centrations at equilibrium is essential for
assessment of the uncertainty in binding
data now available.
PENTTI K. SIITERI
Reproductive Endocrinology Center,
Department of Obstetrics, Gynecology
and Reproductive Sciences,
University of California
San Francisco 94143
References
1. I. M. Klotz, Science 217, 1247 (1982); ibid. 220,
981 (1983).
2. P. J. Munson and D. Rodbard, ibid., p. 979.
3. G. Scatchard, Ann. N.Y. Acad. Sci. 51, 660
(1949).
4. J. F. Tait and S. Burstein, in The Hormones, G.
Pincus, K. V. Thimann, E. B. Astwood, Eds.
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29 June 1983; accepted 27 October 1983
Alcohol-Induced Tolerance in Mitochondrial Membranes
other modulations of membrane structure and function. . . (6). Since Lieber
and his colleagues (1) analyzed only total
fatty acid composition of the mitochondrial membranes, and cardiolipin is a
minor component, they could not detect
this increased saturation of cardiolipin
acyl chains. Nevertheless, their data regarding total fatty acid composition actually confirm our findings of a significant
increase in stearic acid and a decrease in
palmitic acid in both phosphatidylcholine and phosphatidylethanolamine.
Our evidence for increased resistance
to disordering by ethanol is based on
studies with electron paramagnetic resonance (EPR) spin probes (2). The order
parameter, measured by 5-doxyl stearic
13 JANUARY 1984
acid or 12-doxyl stearic acid, is significantly decreased by low concentrations
of ethanol in liver mitochondria from
normal rats but not in ethanol-fed rats.
Similarly, the partition of doxyl-decane
is greatly enhanced by ethanol in control
rats but not ethanol-fed rats. Similar
results were obtained earlier by several other groups in synaptosomal membranes and red blood cells (7) and more
recently by us in liver microsomes (8).
We, therefore, believe that this is a general phenomenon relevant to all membranes in all tissues (9). It is necessary to
explain why Lieber and his colleagues
could not confirm this observation in
their studies. We have found that the
resistance to disordering by ethanol is
observed at high temperature (35°C) but
not at low temperature (15°C) (2). Lieber
and his colleagues measured 12-(9-anthroyloxy) stearic acid (12 AS) fluorescence anisotropy at 28°C, where the difference, if it exists, is expected to be
small. The sensitivity of 12 AS anisotropy to small structural changes is at least
one order of magnitude lower than that
of the EPR technique, particularly in
highly scattering membranes such as mitochondria. In fact, we suspect that scattering artifacts were not properly corrected for in their studies. Vanderkooi
and Chance (10) studied fluorescence
anisotropy of 12 AS in mitochondria at
200 to 45°C (Fig. 3 in 10); their measured
polarization values ranged from 0.125 to
0.1. This value corresponds to an anisotropy range of 0.087 to 0.069. We measured the fluorescence anisotropy of 12
AS in mitochondria and obtained a value
of 0.08 at 28°C. This value is in excellent
agreement with those obtained by Vanderkooi and Chance (10), but is one-third
those reported by Lieber and his colleagues (1). Nevertheless, because of the
low values of fluorescence anisotropy
and the large corrections for light scattering, the effect of low concentrations of
ethanol on membrane fluidity cannot be
easily detected. We suspect that Lieber
and his colleagues, in fact, measured the
Philadelphia, Pennsylvania 19102
References and Notes
Gordon, J. Rochman, M. Arai, C. S.
Lieber, Science 216, 1319 (1982).
2. H. Rottenberg, D. E. Robertson, E. Rubin, Lab.
Invest. 42, 318 (1980); H. Rottenberg, A. J.
Waring, E. Rubin, Science 213, 583 (1981); H.
Rottenberg, T. Ohnishi, E. Rubin, Arch. Biochem. Biophys. 216, 51 (1982).
