Toxic effects of d-limonene in the earthworm Eisenia fetida (Savigny)

Abstract

d-Limonene, a monocyclic monoterpenoid with known insecticidal properties, was assayed (using a standard method of cutaneous exposure) for general lethality effects as well as neurotoxic effects on escape reflex pathways in earthworms, Eisenia fetida (Savigny). Neurotoxicity was assessed by noninvasive electrophysiological techniques involving (a) quantification of the impacts of chronic and acute sublethal exposures on impulse conduction in the worms' medial (MGF) and lateral (LGF) giant nerve fiber pathways, (b) determination of whether such effects were generalized or localized within various body regions, and (c) determination of the reversibility of neurotoxic effects. The LD 50 value for d-limonene alone was 6.0 ppm, and the LT 50 value for exposure to 12.6 ppm was 4.9 hr. Effects on lethality were not synergized significantly by either piperonyl butoxide or sesame oil. Generally, chronic and acute intoxication involved a rapid and predictable cascade of behavioral and morphological symptoms, including increased mucus secretion, writhing, clitellar swelling, and elongation of the body. In addition, chronic d-limonene exposures induced significant weight loss, but there was no effect on MGF and LGF conduction velocities, even though abnormal rebounding of MGF impulses and spontaneous LGF spiking were often evident. Acute exposures, however, induced significant decreases in conduction velocity in both the MGF and LGF, but the effects were regionally specific; for example, LGF velocities were significantly reduced in the posterior half of the body but not in the anterior half. The magnitude of conduction velocity decreases was directly related to both concentration and duration of exposure. Other neurotoxic manifestations of acute exposures included occasional blocking of LGF impulse propagation and reduced amplitude in the muscle potential which normally accompanies each MGF spike. Decreases in conduction velocities after acute exposures were reversed once d-limonene exposure ceased. The time course of this reversibility was related to both exposure concentration and duration.