Summary
Cells were isolated from ∼ 30 d fetal rabbit stomachs and cultured on floating collagen gels. Electron microscopy showed monolayers in which only one cell type persisted. These columnar cells were joined at apical borders by tight junctions and contained an extensive endoplasmic reticular network with an occasional intracellular canaliculus. They also occasionally contained what appeared to be secretory granules (mucus?), and therefore had some characteristics of all the cell types of the intact fetal stomachs, which showed oxyntic, mucous, and undifferentiated cells. In Ussing chambers with Ringer's solution on both sides, cultures developed transepithelial potential (potential difference [PD], mV, mucosa ground)=13, resistance (resistance [R], Ω-cm2)=285, and short-circuit current (I sc , μA/cm2)=45 (n=7), clearly indicating that cellular polarity and junctional integrity were maintained. These transport parameters were somewhat different for intact fetal stomachs (PD=20, R=70, and I sc =220 [n=4]), which may be due to extensive folding of intact fetal stomachs or the presence of only one cell type in culture, or both. Although gastric stimulants histamine, dibutyryl cycle AMP (dbcAMP), and isobutyl-methylxanthine (IMX) (a phosphodiesterase inhibitor) did not elicit H+ secretion or electrophysiological changes in monolayers or intact stomachs, 10−4 M apical amiloride caused a decrease in I sc in cultured monolayers (27%) and intact stomachs (50%). Thus, Na+ transport seems to be a significant fraction of ion transport in both preparations. This culture system may allow the study of oxyntic cell differentiation and the development of H+, Na+, and Cl− transport in the gastric mucosa.
Similar content being viewed by others
References
Blum, A. L.; Shah, G. T.; Wiebelhaus, V. D.; Brennan, F. T.; Helander, H. F.; Ceballos, R.; Sachs, G. Pronase method for isolation of viable cell fromNecturus gastric mucosae. Gastroenterology 61: 189–200; 1971.
Croft, D. N.; Ingelfinger, F. J. Isolated gastric parietal cells; oxygen consumption, electrolyte content and intracellular pH. Clin. Sci. 37: 491–501; 1969.
Kaskebar, D. K.; Blumenthal, G. H. Frog gastric tubular cells: isolation culture, and some properties. Gastroenterology 73: 881–886; 1977.
Romrell, L. J.; Coppe, M. R.; Muro, D. R. Isolation and separation of highly enriched fractions of viable mouse gastric parietal cells by velocity sedimentation. J. Cell Biol. 65: 428–438; 1975.
Soll, A. H. Secretagogue stimulation of [14C] aminopyrine accumulation by isolated canine parietal cells. Am. J. Physiol. 238: G366-G376; 1980.
Walder, A. L.; Lunseth, J. B. A technique for separation of cells of gastric mucosa. Proc. Soc. Exp. Biol. Med. 112: 494–496; 1963.
Cereijido, M.; Robbins, E. S.; Dolan, W. J.; Rotunno, C. A.; Sabatini, D. D. Polarized monolayers formed by epithelial cells on a permeable and translucent support. J. Cell Biol. 77: 853–880; 1978.
Emerman, J. T.; Pitelka, D. R. Maintenance and induction of morphological differentiation in dissociated mammary epithelium on floating collagen membranes In Vitro 13: 316–328; 1977.
Handler, J. S.; Steele, R. E.; Sahib, M. K.; Wade, J. B.; Preston, A. S.; Lawson, N. L.; Johnson, J. P. Toad urinary bladder epithelial cells in culture: Maintenance of epithelial structure, sodium transport, and response to hormones. Proc. Natl. Acad. Sci. USA 76: 4151–4155; 1979.
Ceriani, R. L.; Peterson, J. A.; Abraham, S. The removal of cell surface material by enzymes used to dissociate mammary-gland cells. In Vitro 14: 887–894; 1978.
Bisbee, C. A.; Machen, T. E.; Bern, H. A. Mouse mammary epithelial cells on floating collagen gels: transepithelial ion transport and effects of orolactin. Proc. Natl. Acad. Sci. USA 76: 536–540; 1979.
