cured using small quantities of S6 (final curing agent). The curing agent was contained around the specimen by enclosing the specimens in plastic bags. The plastinated specimens retain their dilated anatomical conformation, and may be used to teach both endoscopic technique and gastrointestinal anatomy.

Anatomy↗
PubMed1997

E12 technique: an aid to study sinuses of psittacine birds.

Infraorbital sinus infections, which form the bulk of upper respiratory tract infections in companion birds, are commonly encountered in clinical practice and often require medical and/or surgical management. The infraorbital sinus with its dorsal drainage into the nasal cavity makes it difficult to treat these infections. The sinus has various compartments throughout the skull, and subcutaneous tissue. As an aid in determining the location and extent of the sinuses of the parrot and macaw, a corrosion cast of the infraorbital sinus was made. Also, computed tomography (CT) images of cadaver heads were completed at 2-mm scans. Prior to sectioning for sheet plastination, the infraorbital sinus of one half of the imaged heads was injected with a mixture of colored epoxy. The specimens were frozen (-25 degrees C) prior to sawing 2-mm-thick sections which corresponded to the CT scans. The sections were dehydrated, impregnated and sheets prepared using the standard E12 technique. The slices were used to identify the compartments of the infraorbital sinuses and to aid identification of such on the CT images.

Animals↗
PubMed1997

Distribution of muscle spindles in the extensor digitorum and hallucis brevis muscles of the macaque as determined by plastination.

The structure-function relationships of the macaque extensor digitorum and hallucis brevis muscle (EDB and EHB), the single intrinsic and four-tendoned muscle of the dorsum of the foot, were investigated in the macaque after preparation of samples by two plastination methods, using gelatin and epoxy resin. These preparative methods clearly revealed the muscle fibers, spindles and myelinated axons in the EDB and EHB. In the more medially located two bellies (the medial two bellies of the EDB), the mean relative proportion of red fibers was larger than in the more laterally located two bellies (the lateral belly of the EDB, and the EHB), while those of intermediate and white fibers were smaller than in the more laterally located two bellies. (The difference was significant between some pairs of bellies.) In the more medially located two bellies, the mean cross-sectional area of each type of fiber, namely, the white, intermediate and red fibers, tended to be smaller than in the more laterally located two bellies. (There were significant differences between many pairs of bellies.) The total number of muscle spindles within an EDB and EHB was 15, and larger numbers of muscle spindles were distributed to the medially located two bellies compared to the laterally located two bellies. (There were significant differences between many pairs of bellies.) These results suggest that the more medially located two bellies of the EDB and EHB are adapted to the relatively precise movements of toes, while the more laterally located two bellies are adapted to the prompt extension and abduction that are related to the arboreal habitat of the macaques.

Animals↗
PubMed1997

Sheet plastination of the larynx for whole-organ histology.

Whole organ serial sections of the larynx are recognized to improve the assessment of laryngeal pathology. The human larynx with its complex anatomy of different types of tissue with their marked differences in hardness complicate the cutting process. The methods established are either very costly, time consuming and prone to artefacts, or sections are too thick for a histological work-up. Prior decalcification causes major shrinkage, thus morphometry is not feasible. We describe a technique of sheet plastination to produce whole-organ sections which allows a full-scale histological investigation without prior decalcification. Deep-frozen larynges are cut into 4-micron-thick sections using an ordinary slicing machine. Slices are dehydrated in acetone and plastinated in Biodur E50/E7/ AE10/E700. These specimens are cut into sections of about 80 microns thickness using a diamond-wire saw. An ultra-milling device reduces the section thickness down to about 10 microns. Surface staining of different kinds like Richardson, methylene blue or Paragon yields sufficient contrast for exact diagnosis. Deplastination and consecutive paraffin-like staining is possible, but results are not yet fully satisfying. Histological sheet plastination produces almost artefact- and shrinkage-free whole-organ sections in a period of about 1 week. The technique is useful for studies of micromorphometry or tumor spread. Theoretically, application of immunohistochemical staining methods seems to be possible. This might lead to mapping the proliferation activity of laryngeal cancer as it is presented on whole-organ sections.

Artifacts↗
PubMed1997

Localization of beta-D-galactosidase activity in semithin epon sections of embryonic tissues using differential interference contrast optics.

In the present study we describe a method for the histochemical demonstration of beta-D-galactosidase activity on tissue sections processed for light microscopy at high resolution. 5-Bromo-indolyl-beta-D-galactopyranoside (Bluo-Gal) was utilized as an indigogenic method for the demonstration of Escherichia coli beta-D-galactosidase reporter gene activity whose expression was studied in a transgenic line where the enzyme, with a nuclear localization signal (nlacZ), is under the transcriptional control of a striated muscle-specific promoter. At the light-microscopic level, by using Differential Interference Contrast (DIC) optics, the reaction product was detected as precipitates in the form of fine birefringent crystals. These were located around and inside the nuclei of beta-gal-expressing cells. This simple method allows an easy and rapid identification of few or even one labeled cell(s) within large microscopic fields (whole embryos) and the labeled cell(s) can be evaluated both morphologically and quantitatively.

Animals↗
PubMed1996

Drug targeting and metabolic investigations of cryoprepared tumor cells with analytical electron energy loss spectroscopy.

Analytical electron microscopy is an ideal tool for holistic data acquisition on biological systems. The use of analytical electron microscopy for both, the investigation of micropharmacokinetic problems and metabolic studies, is becoming more and more important. Depending on the mode of investigation, it is possible to localize drugs and xenobiotics precisely in situ under optical control or to quantify their uptake and distribution in the corresponding target cells without disintegrating the cell or tissue material. In this paper, we present instructive examples for the application of analytical electron energy loss spectroscopy in transmission electron microscopy in order to investigate the cellular uptake and distribution of cisplatin and cyclophosphamide and the metabolic changes induced by an alteration in the extracellular calcium concentration in a holistic manner.

Animals↗