EPA-600/3-77-032
March 1977
Ecological Research Serie:
EFFECTS OF COPPER AND ZINC ON
SMOLTIFICATION OF COHO SALMON
Environmental Research Laboratory
Office of Research and Development
U.S. Environmental Protection Agency
Corvallis, Oregon 97330
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RESEARCH REPORTING SERIES
Research reports of the Office of Research and Development, U.S. Environmental
Protection Agency, have been grouped into five series. These five broad
categories were established to facilitate further development and application of
environmental technology. Elimination of traditional grouping was consciously
planned to foster technology transfer and a maximum interface in related fields.
The five series are:
1. Environmental Health Effects Research
2. Environmental Protection Technology
3. Ecological Research
4. Environmental Monitoring
5. Socioeconomic Environmental Studies
This report has been assigned to the ECOLOGICAL RESEARCH series. This series
describes research on the effects of pollution on humans, plant and animal
species, and materials. Problems are assessed for their long- and short-term
influences. Investigations include formation, transport, and pathway studies to
determine the fate of pollutants and their effects. This work provides the technical
basis for setting standards to minimize undesirable changes in living organisms
in the aquatic, terrestrial, and atmospheric environments.
This document is available to the public through the National Technical Informa-
tion Service, Springfield, Virginia 22161.
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EPA-600/3-77-032
March 1977
EFFECTS OF COPPER AND ZINC ON SMOLTIFlCATION OF COHO SALMON
by
Harold W. Lorz
Barry P. McPherson
Oregon Department of Fish and Wildlife
Corvallis, Oregon 97331
Grant #R 802468
Project Officer
Gary A. Chapman
Western Fish Toxicology Station
Corvallis Environmental Research Laboratory
Corvallis, Oregon 97330
CORVALLIS ENVIRONMENTAL RESEARCH LABORATORY
OFFICE OF RESEARCH AND DEVELOPMENT
U. S. ENVIRONMENTAL PROTECTION AGENCY
CORVALLIS, OREGON 97330
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DISCLAIMER
This report has been reviewed by the Corvallis Environmental Research
Laboratory, U. S. Environmental Protection Agency, and approved for publica-
tion. Approval does not signify that the contents necessarily reflect the
views and policies of the Environmental Protection Agency, nor does mention
of trade names of commercial products constitute endorsement or recommendation
for use.
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FOREWORD
Effective regulatory and enforcement actions by the Environmental
Protection Agency would be virtually impossible without sound scientific data
on pollutants and their impact on environmental stability and human health.
Responsibility for building this data base has been assigned to EPA's Office
of Research and Development and its 15 major field installations, one of which
is the Corvallis Environmental Research Laboratory (CERL).
The primary mission of the Corvallis Laboratory is research on the effects
of environmental pollutants on terrestrial, freshwater, and marine ecosystems;
the behavior, effects and control of pollutants in lake systems; and the de-
velopment of predictive models on the movement of pollutants in the biosphere.
This report describes a potentially adverse effect of pollutants on fish
such as salmon which must migrate from fresh water to sea water, and demon-
strates that under certain conditions exposure to sublethal levels of
pollutant can result in high mortality when fish subsequently enter sea water.
Laboratory test methods are described which should detect this effect in
screening tests and advance knowledge on the effects of pollutants on aquatic
ecosystems.
A.F- Bartsch
Director, CERL
I*
11
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ABSTRACT
The 96-h LC50 values for copper (CuC^) for yearling coho salmon
(Oncorhynchus kisutch) ranged from 74-60 yg/liter, depending on degree of
smol tif ication. The 96-h LC50 of zinc (ZnC^) for yearling coho in April
was 4600 yg/liter. All tests were conducted at 10 or 12 C in water with
alkalinity and hardness ranging from 68-78 rug/liter and 89~99 nig/liter as
CaC03, respectively.
Exposure of yearling coho for 144 h to sublethal concentrations of Zn
in fresh water had little effect on the activity of Na+, K+-activated ATPase
in gill microsomes or on the survival of fish when transferred to sea water.
Exposure of yearling coho to sublethal concentrations of Cu (5~30 yg/liter)
in fresh water (maximum of 172 days) had deleterious effects on downstream
migration in a natural stream, lowered gill ATPase activity and reduced
subsequent survival in sea water. Exposures >10 days had more severe effects
than did 6-day exposures on downstream migration and survival- in sea water,
but not on gill ATPase. Fish ceased feeding after initiation of exposures
to 20 and 30 yg/liter Cu and remained anoretic (loss of appetite) for
several weeks to 4 months with the result that the mean lengths and condition
factors of these fish were significantly lower than the controls at the end
of the test.
Exposure of yearling coho to Cu (20 and 30 yg/liter) in fresh water
affected their ability to maintain normal osmotic pressure and chloride
concentrations in blood plasma. Similarly when these Cu intoxicated fish
were transferred to sea water the plasma osmolality and chloride concentrations
increased significantly compared to control fish. These responses are at-
tributed, at least in part, to the suppression of Na+, K+-activated ATPase
activity in the gills.
Coho yearlings given a "rest" (non-toxicant exposure in fresh water)
following exposure to toxicant showed higher survival when tranferred to
sea water than fish which were transferred immediately.
This report was submitted in fulfillment of Grant R 802468 by the Oregon
Department of Fish and Wildlife under the partial sponsorship of the U.S.
Environmental Protection Agency. This report covers the period October 1, 1973r
to December 31, 1975-
IV
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CONTENTS
Di sclaimer i i
Foreword I i i
Abstract iv
Figures vi
Tab 1 es vi i i
Acknowledgments xi
1. Conclusions 1
2. Recommendations 3
3. Introduction k
k. Methods 6
Experimental fish 6
Exposures to toxicant 6
Tolerance to sea water 7
Downstream migration 10
Gill ATPase activity 10
Osmotic and ionic regulation 11
Assessment of coefficient of condition and
growth 11
Water quality 11
5. Results and Discussion 13
96-h LC50 experiments 13
Effect of copper on growth and survival 17
Effect of copper and zinc on gill ATPase and
survival in sea water 17
Effect of periods of rest following copper
exposure on survival in sea water 35
Effect of copper on osmotic and ionic
properties of blood plasma 40
Effect of copper on downstream migration.... kk
6. References 51
Append i ces 55-68
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FIGURES
Number Page
1 Diagrammatic sketch of flow-through diluter 8
2 Exposure tanks with diluter in background 9
3 Exposure tank with submersible pump for enhancing aeration,
current and mixing 9
k Effect of copper exposure on mortality of yearling coho salmon. 14
5 Median mortality-time of yearling coho salmon exposed to
solutions of copper 15
6 Effect of zinc exposure on mortality of yearling coho salmon... 16
7 Effect of chronic copper exposure on growth of juvenile coho
sa 1 mon 20
8 Effect of chronic copper exposure on coefficient of condition
of j uven i 1 e coho sa 1 mon 21
9 Influence of copper exposure in fresh water on gill microsomal
Na+, K+-activated ATPase activity in juvenile coho salmon 26
10 Influence of copper exposure in fresh water on gill microsomal
Na+, K+-activated ATPase activity in yearling coho salmon
chronically exposed to copper from December 20, 197^ 28
11 Survival of yearling coho salmon in sea water following
exposure to copper in fresh water for 144 h at various times
of the year 29
12 Survival of yearling coho salmon in sea water at various times
of the year following exposure to copper in fresh water
initiated on December 20, 1971* 32
13 Survival curves of yearling coho salmon during exposure to
copper in fresh water and subsequent survival when transferred
to sea water 33
]k Survival curves of yearling coho salmon exposed to copper in
fresh water and their subsequent survival upon transfer to sea
water 3^
vi
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Number Page
15 Effect of copper exposure in fresh water on the osmolality of
the plasma of coho salmon during 144 h of exposure ............ 41
16 Effect of sea water exposure on the osmolality of the plasma
of coho salmon previously exposed to copper (in fresh water)
for 144 h [[[ 43
17 Effect of sea water exposure on the osmolality of the plasma
of coho salmon chronically exposed to copper (in fresh water)
for 792 h in July 1974 ........................................ 45
18 Influence of copper exposure for 3960 h (165 d) in fresh water
on downstream migration of yearling coho salmon ............... 48
19 Influence of copper exposure in fresh water on downstream
migration of yearling coho salmon released April 30, 1975 ..... 49
APPENDIX FIGURES
Number Page
A-l Effect of copper exposure (in fresh water) on gill microsomal
Na+, K+-activated ATPase activity of yearling coho salmon 63
A-2 Effect of copper exposure in fresh water on the chloride ion
concentration of the plasma of coho salmon serially sampled
during 144 h of exposure 64
A-3 Effect of seawater exposure on the chloride ion concentration
of the plasma of coho salmon previously exposed to copper (in
fresh water) for 144 h 65
A-4 Effect of seawater exposure on the chloride ion concentration
of the plasma of coho salmon previously exposed to copper (in
fresh water) for 792 h : 66
A-5 Effect of seawater exposure on the sodium ion concentration
of the plasma of coho salmon previously exposed to copper (in
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TABLES
Number
1 Chemical and physical characteristics of test water (flow-
through and static water systems) 12
2 Percent survival, mean length, weight and coefficient of con-
dition of yearling coho salmon (1972 brood) exposed to copper
for 37 days (880 h) in 1971* 18
3 Percent survival, mean length, weight and coefficient of con-
dition of yearling coho salmon (1973 brood) exposed to copper
for 59 and 165 days in 1975 19
4 Survival and gill ATPase activity of yearling coho salmon
exposed to copper for 144 h in static fresh water and their
subsequent survival and gill ATPase activity after transfer
to sea water 22
5 Survival and gill ATPase activity of yearling coho salmon ex-
posed to zinc for \kk h in static fresh water and their
subsequent survival and gill ATPase activity after transfer
to sea water 2k
6 Survival and gill ATPase activity of yearling coho salmon ex-
posed to copper in fresh water and their subsequent survival
after transfer to sea water 25
7 Gill ATPase activity and survival in sea water of yearling
coho salmon exposed to copper in fresh water for <_lMt h
(tests conducted from March 6 to June 9, 1975) 27
8 Survival and gill ATPase activity of yearling coho salmon ex-
posed to copper in fresh water and their subsequent survival
after transfer to sea water 31
9 Comparison of survival of fish exposed to natural and artifi-
cial sea water following ]k7 and-172 days of copper exposure
in fresh water 35
10 Survival and gill ATPase activity of juvenile coho salmon ex-
posed to copper for \kk h and subsequent survival in sea
water with and without recovery period 3p
Vi i i
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Number
Page
11 Survival of yearling coho salmon in sea water following exposure
to copper for 1600 h or 144 h with or without recovery periods
(February 25-Apri 1 6, 1975) 38
12 Survival of yearling coho salmon in sea water following exposure
to copper for 172 days (4128 h) with or without recovery periods
(June 10-July 2, 1975) 39
13 Effects of freshwater rest or seawater acclimation (20 °/oo
salinity) on survival of yearling coho salmon in 30 °/oo sea
water following 115~125 d exposure to copper in fresh water
(April 14-May 9, 1975) 40
14 Percent migration through July 3, 1975, of coho salmon released
into a small coastal stream following acute and chronic copper
exposures 46
APPENDIX TABLES
Number Page
A-l Chemical and physical characteristics of well water at the
Oregon Department of Fish and Wildlife Laboratory, Corvallis,
Oregon 55
A-2 Effect of copper in fresh water on the survival of yearling coho
salmon (determination of 96-h LC50 values) 56
A-3 Effect of zinc in fresh water on the survival of yearling coho
salmon (determination of 96-h and 144-h LC50 values) 57
A-4 Effect of copper exposure on average length-weight and
coefficient of condition 58
A-5 Activity of microsomal Na+, K+-activated ATPase in gills of coho
following exposure to copper for 144 h and subsequent seawater
exposure 59
A-6 Activity of microsomal Na+- K+-activated ATPase in gills of
coho exposed to copper for various lengths of exposure in 1974. 60
A-7 Activity of microsomal Na+, K+-activated ATPase in gills of
coho exposed to copper for various time periods 61
A-8 Activity of microsomal Nat, K+-activated ATPase in gills of
coho exposed to copper for various time periods in 1975 62
IX
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Number
A~9 Percent migration (to July 3, 1975) of yearling coho salmon re-
leased into a small coastal stream following acute and chronic
copper exposure 68
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ACKNOWLEDGMENTS
This investigation was supported in part by the U.S. Environmental
Protection Agency, Research Grant R-802468, and was funded through the National
Water Quality Laboratory, Duluth Minnesota. Many contributed to this study and
their assistance is gratefully acknowledged: Dr. G. R. Bouck provided valuable
assistance in securing the grant. Dr. W. S. Zaugg and L. R. McLain, Western
Fish Nutritional Laboratory, U. S. Fish and Wildlife Service provided physical
facilities, technical assistance and help in the ATPase analyses run in 197^-
Dr. G. A. Chapman, Project Officer, Western Fish Toxicology Station, EPA
provided technical assistance and made arrangement with CERL for quarterly
chemical water analysis as well as arranged the assistance of Mr. Joel McCrady
(Chemist, WFTS) in analysis of copper and zinc concentrations used. Mr. Glen
Gross, Mr. Will Beidler and Miss Lynda Smart assisted in the laboratory and
field aspects of the work.
The authors gratefully acknowledge Drs. G. A. Chapman, J. D. Mclntyre,
A. V. Nebeker and H. H. Wagner who provided constructive criticism on the
manuscript.
XI
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SECTION 1
CONCLUSIONS
1. The 96-h LC50 of Cu for yearling coho ranged from 60-74 yg/liter. The
96-h LC50 of Zn for yearling coho was 4600 yg/liter. All tests were con-
ducted in water with an alkalinity of 68-78 mg/liter as CaCO,.
2. No deaths were observed when yearling coho were exposed for 6 days to con-
centrations of 20 or 30 yg/liter Cu; however, following exposure to Cu
for 172 days, mortalities of 12 and 38%, respectively, were recorded.
3- Coho yearling exposed to Cu concentrations >JO yg/liter showed anorexia
(loss of appetite) and significantly lower mean lengths and condition
factors than the control fish at the end of the exposure periods in 1974
and 1975 (37 and 172 days, respectively).
4. Gill microsomal Na , K -activated ATPase activity in yearling coho was
decreased proportionally to Cu concentration by exposure to sublethal
concentrations (10-30 yg/liter) whereas ATPase activity appeared unaf-
fected in yearling coho exposed to Zn (100-5000 yg/liter).
5. Exposure of yearling coho to Cu in fresh water affected their ability to
maintain normal blood plasma osmotic pressure and chloride concentration,
and similarly when these Cu intoxicated coho were transferred to sea water
an abnormal increase in plasma osmolality and chloride concentration was
observed.
6. The percent survival of yearling coho in sea water decreased in propor-
tion to the concentration of Cu in fresh water. Death in sea water was
due to loss of osmoregulatory ability, and decreased Na+, K+-activated
ATPase in the gills was probably one .of the factors contributing to this
loss.
7. Coho yearlings given a "rest" (nontoxicant exposure period) betwen Cu
exposure and transfer to sea water showed improved survival over those
transferred immediately to sea water.
8. The survival rate in sea water after a given exposure to Cu in fresh water
was greater as coho became older, larger and transformed into smolts.
9. Long-term exposures to Cu had more severe effects than did short-term
exposures on survival in sea water, but not on gill ATPase activity.
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10. All concentrations of Cu tested (5~30 yg/liter) caused a reduction in
percentage of downstream migrants compared to controls when the coho
yearlings were released into a natural stream.
11. The 0.1 application factor for copper (6.1 x 96-h LC50) probably predicts
"no effect" levels based on growth, direct mortality, latent seawater
mortality, and ATPase activity, but a "no effect" level based on migratory
success would fall below this level.
