Acute phalloidin toxicity in living hepatocytes. Evidence for a possible disturbance in membrane flow and for multiple functions for actin in the liver cell. (2024)

PMCID: PMC1888124

S. Watanabe and M. J. Phillips

Copyright and License information PMC Disclaimer

Abstract

Actin filament based cell functions were examined in freshly isolated hepatocytes using phalloidin as an inhibitor. In particular, cell motility events, namely, surface bleb formation and canalicular contractile movements, were assessed and compared with morphologic changes in the cells. Phalloidin (in 0.1, 1.0, and 10.0 micrograms/ml dosages) was added to the culture medium 4 hours after isolation of the hepatocytes. Cell motility was recorded with time-lapse cinephotomicrography, and the morphologic changes were evaluated by phase-contrast optics and by transmission electron microscopy of serial sections. Membrane-bound pericanalicular cytoplasmic vacuoles appeared first, followed by cytoplasmic protrusions at the cell surface. Vacuolar membrane continuities with the canalicular membrane were noted, and later, with other regions of the cell surface, such that large tortuous irregular membrane-bound canals are seen on serial sectioning to link extracellular and canalicular spaces. These findings suggest a possible disturbance in membrane flow. Canalicular motility was greatly reduced and was dose-dependent. The time-based difference in the changes at the canalicular and sinusoidal surfaces may be indicative of different functions and/or sensitivities of the actin filaments within the liver cell.