3. A. I. Cederbaum, C. S. Lieber, E. Rubin, Arch.
Biochem. Biophys. 165, 560 (1974).
4. W. S. Thayer and E. Rubin, J. Biol. Chem. 256,
6090 (1981).
5. J. Burke and E. Rubin, Lab. Invest. 41, 393
(1979).
6. A. Waring, H. Rottenberg, T. Ohnishi, E. Rubin, Proc. Natl. Acad. Sci. U.S.A. 78, 2582
(1981).
7. J. H. Chin and D. B. Goldstein, Science 196, 684
(1977); D. A. Johnson, N. M. Lee, E. R. Cook,
H. W. Loh, Mol. Pharmacol. 15, 739 (1979).
8. B. C. Ponnappa, A. J. Waring, J. B. Hoek, H.
Rottenberg, E. Rubin, J. Biol. Chem. 257, 10141
(1982).
9. E. Rubin and H. Rottenberg, Fed. Proc. Fed.
Am. Soc. Exp. Biol. 41, 2465 (1982).
10. J. M. Vanderkooi and B. Chance, FEBS Lett.
22, 23 (1972).
2 August 1982; accepted 12 November 1982
1. E. R.
Gordon et al. (1) examined the basis of
challenging theory proposed by Rottenberg and his colleagues (2) that the
"chronic consumption of ethanol induces an adaptation of membrane composition causing increased membrane rigidity (decreased fluidity).... The increased rigidity impairs normal membrane function'. . . but in the presence
of moderate concentrations of ethanol
the membrane becomes sufficiently fluid
to resemble normal membranes (depena
dence)" (2, 3).
To examine this theory, we designed a
controlled nutritional experiment, which
included a group of Chow-fed rats (for
which respiratory functions have been
well defined), ethanol-fed rats, and the
pair-fed controls of the latter. Mitochondrial membranes from the Chow-fed animals contained a larger amount of saturated fatty acids than the mitochondrial
preparations from the pair-fed controls
and were more resistant to the fluidizing
effects of ethanol, although respiratory
functions in the membranes of the two
groups were similar. In contrast, the
193
Downloaded from http://science.sciencemag.org/ on April 25, 2020
Lieber and his colleagues (1) state that
they have found no evidence for a correlation between the structure of membrane phospholipids and function of
mitochondrial membranes from ethanolfed rats, or for resistance to disordering
by ethanol, which we reported earlier
(2). However, extensive studies in our
laboratory show that the decreased rate
of respiration, first described by Cederbaum et al. (3) in intact mitochondria,
can be explained as a direct consequence
of decreased content and activity of individual protein components of the respiratory chain in mitochondrial inner membranes (4). What causes this decrease is
not clear. It might be due to direct inhibition of mitochondrial protein synthesis
(5) or to interference with membrane
assembly. The latter may be influenced
by the phospholipid composition of the
membranes. Moreover, the respiratory
activity indeed may be influenced by the
phospholipid composition. We have
shown that mitochondrial membranes
from ethanol-fed rats display an increased saturation in the acyl chains of
cardiolipin (6), an essential phospholipid component of the electron transport
chain, which may contribute to the regulation of the respiration rate. Therefore,
we suggested that "The phospholipid
composition ... probably plays a role in
considerable effect of alcohol on light
scattering, which is caused by mitochondrial swelling.
In summary, there is sufficient evidence from studies of rat liver mitochondria and other membrane systems to
indicate that chronic alcoholism is associated with changes of membrane structure, composition, and function and that
these changes lead to tolerance to the
acute effects of ethanol.
H. ROTTENBERG
A. WARING
E. RUBIN
Department of Pathology and
Laboratory Medicine, Hahnemann
University School of Medicine,
Alcohol-induced tolerance in mitochondrial membranes
H Rottenberg, A Waring and E Rubin
Science 223 (4632), 193.
DOI: 10.1126/science.6691147
http://science.sciencemag.org/content/223/4632/193.1.citation
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