12. Two types of artificial sea water were found to be satisfactory substi-
tutes for natural sea water in the testing survival of yearling coho
after exposure to Cu.
13- Studies such as this that take into consideration critical points in
the life history of fish (or other aquatic organisms) are essential in
developing good water quality criteria.
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SECTION 2
RECOMMENDATIONS
1. The sensitive techniques and methodology developed in this study are
recommended and should assist EPA in developing more satisfactory water
quality criteria to be used for setting water quality standards.
2. Prudence in applying the results of this study directly and quantitative-
ly to field situations is recommended since variable factors (such as the
proportion of "biologically available" Cu, the presence of other
chemicals, disease, predators and food availability) could alter the
reported response of yearling coho to a given concentration of Cu.
3. Additional research should be conducted to determine if the interference
with smoltification of anadromous salmonids by sublethal levels of pol-
lutants is a general phenomenon and to further validate the methodology.
4. Another area of research potential would be to determine if similar
effects occur in euryhaline and stenohaline fish from estuarine and
marine environments.
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SECTION 3
INTRODUCTION
The problem of toxic effects of metals to fish as a result of water pol-
lution has been recognized and investigated since the early part of this
century. The early literature concerning this problem came primarily from
England where metal pollution of streams occurred because of mining activity.
A large volume of literature on the toxic effects of metals to fish has accu-
mulated in the last fifty years, and there have also been many reviews
prepared, of which Volumes 3 and 5 of the five volume Water Quality Criteria
Data Book (1970-73) and the review of Eisler (1973) and Eisler and Wapner
(1975) are among the most complete.
Considerable research has been done to determine water quality criteria
for copper and zinc based on survival and growth of juvenile salmonids in
fresh water (Chapman 1973; Hodson and Sprague 1975; Lloyd I960; McKim and
Benoit 1971; Sprague 1964; and Sprague and Ramsay 1965)- However, there are
essentially no data available for copper, zinc or any other pollutant which
relates to effects on the migration of anadromous fish into sea water. There
is a particular need for this type of data in the Pacific Northwest where
large runs of anadromous salmonids are a valuable sport and commercial fish-
ery resource and whose well-being is a significant factor in environmental
impact considerations.
The seaward migration of juvenile coho salmon (Onoorhynohus kisutoh)
normally occurs during the spring of their second year of life. They are
fully euryhaline several months earlier (Conte et al. 1966, Otto 1971) pro-
vided that they have achieved a threshold size of 9 cm (Conte et al. 1966).
The experimental transfer of juvenile anadromous salmonids from fresh water
to sea water is followed by a transient but marked disturbance of plasma
water-electrolyte balance (Conte et al. 1966; Miles and Smith 1968). This
osmotic disturbance is caused by the physiological changes necessary to adapt
from freshwater osmoregulation (salt retention, water excretion) to seawater
osmoregulation (salt excretion, water retention). These disturbances are
minimized at the time of normal seaward migration or "parr-smolt transforma-
tion."
Zaugg and Wagner (1973) presented data showing that one of the physio-
logical factors correlated with migratory behavior in steelhead trout (Salmo
gairdneri) is an elevation of activity in Na+, K+-stimulated adenosine
triphosphatase (ATPase) in the microsomes of gills. This enzyme activity
doubles during the parr-smolt transformation of coho salmon and steelhead
trout (Zaugg and McLain 1970; Zaugg and Wagner 1973). ATPase activity in
salmonids increases rapidly during seawater exposure reaching a maximum after
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about 30 days, and is thought to be an important component in maintaining
salt (osmotic and ionic) balance in fish (Epstein et al. 1967; Zaugg and
McLain 1970). Though no data concerning inhibition of Na+, K+-ATPase
activity in fish by heavy metals was found in the literature, several workers
have reported on "in vivo" inhibition of ATPase by chlorinated hydrocarbon
insecticides and polychlorinated biphenyls (Campbell et al. 197^; Koch et al.
1972; Leadem et al. 197*0- Jackim et al. (1970), however, reported on changes
in activities of five liver enzymes after exposing fish or fish liver homoge-
nates to salts of various toxic metals and these authors suggest that enzyme
assay is a valid technique for diagnosing sublethal metal poisoning of fish.
This study was designed to determine the effects of freshwater exposure
to sublethal concentrations of copper or zinc on the subsequent ability of
yearling coho salmon to adapt to sea water, and to determine the effects of
copper on downstream migration of yearling coho salmon. Growth, osmotic and
ionic regulation and microsomal gill adenosine triphosphatase, were monitored
during chronic exposure to copper in fresh water. A sub-objective of the
study was to investigate and recommend methods that could be routinely uti-
lized for measuring the effects of pollutants on the ability of anadromous
salmonid to migrate to the ocean and adapt to sea water.
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SECTION 4
METHODS
Experimental fish
All experiments were conducted on coho salmon between the ages of 10 and
18 months post-hatching. Coho salmon of the 1972 and 1973 year classes a/
originating from Fall Creek Salmon Hatchery (Alsea River, Oregon) were used
in 1974 and 1975 experiments, respectively. Fish were hatched and reared from
fertilized eggs brought into the laboratory and incubated at temperatures that
ranged from 7-8 to 12.3 C. Rearing was done under natural light or simulated
natural photoperiod in well water at a constant temperature of 12.3 C. In
197*1, a constant temperature (12.3 C) was used throughout rearing and exposure
to toxicant. In 1975, experimental fish (stock and exposure groups) were
reared and exposed to toxicant at 12.3 C until February 1. Fish were then
acclimated to 8.6 C over a period of 6 days and exposed to a simulated natural
temperature regime, increasing in semi-monthly increments from 8.6 C on
February 7 to 12.3 C on May 19. The temperature pattern used was based on
mean semi-monthly maximal and minimal temperatures (6-year averages) on the
North Fork of the Alsea River.
Fish were fed a commercially prepared moist pellet generally to reple-
tion. Fish were not fed during the 2k h preceding exposure to toxicant,
during acute (
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All exposures of more than 144 h, as well as some
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oo
Water inlets with
spray heads (well. Float (connected via microswitch to toxicant manifold)
heated or chilled water) ( / /
-Toxicant
Head Box
Distribution pipes
Air stone
Microswitch and
valve (allows
dumping toxicant
water or
electricity fails,
waste
water
To Control Tanks
2.6 m
1.5m
Toxicant
Pump
Covered
Toxicant
Reservoir
Figure 1. Diagrammatic sketch of flow-through diluter.
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Figure 2. Exposure tanks with diluter in background.
Figure 3. Exposure tank with submersible pump for enhancing
aeration, current and mixing.
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(Rila Products of New Jersey) a/ to which water was added to achieve the
desired salinity, and an artificial sea water prepared following the proce-
dure of LaRoche et al. (1970). A batch of each artificial sea water was
mixed daily in a separate container prior to introduction to the test tanks.
During the seawater tests, dissolved oxygen content of the water was main-
tained at 6.4-8.5 mg/liter and pH at 7-4-8.1. The total ammonia ranged from
0.04 to 1.8 mg/liter NH3-N due to the daily exchange of 85% of the water in
the test tanks.
Downstream migration
The effect of Cu on migration was assessed by releasing marked control
and Cu exposed fish into a tributary of the North Fork of the Alsea River
and monitoring their arrival at a trap 6.4 km downstream. The trap was built
into a permanent weir and was usually checked daily. On the day prior to
release, 50 to 100 fish from each exposure tank were anesthetized in MS-222,
weighed, measured, and identified by freeze branding and a fin clip. Parr-
smolt transformation is markedly size dependent and seasonal with wild coho
juveniles normally spending 1 year in the natural stream before migrating
seaward provided they attain a minimal size of 7 cm (Lorz and Mason, unpub-
lished data). Under artificial propagation most coho reach a size of 11-15
cm in less than 1 year. No fish less than 10 cm were released. Seaward mi-
gration of wild juveniles normally begins in April, peaks in late May and
ceases in late June. Releases of Cu exposed fish and their controls were
made between April 8 and June 4, 1975. Trapping was terminated in early July,
one month after the last release.
Gill ATPase activity
The activity of Na+, K+-activated ATPase in microsomes isolated from
gill filaments was assayed by the procedure of Zaugg and McLain (1970). The
activity was measured in three to six fish per metal concentration prior to
each release into the stream and before most seawater tests. Assays of ATPase
were also made on survivors of two seawater tests.
No inhibition of ATPase activity occurred when CuCl2-2H20 was added to
homogenized gill tissue from control fish. Final concentrations of Cu in
the spiked homogenates (250 vg Cu/gm of gill filament tissue) were 100 times
the concentration found in the whole gills of those coho exposed for 6
months to 20 yg/liter Cu in fresh water (Chapman, personal communication).
This suggests that the decreased ATPase activity observed in gills from fish
exposed to Cu was not an in vitro effect of Cu ions liberated during the
homgenization procedure. Any free Cu ions would probably be bound by the
EDTA (5mM) in the homogenizing solution (W.S. Zaugg, personal communication).
a/ Mention of commercial product does not constitute endorsement or recom-
mendation for use.
10
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Osmotic and Ionic regulation
Changes in osmolality and concentrations of Cl~ and Na+ in blood plasma
were followed by sacrificing groups of yearling fish for blood at various
times after the start of toxicant or seawater exposure. The caudal peduncle
was wiped dry, wrapped in a tissue to prevent dilution of blood by water or
mucous and then severed. Blood flowing from the caudal artery was deposited
onto a small piece of ParafilmR and immediately transferred via polyethylene
tubing into a small polyethylene microcentrifuge tube and centrifuged in a
Beckman Microfuge (5500 g) for 1 minute. The supernatant plasma was trans-
ferred into a clean micro-centrifuge tube, centrifuged for 30 seconds and
then frozen until such time as the micro-analysis could be done. The sodium
concentration in plasma was measured by atomic absorption on 1/900 distilled
water dilution of plasma utilizing a Perkin-Elmer Model 306 B atomic absorp-
tion spectrophotometer- Chloride ion was measured directly with a Corning
Model 920 chloride meter. The osmolality of the plasma was measured on a
Wescor Model 5100 vapor pressure osmometer calibrated with standard solutions
of sodium chloride.
Assessment of coefficient of condition and growth
Parr-smolt transformation in several species of Salmo and Oncorhynchus
is characterized by changes in coefficient of condition. A marked decrease
in condition occurs for fish undergoing the transformation, followed by an
increase in condition in fish reverting to a nonmigratory form (Hoar 1939;
Malikova 1974; Vanstone and Markert 1968; Fessler and Wagner 1969; Finder and
Eales 1969; and Wagner 197*0.
Each month, 20 to 50 fish were randomly selected from each exposure tank
after a 24-h starvation period, anesthetized, weighed and measured. Individ-
ual fish >JO cm were weighed to 0.1 g, smaller fish were weighed to 0.01 g,
and fork Tength was determined to 0.1 cm. The coefficient of condition (K)
was determined for each fish in a sample using the formula K = 100 W/L^,
where W denotes weight in grams and L denotes fork length in centimeters
(Nile 1936, Hoar 1939).
Water quality
The source of water for the study was a well (approximately 1 mile east
of Willamette River, Linn County, Oregon) and the chemical characteristics
were measured quarterly by the Corvallis Environmental Research Laboratory
(CERL) of the EPA and the data is presented in Table A-l. In addition
alkalinity, hardness, dissolved oxygen, pH and ammonia in the exposure tanks
were measured routinely at the laboratory and these chemical characteristics
are presented in Table 1. Water temperatures were monitored continuously in
both the static and flow-through systems with continuous recording thermo-
graphs.
11
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Table 1. Chemical and physical characteristics of test water (flow-through and static water systems) a/
Characteristic
Alkalinity
Hardness
Dissolved oxygen
Ammon i a
PH
(static water)
(flow-through)
(static water)
(flow-through)
(static water)
(flow-through)
(static water)
(flow-through)
Year Unit
1974 mg/liter as CaCO?
1975
1974 mg/1 iter as CaC03
1975
1974 mg/liter
197**
1975
1975
1974 mg/liter NH3~N
1975
1974
1974
1975
1975
Mean ±
66
72
95
93
10
9
9
8
0
0
7
7
7
7
.0
.7
.8
.7
.21
+
±
±
±
±
4
+
+
1
387 ±
.34
.41
.30
.26
b/
b/
b/
b/
SD
4
3
3
3
0.
0.
0.
0.
0.
7
3
3
9
183
Range
57
66
87
84
8.1 -
9.2 -
9.4 -
6.2 -
0.124-
0.124-
6.74 -
7.27 -
6.81 -
7.07 -
70
81
100
98
10
10
10
10
0
0
7
7
7
7
.7
.2
.1
.9
.297
.570
.96
.62
.54
.54
Number of
analyses
28
49
30
32
43
18
8
52
2
7
56
19
6
42
a/ Standard Methods for the Examination of Water and Wastewatev, llth Edition 1960.
b/ Median value.
-------
SECTION 5
RESULTS AND DISCUSSION
In the 20 months of study, six static bioassays with Cu and four with Zn
were completed. These included determinations of the 96-h and lM»-h LC50
values, effect of metal ion on gill microsomal ATPase, and ability of juvenile
coho to adapt to sea water following exposure to toxicant in fresh water.
Twelve tests of tolerance to sea water were completed using yearling coho
exposed to Cu in a flowing freshwater toxicant system. In addition two tests
comparing artificial sea salts to natural sea water; four tests of survival
in sea water following periods of freshwater recovery from toxicant exposure;
and one test of seawater survival after toxicant free acclimation to lower
salinity were conducted. During the 1975 migratory period four releases of
coho were made into a small coastal stream and the downstream movement of the
migrants monitored.
96-h LC50 experiments
The 96-h LC50 of Cu for yearling coho was determined at different times
during the parr-smolt transformation. In late November it was 7k yg/liter;
in March it dropped to 70 yg/liter; and by late May had further declined
to 60 yg/liter Cu (Figure k, Table A-2). The greater sensitivity to acutely
lethal effects of Cu in May was probably due to the onset of smoltification
rather than increasing age or size of fish. This physiological transforma-
tion, which preadapts coho to seawater existence apparently increased their
susceptibility to lethal effects of Cu in fresh water. The observed increase
in susceptibility was small and may have been modified slightly in either
direction because the earlier tests were conducted at a higher temperature
(12 C) than the May test (10 C) and the interactions between temperature
and metal toxicity are complex (Hodson and Sprague 1975)-
Amounts of Cu greater than background were found in the controls and
some of the test tanks containing the lower Cu concentrations during the de-
termination of the 96-h LC50 values (Table A-2). The Cu level of the well
water was normally 2 yg/liter but additional contamination from the brass
head of the mixing chamber's water pump and inability to remove all Cu residue
by rinsing the mixing chamber with water between concentration changes
apparently occurred. When the "incipient lethal level" (ILL) was plotted
following the technique of Sprague (1964), the ILL for Cu was close to 75 yg/
liter for the November test and 65-70 yg/liter for the March tests (Figure 5).
As noted by Sprague (1964), a sharp differentiation between lethal and non-
lethal concentrations of Cu occurred and the line relating concentration to
survival time broke and ran nearly parallel to the time axis. The longest
LT50 at any concentration was 58 hours for one test tank of fish in March
Generally the LT50 for Cu occurred from 19 to 35 h.
13
-------
100
0
o
+-
o
w.
+-
C
O)
o
c
o
o
50
«*-
3So
fip
«*
1
# Mar. 7- II, 1974
D Mar. 20-24,1974
X Nov. 20-24,1974
May 5-9, 1975
o May 27-31,1975
10 30 50
Mortality
70
90
99
Figure
Effect of copper exposure on mortality of yearling coho salmon.
