Biochemical and morphological changes in rat muscle cultures caused by 28,000 mol. wt toxin of Bacillus thuringiensis israelensis

The 28,000 mol. wt protein of Bacillus thuringiensis israelensis showed a high degree of toxicity to rat muscle in culture. Application of 1 μg/ml to the culture medium completely inhibited cell fusion. Reversibility of this effect was demonstrated by replacement of the culture medium with fresh medium, and the consequence was that cell fusion was resumed. When differentiated myotubes were treated with 1 μg/ml of the toxin, the spontaneous contractile activity was abolished within 20 min. Cytotoxic effects were observed 1 hr after treatment was initiated, as manifested by creatine kinase (CK) release to the medium. Two hours after toxin was applied to the muscle culture, the myotubes were deteriorated whereas the mononucleated cells were not affected. Six or 7-day-old cultures which were treated by 1 μg/ml of 28,00 mol. wt toxin revealed a change in the levels of Na⁺ and K⁺ within the fibres as analysed by X-ray microanalysis (XRMA). Preincubation of the toxin for 20 min with phospholipids before application to the cells reduced the cytotoxic effect. Phosphatidylinositol and phosphatidylserine were the most efficient inhibitors, whereas phosphatidylcholine, sphingomyelin and phosphatidylethanolamine were less effective in protecting cultures from the cytotoxic effects of the 28,000 mol. wt protein.