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Title:The effects of the tubulin-binding drug, colchicine, on the electrophysiological aspects of phototransduction in toad rod photoreceptors
Author(s):Bert, Robert James
Doctoral Committee Chair(s):Oakley, Burks, II
Department / Program:Biology
Discipline:Biology
Degree Granting Institution:University of Illinois at Urbana-Champaign
Degree:Ph.D.
Genre:Dissertation
Subject(s):Biology, Neuroscience
Biology, Animal Physiology
Biophysics, General
Abstract:Colchicine, known for its antimitotic properties, its effects on neuronal conductances and its use as a treatment for acute gout, was found to affect phototransduction in toad rods during pharmacologic screening of cytoskeletal antagonists. Colchicine superfusion onto intact retinas hyperpolarized rods, increased their membrane resistance and altered the kinetics of voltage responses to light flashes. These effects persisted during blockade of rod inner segment conductances, but colchicine's effects during illumination were minimal. The presence of colchicine, however, did not block rod depolarization evoked by increasing intracellular 3$\sp\prime$,5$\sp\prime$-cyclic guanosine monophosphate, cGMP (the physiologic activator of the light-sensitive conductance, g$\sb{\rm h\nu}$), either physiologically or exogenously. It was concluded that colchicine blocks the light-sensitive current, probably by lowering (cGMP) $\sb{\rm i}$ rather than by acting as a cGMP antagonist at the channel.
The effects of colchicine on rod voltage are similar to the long-known effects of increasing intracellular free calcium, (Ca$\sp{2+}$) $\sb{\rm i}$, so it was hypothesized that colchicine either evoked an increase in (Ca$\sp{2+}$) $\sb{\rm i}$ or acted as a Ca$\sp{2+}$ agonist. The following results supported these hypotheses. Lowering (Ca$\sp{2+}$) $\sb{\rm i}$ either by injecting the Ca$\sp{2+}$ buffer, EGTA, or by lowering (Ca$\sp{2+}$) $\sb{\rm o}$ in the presence of colchicine, reversed the hyperpolarizing effect, but not the desensitizing effects of colchicine. Adding colchicine to low-Ca$\sp{2+}$superfusates reversed the electrophysiologic changes evoked by lowering (Ca$\sp{2+}$) $\sb{\rm o}$ to 10$\sp{-5}{-}$10$\sp{-6}$ M, but colchicine only partially and transiently reversed the desensitization seen when (Ca$\sp{2+}$) $\sb{\rm o}$ was lowered to 10$\sp{-7}{-}$10$\sp{-8}$ M. Colchicine, however, inhibited desensitization when superfused simultaneously with 10$\sp{-7.5}$ M Ca$\sp{2+}$. Colchicine superfusion reversibly attenuated depolarizations evoked by pressure-injections of EGTA.
Measurements of (Ca$\sp{2+}$) $\sb{\rm o}$ during colchicine superfusion with ion-selective electrodes demonstrated colchicine-evoked increases in (Ca$\sp{2+}$) $\sb{\rm o}$. This effect persisted, but was attenuated, when colchicine application was preceded by saturating steps of light, consistent with a colchicine-evoked release of intracellular Ca$\sp{2+}$. It is concluded that colchicine releases Ca$\sp{2+}$ from internal stores, but colchicine's effects on g$\sb{\rm h\nu}$ probably result from a Ca$\sp{2+}$ agonistic effect of the drug on enzymes regulating cGMP synthesis.
Colchicine's effects were discernible over the range 10$\sp{-4}{-}$10$\sp{-2}$ M. Application of 0.5 mM $\beta$-lumicolchicine, a low affinity, low-solubility colchicine analog, produced effects similar to colchicine's at this concentration. Another tubulin-binding drug, vincristine, had effects apparently opposite to those of colchicine. These data are inconsistent with microtubule disruption mediating colchicine's effects, but whether membrane tubulin mediates colchicine's effects on rods, as it does in $\beta$-adrenergic-sensitive heart and brain cells, is inconclusive.
Issue Date:1990
Type:Text
Language:English
URI:http://hdl.handle.net/2142/20601
Rights Information:Copyright 1990 Bert, Robert James
Date Available in IDEALS:2011-05-07
Identifier in Online Catalog:AAI9114176
OCLC Identifier:(UMI)AAI9114176


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