-------
3
o
X
o
w.
o
o
If)
CD
£
100
80
60
40
20
444-
x x o
Nov. 20,
Mar. 7 420, 1974
I I
V 30 50 70 100
Concentration of Metal jug/liter Cu
Figure 5- Median mortality-time of yearling coho salmon exposed to solutions
of copper.
15
-------
Our 96-h LC50 values of Cu for coho salmon are higher than those re-
ported by Sprague (1964) for Atlantic salmon (.ILL of 48 yg/liter) and Lloyd
and Herbert (1962) for rainbow trout (ILL of 50 yg/liter). However, the
alkalinity and hardness of our water, 66 and 95 mg/liter CaCO^ respectively,
were about four times greater, our fish were larger, and were of a different
genus than those used in the cited studies. Chapman (unpublished data) found
the 96-h LC50 of juvenile coho salmon from the same Alsea River stock to be
28-38 yg/liter Cu at the Western Fish Toxicology Station, Corvallis, Oregon,
with a water hardness and alkalinity of 20-25 mg/liter CaCO^.
The 96-h LC50 of Zn for juvenile coho was 4600 yg/liter at 12 C
(Figure 6A, Table A-3). It appeared that there was increasing sensitivity to
Zn as the fish underwent smoltification; however, as only a few Zn concentra-
tions were run in May and June all data were combined for the single estimate.
The 144-h LC50 of Zn for the juvenile coho was 4200 yg/liter at 12 C
(Figure 6B). The longest LT50 in the three tests was 144 h for one test tank
in June 1974. Generally the LT50 for Zn occurred from 30-85 h. Our 96-h
LC50 value for Zn is again higher than that reported by other investigators
(Sprague 1964, Chapman 1973, Hodson and Sprague 1975)- However, our con-
siderably greater alkalinity and water hardness may again account for this
difference. Similarly the solubility of zinc salts decrease markedly as pH
rises above Z.O (Sprague 1964) and, therefore, our higher 96-h LC50 values
could also be related to small pH differences.
§
O April 16-20, 1974
X May 9-13, 1974
A June S- 9, 1974
C
* *
o *
C
o
o
o
O April 16-22,1974
X May 9-15,1974
A June 5-11 , 1974
10 50 7O
Mortality
90
Figure 6. Effect of zinc exposure on mortality of yearling coho salmon,
16
-------
Effect of copper on growth and survival
The longest period of juvenile coho exposure to Cu (in fresh water)
during 1974 was 37 days (880 h) . The exposure occurred in June and early
July at a time after the parr-smolt transformation and majority of juvenile
growth had occurred. The fish exposed to 20 and 30 yg/liter Cu immediately
ceased feeding and only near the end of the 37-day exposure did they again
begin to feed. Consequently, the mean lengths and the mean coefficient of
condition (K) of the fish exposed to 20 and 30 yg/liter Cu were significantly
lower than the control values at the end of the test (Table 2). In the con-
trol through 20 yg/liter Cu groups almost no mortality occurred through 37
days of exposure whereas the 30 yg/liter group incurred about 30% mortality
(Table 2). The mortality in the 30 yg/liter groups occurred after the first
144 h of exposure.
In 1975 the effects of Cu exposure in fresh water on growth, survival
and coefficient of condition of juvenile coho salmon showed a similar corre-
lation with Cu concentration as it did in 1974. The 30 yg/liter groups
showed the poorest survival, growth and coefficient of condition (Table 3).
Survival of juvenile coho in the control through 20 yg/liter groups was
similar after 59 days (I4l6 h) of toxicant exposure although K of the
20 yg/liter coho group was lower. After 6 months of exposure (Dec. -June) the
20 and 30 yg/liter Cu groups had incurred mortalities of 9 and 38%, respec-
tively (Table 3)- A second group of fish exposed for 10 weeks (March-June)
to 30 yg/liter Cu exhibited a mortality of 29%. As noted in 1974, fish in
the 20 and 30 yg/liter Cu groups again ceased feeding upon initiation of
the toxicant exposure. Resumption of feeding in the higher toxicant concen-
tration groups was not resumed until mid-May and then only with reduced
intensity. Feeding was slightly inhibited by exposure to 10 yg/liter Cu.
Fish in the control through 10 yg/liter groups were fed an amount equivalent
to that eaten by the 10 yg/liter fish in an effort to minimize differences in
size of fish among the test concentrations. Fish in the stock rearing tank
exhibited the best growth as they were fed to repletion daily; whereas in
the experimental tanks the controls exhibited the maximum growth and the others
others grew according to the Cu concentration received (Figure 7).
The effect of Cu exposure which was best correlated to Cu concentration
was the coefficient of condition (K) (Figure 8, Table A-4) with the fish
receiving 20 or 30 yg/liter Cu losing condition rapidly after the initiation
of the Cu exposure. The 20 and 30 yg/liter fish evidenced a recovery of
feeding in late April and this was reflected in elevated K's in early June.
The coefficient of condition of a group of juveniles exposed to 30 yg/liter
Cu for 10 weeks (March-June) showed a rapid decline in April, leveled off
in May and showed a very slight increase in June with a resumption of feeding.
Effect of copper and zinc on gill ATPase and survival in sea water
When fish were exposed to Cu for 144 h in static fresh water the
ATPase activity was decreased in proportion to the concentration of Cu. The
percent survival in sea water was also decreased in proportion to the concen-
tration of Cu (Table 4, Table A-5) . The decreased ATPase activity was
17
-------
Table 2. Percent survival, mean length, weight and coefficient of condition of yearling coho salmon
(1972 brood) exposed to copper for 37 days (880 h) in 1971*
Nominal con- Percent survival
centration in toxicant a/ Mean length b/ ± SE Mean weight b/ ± SE Mean condition b/ ± SE
pg/1 iter Cu Rep. 1 Rep. 2 Rep. 1 Rep. 2 Rep. 1 Rep. 2 Rep. 1 Rep. 2
0 100.0 99-5 17.4 ± 0.184 17-5 ± 0.153 56.3 ±1.91 56.0+1.55 1.057 ±0.0096 1.042 ±0.0078
5 100.0 100.0 17.5 ± 0.184 17.4 ± 0.211 56.9 ±2.17 55.6+2.21 1.041+0.0092 1.034+0.0088
10 100.0 100.0 17.6 + 0.181 17-7 ± 0.189 55-1 ± 1.80 57.2 ± 1.82 1.004 ± 0.0085 1.023 ± 0.0087
20 100.0 99.0 17.1 ± 0.212 16.9 ± 0.203 49.5 ± 1.93 48.4 ± 1.94 0.967 + 0.0092 0.988 + 0.0096
30 68.3 73.9 16.3 ± 0.180 16.2 ± 0.180 38.8 ± 1.55 38.6 ± 1.60 0.884 + 0.0126 0.893 1 0.0139
a/ Originally placed 225-250 fish in each tank. Survival calculated on number of fish remaining in tank following
~ removal of fish for survival tests in sea water after exposures to toxicant of 144, 535, 880 h. Average survival
calculated as the product of the percent survival during each time period.
CO
b/ Sample size of 40 fish per test tank.
-------
Table 3. Percent survival, mean length, weight and coefficient of condition of yearling coho salmon
(1973 brood) exposed to copper for 59 and 165 days in 1975
Nominal con- Percent survival
centration in toxicant a/ Mean length ± SE Mean length ± SE Mean condition ± SE
ug/liter Cu Rep. 1 Rep. 2 Rep. 1 Rep. 2 Rep. 1 Rep. 2 Rep. 1 Rep. 2
A. 1416 h Cu exposure (59 d, December 20, 1974-February 17,. 1975)
0 99.6 99.9 15-1 ± 0.361 b/ 15-3 ± 0.276 b/ 40.9 ± 2.87 41.6 ± 2.30 1.160 ± 0.0174 1.149 i 0.0175
5 99.9 100.0 14.7 ± 0.303 V 15.3 ± 0.417 &/ 35-5 ± 2.26 41.5+3.46 1.102 ±0.0145 1.118 ±0.0185
10 99.9 99.6 14.5 ± 0.434 b/ 14.3 ± 0.205 b/ 36.6 ± 4.09 33.6 ±2.10 1.132+0.0248 1.127 ±0.0330
20 98.6 99.5 14.6 ± 0.290 bj 13.7 ± 0.315 b/ 33.0 ± 2.28 26.0 + 1.90 1.028 + 0.0199 0.988 ± 0 016?
30 83.7 74.3 13-3 ± 0.247 a/ 12.8 ± 0.261 c_/ 22.7 + 1.619 19.4 + 1.371 0.957 ± 0.0227 0.915 ± 0.0213
B. 3960 h Cu exposure (165 d, December 20, 1974-June 3, 1975)
0 99.4 99.4 17.2 ± 0.224 d/ 17.9 ± 0.289 d/ 56.2 ± 2.41 64.6 ± 3-51 1.083 + 0.0099 1.096 + 0.0120
5 97.0 96.8 17.1 ± 0.303 £/ 17.4 ± 0.334 ej 54.1 ± 3.81 57-5 ± 3.81 1.021 ± 0.0119 1.031 ± 0.0184
10 98.4 97.9 16.1 + 0.297 ej 15-5 ± 0.299 £/ 43.1 + 3-26 38.3 ± 2.85 0.959 ± 0.0209 0.953 + 0.0216
20 90.8 92.3 15-3 ± 0.269 d/ 14.6 + 0.286 d/ 36.0 ± 2.50 31-5 ± 2.64 0.959 + 0.0262 0.943 ± 0.0294
30 61.9 71.2 £/l4.4 ± 0.216 e/ 16.7 + 0.253 £/ 26.8 + 1.75 42.5 H- 2.12 0.851 + 0.0247 0.900 ± 0.0207
a/ Each tank originally contained 750-770 ooho. Survival calculated on number of fish remaining in tank following removal
for eeawater tests or length-weight data.
b/ Sample size 15 fish.
oj Sample size 10 fish.
d/ Sample eize 30 fish.
e/ Sample size 40 fi. h.
f_/ Fish exposed for 1632 h (68 d).
-------
NJ
o
REARING.
ig/littr
**- « * 10 jug / li t«r
.---"* ~*
a)
Dec.
Jan.
Feb.
March April
Date (1975)
May
June
Figure 7- Effect of chronic copper exposure on growth of juvenile coho salmon.
(a/ Coho exposed to 30 v.gfi-Ltey on Mar>oh 273 1975).
-------
1.250 -
5/jg/liter
_IOyu/ liter
Dec.
Jan.
Feb. March April
Date (1975)
May
June
Figure 8. Effect of chronic copper exposure on coefficient of condition of juvenile coho salmon.
(a/ Coho exposed to SO yg/liter on March 2?, 1975).
-------
Table 4. Survival and gill ATPase activity of yearling coho salmon exposed
to copper for 1M h in static fresh water and their subsequent
survival and gill ATPase activity after transfer to sea water
Kominal Measured con- Percent
concentration centration ± SO survival
yg/liter Cu in copper a/
A. March 20-April 8, 1974 e/
0 13-7 ± 6.1 100
20 27.6 ± 3-1 100
30 34.7 ± 4.2 100
50 51.8 ± 4.1 95
60 60.7 ± 5-3 85
80 81. A ± 5.5 25
B. April 16-May 6, 1974 £/
0 15.7 ± 3-4 100
5 16.2 ± 2.4 100
10 22.1 ± 5-3 100
20 32.3 ± 2.7 100
30 42.8 ± 4.1 100
60 75.1 ± 3.9 30
C. May 9-31, 1974 h/
0 7-9 ± 2.5 100
5 11.5 ± 2.3 100
10 15-4 ± 2.6 100
20 23-1 ± 4.4 100
60 62.9 ± 7.4 30
D. June 5-26, 1974 h/
0 14.4 ± 9-9 100
5 16.3 ± 6.8 100
10 22.0 ± 6.0 100
20 28.0 ± 7-7 95
30 37.4 ± 4.6 100
50 52.7 ±8.1 40
60 61.2 ± 3-0 35
a/ Twenty fish exposed per oonaentration.
b/ Gill microsomal Na+> K^-activated ATPase;
of four fish at the end of exposure.
ATPase
activity
(fresh water) b/
12.9
7.1
5.6
5.0
25-5
16.3 f_/
9.8 f_/
6.6 £/ .
6.5 f_/
4.0 £/
48.4
24.0
14.0
10.3
6.9
24.8
15-6
11.1
7.2
6.8
5.8
Percent ATPase
survival activity
in sea water (sea water) b/
94.0 51.4
6.0 63.4 d/
0.0
0.0
0.0
0.0
100.0 57-3
94.0 58.6
59-0 57.8
24.0 53-7 f_/
6.0 70.2 d/
0.0
100.0
100.0
82.0
12.5
0.0
100.0
94.1
64.7
37-5
11.8
0.0
0.0
\moles ATP hydrolyzed/mg protein/h; mean
o/ Three hundred and twelve h seauater exposure.
dj Only one fish.
ej Three hundred and thirty-six h seauater exposure.
f_/ Mean of three fish.
a/ Mean of tao fish.
hj Three hundred and sixty h seawater exposure.
22
-------
probably one of the factors leading to loss of osmoregulatory ability and
death in sea water. The ATPase activity of fish that survived exposure to
sea water (Table 4, Table A-5) was increased two to threefold over the values
obtained for control coho in fresh water, corroborating the results of Zaugg
and McLain (1970).
Exposure of yearling coho for 144 h to sublethal levels of Zn (<2000
yg/liter) in static fresh water did not seriously affect survival in~sea water
(Table 5). The percent survival in sea water was not consistently related
to the Zn concentration, perhaps reflecting the tendency of fish to become
variably hypersensitive and hyperactive to stimuli during and after exposures
of >_1000 yg/liter Zn. These hyper-responsive episodes were sometimes fol-
lowed by tetanic spasms and death. In another study Chapman (personal
communication) exposed sockeye salmon to Zn for 19 months (egg through smolt)
and then transferred the smolts to sea water and observed no difference in
seawater survival between Zn exposed and control groups.
Gill ATPase was also inconsistently affected by Zn. It appeared to be
stimulated by all concentrations in March but not by any concentrations in
April or May (Table 5). The stimulation in March may have been an artifact
of the fish analyzed as controls; the fish chosen may not have been represen-
tative, in addition to being subjected to a low Cu contamination
(13-7 yg/liter). The difference in effects of Cu and Zn on gill ATPase and
subsequent survival in sea water indicates that the two metals probably do
not have a common mode of action.
The gill microsomal Na"1", K+-activated ATPase activity of juvenile coho
exposed to Cu in the flowing bioassay in June and July 1974 was reduced in a
concentration dependent manner except that the 5 yg/liter group produced a
slight stimulation in ATPase activity (Table 6, Table A-6) and was generally
similar to that noted for exposures in the static tests (Table 4). The
effects of Cu on seawater tolerance were greater after exposures to toxicant
in the static water system (Table 4) than after exposures in the flowing
water system (Table 6). This was probably due to the manual water changes
(once/day) which caused excitation and stressing of the fish and fluctuations
in water chemistry (Table 1) during the static exposures to toxicants.
The ATPase activity of coho exposed to 30 ug/liter Cu declined rapidly
following the introduction of the toxicant (Figure 9, Table A-7). The
ATPase activity declined to 72.6% of the control value after 56 h of toxicant
exposure and continued to decline until 310 h when activity leveled off at
24.0% of the control value. Yearling coho exposed for as little as 24 h to
30 yg/liter Cu incurred some deaths upon transfer to sea water but the effect
reached a maximum in those coho previously exposed for 96 to 120 h at
30 yg/liter Cu (Table 7). Additional exposure time to 30 yg/liter Cu did not
appear to result in more deaths when the coho were transferred to sea water.
