§ 79.66 Neuropathology assessment.
(a) Purpose. (1) The histopathological and biochemical techniques in this guideline are designed to develop data in animals on morphologic changes in the nervous system associated with repeated inhalation exposures to motor vehicle emissions. These tests are not intended to provide a detailed evaluation of neurotoxicity. Neuropathological evaluation should be complemented by other neurotoxicity studies, e.g. behavioral and neurophysiological studies and/or general toxicity testing, to more completely assess the neurotoxic potential of an exposure.
(2) [Reserved]
(b) Definition. Neurotoxicity (NTX) or a neurotoxic effect is an adverse change in the structure or function of the nervous system following exposure to a chemical substance.
(c) Principle of the test method. (1) Laboratory rodents are exposed to one of several concentration levels of a test atmosphere for at least six hours daily over a period of 90 days. At the end of the exposure period, the animals are anaesthetized, perfused in situ with fixative, and tissues in the nervous system are examined grossly and prepared for microscopic examination. Starting with the highest dosage level, tissues are examined under the light microscope for morphologic changes, until a no-observed-adverse-effect level is determined. In cases where light microscopy has revealed neuropathology, the NOAEL may be confirmed by electron microscopy.
(2) The tests described herein may be combined with any other toxicity study, as long as none of the requirements of either are violated by the combination. Specifically, this assay may be combined with a subchronic toxicity study, pursuant to provisions in § 79.62.
(d) Limit test. If a test at one dose level of the highest concentration that can be achieved while maintaining a particle size distribution with a mass median aerodynamic diameter (MMAD) of 4 micrometers (µm) or less, using the procedures described in paragraph (a) of this section, produces no observable toxic effects and if toxicity would not be expected based upon data of structurally related compounds, then a full study using three dose levels might not be necessary. Expected human exposure though may indicate the need for a higher dose level.
(e) Test procedures—(1) Animal selection—(i) Species and strain. Testing shall be performed in the species being used in other NTX tests. A standard strain of laboratory rat is recommended. The choice of species shall take into consideration such factors as the comparative metabolism of the chemical and species sensitivity to the toxic effects of the test substance, as evidenced by the results of other studies, the potential for combined studies, and the availability of other toxicity data for the species.
(ii) Age. Animals shall be at least ten weeks of age at the start of exposure.
(iii) Sex. Both sexes shall be used unless it is demonstrated that one sex is refractory to the effects of exposure.