Effect of pneumolysin on rat brain ciliary function: comparison of brain slices with cultured ependymal cells.
This study compares two models for examining ependymal ciliary function: rat brain slices cut from the fourth ventricle and primary ependymal cells in culture. The cilia from both preparations were very reproducible; each preparation had cilia beating at a constant frequency of between 38 and 44 Hz. With the brain slices, ciliary stasis occurred after 5 d in culture. However, ependymal cells had fully functional cilia for up to 48 d in culture. The pneumococcal toxin, pneumolysin, caused a dose-dependent inhibition of cilia beat frequency within 15 min in both models. There were no significant differences in the mean log 50% inhibitory concentration (pIC50) slice = 0.65 +/- 0.05, equivalent to 4.4 hemolytic units (HU)/mL; cells = 0.57 +/- 0.14, equivalent to 3.7 HU/mL. There were also no significant differences in the mean Hill slope factors for the curves (slice = 1.4 +/- 0.05; cells = 1.6 +/- 0.4). These data demonstrate that both models can be used to examine the acute (15-min) effects of pneumolysin on cilia beat frequency. The main advantage of the primary ependymal culture model is that considerably more cultured ependymal cells (approximately 70%) are available, compared with the number of ependymal cells on the brain slices (approximately 2%), thus reducing the number of animals used. A pure ependymal culture was not achieved (approximately 30% of the cells were not ciliated). The increased survival time of the ependymal cells compared with the brain slices make cultured ependymal cells more useful for examining long-term ciliary function, whereas brain slices may be more useful for examining the interactions between ependymal and other nearby cells.
Hirst
R A
RA
Department of Child Health, Leicester Royal Infirmary, University of Leicester, England, UK.
Rutman
A
A
Sikand
K
K
Andrew
P W
PW
Mitchell
T J
TJ
O'Callaghan
C
C
D000818:Animals; D001426:Bacterial Proteins; D002478:Cells, Cultured; D002531:Cerebellum / Q000187:drug effects* / Q000502:physiology / Q000648:ultrastructure; D002923:Cilia / Q000187:drug effects* / Q000502:physiology; D004805:Ependyma / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D066298:In Vitro Techniques; D008855:Microscopy, Electron, Scanning; D051381:Rats; D017208:Rats, Wistar; D013301:Streptolysins / Q000494:pharmacology*
D001426:Bacterial Proteins; D013301:Streptolysins; C032290:plY protein, Streptococcus pneumoniae