BIOLOGY
OF
REPRODUCTION
47,
Stress
1140-1150
Reduces
the Quality
of Biology Institute
of Gametes
CAMPBELL,2’3 T.G.
P.M.
Department
(1992)
Produced
POTFINGER,4
and
by Rainbow
J.P.
Trout1
SUMPTER3
and Biochemistry,3 Brunel University, Uxbridge, of Freshwater Ecology,4 Windermere Laboratory, Cumbria LA22 OLP United Kingdom
Middlesex UB8 3PH, Far Sawrey, Ambleside
United
Kingdom
ABSTRACT In this gamete
study
9 mo
the
we
quality
carried
prior
out
with
mill
from
in somatic acute nificantly fish
Time
milt
the
males
weight
stress
from subjected
compared
of ovulation, male
the
from
regime.
groups
two
at the
perhaps control
Hence,
fish.
to adversely affect a range of reproductive ing a suppressive effect on reproductive [1, 2], mammals
phibians
[12-14],
evidence pothalamus
that
fish
the
the deal
suppressive of work
mechanisms
success
believed
have,
There
includin
[9-11],
effects has been
amhygo-
of stress directed
to mediate
received
less
parameters
such
as sperm
counts
and
motility,
at-
Accepted
August
Received This
May work
‘Correspondence.
was
18, 10,
by the
parents.
In humans,
female
egg
to repeated
in females,
for progeny
advancing
from
and
trout.
it has
thus
of the
study
been
reproductive failure rather endometrial
failure [25]. The parameters sperm quality are still not well
the
sig-
stressed
by rainbow
age
factors
that
a
affect
estab-
results from inadequacy that deterdefined and
gamete
quality
are
and
in vitro
fertilization).
parameters
that
However, correlate
the
best
identification
with
fertility
and
gamete
quality
in these
number of tiny can be obtained,
be removed thermore,
from a number there are subsequent
embryonic
survival
takes
in utero
place
and
large
species
have used the rainbow
is egg
eggs, often and these
of different difficulties
development,
or within
scale studies reThe limiting factor
individuals. Furin monitoring
because
an opaque
this
egg.
trout as a model
animal
of stress in terms
quality
its
quantity,
because
mode
usually
In this study
the biological consequences and
produc-
at different must often
to study
of gamete
of reproduction
of-
fers
many advantages. Female rainbow trout undergo a very pronounced annual reproductive cycle, during which two large ovaries develop. Immediately prior to spawning, the ovaries together
fer-
comprise approximately 20% of the body weight. At ovulation 2000-3000 eggs per kg body weight, each measuring approximately 5 mm, are synchronously released into the body cavity [26]. Following ovulation, the eggs can be man-
by NERC. (0895)
trout
produced
with
differences
and implantation mine egg and
we
1992. FAX:
of gametes
age-related oocyte quality
tion; only a small stages of maturation,
1992.
funded
rates
were
fertilized
no
size
egg
during
fertilized
were
of rainbow reduced
survival
with
were
There
exposure
were
of
terms
counts
sperm
females
female
this
for studying
tion of viable offspring which attests to any animal’s full reproductive capability, and it is the effect of stress on this ultimate reproductive parameter which is most important. The successful fertilization of eggs and subsequent development of offspring will depend greatly on the quality produced
each
monitored. and
and
ovulated
little possibility of performing the quired to establish these relationships.
size, and chemical composition, etc., in an attempt to assess the inhibitory influence of acute and chronic stress on reproductive function. However, it is the successful produc-
of gametes
quality
stress
offspring survivorship and the factors that affect gamete quality are difficult to achieve in most animals, especially in species such as mammals and humans where there is
tention. It is possible to measure the concentrations of a wide variety of hormones and neurotransmitters involved in coordinating the reproductive process, and other reproductive
lower
declines
of gamete
stress-in-
much
the
lished that diminished
nation
on at
functions and the anatomtake place [for review, see of stress in terms of overall
however,
but
from
in
acute
of considerable interest, not only from a fundamental point of view, but also because a better understanding would undoubtedly improve the success of the many different assisted reproduction techniques (such as artificial insemi-
is considerable
acting at the level of the gland [20, 211, and the
duced inhibition of reproductive ical sites at which these effects 24]. The ultimate consequences reproductive
indices, endocrinology,
[8], reptiles
[15-18].