The decline in ATPase activity with time of Cu exposure during the first
144 h correlated with subsequent mortality observed following transfer of the
fish to sea water (Table 7)
23
-------
Table 5- Survival and gill
to zinc for \kk h
survival and gill
ATPase activity of yearling coho salmon exposed
in static fresh water and their subsequent
ATPase activity after transfer to sea water
Nominal Measured con-
concentration centration ± SD
pg/1 i ter Zn
A. March 20-April 8, 1974 e/
0 29 ± 17
100 115 ± 12
300 299 ± 9
600 555 ± 20
1000 924 ± 32
2000 1772 ± 46
2500 22/1 ± 57
B. April 16-May 6, 1974 e/
0 37 ± 29
1000 937 ± 46
2500 2495 ± 178
4000 3803 ± 207
5000 4833 ± 369
6000 5611 ± 172
C. May 9-31, 1974
0 33 ± 30
4000 4261 ± 156
5000 5245 ± 144
6000 6158 ± 172
D. June 6-26, 1974
0 18 ± 13
1000 1214 ± 62
2500 2620 ± 56
4000 4062 ± 76
5000 4980 ± 231
Percent
ATPase
survival activity
in zinc
100
100
100
100
100
100
100
100
95
85
75
65
10
100
45
5
0
100
100
100
55
20
a/ (fresh water)
12.9 d/
22.2
25.5
24.4
29.8
25.8
25-5 d/fj
27-1 £/
21.6£/
21.8 £/
20.2 £/
48.4 d/
32.0
--
--
Percent ATPase
survival activity
in sea water (sea water) b/
100.0
94.0
100.0
100.0
81.0
81.0
87,0
100.0
100.0
100.0
67.0
80.0
50. 0,
100.0
33-0
0.0
100.0
100.0
100.0
70.0
66.7
51.4
70.1
65-7
76.9
95-0
70.5
66.2
57-3
50.9
48.6 £/
46.5 £/
58.3 £/
a/ Twenty fish exposed per concentration.
b/ Gill microsomal Na+, K^-activated ATPase; vmoles ATP hydrolyzed/mg protein/h;
four fish at the end of exposure.
mean of
c/ Three hundred and twelve h seawater exposure.
d/ Control also served for copper
copper levels) .
e/ Three hundred and thirty-six h
£/ Mean of three fish.
exposed
fish concurrently
being run (see Table
4 for
seawater exposure.
-------
Table 6. Survival and gill ATPase activity of yearling coho salmon exposed
to copper in fresh water and their subsequent survival after
transfer to sea water
Nominal Measured con-
concentration centration ± SD (n)
A. June
0
5
10
20
30
B. June
0
5
10
20
30
C. July
0
5
10
20
30
D. July
0
5
10
20
30
ug/1 iter
Cu
Percent
survival
in copper a/
ATPase
activity
(fresh water) b/
Percent
survival in
sea water c/
5-26, 1974 (144 h exposure)
1.8
8.0
9.9
18.2
31.3
27-July 10,
1.4
7-7
10.6
20.3
31.5
10-24, 1974
1.4
7.4
10.8
20.3
31.3
16-August 6,
3.6
9.7
13-2
26.0
33.9
± 1.0
± 3-9
± 2.7
± 2.9
± 4.0
(10)
(18)
(18)
(18)
(15)
100.
100.
100.
100.
100.
0
0
0
0
0
24.
26.
18.
11.
8.
0
5
2
4
8
(5)
(6)
(6)
(6)
(6)
100
100
97
70
35
.0
.0
5
.0
.0
(40)
(40)
(40)
(40)
(40)
1974 (525 h exposure)
± 0.9
± 3.9
± 3-3
± 3.6
± 3.8
(880 h
± 0.9
£ 3-5
± 3.1
± 3-5
± 3.4
1974
± 1.8
± 3.5
± 2.7
± 4.3
± 2.0
(16)
(3D
(32)
(32)
(28)
exposure)
(24)
(39)
(40)
(42)
(36)
99.
100.
100.
99.
81.
99.
100.
100.
99.
71.
7
0
0
5
9
7
0
0
5
1
32.
36.
25.
17-
14.
-_
--
3
0
9
5
5
(6)
(8)
(8)
(8)
(6)
100
100
100
96
35
100
100
97
67
10
.0
.0
.0
.3
.0
.0
.0
.5
5
.0
(80)
(80)
(80)
(80)
(80)
(40)
(40)
(40)
(40)
(40)
(144 h exposure) d/
(6)
(8)
(8)
(7)
(6)
100.
100.
100.
100.
100.
0
0
0
0
0
26.
33-
18.
10.
7.
1
0
8
4
5
(8)
(10)
(10)
(10)
(10)
97
100
60
0
0
.5
.0
.0
.0
.0
(40)
(40)
(40)
(40)
(40)
a/ Orginally placed 225-250 coho into each tank. Survival calculated on number of fish
remaining in tank following removal of fish for survival tests in sea water.
b/ Gill microsomal Na+> K^-activated ATPase; vmoles ATP hydrolyzed/mg protein/h; sample
size given in parenthesis.
c/ Three hundred and thirty to three hundred and sixty h seawater exposure; sample size
given in parenthesis.
d/ Placed 26-30 coho into each tank.
25
-------
N)
600
1200
1800
Figure 9.
EXPOSURE TIME IN COPPER (hours)
Influence of copper exposure in fresh water on gill microsomal Na+, K+-activated ATPase
activity in juvenile coho salmon. Each point represents mean of four to six fish (ymoles
ATPase hydrolyzed/mg protein/h ± SD).
-------
Table 7' Gill ATPase activU, and survival In sea water of yearling coho
salmon exposed to copper in fresh water for <_144 h (tests con-
ducted from March 6 to June 9, 1975) ~~
Nominal
concentration
(yg/1 iter Cu)
0
10
20
30
30
30
30
30
30
30
Exposure
time
(h)
144
144
144
144
120
96
69
56
41
24
ATPase acti vi ty
(fresh water)
(mean ±
46.3 ±
39.3 ±
--
13.4 ±
15.4 ±
17.2 ±
--
30.2 ±
SD)
8.8
8.0
3-1
1.5
4.6
13-5
a/b/
(18)
(4)
(8)
(4)
(4)
(4)
Percent
survival
(240-312 h
100.0
98.8
60.7
10.1
9-5
45-0
61.0
78.0
75.6
exposure) b/
(103)
(83)
(107)
(99)
(21)
(20)
(59)
(41)
(41)
a/ Gill microsomal Na+3 K+-aativated ATPase; \imoles ATP hydrolyzed/mg
~~ protein/h.
b/ Hwrbevs of fish in parentheses.
The ATPase activity of juvenile coho chronically exposed to Cu starting
December 20, 1975, is shown in Figure 10, (Table A-8). A peak in activity
occurred in the control fish during April and May, corroborating the data of
Zaugg and McLain (1970) that ATPase activity is highest during the parr-smolt
transformation. Gill ATPase activity was suppressed by Cu (Figure A-l)
and generally reflected decreased seawater tolerance but did not show any
increase during the time an increasing tolerance of Cu-exposed coho juveniles
to sea water occurred. The values for gill ATPase activity in 1974 (Tables 4
and 6) are lower than those in 1975 (Table 7) but may not represent biolog-
ical differences. The discrepancies may also be due to the analysis being
conducted at the Western Fish Nutrition Laboratory, Willard, WA in 1974 and
at our laboratory in 1975-
t
f
The survival of juvenile coho in sea water following exposure to Cu in
fresh water for 144 h at different times of the year (1975) is showr in
Figure 11. The percent survival was inversely related to Cu concentration
but there was a trend toward reduced effect with the onset of smoltification
in the fish. Disturbance of osmotic balance in coho upon transfer to sea
water is minimized during the smelting process (Conte et al. 1966). This
physiological process of smolting similarly may have reduced the deleterious
effects of Cu on survival of smolts in sea water, although increasing size and
age may also have contributed.
From June to early August 1974, four survival tests in sea water were run
on coho from the flowing toxicant system after 6 to 37 days (144 to 880 h) of
27
-------
60
00
50
>»
>
40
0>
U)
o
0- 30
10
Dec.
Figure 10.
Jan.
Feb.
March
Apr! I
May
June
DATE (1975)
Influence of copper exposure in fresh water on gill microsomal Na"1", K+-activated ATPase
activity in yearling coho salmon chronically exposed to copper from December 20, 197^.
Each point represents mean of four to six fish (ymoles ATPase hydrolyzed/mg protein/h
± SD.
-------
LJ
70
to
5 60
^ 50
40
W 30
»-
Z 20
UJ
O
£ 'o
a
rr
D
jug/1
,' 20;ug/l
Pec.'74 Jon. '75
Feb.'75 March '75
April '75
May '75
June '75
Figure 11. Survival of yearling coho salmon in sea water following exposure to copper in fresh water
for 144 h at various times of the year. (Each point represents 16-^5 fish).
-------
Cu exposure (Table 6). The group receiving 30 yg/liter Cu suffered 29%
mortality during the 37~day exposure, and 90% mortality of the toxicant sur-
vivors occurred upon transfer to seawater (Table 6). In the groups of coho
that received 20 and 30 yg/liter Cu for 144 h in mid-July no mortality oc-
curred during the freshwater exposure; however, total mortality occurred in
sea water (Table 6). This higher mortality in sea water as compared to that
observed earlier (Table 6) is probably due to desmoltification and the higher
salinity of the sea water used. In the two prior tests the salinity averaged
about 30 °/oo whereas in the mid-July test the salinity was 31-5 to 33 °/oo.
From mid-December 1974 to early June 1975, five survival tests in sea
water were carried out on coho exposed to Cu for 6 to 172 days (144 to 4128 h,
Table 8). Mortality during the 6 months of toxicant exposure in fresh water
was 39% at 30 yg/liter, 12% at 20 yg/liter, less than 5% for the 5 and
10 yg/liter concentrations and less than 1% of the controls (Table 8). A
group of fish exposed to 30 yg/liter Cu starting in late March showed a 23%
mortality in fresh water following Ik days of exposure (Table 8). Data show-
ing the survival in sea water of the various chronic exposure groups in 1975
are shown in Figure 12. A steady increase in tolerance to sea water occurred
from February through June, and was similar to the increased tolerance ob-
served for coho exposed to Cu for only 144 h (Figure 11) at various times
throughout this period. However, the chronically exposed fish generally had
higher mortality in sea water than did the 144-h exposed fish (Figure 13).
This was partly due to the poorer condition and smaller size of the chron-
ically exposed fish resulting from a Cu-induced concentration dependent
suppression of feeding. These same factors precipitated a faster onset of
mortality following transfer to sea water (Figure 13B).
A marked recovery of ability to survive in sea water was observed during
May and June in the groups chronically exposed to 30 yg/liter Cu (Figure 10 and
and 14C). Deaths during exposure to 30 yg/liter Cu in fresh water began to
occur after 144 h and reached an asymptote after about 1800 h (Figure 13B).
The fish that remained were Cu-tolerant and subsequently began to feed. Re-
sumption of feeding led to improved condition and nutritional states, and
consequently improved tolerance to sea water. In both groups chronically
exposed to 30 yg/liter Cu, recovery of feeding began to occur in May, even
though one group had been exposed for over 140 days and the other for less
than 50 days. Therefore, during the April, May and June tests of survival
in sea water, fish in the group exposed to Cu for the shorter time were
larger and in better condition as well as having begun parr-smolt transforma-
tion before their exposure to Cu on March 27. Probably a combination of
these factors resulted in the greater recovery of tolerance to sea water in
this 30 yg/liter group (Figure 14C).
A comparison of survival in natural and artificially prepared sea water
was also carried out with coho juveniles chronically exposed to Cu. The
survival was generally comparable in natural and artificial sea water for all
groups tested (Table 9)
30
-------
Table 8. Survival and gill ATPase activity of yearling coho salmon exposed
to copper in fresh water and their subsequent survival after
transfer to sea water
Nominal
concentration
A.
B.
C.
D.
E.
Dec.
0
5
10
20
30
Feb.
0
5
10
20
30
Apri
0
5
10
20
30
30
May
0
5
10
20
30
30
June
0
5
10
20
30
30
ug/U
Measured con- Percent
centration ± SD survival
ter Cu
in copper
ATPase
activity
a/ (fresh
water) b/
20, 1974-Jan. 5, 1975 (6 d exposure)
2.8 ± 2.1 (12) 99-87
6.5 ± 1.3 100.00
11.4 ± 1.6 99-93
21.2 ± 3.0 99-87
31.3 ± 3.8 100.00
25, 1975-Mar. 7, 1975 (67 d
1 14-24
(18 d)
16-26,
(49 d)
10-20,
(74 d)
3-1 ± 1-5
7-3 ± 1.4
11.9 ± 1.4
21.7 ± 2.6
31.7 ± 2.5
, 1975 (115
2.6 ± 1.5
7.0 ± 1.5
11.6 ± 1.5
21.5 ± 2.8
31.4 ± 2.8
1975 (147 d
2.6 ± 1.5
7.1 ± 1.4
11.7 ± 1-5
21.7 ± 2.7
31.6 ± 2.8
1975 (172 d
2.6 ± 1.6
7-1 ± 1.5
11.7 ± 1.5
21.6 ± 2.6
31.7 ± 2.9
(35)
Percent
survival in
sea
100.
100.
82.
0.
0.
water c/
0
0
5
0
0
(40)
(40)
(41)
exposure)
99-73
99-93
99.72
98.97
77-90
34.
27.
30.
15.
12.
5
3
2
1
7
(6)
(6)
(6)
(6)
(5)
100.
92.
75-
11.
0.
0
7
0
6
0
(41)
(41)
(44)
(43)
(43)
d exposure)
(55)
exposure)
(61)
exposure)
(68)
99-51
99.46
99-56
97-99
72.02
92.82
99.41
97-92
99.18
94.65
67-70
73-50
99.41
96.37
97.53
88.30
61.52
71.18
a/ Originally placed 750-770 fish into each
fish
remaining in tank
following
removal
44.
--
24.
16.
10.
10.
46.
--
41.
17.
13.
14.
33.
--
29.
15.
8.
8.
2
0
1
4
6
9
3
5
0
2
9
0
3
1
4
(6)
(5)
(4)
(4)
(5)
(5)
(4)
(4)
(3)
(3)
(5)
(4)
(4)
(3)
(3)
100.
100.
100.
25.
0.
0.
100.
92.
87.
47.
4.
17.
100.
92.
90.
55.
20.
63-
0
0
0
0
0
0
0
5
5
5
4
5
0
3
2
0
0
6
(40)
(41)
(41)
(40)
(40)
(40)
(21)
(40)
(40)
(41)
(45)
(50)
(20)
(39)
(41)
(40)
(20)
(11)
tank. Survival calculated on number of
for survival
tests in sec.
water.
miqration, etc.
L/
£/
Gill
microsomal Na+, K* '-activated ATPase;
sample size
Two hundred
in parenthesis, analysis, run
and forty h seawater exposure
\moles ATP hydroly?.ed/mg
within a week
, sample size
of
in
protein/hr;
seawater tests
parenthesis.
*
-
31
-------
10
cr 100
UJ
< 90
LJ
tn
80
70
60
cr 40
D
20
10
X
(1776)
20/ig/l ,
^- ~
L-fri^:
I
144
Dec'74
Jon '75
1600
Feb'75
EXPOSURE
.--0'
2760
March'75 April '75
TIME IN COPPER (hours)
4128
June '75
Figure 12. Survival of yearling coho salmon in sea water at various times of the year following
exposure to copper in fresh water initiated on December 20, 197^- (Each point represents
20-^*5 fish. Numbers in parenthesis indicate exposure hours of coho placed in 30 yg/liter
copper on March 27, 1975).
