Abstract
The axial skeleton represents one product of the metameric segregation of the mesoderm in the developing embryo. The mechanisms underlying this pattern formation remain poorly understood. Genetic alterations, either resulting from spontaneous mutation or as a result of xenobiotic exposure, may disrupt this patterning and lead to a variety of skeletal alterations. Chemical agents including valproic acid (1), retinoic acid (2), salicylate (3), and acetazolamide (4) have been shown to cause supernumerary ribs in rodents. Fewer agents cause a reduction in the number of ribs or vertebrae. Agents or conditions in the latter category include boric acid (5,6), arsenate (7), methanol (8), 2-chlorodeoxyadenosine (9), and hyperthermia (10). Effects on axial development have also been associated with changes in homeotic gene expression (11–13) as well as deletion of the bmi-1 proto-oncogene (14). Posteriorization of Hoxa10 expression has been associated with lumbar ribs in mice following prenatal exposure to sali-cylate (15) as well as retinoic acid (2). Careful characterization of the morphology of the axial skeleton is critical in identifying homeotic shifts and other changes produced by xenobiotics or altered gene expression. Here we present our methods for preparing and examining rodent skeletons, including anatomical landmarks and a brief discussion of methods for analyzing and presenting these data.
This document has been reviewed in accordance with U.S. Environmental Protection Agency policy and approved for publication. Mention of trade names or commercial products does not constitute endorsement or recommendation for use.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Narotsky, M. G., Francis, E. Z., and Kavlock, R. J. (1994) Developmental toxicity and structure-activity relationships of aliphatic acids, including dose-response assessment of valproic acid in mice and rats. Fundam. Appl. Toxicol. 22, 251–265.
Kessel, M. (1992) Respecification of vertebral identities by retinoic acid. Development 115, 487–501.
Wickramaratne, G. A. de S. (1988) The postnatal fate of supernumerary ribs in rat teratogenicity studies. J. Appl. Toxicol. 8, 91–94.
Beck, S. L. (1983) Assessment of adult skeletons to detect prenatal exposure to acetazolamide in mice. Teratology 28, 45–66.
Heindel, J. J., Price, C. J., and Schwetz, B. A. (1994) The developmental toxicity of boric acid in mice, rats, and rabbits. Environ. Health Perspect. 102(Suppl 7), 107–112.
Narotsky, M. G., Schmid, J. E., Andrews, J. E., and Kavlock, R. J. (1998) Effects of boric acid on axial skeletal development in rats. Biol. Trace Element Res. 66, 373–394.
Beaudoin, A. R. (1974) Teratogenicity of sodium arsenate in rats. Teratology 10, 153–158.
Connelly, L. E. and Rogers, J. M. (1997) Methanol causes posteriorization of cervical vertebrae in mice. Teratology 55, 138–144.
Narotsky, M. G., Best, D. S., Guidici, D. L., Hamby, B. T., Knudsen, T. B., Kavlock, R. J., et al. (1998) 2-Chlorodeoxyadenosine-induced lumbar hernia in rats. Teratology 57, 245 [abstract].
Kimmel, C. A., Cuff, J. M., Kimmel, G. L., Heredia, D. J., Tudor, N., Silverman, P. M., et al. (1993) Skeletal development following heat exposure in the rat. Teratology 47, 229–242.
Kessel, M., Balling, R., and Gruss, P. (1990) Variations of cervical vertebrae after expression of a Hox-1.1 transgene in mice. Cell 61, 301–308.
Small, K. M. and Potter, S. S. (1993) Homeotic transformations and limb defects in Hox A11 mutant mice. Genes Dev. 7, 2318–2328.
Charité, J. W., de Graaff, W., Shen, S., and Deschamps, J. (1994) Ectopic expression of Hoxb-8 causes duplication of the ZPA in the forelimb and homeotic transformation of axial structures. Cell 78, 589–601.
van der Lugt, N. M., Domen, J., Linders, K., van Roon, M., Robanus-Maandag, E., te Riele, H., et al. (1994) Posterior transformation, neurological abnormalities, and severe hematopoietic defects in mice with a targeted deletion of the bmi-1 proto-oncogene. Genes Dev. 8, 757–769.
Daston, G. P. and Overmann, G. J. (1996) Lumbar ribs associated with posteriorization of Hox a10 expression in salicylate-treated mouse embryos. Teratology 53, 85 [abstract].
National Fire Protection Association (1996) NFPA 30: Flammable & Combustible Liquids Code, Quincy, MA.
Selby, P. B. (1987) A rapid method for preparing high quality alizarin stained skeletons of adult mice. Stain Technol. 62, 143–146.
Dawson, A. B. (1926) A note on the staining of the skeleton of cleared specimens with alizarin red S. Stain Technol. 1, 123–124.
Kawamura, S., Hirohashi, A., Kato, T., and Yasuda, M. (1990) Bone-staining technique for fetal rat specimens without skinning and removing adipose tissue. Cong. Anom. 30, 93–95.
Staples, R. E. and Schnell, V. L. (1964) Refinements in rapid clearing technic on the KOH-alizarin red S method for fetal bone. Stain Technol. 39, 61–63.
Inouye, M. (1976) Differential staining of cartilage and bone in fetal mouse skeleton by alcian blue and alizarin red S. Cong. Anom. 16, 171–173.
Kimmel, C. A. and Trammell, C. (1981) A rapid procedure for routine double staining of cartilage and bone in fetal and adult animals. Stain Technol. 56, 271–273.
Whitaker, J. and Dix, K. M. (1979) Double staining technique for rat foetus skeletons in teratological studies. Lab. Anim. 13, 309–310.
Miller, D. M. and Tarpley, J. (1996) An automated double staining procedure for bone and cartilage. Biotech. Histochem. 71, 79–83.
Greene, E. C. (1949) Gross anatomy, in The Rat in Laboratory Investigation (Farris, E. J. and Griffith, J. Q., Jr., eds.), Hafner, New York, pp. 24–50.
Yasuda, M. and Tsunetsugu, Y. (1996) Color Atlas of Fetal Skeleton of the Mouse, Rat and Rabbit. Ace Art Co., Ltd., Osaka, Japan, pp. 10, 26.
Kimmel, C. A. and Wilson, J. G. (1973) Skeletal deviations in rats: Malformations or variations. Teratology 8, 309–316.
Wise, L. D., Beck, S. L., Beltrame, D., Beyer, B. K., Chahoud, I., Clark, R. L., et al. (1997) Terminology of developmental abnormalities in common laboratory mammals (version 1). Teratology 55, 249–292.
Narotsky, M. G., Hamby, B. T., Best, D. S., and Kavlock, R. J. (1996) Effects of singleday boric acid treatment on axial skeletal development in rats. Teratology 53, 101 [abstract].
Haseman, J. K. and Kupper, L. L. (1979) Analysis of dichotomous response data from certain toxicological experiments. Biometrics 35, 281–293.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2000 Humana Press Inc., Totowa, NJ
About this protocol
Cite this protocol
Narotsky, M.G., Rogers, J.M. (2000). Examination of the Axial Skeleton of Fetal Rodents. In: Walker, J.M., Tuan, R.S., Lo, C.W. (eds) Developmental Biology Protocols. Methods in Molecular Biology™, vol 135. Humana Press. https://doi.org/10.1385/1-59259-685-1:139
Download citation
DOI: https://doi.org/10.1385/1-59259-685-1:139
Publisher Name: Humana Press
Print ISBN: 978-0-89603-852-3
Online ISBN: 978-1-59259-685-0
eBook Packages: Springer Protocols