The length of heat in the albino rat as determined by the copulatory response.

T H E LENGTH O F HEAT I N T H E ALBINO RAT AS
DETERMINED BY T H E COPULATORY RESPONSE
RICHARD J. BLANDAU, JOHN L. BOLING AND WILLIAM C. YOUNG
Arnold Biological Laboratory, Brown Univer.Pity, Providence, Rho& Island, and
Laboratories of Primute Biology, Yale University School of Medicine,
New Haeen. