sq-23377 and Peripheral-Nervous-System-Diseases

Cytosine arabinoside (Ara C) is a useful chemotherapy agent, used for treating acute myeloid leukaemia, although it may be associated with side effects including painful neuropathy. It is also used for in vitro neuronal studies to limit the proliferation of non-neuronal cells and thereby select nondividing neuronal cells. We studied the effects of Ara C on human dorsal root ganglion (DRG) neurons, especially the expression and sensitivity of the ion channel TRPV1, which responds to noxious heat and capsaicin and is a key mediator of neuropathic pain.. Human DRG neurons were cultured with or without Ara C for 2 weeks, after which Ara C was discontinued. Double immunostaining for the regenerative neuronal marker Gap43 and the capsaicin receptor TRPV1 showed that the normal membrane-bound localisation of TRPV1 was absent in neurons with Ara C treatment, and as expected there was massive diminution of dividing non-neuronal cells. Calcium imaging studies showed that during exposure to Ara C the neurons lost responsiveness to capsaicin, although ionomycin responses were intact, indicating general cell viability and responsiveness. Between 2 days and up to 3 weeks after removal of Ara C, the neuronal responses to capsaicin were regained and were observed to be four times (P = 0.0008, Student's t-test) that of controls, but there was only a gradual recovery of non-neuronal cells. Three to six weeks after Ara C removal, capsaicin responses were comparable to controls.. It is postulated that Ara C treatment blocked insertion of TRPV1 in the cell membrane, resulting in accumulation of the receptors in the cytoplasm, loss of capsaicin sensitivity, and membrane-bound immunostaining, which was restored with a rebound on withdrawal of Ara C. The observed pattern of loss of capsaicin sensitivity, followed by hypersensitivity and recovery, appears to reflect some of the features observed in chemotherapy-induced neuropathy, and may provide a model for developing new treatments and prophylaxis.

Topics: Adult; Antimetabolites, Antineoplastic; Calcium; Capsaicin; Cell Division; Cell Membrane; Cell Survival; Cells, Cultured; Cytarabine; Dose-Response Relationship, Drug; Ganglia, Spinal; GAP-43 Protein; Humans; Ionomycin; Neurons, Afferent; Peripheral Nervous System Diseases; Sensory System Agents; Time Factors; TRPV Cation Channels

Incubation of nerve with high concentrations of the divalent cation ionophore A23187 produces myelin vesiculation (Schlaepfer 1977). This observation has now been extended using segments of rat ventral or dorsal root incubated with high (19 microM, 10 micrograms/ml) or low (1-1.5 microM) concentrations of A23187, or another divalent ionophore, ionomycin. Low concentrations of A23187 induced no vesiculation within a 2-h period. However, subsequent incubation of these roots in fresh, ionophore-free medium for 20 h, resulted in a prominent vesicular demyelination at the Schmidt-Lanterman incisures and paranodes of many fibres. At this time (22 h) the Schwann cells associated with some demyelinating internodes appeared vital upon ultrastructural examination: the cells also excluded the nuclear dye nigrosin. High concentrations of A23187 induced a similar vesicular demyelination in affected fibres within only 15-20 min. While the Schwann cells continued to exclude nigrosin for a further 4 h, their ultrastructural appearance indicated that they were probably in the early stages of necrosis. Incubation of moribund root with the ionophore produced no myelin vesiculation. At all ionophore concentrations, the myelin vesiculation was dependent upon the presence of extracellular Ca2+, and could be modulated in severity by varying this concentration. Other divalent cations (Ba2+, Co2+, Mg2+, Mn2+, Ni2+, Sr2+) could not substitute for Ca2+. The vesiculation induced by A23187 could be entirely prevented by the addition of Zn2+ (greater than or equal to 1 microM), Ni2+ (greater than or equal to 1-10 microM), Co2+ (greater than or equal to 100 microM) or Mn2+ (greater than or equal to 100 microM) to the bathing medium. A23187 applied to only part of an isolated internode resulted in a localization of the myelin disruption to that region. Ionomycin (greater than or equal to 1 microM), an ionophore with a greater selectivity for Ca2+ than A23187, also induced a prompt Ca2+-dependent myelin vesiculation. We conclude that vesicular demyelination can be initiated in vital Schwann cells by a raised intracellular Ca2+ concentration. Such demyelination does not necessarily lead to Schwann cell death. The possible relevance of the findings to vesicular demyelinating neuropathies is discussed, and a hypothesis regarding the mechanism of demyelination is advanced.

Topics: Animals; Calcimycin; Calcium; Culture Techniques; Demyelinating Diseases; Ethers; Ionomycin; Male; Peripheral Nerves; Peripheral Nervous System Diseases; Rats; Rats, Inbred Strains; Schwann Cells