-------
oc
o
cr
LU
CL
100
90
80
70
60
50
40
30
20
10
i A- -D--x--x--x--its=-
-H x x x x x
Opg/ i
I
P W ^W I
April 8 -Z> April 14 ±>April 24,1975
I I
I 1
24
72
120
24
96
168
240
__ x x x x X
1000 2000
Fresh water
240
Figure 13-
EXPOSURE TIME (hours)
Survival curves of yearling coho salmon during exposure to copper
in fresh water and subsequent survival when transferred to sea
water. A. represents exposure to copper for 144 h; B. exposure
to copper for 2760 h (also includes coho exposed to 30 yg/liter
for 430 h prior to seawater test).
33
-------
IOO
9O
CO
TO
60
SO
40
30
20
10
Otc 20 Ht-Dtc 26 20+ Jon S.I|9T5
20
-------
Table 9- Comparison of survival of fish exposed to natural and artificial
sea water following 147 and 172 days of copper exposure in
fresh water
NominalPercent survival for 240 hours of exposure a/
concentration Natural Artificial sea water
pg/1 iter Cu sea water A b/ B c/
A. 3528 h of copper exposure (147 d) May 16-26, 1975
0 100.0 (21) 100.0 (21)
5 92.5 (40) 95.2 (21) 94.8 (58)
20 47.5 (41) 45.2 (42) 50.0 (16)
30 4.4 (45) 2.3 (44)
B. 4128 h of copper exposure (172 d) June 10-20, 1975
0 100.0 (20) 100.0 (20) 100.0 (20)
5 92.3 (39) 100.0 (20)
20 55-0 (40) 28.6 (42) 60.0 (20)
30 20.0 (20) 39-4 (33) 10.0 (20)
a/ Niaiber of fish tested in parenthesis.
b/ LaRoche, G.3 B. Eisler3 and C. M. Tarzaell. 1970. Bioassay procedures
for oil and oil dispersant toxicity evaluation. J. Water. Poll. Cont.
Fed. 42(11):1982-1989.
c/ Rila Marine Mix. Rila Products, P. 0. Box 114, Teaneck* N. J. 07666.
Effect of periods of rest following copper exposure on survival in sea water
i
Cu-exposed yearling coho held in clean fresh water for 5 days prior to
their transfer to sea water showed better survival (Table 10) than those
transferred immediately to sea water following toxicant exposure. The sur-
vival in sea water of all static Cu-exposed groups was higher in rested fish
than groups transferred to sea water immediately after toxicant exposure
(Table 10). The increased survival may in part be accounted for by a two to
fivefold increase in gill ATPase activity which occurred during the 5-d
recovery period in clean fresh water.
Coho juveniles given a 5~d rest following toxicant exposure of
30 yg/liter Cu in the flowing system showed only a 10% mortality in sea water
(Table 10) compared to 65% for those transferred immediately to sea water.
35
-------
Table 10. Survival and gill ATPase activity of juvenile coho salmon
exposed to copper for '\bk h and subsequent survival in sea
water with and without recovery periods
Nominal Measured
concentration central ion
con-
4 SD
ug/1 i ter Cu
Percent
survival
in copper
ATPase
activity a/
(fresh wateT)
Percent survival b/
Fresh water Sea
water
A. Static toxicant exposure c/
\. Direct transfer (April 16-May 6 1974)
0
10
30
60
2. Five
0
10
30
50
B. Flowing
15-
16.
42.
75-
7 ±
2 4
8 ±
1 ±
3
2
4
3
day recovery
13-
23-
44.
65.
toxicant
3 ±
5 4
1 4
4 4
3
3
3
2
.4
.4
.1
.9
(April
.1
.9
.1
.7
100
100
100
30
. 25-May 21, 1974)
100
100
95
75
25
9
6
4
32
20
15
19
.5
.8
5
.0
.7
.3
.4
3
w
(3)
(3)
(2)
(3)
(3)
(3)
(3)
100
59
6
0
100 (6) 100
100 (6) 90
83 (6) 10
45
(16)
(17)
(17)
(4)
(10)
(10)
(10)
(11)
exposure d/
1. Direct transfer (J
0
10
30
2. Five
0
10
30
1.
9-
31.
8 ±
9 4
3 4
1
2
4
day recovery
1.
9-
31.
8 ±
9 4
3 4
1
2
4
une 5-26, 1974)
.0
-7
.0
(June
.0
.7
.0
100
100
100
5-July 1, 1974)
100
100
100
24
18
8
_
-
~
.0
.2
.8
_
-
~
(5)
(6)
(6)
100
97-5
35-0
100
100 (20) 100
100 (20) 90
do)
40)
(40)_
(20)
(20)
(20)
a/ cm microsomal Na+, ^-activated ATPase; vmoles ATP hydrolyzed/mg protein/h; sample
size in parenthesis.
b/ Three hundred thirty-six to three hundred and sixty h seawater exposure; sample size
in parenthesis.
£/ Twenty fish exposed per concentration.
d_/ Originally placed 225-250 fish in each exposure tank.
36
-------
As the period of Cu exposure was considerably greater in 1975 than in
197^ i't was important to again check survival in sea water following periods
of rest or reduced toxicant exposure. The seawater survival of fish
previously exposed for 1600 h to 20 and 30 yg/liter Cu was 12 and 0%,
respectively (Table 11). When the fish were given a 5-d recovery or reduced
Cu concentration exposure prior to challenge with sea water the survival of
the 20 and 30 yg/liter Cu groups improved. However, the survival in sea water
was lower than observed in 197**. This reduced survival may be a result of:
(a) the poor condition of the fish following 1600 h of Cu exposure; (b) the
coho had not reached smolt condition and the enzyme activity was still low
making it more difficult for them to adapt to the osmotic stress; and (c) com-
bination of the above plus other unknown factors important in adjustment to
sea water. In the coho given a 15~d recovery prior to transfer to sea water
the survival was almost equivalent to the control group (Table 11). Groups
that received an exposure to reduced Cu concentration for 5 days and then
10 days of rest in fresh water prior to seawater challenge, incurred a
mortality about 20-25% greater than the control fish (Table 11).
Coho given a 5-d recovery following 144-h exposure to 30 yg/liter Cu
showed improved survival in sea water over the group challenged directly with
sea water (Table 11). When the rest period was extended to 15 days the sea
water survival improved threefold and was comparable to the control fish
(Table 11). The reason for the longer recovery time required to achieve sur-
vival equivalent to the control groups in the 1975 tests is not known but
might be related to lower ATPase levels at this time of year compared to fish
tested in June
Coho yearlings given a 10 or 15 d-rest in fresh water following exposure
to Cu for 172 d (A128 h) survived better in sea water than did fish chal-
lenged immediately or given a 5 d rest (Table 12). The lower survival of
20 yg/15ter Cu following 5 days of rest is unknown but fish chosen for this
seawater test might have been in poorer condition than others in the popula-
tion. Lower seawater survival was also noted in the groups (20 and 30 yg/
liter Cu) that were given a 10 d rest prior to testing. Coho held in sub-
tanks for 5 days (12.3 C) and then transferred to test tanks but held another
5 days in static fresh water (10 C) prior to seawater exposure showed lower
survival than fish rested in the test tanks (10 C) for the entire 10 days
prior to seawater tests. The extra handling, different temperature and the
low number of fish tested may account for the 20% differences in survival.
Coho given a 5 d rest in fresh water or acclimated to sea water of
20 °/oo after long exposure to Cu showed better survival when transferred to
sea water (30 °/oo) than a group transferred immediately (Table 13). Fish
from a group exposed to 20 yg/liter Cu showed nearly equal survival in sea
water (30 °/oo) after being given either a 5-d rest in fresh water or in sea
water of 20 °/oo. Fish from the 30 yg/liter Cu group, however, had better
survival when rested in 20 °/oo sea water (Table 13). Similarly the coho
exposed for 670 h to 30 yg/liter Cu showed almost a threefold better survival
than those exposed for 3000 h. The exposure to 20 °/oo sea water should have
been less stressful than fresh water to the fish as this salinity is closer
to being isomotic with the blood and may have resulted in the better survival
observed of the 30 yg/liter Cu groups.
37
-------
Table 11. Survival of yearling coho salmon in sea water following exposure to
copper for 1600 h or 144 h with or without recovery periods
(February 25-April 6, 1975)
Norn i na 1
concentration
yg/1 iter Cu
1. 1600 h of copper exposure
A. Direct transfer to sea water
0
20
30
B. Five-day rest prior to
seawater transfer
20
20
30
30
C. Fifteen-day rest prior to
seawater transfer
20
20
30
30
1. 144 h of copper exposure
A. Direct transfer to sea water
0
30
B. Five-day rest prior to
seawater transfer
30
30
C. Fifteen-day rest prior to
seawater transfer
30
30
Rest period
concentration Percent
yg/1 iter Cu in sea
100.0
11.6
0.0
0 54.5
10 54.5
0 7.1
10 4.9
0 100.0
10 - 5 d, 0 - 10 d 78.6
0 95-3
10 - 5 d, 0 - 10 d 77-3
100.0
7-3
0 30.0
10 25.0
0 100.0
10 5 d, 0 10 d 90.0
survival
water a/
(41)
C43)
(43)
(33)
(33)
(42)
(41)
(15)
(15)
(22)
(22)
(42)
(41)
(20)
(20)
(9)
(10)
a/ Tuo hundred and forty h seawatev exposure; sample size in parenthesis.
38
-------
Table 12. Survival of yearling coho salmon in sea water following exposure
to copper for 172 days (4128 h) with or without recovery periods
(June 10-July 2, 1975)
Nominal
concentration
pg/1iter Cu
Rest period
concentration
yg/1iter Cu
Percent survival
in sea water a/
A. Direct transfer to sea water
0
20
30
B.
C.
Five-day rest prior to seawater
transfer
20
30
Ten-day rest prior to seawater
transer
20
20
30
0 @ 12°
0 @ 12°
0 @ 10° b/
0 - 5 d @ 12°
0 - 5 d @ 12
5 d @
5 d @
10°
10°
D.
Fifteen-day rest prior to seawater
transfer
100.0 (20)
55.0 (40)
20.0 (20)
38.7
34.5
(3D
(29)
85.7 (30)
61.5 (13)
67.7 (12)
20
30
30
0
0
0
§
e
§
12
12
12
o
o
rt
£/
94
90
-7
.0
(19)
(20)
(8)
a/ Tuo hundred and forty h seawater exposure; sample size in parenthesis,
b/ Two fish (6. 7%) died during freshwater holding phase.
q/ Eight fish held for 25 d in fresh water without mortalities.
39
-------
Table 13- Effects of freshwater rest or seawater acclimation (20 °/oo
salinity) on survival of yearling coho salmon in 30 °/oo sea
water following 115-125 d exposure to copper in fresh water
(April 14-May 9, 1975)
Nominal
concent rat ion
pg/1iter Cu
Percent
Rest survival Percent
med i a in sea water a/ survival b/
2760-h copper exposure (115 d)
A. Direct transfer to sea water
0
20
30
30 o/ (430 h)
3000-h copper exposure (125 d)
B. Five days rest in fresh water or
100.0
25-0
0.0
0.0
(40)
(40)
(40)
(40)
20 °/oo sea water
0
20
30
30 a/ (670 h)
20
30
20 o/oo
20 °/oo
20 °/oo
20 °/00
Fresh water
Fresh water
100.0
77-5
33.3
90.0
88.9
5.0
100.0
75.6
16.7
42.9
80.0
5-0
(21)
(41)
(42)
(21)
(20)
(20)
a/ Two hundred and forty h seawater exposure.
b/ Survival based on total number of fish used; some died during the rest
period in fresh water (<_20%) and 20 o/oo sea water (@ 50%
mortality in each of the 30 vg/liter groups); sample size in parenthesis.
c/ Hours of exposure to copper.
Effect of copper on osmotic and ionic properties of blood plasma
Exposure of yearling coho to Cu in fresh water rapidly affected their
ability to maintain normal osmolality and chloride concentrations in blood
plasma (Figure 15, Figure A-2, respectively). Although the osmolality
of plasma from control fish decreased 7% in the first 24 h (probably due to
handling) and remained depressed, decreases in plasma osmolality of coho
exposed to 10 and 30 yg/liter Cu were even greater during the 144 h test
(9 and ]6%, respectively). After 144 h of exposure, analysis of variance
showed that the difference in osmolality of plasma between control fish and
fish exposed to 30 ug/liter Cu was highly significant (P <.01) but the
reduction caused by 10 yg/1fter Cu was not significant (P >.05). The effect
of 30 yg/liter Cu in reducing plasma osmolality below that of control fish
was highly significant after only 8 h of exposure.
40
-------
340
T 0^/ll*^5!l *
" " "" ["~\OtW/ liter CjLjft
" '
30 jug /liter Cu , .
8 16 24 32 40 48 56 72 96
Exposure time (hours)
120
144
Figure
Effect of copper exposure in fresh water on the osmolality of the plasma of coho salmon
during 1M h of exposure.
-------
Chloride concentration (m Eq/liter) in the plasma followed g similar
trend to osmolality (Figure A-2) which is supported by the. highly significant
correlation coefficient of 0..741 found when osmolality was regressed on
Cl- concentration for fish exposed to 30 yg/liter Cu. The correlation
coefficient was least for control fish as a result of the small range of
osmolality and Cl- values but was still 0.550 (P <.0l) for all groups com-
bined.
A similar effect of sub-lethal levels on plasma osmolality and chloride
concentration was observed in brook trout CSalveltnus fontinalis) by McKim,
Christensen and Hunt (1970). They found 5 to 8% reductions in osmolality
after 6 days (144 h) of exposure to 2k and 39 yg/liter Cu and 2 to 13%
decreases after 21 days. Lewis and Lewis (1971) presented data showing that
exposures of Cu and Zn to golden shiners and juvenile channel catfish for a
maximum of Sk h reduced the plasma osmolality and was one of the probable
causes of death.
The effects of 144-h exposure to Cu in fresh water on plasma osmolality
and chloride concentration in coho smolts transferred to sea water are shown
In Figure 16 and Figure A~3, respectively. Plasma osmolality was signif-
icantly lower for the coho exposed to 10 and 20 yg/liter Cu compared to con-
trol fish following the 144 h of toxicant exposure (zero time, Figure 16).
Once in sea water, plasma osmol'ality of fish previously exposed to 20 yg/liter
Cu increased rapidly so that after 2k h it was higher than that of control fish
when tested at the 0.01 probability level. A mean maximum value of 432 mOsm
(143% of the control) was reached after 120 h of exposure to sea water.
Fish began to die in this 20 yg/liter Cu group after 98 h in sea water
and the last death occurred before 193 h, similar to the pattern shown in
Figure 13A for mortality in sea water of another group previously exposed to
Cu for T44 h. The mean osmolality of plasma from survivors sampled after
240 h in sea water was only 111% of control fish sampled at the same time,
correlated with the leveling-off of mortality.
Exposure of coho to 10 yg/liter Cu in fresh water for 144 h did not
strongly affect their adaptation to sea water in the May 1975 test. Plasma
osmolality and chloride concentrations were not significantly higher than
control fish during the test, although they increased during the first 24 h
while control values decreased (Figure 16 and Figure A-3). No deaths occurred
nor were there deaths in the April and June seawater tolerance tests of smolts
exposed to 10 yg/liter Cu for 144 h (Figure 11).
As observed for coho in fresh water, a close relationship between plasma
osmolality and Cl- concentration was also found fn -fish exposed to sea water.
The correlation coefficient was again lowest for control fish and highest
for the group exposed to the highest concentration of Cu (20 yg/liter). The
correlation coefficients were 0.074 and 0.948, respectively, and 0.936
(P <.01) for all g'roups "combined.
42
-------
460
440
£ 420
O
£ 400
^_x
0 380
£
S 36°
"a.
340
«*-
O
1O^\ i
_ 3ZO1
2*
o 30C5
0 '
£ 280
(/)
O
260
^
^
-
~ *
/
/
L y
- ,A
' £
'V rf
\ '
rt\ ' 'V* , v Y
*
-
F
1 1 1 1 1
<
, I
I
[481 1
/
i
r""*
) .. ..o
3
i
1520
1
\
%
\
\
,
%
\
\
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\
c
^^
[
/'
I/'
L"t^H
y- l
i
. 4
t ^
i
i
1 C
1
21
..
