AJP - Regulatory, Integrative and Comparative Physiology, Vol 267, Issue 3 695-R704, Copyright © 1994 by American Physiological Society
Nocodazole inhibition of organic anion secretion in teleost renal proximal tubules
D. S. Miller and J. B. Pritchard
Intracellular Regulation Sections, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709.
The impact of the microtubule-disrupting drug nocodazole on renal tubular
secretion of organic anions was examined in vitro using proximal tubular
masses from teleost fish. Nocodazole reversibly inhibited 20-30% of the
tubular accumulation of two model organic anions, p-aminohippurate and
fluorescein (FL), by winter flounder tubular masses. However, the drug had
no effect on the initial rate of organic anion uptake. Thus it did not
reduce transport into the cells at the basolateral membrane, either
directly by affecting basolateral organic anion transport proteins or
indirectly by altering metabolism or ion gradients. Instead,
epifluorescence video microscopy and digital image analysis of killifish
tubules showed that nocodazole greatly reduced luminal accumulation of FL
and had a smaller effect on cellular dye accumulation. Luminal FL
accumulation returned to control levels when tubules were incubated in
drug-free medium. Confocal fluorescence microscopy confirmed the marked
reduction in luminal FL concentration and demonstrated that intracellular
punctate FL accumulation was also markedly reduced. Finally,
immunohistochemistry with an anti-tubulin antibody showed that the
concentrations of nocodazole used in the above experiments reversibly
disrupted microtubules within renal epithelial cells. These data indicate
that a component of organic anion secretion in teleost proximal tubule is
dependent on an intact microtubular network.