corticosteroids, [3, 13, 19], pituitary
nads [5, 22, 23] mediate reproduction. A great elucidating
birds
13-7],
and
reduces
tility
then
in ovulation
significantly
stress
Stress is a ubiquitous feature of life and reproduction is one physiological process that is particularly sensitive to its disruptive effects. Acute and chronic stress have been shown
eggs
was
experiment, delay
importantly,
INTRODUCTiON
humans
of the
eggs
females,
from
Eggs
of stress
to repeated
in mature
300
fertilized
consequences
subjected
groups.
Approximately end
biological were
recorded
control
in a significant
most
the trout
were
and
of the
resulted
to study rainbow
egg size
stressed
development
and,
unstressed
female
and
the
both
development in males,
and
treatment
same
animal
as a model male
fecundity,
from
Subsequent
between
Counts
to progeny
fish,
to the
reproductive
sperm
trout
of 30 mature
in diluent.
or length
during lower
Groups
to spawning.
of iO
rainbow
the
used
quantity.
Ott the
dilution
sperm
have
and
274348.
1140
STRESS
ually dures
recovered from females and fertilized externally
from and
it
the males. is possible
by routine with milt
REDUCES
stripping similarly
Eggs are then placed to monitor fertilization
THE
ternal sequent
be
fertilization development
from
each
10
eye. Thus system for
the
8
ti
since stage
female,
is readily manipulated, of the eggs can be very
C,) C,) C,)
I-
ex-
and the subeasily mon-
6-
I-
c)
itored. Previous
work
shown
acute
that
evation nology,
stage
obtained
12
in incubation trays, success and sub-
the study of the factors that determine gamete quality, a very large number of eggs-all at exactly the same
using and
this
animal
chronic
stress
have a suppressive effect resulting in significantly
one levels in males and steroids and vitellogenin this study induced
model, and
the
plasma
has
cortisol
el-
endocritestoster-
significantly reduced levels in females [17].
was to establish the endocrine dysfunction
trout,
on reproductive reduced plasma
consequences on gamete
1141
GAMETES
OF
proceremoved
sequent development of the eggs by the naked the rainbow trout constitutes an excellent model
of maturation-can
QUALITY
4.
C 2’
0
z
plasma sex The aim of 0
of this stressquality and
5
10
15
20
25
WEEK
30
35
40
No.
quantity.
MATERIALS Animals
and
AND
FIG. 1. Experimental design. The experiment began in mid-April and ended in January 1991. The Stress was applied randomly throughout the 38 wk. At the beginning, the stress was applied, on average, once a week; later in the experiment, the fish were stressed more frequently. The interval between each stress and the time of day the stress was applied varied ran-
METHODS
Treatment
domly.
Three-year-old rainbow trout (Annan strain) were maintained at a density of 30 fish (mean weight 1462 ± 55.8 g, n = 90)/tank in 1500-L outdoor fiberglass tanks, each supplied fish
with were
a constant fed
once
flow daily
of lake
with
1% body weight per day. To peated acute stress on a variety in both maturing
male and females,
water
(35
commercial
L/min).
feed
determine the of reproductive
female rainbow all of which had
The
at a rate
of
effects of reparameters
trout, two groups of 30 spawned in the previous
year, and two groups of 30 maturing males were subjected to repeated acute stress during the 9 mo prior to spawning. A similar number of control fish were maintained in identical tanks but were not subjected to the stress. The fish were stressed by exposure to a brief period of emersion; this involved allowing the water to drain completely from the mm have
tanks, before been
stressful [30], and
leaving
the
experiment, fish were
exposed
to the
stimulus [27] as have sheep birds [31]; hence the emersion
at random intervals To assess whether procedure, and blood
fish
air
the tanks were slowly allowed shown to acclimate to repeated
to prevent emersion
a subsample was sampled 4 mo removed
prior with
about
3
Trout to a
to spawning. During as little disturbance
Collection
each
ice
the to
for
fecundity,
females, males from
female
and
and both
ovulated
on
resultant
blood
for cortisol, [32].
using
from
ovulation. The
ovulated
female
into
egg
sperm stress
size
1-2
a dry
OF
REPRODUCTION
47,
Stress
1140-1150
Reduces
the Quality
of Biology Institute
of Gametes
CAMPBELL,2’3 T.G.