-------
The effects of chronic exposure to a sublethal concentration of Cu on the
ability of coho to regulate osmotic and ionic properties of (Cl- and Na+)
plasma during exposure to sea water were studied in July 1974 (Figure 17,
Figure A-4 and Figure A-5). The fish were exposed for 792 h (33 days) to 0
and 20 yg/liter Cu before transfer to sea water. The sea water was only
28 °/oo during the first 2k h and ranged from 31.8 to 32.6 °/oo during the
rest of the experiment.
The mean osmolality of plasma in control fish increased by J% to 351 mOsm
after 12 h and did not return to zero hour values until 36 h (Figure 17),
probably a reflection of the poorer ability of these post-smolt coho to adapt
to sea water as compared to smolting fish in the May test. Although the group
exposed to 20 yg/liter Cu did not regulate osmolality, C1-, or Na+ as well
as the control fish, the values of osmolality and Cl- concentration did not
reach the levels they did for fish exposed to 20 yg/liter Cu for 144 h
(Figure 16, Figure A-3). This may indicate a lower tolerance to hyperosmotic
or hyperionic blood condition in the chronically exposed fish. Deaths in
sea water occurred as early as 24 h in fish chronically exposed to 20 yg/liter,
at which time the mean osmotic and Cl- concentrations were well below the
values observed in the 144-h exposed fish when they began dying after 98 h
in sea water. The overall survival of the control and 20 yg/liter Cu groups
of 100% and 76.7% compares equally with the survival of other chronically
exposed groups (37 d) presented in Table 6.
These data show that exposure to Cu decreases the osmoregulatory ability
of juvenile coho salmon in a concentration dependent manner leading to death
in sea water. The data on effects of Cu upon the activity of gill microsomal
Na+, K+-activated ATPase correlates with the osmoregulatory data. Suppression
of this enzyme activity by Cu must be responsibile, in part, for the loss of
osmoregulatory ability.
Effect of copper on downstream migration
Four releases of coho exposed to Cu since late December 1974 and two
releases of coho exposed to Cu for 144 h were made into Crooked Creek, a
tributary of the Alsea River. The releases were made during April, May and
June 1975, during the normal period of coho smolt migration. The percent
of released fish migrating to the trap was always greatest in control fish
and percent migration was inversely related to Cu concentration (Table 14).
Even the lowest concentration (5 yg/liter Cu), which had no measurable effect
on gill ATPase activity or survival in sea water in the 1975 experiments,
reduced the percentage of downstream migrants. The inhibitory effect of Cu
on downstream migration was directly related to exposure time with acutely
exposed fish having a higher percent migration than chronically exposed
fish at any given Cu concentration (Table 14). Exposure to 30 yg/liter Cu
for as little as 72 h caused a considerable reduction in migration as compared
to the control fish (52% and 93%, respectively, Table 14).
44
-------
460 h
-e-
vn
24 36 48
192
216 240
72 96 120 144 168
Seawater exposure time (hours)
336
Figure 17.
Effect of sea water exposure on the osmolality of the plasma of coho salmon chronically
exposed to copper (in fresh water) for 792 h in July
-------
Table 14. Percent migration through July 3, 1975, of coho salmon released
into a small coastal stream following acute and chronic copper
exposures
Nominal
concentration
pg/liter Cu April 8, 1975 a/ April 30,
A. Chronic exposures
0 41.6 51.2
5 35-3 29.8
10 36.0 35.1
20 21.7 25-0
30 6.0 10.3
30 e/ 10.9 (11) 32.1
B. Acute exposures (144 h or less) f_/
0 144 55. V 68.0
5 144 46.7 62.6
10 144 52.4 58.6
20 144 34.0 48.4
30 e/ (6) 14.6 17-5
30 e/ (4)
30 £/ (3)
aj One hundred and twenty to one hundred and
b/ One hundred and twenty to one hundred and
a/ One hundred and taenty to one hundred and
exposure).
d/ One hundred and twenty to one hundred and
exposure).
&/ Days of exposure to copper in parentheses
Release dates
1975 b/ May 14, 1975 c/ June 4, 1975 d/
28.5
23.4
18.5
11.5
4.0
(33) 8.9 (47)
No release
88.3
72.2
61.6
44.2
29.1
60.3 (68)
92.7 a/
65.9 g_/
44.4 g_/
52.0 g/
fifty fish per concentration (108-d exposure)
fifty fish per concentration (130-d exposure)
seventy fish per concentration
seventy fish per concentration
.
(145-d
(165-d.
f_/ Ninety-one to one hundred and five fish per concentration (6-d exposure).
g/ Forty-five to fifty-five fish per concentration.
-------
The migrational pattern of fish released in early June is shown in
Figure 18. Similar trends of movement occurred in the acute and chronic
exposure groups released earlier (Figure 19)- The migration of the May ]k
release was poorer than that observed for either earlier or later releases
and is probably related to the low water flow and vandalism at the weir
(Table 14). Most downstream migration occurred within the first 7 days of
release; however, coho that had received 20 or 30 yg/liter Cu appeared to
lag a little in their migration the first 2-3 days post-release but recovered
rapidly and their movement was usually complete within 10 days. Only a few
fish showed delayed migration and arrived at the weir after the 10th day
following release (Figures 18, 19, Table A~9) A large percentage of the fish
that did not migrate by early July, including control fish, are presumed to
have died. In various migrational studies on this stream in prior years,
electro-fishing above the weir following the normal migrational period (July)
accounted for <$% of the fish that failed to migrate.
There is a possibility that the coho juveniles released in early June
were in a better condition and nutritional state (because of resumed feeding
in May than their counterparts released earlier. This improved condition was
postulated as one of the factors important in the improved survival in sea
water of the 30 pg/liter groups (Figures 12 and ]kC, June test) and may also
be the factor responsible for the observed increase in migrational tendency
of the 30 pg/liter groups released June k, 1975 (Table 14).
In conclusion, exposure of coho salmon yearlings to sublethal concentra-
tions of Cu in fresh water resulted in significant mortality after these fish
were placed in sea water, reduction of the migrational urge, and suppression
of gill Na, K-ATPase thought to be important in sea water survival and an
indicator of migrational readiness. However, caution is required in applying
the results directly to field situations. The proportion of a total Cu
concentration that is "biologically available" may vary among streams due to
natural chelating agents, and there may be other antagonistic chemicals or
factors that protect fish from Cu toxicity.
In the tests we conducted with freshwater recovery period of 5 to 15 days
between Cu exposure and 30 °/oo seawater exposure, we noted better survival in
the fish given a recovery than in Cu exposed fish that were transferred
immediately to 30 °/oo sea water. Thus, fish that are affected by Cu pollu-
tants in their headwater streams and have an extensive downstream migration
before reaching the ocean may have a higher survival rate (provided fish
migrate and have additional streams entering their waterways to dilute the
pollutant) than fish subjected to Cu pollutants on short coastal streams.
Also, Cu-induced seawater mortality would depend on rate of transition from
fresh water to sea water. The exposure of sublethal levels of Cu for only
several days just before the fish enter sea water could produce significant
seawater mortality.
A stream may also have chemicals or other factors (such as temperature or
disease) which increase the sensitivity of fish to Cu. Fish which survived
and grew on a hand-fed diet in the laboratory might be incapable of foraging
efficiently for natural foods. Therefore, we again recommend caution in
-------
oo
too
^ 90
^ 80
Z
2 70
O
60
50
40
UJ
or so
Z 20
Q 10
0/ug/l
;.' 30/ufl/
30/jg/l A -A
A A
29
DAYS POST RELEASE
Figure 18. Influence of copper exposure for 3960 h (165 d) in fresh water on downstream migration
of yearling coho salmon. Includes coho exposed to 30 yg/liter for 1632 h (68 d). Each
line represents a release of 78-172 fish on June 4, 1975.
-------
cr
CD
ui
o:
h-
cn
o
o
2 4 6 6 10 12 14 16 18 20
49
100
90
80
70
6O
B
,--*
w-X
-//--x
2 4 6 8 10 12 14 16 18 20 49
DAYS POST RELEASE
Figure 19- Influence of copper exposure in fresh water on downstream
migration of yearling coho salmon released April 30, 1975-
A. represents coho salmon exposed to copper for \kk h (6 d);
B. represents copper exposure for 3120 h (130 d) and includes
coho exposed to 30 pg/liter (v) for 792 h (33 d).
-------
applying laboratory results to natural situations because a given concentra-
tion of Cu in different streams could have somewhat different effects on
migration and subsequent survival in sea water than the same concentration
studied in the laboratory.
We conclude that studies such as this that take into consideration the
life history of the fish are critical to the setting of water quality
standards.
50
-------
SECTION 6
REFERENCES
1. Campbell, R.D., T.P. Leadem and D.W. Johnson. 1974. The "In Vivo" effect
of p, pi DDT on Na+ - K+-Activated ATPase activity in rainbow trout
(Salmo gairdneri). Bull. Environ. Contam. SToxicol. 11 (5):425-1*28.
2. Chapman, G.A. 1973- Effect of heavy metals on fish. In: Heavy Metals
in the Environment. Water Resources Research Institute, Oregon State
University, p. 141-162.
3. Conte, P.P., H.H. Wagner, J. Fessler and C. Gnose. 1966. Development of
osmotic and ionic regulation in juvenile coho salmon Oncorhynchus kisutch.
Comp. Biochem. Physiol. 18:1-15.
4. Eisler, R. 1973- Annotated bibliography on biological effects of metals
in aquatic environments. Environmental Protection Agency R3~73~007-
287 p.
5- Eisler, R. and M. Wapner. 1975. Second annotated bibliography on
biological effects of metals in aquatic environments. Environmental
Protection Agency R3-75-023-
6. Epstein, F.H., A.I. Katz and G.E. Pickford. 1967- Sodium and potassium
activated adenosine triphosphatase of gills: role in adaptation of
teleosts to saltwater. Science. 156:1245-1247
7- Fessler, J.L. and H.H. Wagner. 1969- Some morphological and biochemical
changes in steelhead trout during the parr-smolt transformation. J. Fish.
Res. Bd. Canada. 26(11):2823-284l.
8. Granmo, A. and S.O. Kollberg. 1972. A new simple water flow system for
accurate continuous flow tests. Water Res. 6:1597-1599-
9. Hile, R. 1936. Age and growth of the cisco, Leueichtkys artedi
(Le Sueur), in the lakes of northeastern highlands, Wisconsin. Bull,
U.S. Bur. Fish. 19:211-317-
10. Hoar, W. S. 1959. The weight-length relationship of the Atlantic salmon.
J. Fish. Res. Bd. Canada 4:441-460.
11. Hodson, P.V. and J.B. Sprague. 1975- Temperature-included changes in
acute toxicity of zinc to Atlantic salmon (Salmon salar). J. Fish. Res.
Bd. Canada. 32(l):l-10.
51
-------
12. Jackim, E., J.M. Hamlin and S. Sonjs, 1970. Effects of metal poisoning
on five liver enzymes in the killtfish CFundulus heteroelitus). J. Fish.
Res. Bd. Canada 27(2):383~390.
13. Koch, R.B., D. Desaiah, H.H. Yap and K.L. Cutkomp. 1972. Poly-
chlorinated biphenyls: effect of long-term exposure on ATRase activity
in fish, Primephales promelas. Bull. Environ. Contarn. & Toxicol.
7:87-92.
14. LaRoche, G., R. Eisler and C.M. Tarzwell. 1970. Bioassay procedures for
oil and oil dispersant toxicity evaluation. J. Wat. Poll. Cont. Fed.
42(11):1982-1989.
15. Leadem, T.P., R.D. Campbell and D.W. Johnson. 1974. Osmoregulatory
responses to DDT and varying salinities in Salmo gairdneri, - I. Gill
Na-K-ATPase. Comp. Biochem. Physiol. 49A:197-205.
16. Lewis, S.D. and W.M. Lewis. 1971- The effect of zinc and copper on the
osmolality of blood serum of the channel catfish Ictalurus punctatus
Rafinesque and golden shiner Notemigonus crysoleucas Mitchell. Trans.
Amer. Fish. Soc. 100(4):639-643.
17. Lloyd, R. I960. The toxicity of zinc sulphate to rainbow trout. Ann.
Appl. Biol. 48(l):84-94.
18. Lloyd. R. and D.W.M. Herbert. 1962. The effect of the environment on
the toxicity of poisons to fish. J. Inst. Public Health Eng. 61:132-145-
19. Malikova, E.M. 1957- Biokhimicheskaia otsenka molodi lososia pri
perekhode v sostoianie, blizkoe k pokatnomu, i pri zadershke serebrianok
v presnoi vode. (Biochemical analysis of young salmon at the time of
their transformation to a condition close to the smolt stage, and
during retention of smolts in freshwater.) (Fish. Res. Board Transl.
Ser. 232). Tr. Latv. Otd. VNIRO 2:241-245.
20. McKim, J.M., C.M. Christensen and E.P. Hunt. 1970. Changes in the blood
of brook trout (Salvelinus font-inalis) after short-term and long-term
exposure to copper. J. Fish. Res. Bd. Canada. 27(10):1883-1889.
21. McKim, J.W. and D.A. Benoit. 1971. Effects of long-term exposures to
copper on survival, growth and reproduction of brook trout (.Salvelinus
font-inaUs). J. Fish. Res. Bd. Canada 28(5):655-662.
22. Miles, H.M. and L.S. Smith. 1968. tonic regulation in migrating juve-
nile coho salmon Oncorhynchus kisutoh. Comp. Biochem. Physiol. 26:
381-398.
23. Otto, R.G. 1971- Effects of salinity on the survival and growth of pre-
smolt coho salmon (Oncorhynchus kisutoh). 28(3):343~349.
52
-------
2k. Finder, L.J. and J.G. Eales. 1969. Seasonal buoyancy changes in
Atlantic salmon (Salmo solar) parr and smolt. J. Fish. Res. Bd.
Canada. 26(8):2093-2100.
25. Sprague, J.B. 1964. Lethal concentrations of copper and zinc for young
Atlantic salmon. J. Fish. Res. Bd. Canada 21(l):17-26.
26. Sprague, J.B. 1969- Measurement of pollutant toxicity to fish. I Bio-
assay methods for acute toxicity. Water Res. 3:793-821.
27. Sprague, J.B. and B.A. Ramsay. 1965. Lethal levels of mixed copper-zinc
solutions for juvenile salmon. J. Fish. Res. Bd. Canada 22(2):425-432.
28. Vanstone, W.E. and J.R. Markert. 1968. Some morphological and bio-
chemical changes in coho salmon, Oncorhynchus kisutch, during parr-smolt
transformation. J. Fish Res. Bd. Canada 25 (ll):2403-24l8.
29. Wagner, H.H. 1974. Photoperiod and temperature regulation of smolting
in steelhead trout (Salmo gairdnevi). Can. J. Zool. 42(2) :219-234.
30. Wagner, H.H. 1974. Seawater adaptation independent of photoperiod in
steelhead trout (Salmo gairdneri). Can. J. Zool. 52(7):805~8l2.
31. Water Quality Criteria Data Book. 1970. t. Organic chemical pollution
of freshwater- Water Poll. Control Res. Ser. 18010 DPV, EPA. Wash. D.C.
32. Water Quality Criteria Data Book. 1971- I I. Inorganic chemical pollu-
tion of freshwater. Water Poll. Control Res. Ser. 18010 DPV, EPA.
Wash. D.C.
33. Water Quality Criteria Data Book. 1971. III. Effects of chemicals on
aquatic life. Water Poll. Control Res. Ser. 18050 GWV, EPA. Wash. D.C.