Images in this article

• Wieland T. Modification of actins by phallotoxins. Naturwissenschaften. 1977 Jun;64(6):303–309. [PubMed] [Google Scholar]
• Oda M, Price VM, Fisher MM, Phillips MJ. Ultrastructure of bile canaliculi, with special reference to the surface coat and the pericanalicular web. Lab Invest. 1974 Oct;31(4):314–323. [PubMed] [Google Scholar]
• French SW, Davies PL. Ultrastructural localization of actin-like filaments in rat hepatocytes. Gastroenterology. 1975 Apr;68(4 Pt 1):765–774. [PubMed] [Google Scholar]
• Imanari H, Kuroda H, Tamura K. Microfilaments around the bile canaliculi in patients with intrahepatic cholestasis. Gastroenterol Jpn. 1981;16(2):168–173. [PubMed] [Google Scholar]
• Namihisa T, Tamura K, Saif*cku K, Imanari H, Kuroda H, Kanaoka Y, Okamoto Y, Sekine T. Fluorescent staining of microfilaments with heavy meromyosin labeled with N-(7-dimethylamino-4-methylcoumarinyl) maleimide. J Histochem Cytochem. 1980 Apr;28(4):335–338. [PubMed] [Google Scholar]
• Holborow EJ, Trenchev PS, Dorling J, Webb J. Demostration of smooth muscle contractile protein antigens in liver and epithelial cells. Ann N Y Acad Sci. 1975 Jun 30;254:489–504. [PubMed] [Google Scholar]
• Elias E, Hruban Z, Wade JB, Boyer JL. Phalloidin-induced cholestasis: a microfilament-mediated change in junctional complex permeability. Proc Natl Acad Sci U S A. 1980 Apr;77(4):2229–2233. [PMC free article] [PubMed] [Google Scholar]
• Gabbiani G, Montesano R, Tuchweber B, Salas M, Orci L. Phalloidin-induced hyperplasia of actin filaments in rat hepatocytes. Lab Invest. 1975 Nov;33(5):562–569. [PubMed] [Google Scholar]
• Dubin M, Maurice M, Feldmann G, Erlinger S. Phalloidin-induced cholestasis in the rat: relation to changes in microfilaments. Gastroenterology. 1978 Sep;75(3):450–455. [PubMed] [Google Scholar]
• Oshio C, Phillips MJ. Contractility of bile canaliculi: implications for liver function. Science. 1981 May 29;212(4498):1041–1042. [PubMed] [Google Scholar]
• Wu PC, Lai CL, Lam KC, Ho J. Prednisolone in HBsAg-positive chronic active hepatitis: histologic evaluation in a controlled prospective study. Hepatology. 1982 Nov-Dec;2(6):777–783. [PubMed] [Google Scholar]
• Watanabe S, Miyairi M, Oshio C, Smith CR, Phillips MJ. Phalloidin alters bile canalicular contractility in primary monolayer cultures of rat liver. Gastroenterology. 1983 Aug;85(2):245–253. [PubMed] [Google Scholar]
• Weiss E, Sterz I, Frimmer M, Kroker R. Electron microscopy of isolated rat hepatocytes before and after treatment with phalloidin. Beitr Pathol. 1973 Dec;150(4):345–356. [PubMed] [Google Scholar]
• Phillips MJ, Oda M, Mak E, Fisher MM, Jeejeebhoy KN. Microfilament dysfunction as a possible cause of intrahepatic cholestasis. Gastroenterology. 1975 Jul;69(1):48–58. [PubMed] [Google Scholar]
• Phillips MJ, Oshio C, Miyairi M, Smith CR. Intrahepatic cholestasis as a canalicular motility disorder. Evidence using cytochalasin. Lab Invest. 1983 Feb;48(2):205–211. [PubMed] [Google Scholar]
• Laishes BA, Williams GM. Conditions affecting primary cell cultures of functional adult rat hepatocytes. 1. The effect of insulin. In Vitro. 1976 Jul;12(7):521–532. [PubMed] [Google Scholar]
• McDowell EM, Trump BF. Histologic fixatives suitable for diagnostic light and electron microscopy. Arch Pathol Lab Med. 1976 Aug;100(8):405–414. [PubMed] [Google Scholar]
• Sato T. A modified method for lead staining of thin sections. J Electron Microsc (Tokyo) 1968;17(2):158–159. [PubMed] [Google Scholar]
• Wehland J, Stockem W, Weber K. Cytoplasmic streaming in Amoeba proteus is inhibited by the actin-specific drug phalloidin. Exp Cell Res. 1978 Sep;115(2):451–454. [PubMed] [Google Scholar]
• Wehland J, Osborn M, Weber K. Phalloidin-induced actin polymerization in the cytoplasm of cultured cells interferes with cell locomotion and growth. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5613–5617. [PMC free article] [PubMed] [Google Scholar]
• Petzinger E, Frimmer M. Comparative studies on the uptake of 14C-bile acids and 3H-demethylphalloin in isolated rat liver cells. Arch Toxicol. 1980 Mar;44(1-3):127–135. [PubMed] [Google Scholar]
• Petzinger E. Competitive inhibition of the uptake of demethylphalloin by cholic acid in isolated hepatocytes. Evidence for a transport competition rather than a binding competition. Naunyn Schmiedebergs Arch Pharmacol. 1981 Jul;316(4):345–349. [PubMed] [Google Scholar]
• Jones AL, Schmucker DL, Mooney JS, Ockner RK, Adler RD. Alterations in hepatic pericanalicular cytoplasm during enhanced bile secretory activity. Lab Invest. 1979 Apr;40(4):512–517. [PubMed] [Google Scholar]
• Kane AB, Young EE, Schanne FA, Farber JL. Calcium dependence of phalloidin-induced liver cell death. Proc Natl Acad Sci U S A. 1980 Feb;77(2):1177–1180. [PMC free article] [PubMed] [Google Scholar]
• Russo MA, Kane AB, Farber JL. Ultrastruct pathology of phalloidin-intoxicated hepatocytes in the presence and absence of extracellular calcium. Am J Pathol. 1982 Nov;109(2):133–144. [PMC free article] [PubMed] [Google Scholar]
• Simons TJ. A method for estimating free Ca within human red blood cells, with an application to the study of their Ca-dependent K permeability. J Membr Biol. 1982;66(3):235–247. [PubMed] [Google Scholar]
• Jewell SA, Bellomo G, Thor H, Orrenius S, Smith M. Bleb formation in hepatocytes during drug metabolism is caused by disturbances in thiol and calcium ion homeostasis. Science. 1982 Sep 24;217(4566):1257–1259. [PubMed] [Google Scholar]
• Godman GC, Miranda AF, Deitch AD, Tanenbaum SW. Action of cytochalasin D on cells of established lines. III. Zeiosis and movements at the cell surface. J Cell Biol. 1975 Mar;64(3):644–667. [PMC free article] [PubMed] [Google Scholar]