P.M.
Department
(1992)
Produced
POTFINGER,4
and
by Rainbow
J.P.
Trout1
SUMPTER3
and Biochemistry,3 Brunel University, Uxbridge, of Freshwater Ecology,4 Windermere Laboratory, Cumbria LA22 OLP United Kingdom
Middlesex UB8 3PH, Far Sawrey, Ambleside
United
Kingdom
ABSTRACT In this gamete
study
9 mo
the
we
quality
carried
prior
out
with
mill
from
in somatic acute nificantly fish
Time
milt
the
males
weight
stress
from subjected
compared
of ovulation, male
the
from
regime.
groups
two
at the
perhaps control
Hence,
fish.
to adversely affect a range of reproductive ing a suppressive effect on reproductive [1, 2], mammals
phibians
[12-14],
evidence pothalamus
that
fish
the
the deal
suppressive of work
mechanisms
success
believed
have,
There
includin
[9-11],
effects has been
amhygo-
of stress directed
to mediate
received
less
parameters
such
as sperm
counts
and
motility,
at-
Accepted
August
Received This
May work
‘Correspondence.
was
18, 10,
by the
parents.
In humans,
female
egg
to repeated
in females,
for progeny
advancing
from
and
trout.
it has
thus
of the
study
been
reproductive failure rather endometrial
failure [25]. The parameters sperm quality are still not well
the
sig-
stressed
by rainbow
age
factors
that
a
affect
estab-
results from inadequacy that deterdefined and
gamete
quality
are
and
in vitro
fertilization).
parameters
that
However, correlate
the
best
identification
with
fertility
and
gamete
quality
in these
number of tiny can be obtained,
be removed thermore,
from a number there are subsequent
embryonic
survival
takes
in utero
place
and
large
species
have used the rainbow
is egg
eggs, often and these
of different difficulties
development,
or within
scale studies reThe limiting factor
individuals. Furin monitoring
because
an opaque
this
egg.
trout as a model
animal
of stress in terms
quality
its
quantity,
because
mode
usually
In this study
the biological consequences and
produc-
at different must often
to study
of gamete
of reproduction
of-
fers
many advantages. Female rainbow trout undergo a very pronounced annual reproductive cycle, during which two large ovaries develop. Immediately prior to spawning, the ovaries together
fer-
comprise approximately 20% of the body weight. At ovulation 2000-3000 eggs per kg body weight, each measuring approximately 5 mm, are synchronously released into the body cavity [26]. Following ovulation, the eggs can be man-
by NERC. (0895)
trout
produced
with
differences
and implantation mine egg and
we
1992. FAX:
of gametes
age-related oocyte quality
tion; only a small stages of maturation,
1992.
funded
rates
were
fertilized
no
size
egg
during
fertilized
were
of rainbow reduced
survival
with
were
There
exposure
were
of
terms
counts
sperm
females
female
this
for studying
tion of viable offspring which attests to any animal’s full reproductive capability, and it is the effect of stress on this ultimate reproductive parameter which is most important. The successful fertilization of eggs and subsequent development of offspring will depend greatly on the quality produced
each
monitored. and
and
ovulated
little possibility of performing the quired to establish these relationships.
size, and chemical composition, etc., in an attempt to assess the inhibitory influence of acute and chronic stress on reproductive function. However, it is the successful produc-
of gametes
quality
stress
offspring survivorship and the factors that affect gamete quality are difficult to achieve in most animals, especially in species such as mammals and humans where there is
tention. It is possible to measure the concentrations of a wide variety of hormones and neurotransmitters involved in coordinating the reproductive process, and other reproductive
lower
declines
of gamete
stress-in-
much
the
lished that diminished
nation
on at
functions and the anatomtake place [for review, see of stress in terms of overall
however,
but
from
in
acute
of considerable interest, not only from a fundamental point of view, but also because a better understanding would undoubtedly improve the success of the many different assisted reproduction techniques (such as artificial insemi-
is considerable
acting at the level of the gland [20, 211, and the
duced inhibition of reproductive ical sites at which these effects 24]. The ultimate consequences reproductive
indices, endocrinology,
[8], reptiles
[15-18].
