Cytotoxic Action of Palytoxin in Cultured Aortic Smooth Muscle Cells
Abstract
Palytoxin (PaTX) produced by coelenterate species (genus Palythoa) is a potent marine toxin which induces intense vasoconstriction and hemorrhage which contribute to lethality. The A7r5 clonal cell line, derived from fetal rat aorta, was used to study the mechanism of PaTX toxicity. A7r5 cells exposed to nM concentrations of PaTX for 15 min at room temperature followed by wash and incubation in culture medium at 37 dec C (> or = 30 min) developed swelling of endoplasmic reticulum, clumping of cytoplasm, blebbing and shrinking of heterochromatin. These cells were nonviable as confirmed by inclusion of trypan blue and release of LDH. The ED50 for PaTX cytotoxicity was 7.1 nM using the vital dye exclusion and 4.1 nM for LDH release. Whole-cell Patch clamp recordings from single A7r5 cells showed that PaTX (3-10 nM, 25 deg C, 15 min) depolarized cells and increased membrane conductance to Na(+) and K(+) > 5-fold. Neither the PaTX-induced cytotoxicity nor the increased ionic permeability could be reversed by washing. However, prior incubation of cells with ouabain (10-4M) or replacement of extracellular Na(+) with choline(+) or K(+) after exposure to PaTX did reduce the cytoxicity. Removal of extracellular Ca2+ reduced PaTX- induced cytotoxicity, but treatment with 10 to 50 uM verapamil did not, indicating that voltage-gated calcium channels were not involved. The mechanism of PaTX cytotoxicity in A7r5 cells seems to involve intracellular Na(+) overload and can be duplicated by other Na(+) ionophores such as nystatin. In light of the involvement of smooth muscle in PaTX poisoning, A7r5 cells could serve as a useful model to test specific drugs for protection against PaTX acute and delayed toxicity.
Document Details
- Document Type
- Technical Report
- Publication Date
- May 13, 1993
- Accession Number
- ADP008881
Entities
People
- B. F. Doxzon
- R. E. Sheridan
- S. S. Deshpande
Organizations
- United States Army Medical Research Institute of Chemical Defense