Staurosporine does not prevent adrenergic-induced situs inversus, but causes a unique syndrome of defects in rat embryos grown in culture.

Staurosporine, an alkaloid isolated from Streptomyces species, is commonly used as a protein kinase C (PKC) inhibitor in animal investigations. In the present study, we used this compound to determine whether α1-adrenergic stimulation-induced situs inversus in rats is mediated by PKC. Embryos were explanted at 8 A.M. on day 9 of gestation. Those with a neural groove but with no visible neural folds (Stage 11a) were selected and were cultured in medium containing various concentrations of staurosporine with or without 50 μM of phenylephrine, an α1-adrenergic agonist. At 10 A.M. On day 11 of gestation, embryos were examined for situs inversus and other abnormalities. Staurosporine, tested at 0.0001, 0.001, 0.01, 0.1, 0.375, and 0.5 μM (lethal concentration), did not block phenylephrine-induced situs inversus at any concentration. However, staurosporine alone produced situs inversus at concentrations above 0.1 μM. At 0.5 and 1.0 μM, staurosporine also caused cyst-like lesions projecting dorsally from the mesencephalon that we named “mesencephalic vesicles” and the formation of secondary somites. To confirm and further examine these unique effects of staurosporine both grossly and histologically, we conducted additional experiments using staurosporine from another source. Embryos were explanted between 6 A.M. and 9 P.M. on day 9 of gestation and were placed in one of the following groups according to their stage of development: 10b, 11a, 11b, 11c, 12/sl–2, 12/s3–4, and 12/s5–6. Embryos were then cultured with various concentrations of staurosporine. Those cultured from Stage 11a exhibited similar lesions to those seen in the initial experiment but at somewhat higher concentrations of staurosporine. Embryos cultured from Stage 10b showed a similar pattern of lesions as seen at Stage 11a, except that higher concentrations of staurosporine were required to cause mesencephalic vesicles and secondary somites formation. Embryos cultured from Stage 11b showed similar effects to those cultured from younger stages except that maximum incidences of situs inversus were much lower. Those cultured from Stage 11c showed similar dose–response to those cultured from Stage 11b except that the incidence of secondary somites formation was much higher. In addition, in approximately 40% (n = 25) of embryos treated with greater than 1. 0 μM of staurosporine, the growing end of the allantois did not reach the chorion and remained unattached in the exocoelomic cavity. Embryos cultured from Stage 12/s1–2 showed similar effects to those cultured from Stage 11c except that incidences of failure of chorion—allantois fusion were somewhat higher. Embryos cultured from Stages 12/s3–4 and 12/s5–6 were not viable after exposure to 1. 0 μM staurosporine. At 0. 5 μM, approximately 15% of embryos showed some kind of abnormality but no mesencephalic vesicles were observed. Histological examinations on the embryos cultured from Stage 11b with 1. 5 μM staurosporine revealed that the first sign of the mesencephalic vesicles was abnormal variable thickness and convolutions of the neuroepithelium lateral to the neural groove. These results suggest that situs inversus induced by stimulation of α1-adrenergic receptors is not mediated through PKC. They also show that staurosporine causes situs inversus and various kinds of unique abnormalities in a dose-dependent and stage-specific manner. © Wiley-Liss, Inc.