corticosteroids, [3, 13, 19], pituitary
nads [5, 22, 23] mediate reproduction. A great elucidating
birds
13-7],
and
reduces
tility
then
in ovulation
significantly
stress
Stress is a ubiquitous feature of life and reproduction is one physiological process that is particularly sensitive to its disruptive effects. Acute and chronic stress have been shown
eggs
was
experiment, delay
importantly,
INTRODUCTiON
humans
of the
eggs
females,
from
Eggs
of stress
to repeated
in mature
300
fertilized
consequences
subjected
groups.
Approximately end
biological were
recorded
control
in a significant
most
the trout
were
and
of the
resulted
to study rainbow
egg size
stressed
development
and,
unstressed
female
and
the
both
development in males,
and
treatment
same
animal
as a model male
fecundity,
from
Subsequent
between
Counts
to progeny
fish,
to the
reproductive
sperm
trout
of 30 mature
in diluent.
or length
during lower
Groups
to spawning.
of iO
rainbow
the
used
quantity.
Ott the
dilution
sperm
have
and
274348.
1140
STRESS
ually dures
recovered from females and fertilized externally
from and
it
the males. is possible
by routine with milt
REDUCES
stripping similarly
Eggs are then placed to monitor fertilization
THE
ternal sequent
be
fertilization development
from
each
10
eye. Thus system for
the
8
ti
since stage
female,
is readily manipulated, of the eggs can be very
C,) C,) C,)
I-
ex-
and the subeasily mon-
6-
I-
c)
itored. Previous
work
shown
acute
that
evation nology,
stage
obtained
12
in incubation trays, success and sub-
the study of the factors that determine gamete quality, a very large number of eggs-all at exactly the same
using and
this
animal
chronic
stress
have a suppressive effect resulting in significantly
one levels in males and steroids and vitellogenin this study induced
model, and
the
plasma
has
cortisol
el-
endocritestoster-
significantly reduced levels in females [17].
was to establish the endocrine dysfunction
trout,
on reproductive reduced plasma
consequences on gamete
1141
GAMETES
OF
proceremoved
sequent development of the eggs by the naked the rainbow trout constitutes an excellent model
of maturation-can
QUALITY
4.
C 2’
0
z
plasma sex The aim of 0
of this stressquality and
5
10
15
20
25
WEEK
30
35
40
No.
quantity.
MATERIALS Animals
and
AND
FIG. 1. Experimental design. The experiment began in mid-April and ended in January 1991. The Stress was applied randomly throughout the 38 wk. At the beginning, the stress was applied, on average, once a week; later in the experiment, the fish were stressed more frequently. The interval between each stress and the time of day the stress was applied varied ran-
METHODS
Treatment
domly.
Three-year-old rainbow trout (Annan strain) were maintained at a density of 30 fish (mean weight 1462 ± 55.8 g, n = 90)/tank in 1500-L outdoor fiberglass tanks, each supplied fish
with were
a constant fed
once
flow daily
of lake
with
1% body weight per day. To peated acute stress on a variety in both maturing
male and females,
water
(35
commercial
L/min).
feed
determine the of reproductive
female rainbow all of which had
The
at a rate
of
effects of reparameters
trout, two groups of 30 spawned in the previous
year, and two groups of 30 maturing males were subjected to repeated acute stress during the 9 mo prior to spawning. A similar number of control fish were maintained in identical tanks but were not subjected to the stress. The fish were stressed by exposure to a brief period of emersion; this involved allowing the water to drain completely from the mm have
tanks, before been
stressful [30], and
leaving
the
experiment, fish were
exposed
to the
stimulus [27] as have sheep birds [31]; hence the emersion
at random intervals To assess whether procedure, and blood
fish
air
the tanks were slowly allowed shown to acclimate to repeated
to prevent emersion
a subsample was sampled 4 mo removed
prior with
about
3
Trout to a
to spawning. During as little disturbance
Collection
each
ice
the to
for
fecundity,
females, males from
female
and
and both
ovulated
on
resultant
blood
for cortisol, [32].
using
from
ovulation. The
ovulated
female
into
egg
sperm stress
size
1-2
a dry