Misfeldt, D. S.; Hamamoto, S. T.; Pitelka, D. R. Transepithelial transport in cell culture. Proc. Natl. Acad. Sci. USA 73: 1212–1216; 1976.
Michalopoulas, G.; Pitot, H. C. Primary culture of parenchymal liver cells on collagen membranes. Exp. Cell Res. 94: 70–78; 1975.
Emerman, J. T.; Enami, J.; Pitelka, D. R.; Nandi, S. Hormonal effects on intracellular and secreted casein in cultures of mouse mammary epithelial cells on floating collagen membranes. Proc. Natl. Acad. Sci. USA 74: 4466–4470; 1977.
Hayward, A. F. The ultrastructure of developing gastric parietal cells in the fetal rabbit. J. Anat. 101: 69–81; 1967.
Hayward, A. F. The fine structure of gastric epithelial cells in the suckling rabbit with particular reference to parietal cell. Z. Zellforsh. 78: 474–483; 1967.
Kataoka, K. Electron microscopic observations on cell proliferation and differentiation in the gastric mucosa of the mouse. Arch. Histol. Jpn. 32: 251–273; 1970.
Krause, W. J.; Cutts, J. H.; Leeson, C.R. The postnatal development of the alimentary canal in the opposum. II. Stomach. J. Anat. 122: 499–519; 1976.
Nomura, Y. On the submicroscopic morphogenesis of parietal cells in the gastric gland of the human fetus. Z. Anat. Entwicklungsgech 125: 316–356; 1966.
Helander, H. F. Ultrastructure and function of gastric parietal cells in the rat during development. Gastroenterology 56: 35–52; 1969.
Helander, H. F. Ultrastructure and function of gastric mucoid and zymogen cells in the rat during development. Gastroenterology 56: 53–69; 1969.
Menzies, G. Observations on the developmental cytology of the fundic region of the rabbit's stomach, with particular reference to the peptic cells. Q. J. Microscop. Sci. 99: 485–496; 1958.
Menzies, G. Observations on the development of certain cell types in the fundic region of the rabbit's stomach. Q. J. Microscop. Sci. 105: 449–454; 1964.
Kendal, A.; Wright, G. H. Active transport of ions by the gastric mucosa of the rabbit foetus. J. Physiol. 190: 531–540; 1967.
Wright, G. H. Net transfers of water, sodium chloride and hydrogen ions across the gastric mucosa of the rabbit foetus. J. Physiol. 163: 281–293; 1962.
Farquhar, M. G.; Palade, G. E. Junctional complexes in various epithelia. J. Cell Biol. 17: 375–412; 1963.
Miller, L. R.; Jacobsen, E. D.; Johnson, L. R., Effect of pentagastrin on gastric mucosal cells grown in tissue culture. Gastroenterology 64: 264–267; 1973.
Matsuyama, M.; Suzuki, H. Differentiation of immature mucous cells into parietal, argyophil, and chief cells in stomach grafts. Science 169: 385–387; 1970.
Fromter, E.; Diamond, J. Route of passive ion permeation in epithelia. Nature 235: 9–13; 1972.
Forte, J. G.; Machen, T. E. Transport and electron phenomena in resting and secreting piglet gastric mucosa. J. Physiol. 244: 33–51; 1975.
Machen, T. E.; Silen, W.; Forte, J. E. Na+ transport by mammalian gastric mucosa. Am. J. Physiol. 234: E228-E235; 1975.
Author information
Authors and Affiliations
Additional information
This work was supported by NIH Grant AM 19520. The electron microscope was purchased in part by NSF Grant PM 76-80300. C. Bisbee was supported by National Cancer Institute Grants CA-05388 and CA-09041. C. Logsdon received support from the Systems and Integrative Biology Training Grant.
Rights and permissions
About this article
Cite this article
Logsdon, C.D., Bisbee, C.A., Rutten, M.J. et al. Fetal rabbit gastric epithelial cells cultured on floating collagen gels. In Vitro 18, 233–242 (1982). https://doi.org/10.1007/BF02618576
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF02618576