Quantitative Techniques in Neuropathology

Abstract

In the future, quantitative techniques will probably be used in industry as part studies in the evaluation of compounds for neurotoxicity (1). These studies can be used to determine dose-responses or no-effect levels. Various factors need consideration in the design of a quantitative study (10,28). These include the quality of the samples, the standardization of techniques for sample preparation, the methods of measurement, the sampling procedures and the statistical analyses that will be used to evaluate the data.This study was undertaken to evaluate various parameters in a cross section of rat sural nerve using 3 different sampling methods. The sural nerve is studied frequently in human peripheral nerve disease (23) and determinations of fiber counts and measurements of fiber size are used as indicators of pathological change. The sural nerve is one of the peripheral nerves that is required to be evaluated in rats as part of an evaluation of the potential neurotoxicity ofchemical compounds (1 2). Little quantitative data has been published on the peripheral nerve fibers of rats in cross section (18). As nerve fibers of different sizes may not be homogeneously distributed within a nerve (9), the method of sampling may be critical for a definitive quantitative study. MATERIALS AND h h k i O D S Tissue PreparationAn 8-week-old male Sprague-Dawley rat (Charles River Canada Inc., St. Constant, Quebec, Canada) weighing 310 g was anesthetized by an intraperitoneal injection of sodium pentobarbital and perfused with a mixture of 3% glutaraldehyde, 3% formaldehyde, 0.05% calcium chloride and 0.1% picric acid in 0.1 M cacodylate buffer. A 3 mm section of the sural nerve proximal to the proximal end of the gastrocnemius muscle was removed, rinsed in cacodylate buffer, placed in 1.5% osmium tetroxide for 2 hr, rinsed in buffer and stained with a 1% aqueous solution of uranyl acetate for 2 hr. The tissue was then rinsed in distilled water, dehydrated in ascending concentrations of ethyl alcohol cleared in propylene oxide and embedded in a mixture of Jembed and Araldite. Semi-thin epoxy sections (0.5 pmj'were obtained'with a glass knife and stained with borate-buffered 1% toluidine blue. Thin sections were cut with a diamond knife and placed on slotted grids covered by a celloidin film and stained with 4% aqueous uranyl acetate and 0.4% lead citrate- Myelitrated Nerve Fiber hleasuretnetrtsBlack and white photomicrographs of the entire semi-thin cross section of the sural nerve were taken

Clinically Translatable Formulation Strategies for Systemic Administration of Nerve‐Specific Probes

“…Tissue sections were analyzed for any pathological change in axons, neurons, glia, blood vessels, and muscle tissue including inflammation, demyelination, swelling, hemorrhage, and atrophy by the department of pathology at Dartmouth‐Hitchcock Medical Center. [ 86,87 ] The TAD was designated as the MTD because mice demonstrated healthy weight gain and minimal change in blood chemistry metrics over the 14 days study period.…”

The Society of Toxicologic Pathology: Advances and Adventures in the First 50 Years

“…For example, STP members were key leaders in evaluating the biological effects of dietary restriction on animal viability, as well as the incidences and severities of chronic diseases and neoplasms during lifetime rodent carcinogenicity bioassays. [16][17][18][19] Members of STP were also influential discussants in defining appropriate practice standards for such routine toxicologic pathology tasks as clinical pathology interpretation, [20][21][22] pathology peer review, [23][24][25][26] quantitative analysis, 27 and collection and trimming of unusual organs (eg, inner ear, 28 larynx 29 ). The Society's dedication to collaborative science in the field was affirmed by the EC decision in 1999 to support the creation of 2 toxicologic pathology groups.…”

Modern Pathology Methods for Neural Investigations

“…High-quality peripheral nerve sections for light microscopic evaluation are best achieved by post-fixation with osmium, embedding in epoxy resin, sectioning at 1 mm, and staining with toluidine blue (Aparicio and Marsden 1969, with many other references in print and online). Morphometric methods can be easily employed on resulting cross-sections to determine nerve fiber sizes and myelin thickness (Broxup et al 1990). A promising technique adapted from clinical medicine that is used to evaluate neuropathies of nonmyelinated nociceptor fibers is to count fiber numbers in specially stained skin biopsies.…”