34. Water Quality Criteria Data Book. 1972. IV. An investigation into
recreational water quality. Water Poll. Control Res. Ser. 18040 DAZ,
EPA. Wash. D.C.
35. Water Quality Criteria Data Book. 1973- V. Effects of chemicals on
aquatic life. Water Poll. Control Res. Ser. 18050 HLA, EPA. Wash. D.C.
36. Wedemeyer, G. and K. Chatterton. 1971- Some blood chemistry values for
the juvenile coho salmon (Oncorhynchus kisutch). J. Fish. Res. Bd.
Canada 28(4):6o6-608.
37. Wedemeyer, G. 1972. Some physiological consequences of handling stress
in the juvenile coho salmon (Oncorhynchus kisutoh] and steelhead trout
(Salmo gairdneri). J. Fish. Res. Bd. Canada. 29(12):1780-1783.
38. Zaugg, W. S. and L. R. McLain. 1970. Adenosinetriphosphatase activity
in gills of salmonids: seasonal variations and saltwater influence in
coho salmon, Oncorhynchus kLsutch. Comp. Biochem. Physiol. 35:587~596.
53
-------
39- Zaugg, W. S. and L. R. McLain. 1972. Changes In gill adenosinetriphos-
phatase activity associated with parr-smolt transformation in steelhead
trout, coho, and spring chinook salmon. J. Fish. Res, Bd. Canada 29(2):
167-171.
40. Zaugg, W.S. and H.H. Wagner. 1973- Gill ATPase activity related to
parr-smolt transformation and migration in steelhead trout (_Sa1mo
gairdneri): influence of photoperiod and temperature. Comp. Biochem.
Physiol. k5(8):955-965.
-------
APPENDICES
-------
\n
Table A-l. Chemical and physical characteristics of well water at the Oregon Department of Fish
and Wildlife Laboratory a/, Corvallis, Oregon
Parameter Units
Cadmium yg/1 i ter
Chromium "
Cobalt
Copper
Iron "
Lead "
Manganese '
Mercury
Nickel
Zinc
Alkalinity as CaC03 mg/liter
Calcium
Chloride
Dissolved oxygen
Hardness as CaCOj
Magnesium
Nitrogen
Free ammonia as N
Nitrate as N
Nitrite as N "
Potassium "
Sod i urn
Solids
Dissolved '
Suspended '
Sulfate
pH
Turbidity JKSN
10/18/73
ii's
4^0
48.0
10.0
8.0
<0.5
1.0
4.0
76.0
2UO
6.0
7.0
101.0
12.0
<0.060
3-1
<0.001
0.9
6.9
173-0
<1 .0
7-0
-_
3/1/74 3/15/74
<0-5
24.0
6.0
--
85.0
13.0
<0.04o
8.6
<0.001
1.1
6.1
174.0
1.0
10.0
6.6
5.0
--
--
61.0
27.0
6.0
95.0
14.0
<0.045
7-5
<0.001
1.0
8.7
180.0
2.0
12.0
6.8
3.0
5/10/74 6/20/74
1.0
<2.0
2.0
22.0
5.0
<1 .0
12.0
<2.0
5.0
22.0
6.0
--
93.0
11.0
<0.090
7.8
0.004
0.95
6.6
203.0
1.0
15.0
6-7
5.0
3.0
2.0
2.0
20.0
<5.0
1.0
<0.5
1.0
1..0
66.0
16.3
7.0
95.0
10.7
<0.005
6.8
<0.001
1.0
7.2
176.0
<1 .0
13.0
6.7
6.0
8/8/74
<1 .0
6.0
16.0
<5.0
1.0
2.0
4.0
71.3
17-0
7.0
8.0
100.0
10.4
<0.085
6.72
<0.001
1.0
6.4
192.0
2.0
12.0
6.7
5.0
11/7/74
13.0
4!o
110.0
<1 .0
5.0
1.1
10.0 '
1.0
65.0
24.0
7.7
6.8
99.0
18.0
<0.005
6.1
<0.001
1.1
10.0
6.61
6.0
2/4/75
<2.0
<1 .0
3.0
28.0
8.0
3-0
2.0
71.5
92.0
6.58
6/3/75
<3.0
<3.0
<3.0
21.0
6.0
<10.0
2.1
2.5
< 3-0
73.5
12.6
7.0
96.0
10.2
<0.005
1.0
8.7
12.0
6.80
23.0
Mean :
0.6 b/
2.8 b/
0.9 b/
3-2
38
3.1 b/
2.7 b/
2.9
2.6
69.0
20.0
7-0
7.3
95-0
12.0
0.022
6.7
0.001
1 .0
7.6
183.0
1.2
12.0
6.7
8.0
b SD
-
± 1.6 b/
±34
-
-
± 3.2 b/
± 1.7 V
± 5-0
± 5.0
± 1.0
± 0.5
± 5.0
± 3-0
- b/
± T.8
- b/
± 0.1
± 1.4
±12.0
- b/
± 2.0
J/
± 7.0
Range
<|:S
2.0
16.0
-------
Table A-2. Effect of copper in fresh water on the survival of yearling coho
salmon (determination of 9&-h LC50 values)
Nominal
concentration
ug/1 iter Cu
a)
b)
c)
I/
March 7-1 1,
0
30
60
80
100
130
March 20-24
0
20
30
50
60
80
Measured concent rat
jjg/1 iter Cu
Mean ±
1974 a/
5
30
58
71
87
108
, 1974 a/
11
27
35
50
61
80
.1
.0
.5
.7
.8
.8
.7
.4
.2
.9
.0
.5
+
+
±
+
±
+
±
±
+
±
+
+
SD
1.4
4.3
4.7
5-7
2.9
5.0
7-2
3.9
5.0
4.1
6.6
6.5
ion
Range
(3
(26
(53
(64
(86
(103
(2
(21
(28
(45
(53
(71
.0-
.0-
.5-
.0-
.0-
.0-1
.4-
.7-
.5-
.5-
3-
.3-
7.0)
34.5)
67.8)
78.8)
92.0)
14.5)
22.3)
33.5)
44.0)
57.0)
73.5)
90.8)
Percent mortal i ty
n
7
8
8
8
4
4
6
8
8
8
8
8
(Ranae)
0
0
50
75
100
100
0
0
0
5
15
75
(40-60)
(60-90)
(0-10)
(10-20)
(70-80)
November 20-24, 1974 b/
0
30
50
60
70
80
100
May 27-31 ,
0
45
50 c/
55
65
70 c/
80
100 o/
7
36
56
62
75
81
102
1975 b/
19
54
58
57
65
65
81
83
.8
.3
7
.7
5
.8
.9
.1
.3
.6
.5
.6
.0
.2
.3
+
±
±
±
+
+
+
±
+
+
+
+
+
+
+
2.3
4.5
4.6
3.0
8.0
6.3
6.7
11.5
3.6
14.6
2.6
2.2
10.1
1.5
7.5
(5
(32
(52
(58
(62
(76
(96
(4
(49
(39
(53
(60
(51
(79
(76
.0-
.2-
.1-
.0-
5-
.0-
.0-1
.0-
.2-
3-
.6-
.8-
.6-
.8-
.8-
12.0)
45.5)
65.5)
67.1)
85-0)
96.5)
12.0)
35.4)
58.4)
69.8)
60.4)
67-2)
76.8)
83-4)
93.4)
8
8
8
8
/ 8
/ 8
4
8
8
6
8
8
6
4
4
0
0
5
20
35
75
100
0
20
45
45
63.
95.
100
100
(0-10)
(30-40)
(70-80)
(10-30)
(40-50)
(40-50)
2 (40-88.9)
0 (90-100)
a/ Test man. on odho of 1972 brood year.
b/ Test Pun on coho of 1973 brood year.
a/ Test mm May 5, 1975; contamination of control with copper.
56
-------
Table A-3. Effect of zinc in fresh water on the survival of yearling coho
salmon (determination of 96-h and iM-h LC50 values)
Nominal
concentration
yg/1 Iter Zn
Measured concentration
pg/liter Cu
Mean ± SD (Range) n
Percent mortal i ty
Hean Range
96-h exposures
a) April 16-20, 1974
b)
c)
0
1000
2500
4000
5000
6000
May 9-13, 1974
0
4000
5000
6000
June 5-9, 1974
0
1000
2500
4000
5000
48 ±
933 ±
2426 ±
3761 ±
4837 ±
5673 ±
38 ±
4286 ±
5243 ±
6158 ±
12 ±
1230 ±
2637 ±
4051 ±
4953 ±
144-h exposures (continuation
a)
b)
c)
d)
March 20-26, 1974
0
100
300
600
1000
2000
2500
April 16-22, 1974
0
1000
2500
4000
5000
6000
May 9-15 1974
0
4000
5000
6000
June 5-11, 1974
0
1000
2500
4000
5000
29
115
299
555
924
1772
2271
37
937
2495
3803
4833
5611
33 ±
4261 ±
5245 ±
6158 ±
18 ±
1214 ±
2620 ±
4062 ±
4980 ±
31
56
186
253
468
231
34
170
169
172
4
66
47
63
260
of 96-h
17
12
9
20
32
46
57
29
46
178
207
369'
172
30
156
144
172
13
62
56
76
23
(10- 109)
(859-1040)
(2150-2650)
(3480-4025)
(4440-5750)
(5465-6000)
(9- 94)
(4100-4550)
(5050-5500)
(5900-6350)
(6- 17)
(1170-1340)
(2570-2695)
(3990-4140)
(4465-5125)
run)
(8- 56)
(99- 139)
(286- 314)
(530- 605)
(905-1018)
(1718-1870)
(2168-2361)
(6- 109)
(859-1040)
(2150-2700)
(3480-4025)
(4440-5750)
(5465-6000)
(9- 94)
(4100-4550)
(5050-5500)
(5900-6350)
(6- 40)
(1150-1340)
(2530-2695)
(3990-4180)
(4465-5230)
7
7
7
7
7
4
6
7
7
6
6
5
5
5
6
10
12
11
12
11
12
12
11
11
11
11
11
8
8
9
10
6
8
7
7
7
9
0
0
10
15
25
75
0
30
75
100
0
0
0
0
60
0
5
15
25
35
90
0
55
95
100
0
0
0
45
80
(10-20)
(20-30)
(70-80)
(10-50)
(70-80)
(0-10)
(10-20)
(10-40)
(30-40)
(80-100)
(30-80)
(90-100)
(40-50)
57
-------
Table A-4. Effect of copper exposure on average length-weight and coefficient
of condition
Date of
samp 1'e
12/18/74
1/17/75
2/17/75
3/25/75
VI 7/75
4/29/75
5/13/75
6/3/75
Days
of Cu
exposure
0
0
0
0
0
28
28
28
28
28
58
58
58
58
58
0
108
108
108
108
108
11
130
130
130
130
130
33
144
144
144
144
144
47
165
165
165
165
165
68
Norn i na 1
concentration
yg/1 iter Cu
0
5
10
20
30
0
5
10
20
30
0
5
10
20
30
30
0
5
10
20
30
30
0
5
10
20
30
30
0
5
10
20
30
30
0
5
10
20
30
30
Fork length
cm
13.
14.
13.
H.
14.
14.
14.
13-
13.
13-
15.
15.
14.
14.
13.
16.
16.
15-
15-
14.
13.
16.
16.
15.
15-
14.
13.
16.
17-
16.
15.
14.
13.
16.
17.
17.
15.
14.
14.
16.
+
8
0
7
1
1
7
3
7
8
5
2
0
4
1
0
3
1
4
1
8
5
4
6
9
2
5
8
0
0
4
8
6
5
0
6
3
8
9
4
7
SE
0.160
0.196
0.162
0.153
0.172
0.274
0.232
0.165
0.197
0.184
0.224
0.257
0.236
0.228
0.183
0.127
0.186
0.145
0.170
0.183
0.171
0.237
0.205
0.188
0.164
0.203
0.208
0.203
0.170
0.206
0.173
0.194
0.155
0.234
0.187
0.225
0.213
0.201
0.216
0.253
Weight
q ±
31.0
32.3
30.7
32.3
33-8
39.6
34.4
28.8
28.0
25-0
41.3
38.4
35.1
29-5
21.1
47.8
46.3
38.5
36.9
33.8
21.0
47.4
49.3
41.6
36.1
28.9
22.9
37.7
54.3
46.3
41.2
30.3
20.0
37-5
60.4
55.8
40.7
33.7
26.8
42.5
SE
1
1
1
1
1
2
1
1
1
1
1
2
2
1
1
1
1
1
I
1
0
2
.197
.342
.147
.150
.286
.316
.741
.110
.301
.176
.810
.080
.276
587
Coefficient
of condition
K ±
.156
.163
.158
.145
.188
.196
.128
.105
.039
.007
.155
.109
.129
.008
.108 0.937
.200
.670
.205
.451
.460
1.084
1.091
1.036
.027
1. 012
.922 0.825
.247
.895
1.041
1.058
.625 0.992
.396 0.992
.471 0.890
.509 0.829
.656 0.905
1
1
1
1
0
1
2
2
2
1
1
2
.766
.783
1.084
1.009
.728 0.993
785 0.907
.975 0.782
.781 0.889
.179
.539
1.080
1.026
.169 0.956
.824 0.956
.747 0.851
.188 O.goo
SE
0.00990
0.02551
0.00904
0.01456
0.01759
0.01102
0.01116
0.01194
0.01103
0.00975
0.01217
0.01156
0.02029
0.01330
0.01593
0.00574
0.00735
0.00720
0.00989
0.01647
0.01126
0.01075
0.00770
0.00990
0.01059
0.01449
0.01952
0.00823
0.00787
0.00936
0.01182
0.01913
0.01785
0.00697
0.00776
0.01091
0.01495
0.01955
0.02472
0.02068
n
50
50
50
50
50
39
40
40
40
30
30
31
30
32
21
120
60
80
80
60
40
40
60
80
80
60
40
40
60
80
80
bO
40
40
60
80
80
60
40
30
58
-------
VJl
Table A-5. Activity of microsomal Na+, K+-activated ATPase in gills of coho following exposure to
copper for I*f4 h and subsequent seawater exposure
Concentration
pg/liter Cu Exposure time
Nominal
0
20
30
50
0
5
10
20
30
60
0
5
10
20
60
0
5
10
20
30
50
Measured
13-7
27.6
34.7
51.8
15-7
16.2
22.1
32.3
42.8
75.1
7-9
11.5
15-5
23-1
62.9
14.4
16.3
22.0
28.0
37- A
52.7
in copper
144
144
144
144
144
144
144
144
144
144
144
144
144
144
144
144
144
144
144
144
144
Date
3/26/74
3/26/74
3/26/74
3/26/74
4/22/74
4/22/74
4/22/74
4/22/74
4/22/74
4/22/74
5/15/74
5/15/74
5/15/74
5/15/74
5/15/74
6/11/74
6/11/74
6/11/74
6/11/74
6/11/74
6/11/74
ATPase activity in toxicant Exposure time
Mean ± SD
12.9 ± 5.2
7.1 ± 1.0
5.6 + 1.1
5.0 ± 0.7
25.5 ± 5.7
16.3 ± 4.6
9.8 ± 2.1
6.6 ± 2.1
6.5 ± 1.3
4.0
48.0 ± 7-3
23.9 ± 4.8
14.4 ± 6.9
10.3 ± 2.0
6.9 ± 0.3
24.8 ± 7-9
15-6 + 2.8
11.1 ± 2.2
7.2 ± 1.5
6.8 ± 0.6
5-8 ± 1.3
Range
9-3
6.3
5-1
4.1
18.2
11.4
7.9
4.6
5.7
3-2
40.0
16.7
9.6
8.1
6.7
16.2
12.6
8.6
5.5
6.3
4.6
- 20.3
- 8.2
- 7.1
- 5-7
- 32.2
- 20.6
-'12.0
- 8.8
- 8.0
- 4.8
- 57-3
- 26.5
- 24.6
- 12.9
- 7-3
- 31.8
- 18.1
- 12.5
- 8.5
- 7-4
- 7.2
n in sea water Date
4
4
4
4
4
3
3
3
3
2
4
4
4
4
3
3
3
3
3
3
3
312
312
No
No
336
336
336
336
336
No
4/8/74
4/8/74
survivors
survivors
5/6/74
5/6/74
5/6/74
5/6/74
5/6/74
survivors
ATPase activity in sea water
Mean ± SD Range n
51.4 ± 14.7 26.7 - 72.2 5
63.4 1
57.3 ± 6.3 52.0 - 65.3 4
58.6 ± 6.8 53-5 - 68.4 4
57.8 ± 9.6 44.9 - 66.7 4
53.7 ± 1.1 52.9 - 54.9 3
70.2 1
-------
Table A-6. Activity of microsomal Na+, K+-activated ATPase in gills of coho
exposed to copper for various lengths of exposure in
Concentration
yg/1 iter Cu
Nominal Measured
0
5
10
20
30
0
5
10
20
30
0
5
10
20
30
1.8
8.0
9-9
18.2
31-3
1.4
7.7
10.6
20.3
31-5
3-6
9-7
13-2
.26.0
33-9
Exposure time
in copper
144
144
144
144
144
645
645
645
645
645
144
144
144
144
144
ATPase activity in toxicant
Date
6/11/74
6/11/74
6/11/74
6/11/74
6/11/74
6/11/74
6/11/74
6/11/74
6/11/74
6/11/74
7/22/74
7/22/74
7/22/74
7/22/74
7/22/74
Mean ± SD
24.0 ± 5-8
26.5 ± 3.8
18.2 ± 6.4
11.4 ± 1.3
8.8 ± 1.6
32.3 ± 8.2
36.0 ± 8.8
25.9 ± 5-2
17-5 ± 2.9
14.5 ± 4.5
26.1 ± 5-6
33.0 ± 9-7
18.8 ± 3.5
10.4 ± 2.3
7.5 ± 7-9
Range
19.0
20.1
11.2
10.0
6.4
22.7
17-2
17-9
13.1
8.0
17-6
25.0
15-1
7.2
5.1
32.5
- 31.4
29.9
13.6
- 10.7
- 47.1
- 45.6
- 33.4
22.7
- 21.6
- 33-3
58.2
- 24.9
- 13.2
- 10.7
n
5
6
6
6
6
6
8
8
8
8
8
10
10
10
1.0
60
-------
Table A-7- Activity of microsomal Na+, K+-activated ATPase in gills of coho
exposed to copper for various time periods
Concentration
Mg/1 iter Cu E
Nominal
0
30
0
30
0
30
0
30
0
30
0
30
0
30
30
30
0
30
0
30
Measured 5
3
31
3
31
2
31
2
31
2
31
2
31
2
31
31
31
2
31
31
.0
.5
.1
.7
.8
.5
.6
.6
.6
.6
.6
.6
.6
.7
7
.7
.6
7
Rearing
7
:xposure time
n copper (h)
744
744
1416
1416
1968
1968
2640
312
144
144
3144
816
3648
56
96
118
3984
1656
tank
ATPase activity in toxicant
Date
1/20/75
1/20/75
2/17/75
2/17/75
3/12/75
3/12/75
4/9/75
4/9/75
4/25/75
4/25/75
4/30/75
4/30/75
5/21/75
5/21/75
5/21/75
5/21/75
6/4/75
6/4/75
6/25/75
6/25/75 '
Mean ± SD
29-
10.
