A rapid silver-free method for staining the myenteric plexus.

A method is presented for the relatively rapid demonstration of the myenteric plexus. Saturated Sudan black B in 70% ethanol followed by 0.01% aqueous buffered thionein were used on intestinal peels (whole-mounts) to stain myelinated and unmyelinated fibers and neuron cell bodies, respectively. In contrast to accepted silver methods, these two kinds of fibers were distinguished clearly; Schwann cell nuclei and nodes of Ranvier were visible. Preparations had the following attributes: relatively low optical density coupled with high visual contrast, freedom from metallic "mirroring," low background staining of subjacent muscle fibers, and presentation of a polychromatic picture. The entire procedure was under the complete and repeatable control of the operator. Perikaryon and nuclear morphology were clearly demonstrated. The limitations of this method are that it does not provide good visualization of individual unmyelinated neuronal processes and does not permit preparation of permanent slides.

Electron microscopic serial section analysis of nodes of Ranvier in lumbar spinal roots of the cat: a morphometric study of nodal compartments in fibres of different sizes.

Serially sectioned nodes of Ranvier from nerve fibres 2-20 micron in diameter of feline ventral and dorsal spinal roots were examined electron microscopically, reconstructed to scale and analysed morphometrically. The assumed 'fresh-state' value of several structural variables, considered to be of functional significance, were calculated by the use of compensation factors. The compensated data were plotted against fibre and axon diameters. It was calculated that the membranous area of the 'fresh-state' nodal axon segment increased more or less exponentially from less than 5 micron2 to 30 micron2 with increasing fibre diameter (D). Most variables associated with the nodal gap and the Schwann cell initially increased rapidly with D and then levelled out or even decreased in fibres with a D value greater than 8-12 micron. The area open for communication between the nodal axolemma and the endoneurial space was 30-100 times smaller than the membrane area of the nodal axolemma. The volume of the extracellular space in the nodal gap, outside the nodal axolemma, increased linearly from less than 0.1 micron3 to about 0.6 micron3 with increasing fibre size. The Schwann cell membrane area facing the nodal gap outnumbered the membrane area of the nodal axon by 10-15 times in nerve fibres with a D value between 5 and 15 microns. Some functional implications of the 'fresh-state' nodal model are discussed.

How is the exact length of an internode determined.

(1) The length of the human ventral spinal roots in the newborn and in the adult were compared with the length of the internodes in these roots. Internodes of fibers of a given caliber were more than twice as long in the adult sacral roots than in the cervical roots. The factors of root elongation corresponded closely to the factors of internode elongation. (2) Internode length was in a statistically significant linear relation with fiber caliber in individual roots. The slope of the regression lines differed among roots, and were steeper caudally. If data from roots having different elongation factors were pooled there was no longer a linear relationship between fiber caliber and internode length. Internode length, therefore, correlated directly and quantitatively with root elongation; it was not directly linked to fiber caliber. (3) The total Schwann cell population per fiber was nearly stable from the newborn to the adult, but the sacral roots had 5 times as many Schwann cells as the cervical roots. Calculations based on the number of internodes and on the length of fetal roots at the onset of myelination showed that the mean "initial length" of the Schwann cell at the onset of myelination was 187 micrometer, the same for all roots. (4) The Schwann cell population per fiber is determined by 2 reciprocal growth phases: elongation of the fetal fiber up to the onset of myelination corresponds to a multiplication of Schwann cells each having a given initial length. Schwann cell populations stabilize after the onset of myelination; from then on the geometry of each internode is determined by its passive elongation. The timing of myelination is critical for determining the ultimate length of the Schwann cell. If myelination does not begin before a fiber system has attained approximately one eighth of its definitive size, the longest internodes of that system will average 1.5 mm and most will remain below 2 mm, regardless of body size.

The precise geometry of large internodes.

The interrelations, in each given internode, between internodal length, axon caliber, axoplasmic volume and several parameters of sheath thickness were studied in a set of 37 large internodes from the sciatic nerve of the German shepherd dog, using multiple electron-microscopic measurements of isolated fibers. Internodal length varied in a nonlinear relation with axon caliber. Nonlinear relations were also found for the number of lamellae in the sheath as related to either axon caliber or to internodal length, particularly for large internodes: the deviations observed for axon caliber (thick fibers had fewer myelin lamellae than would correspond to axon caliber) were the opposite of for internodal length (thick fibers had more lamellae than would correspond to internodal length). A near linear relationship was found if the volume of myelin per internode was related to both the length and the circumference of the axon segment ensheated by it, i.e., axolemmal surface area. The deviations in the proportions of large internodes probably indicate the existence of upper limits of internodal growth conditioned by the enormous metabolic demands placed on the individual Schwann cell by the growth of internodes beyond 1.5--2 mm length or 100 thousand micrometer3 of myelin, respectively. The demands on myelin metabolism from unabated internodal growth and the resultant logistic problems of the Schwann cells are demonstrated from measurement of observed internodes and calculations of extrapolated growth.

The relationship between internodal length and fibre diameter in the spinal cord of the cat.

The relationship between internodal length and external fibre diameter in the spinal cord was examined on glutaraldehyde perfused material from a number of cats. The relationship in all samples was highly significant and there was no significant difference either between animals or between the lateral and dorsal columns. However, there was considerable variability, some internodes being twice and others less than half as long as the length predicted by the regression equation. Our results showed that the relationship between internodal length and fibre diameter was essentially a linear function. No internodes shorter than 100 micrometers were seen.