34.
12.
35-
17.
44.
10.
53-
12.
46.
14.
41.
30.
17.
15-
33-
8.
34.
9-
9 ±
4 ±
5 ±
7 ±
6 ±
7 ±
2 ±
6 ±
3 ±
1 ±
9 ±
2 ±
6 ±
2 ±
2 ±
4 ±
9 ±
4 ±
6
3 ±
3.2
4.3
9-5
1.9
17.2
5.4
11.7
1.9
12.4
3.6
5.4
3-3
5.4
13.5
4.6
1.5
9.5
2.4
0.8
Range
26.4
6.3
23.4
10.6
17.8
12.4
32.3
8.6
42.9
9.4
41.9
11.1
33.6
13.5
12.6
14.1
22.9
6.4
34.4
8.6
33
- 15
- 46
- 15
58
- 23
- 59
- 13
69
- 17
- 56
- 17
- 44
42
22
17
45
11
- 34
- 10
.8
.7
.1
.6
.8
.6
.1
.2
.6
.2
.0
.7
.5
.7
.2
.6
.9
.1
.8
.1
n
5
4
6
5
4
4
6
5
4
4
5
3
4
4
4
4
5
3
2
3
61
-------
Table A-8. Activity of microsomal Na+, K+-activated ATPase in gills of coho
exposed to copper for various time periods in 1975
Concentration
yg/liter Cu
Nominal
0
no
20
30
0
10
20
30
0
30
0
0
10
20
30
0
10
20
30
0
0
0
10
20
30
Measured
3.0
11.6
21.3
31.5
3-1
11.9
21.7
31-7
2.8
31.5
2.6
2.6
11.6
21.6
31.6
2.6
11.7
21.7
31.6
2.6
2.6
2.6
11.7
21.6
31-7
Exposure time
in copper (h) Date
744
744
744
744
1416
1416
1416
1416
1968
1968
2232
2640
2640
2640
2640
3144
3144
3144
3144
3648
3816
3984
3984
3984
3984
1/20/75
1/20/75
1/20/75
1/20/75
2/17/75
2/17/75
2/17/75
2/17/75
3/12/75
3/12/75
3/23/75
4/9/75
4/9/75
4/9/75
4/9/75
4/30/75
4/30/75
4/30/74
4/30/75
5/21/75
5/28/75
6/4/75
6/4/75
6/4/75
6/4/75
ATPase activity in toxicant
Mean :
29-9 ±
35-7 ±
19.5 ±
10.4 ±
34.5 ±
30.2 ±
15.1 +
12.7 +
35.6 t
17-7 -
32.8 +
44.2 +
24.0 +
16.1 +
10.4 ±
46.9 ±
41.3 ±
17-5 ±
13-0 ±
41.6 ±
36.2 ±
33.9 ±
29-0 ±
15-3 ±
8.1 ±
t SD
3.2
11.3
3-5
4.3
9.5
6.1
3.6
1.9
17.2
5.4
13-3
11-7
2.7
4.3
2.4
5.9
8.0
3.5
3-8
5.4
11.1
9.5
13-9
5-2
2.9
Range
26.4
24.4
16.8
6.3
23.4
21.6
10.2
10.6
17-8
12.4
19.7
32.3
19.8
12.4
8.0
41.9
42.8
14.4
9.8
33-6
25-3
22.9
19.2
10.7
6.4
33-8
50.2
24.6
15-7
46.1
37-2
19.2
15.6
58.8
23.6
44.4
59-1
27-1
22.2
13-3
- 56.0
54.0
22.5
17-2
45.5
51.6
45-9
49-5
20.6
11.5
n
5
4
4
4
6
6
6
6
4
4
4
6
5
4
4
5
4
4
3
4
4
5
4
4
3
62
-------
50
30
T3
CD
10
2/17/75 (1416 h)
D 4/9/75 (2640H)
O 4/30/75 (3l44h)
6/4/75 (3984h)
10 15 20 25
Measured jug /liter Cu
30
35
Figure A-l. Effect of copper exposure (in fresh water) on gill microsomal Na+, K+-activated ATPase
activity of yearling coho salmon.
-------
130
\_
£ 120
^^W > f\
1 10
pr
x^
UJ
c 100
E
g 80
Q.
70
C
'"" ^
n 1
J^j
i I i
t if A - X
*'*o\. \^M\
^%Jt₯"T^ft^
- v T ^K^
-
-
>
i i i
8 16 24 :
. 1
T ,
KJ- I S
3>-4^j
r- ^ i
S
t
i i i i
52 40 48 56
r
. _J
. i
L.... '
* <
^
i i
64 7
: ^
:
r-4-H
iii i
2 80 88 96 I2C
0 jug/ liter Cu i
-J
L J
10 /jg/ liter Cu]
J
30 AJg/ liter Cu
1
) 144
Exposure time (hours)
Figure A-2. Effect of copper exposure in fresh water on the chloride ion concentration of the plasma of
coho salmon serially sampled during 1^(4 h of exposure.
-------
220
210
^ 200
»_
0)
.t: '90
^^
^
O"
UJ
^ 170
O
E
150
O
Q.
140
c
1 1 I3°
M
^
1
T J?
T *
_ /
T*
- S.*-^*«. i
/^* /iV
e T i \<
f^^
li
I20»-
Ilo4-
4i iiii
<
r
«
J[
/
/
. /
^J
^^
/
229 -I
^
1
X
T
1
i
*
..(^--
^ '
'^<<
_
3
\
\
\
\
\
l
' J
5- -0*x^--<
1 ^3
^
l^'
J
l .^
*
[ **"9 i1
f i " *
1 -C
>
1
20\jjg/ liter Cu
\
vl
v
10 jjg/lliter Cu
r .-6
K^y1-" '|°
Opg / iter Cu
III I I I I I
4 12 24 36 48 72 96 120 144 163 192 216 240
Figure A-3.
Seawater exposure time (hours)
Effect of seawater exposure on the chloride ion concentration of the plasma of coho salmon
previously exposed to copper (in fresh water) for 1M h.
-------
1801-
CT>
96 120
Seawater
144 168
exposure
192 216 240
time (hours)
336
Figure A-4.
Effect of seawater exposure on the chloride ion concentration of the plasma of coho salmon
previously exposed to copper (in fresh water) for 792 h (x - control,Q- 20 ug/liter Cu).
-------
2001-
140
130
1206
_L
_L
_L
_L
12 24 46 72 96 120 144 168 192
Seawater exposure time
216
240
336
Figure A-5- Effect of seawater exposure on the sodium ion concentration of the plasma of coho salmon
previously exposed to copper (in fresh water) for 792 h (x - control, O - 20 yg/liter Cu)
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Table A-9. Percent migration (to July 3,
released into a small coastal
chronic copper exposure
1975) of yearling coho salmon
stream following acute and
Norn i na 1
concentration
gg/1 iter Cu
A.
B.
C.
D.
a/
b/
c/
1-5 6-10
Chronic exposures
0 39.2 40.8
5 30.7 33-3
10 28.7 30.6
20
30
30
£/
Acute exposure
0
5
10
20
30
15.8 18.3
3.0 4.0
6.8 10.9
(144 h) d/
54.5 54.5
40.9 44.8
49-5 49-5
31.1 33-0
7.8 10.7
1-5 6-10
Apri
1 8,
11-20
41
34
34
19
5
10
55
46
52
34
12
May
.6
.0
.0
.2
.0
.9
.4
.7
.4
.0
.6
14,
11-20
1975 a/
Percent migration
Days post release
21-30 31-40 41+
34.6
36.0
21.7
5.0
12.6
1975 e/
35.3
6.0
-- (lid)
__
13-6 14.6
Days post
21-30 31-40
April 29, 1975 b/
1-5 6-10
50.4 51
28.5 29
33-1 34
19-2 21
5.2 7
24.4 30
68.0 68
60.4 62
57-6 58
40.9 47
13.4 17
release
.2
.1
.4
.7
.2
.8
.0
.6
.6
.3
.5
1-5
11-20
29.8
34.4
24.2
7-2
30.8
_-
48.4
--
June
6-10
21-30 31-40 41+
34.4 34.4 35.1
25.0
7.2 10.3 ~
30.8 30.8 32.1 (33d)
4, 1975 //
11-20 21-30
Chronic exposures
0
5
10
20
30
30
£/
26.8 28.5
20.4 22.2
15-9 17.9
3-3 4.9
1.0 2.0
2.5 6.3
-
23
18
8
3
7
-
.4
.5
.2
.0
.6
10.6
3.0
8.9
11.5
4.0
(47d)
79-
60.
55-
32.
18.
38.
2
4
8
5
4
5
88.3
71.6
61.0
40.0
26.2
57-7
__ -.-
72.2
61.6
43-3 44.2
27-2 29.1
57-7 60.3 (68d)
Acute (144 h or less) aj
0
30
30
30
One
One
(72h)
(96h)
(I44h)
No release
~ ~
hundred and taenty to
hundred and taenty to
Days of copper
one
one
hundred and
hundred and
exposure in parents
tests.
fifty fish
fifty fish
89-
36.
20.
27-
1
0
0
7
92.7
42.2
65.9
__
__
44.4
per concentration (108-d copper exposure).
per concentration (130-d copper exposure).
Ninety-one to one hundred and five fish per concentration (6-d copper exposure).
One hundred and twenty to one hundred and seventy fish per concentration (145-d copper exposure).
_V One hundred and taenty to one hundred and seventy fish per concentration (165-d copper exposure).
oj forty-five to fifty-five fish per concentration.
68
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TECHNICAL REPORT DATA
(Please read Instructions on the reverse before completing)
1. REPORT NO.
EPA-600/3-77-032
2.
3. RECIPIENT'S ACCESSIOI*NO.
4. TITLE AND SUBTITLE
Effects of Copper and Zinc on Smoltification of
Coho Salmon
5. REPORT DATE
March 1977
6. PERFORMING ORGANIZATION CODE
7. AUTHOR(S)
Harold W. Lorz and Barry P. McPherson
8. PERFORMING ORGANIZATION REPORT NO.
9. PERFORMING ORGANIZATION NAME AND ADDRESS
Oregon Department of Fish and Wildlife
Corvallis, Oregon 97331
10. PROGRAM ELEMENT NO.
1BA608
11. CONTRACT/GRANT NO.
R802468
12. SPONSORING AGENCY NAME AND ADDRESS
U. S. Environmental Protection Agency -Corvallis, OR
Corvallis Environmental Research Laboratory
Corvallis, Oregon 97330
13. TYPE OF REPORT AND PERIOD COVERED
Final
14. SPONSORING AGENCY CODE
EPA/600/02
15. SUPPLEMENTARY NOTES
16.ABSTRACTMany species of trout and saimon spend their early life in freshwater and
then migrate to the sea. Transition from freshwater to marine existence requires
physiological changes which are involved in the development of the migratory smolt
stage. Sublethal exposure to pollutants in freshwater could theoretically disrupt
smoltification and indirectly cause the dealth of smolts.
In this study, exposure of smolt age coho to sublethal levels of copper in
freshwater interfered with normal osmotic and ionic control in blood plasma; when the
copper exposed fish were transferred to seawater the plasma osmolality and chloride
concentrations increased significantly, compared to controls, and many+died. These
responses were attributed in part to an observed suppression of Na+, K - activated
ATPase activity in the gills of copper exposed fish. The most sensitive latent effect
of exposure to sublethal levels of copper was the failure of copper exposed coho smolts
to migrate successfully following release into a natural stream.
All copper concentrations tested (5-30 ^ig/1) produced adverse effects and were
well below the 96-hr LC50 (60-74 jug/1). Exposure to sub-lethal levels of zinc
produced no similar adverse effects.
17.
KEY WORDS AND DOCUMENT ANALYSIS
DESCRIPTORS
b.lDENTIFIERS/OPEN ENDED TERMS
c. COSATI Field/Group
Salmon, Copper, Zinc
Smoltification,
Migration
Osmoregulation
06/A,F,T
18. DISTRIBUTION STATEMENT
Release unlimited
19. SECURITY CLASS (ThisReport)'
Unclassified
21. NO. OF PAGES
84
20. SECURITY CLASS (Thispage)
Unclassified
22. PRICE
EPA Form 2220-1 (9-73)
69
U.S. GOVERNMENT PRINTING OFFICE: 1977797-5871104 REGION 10
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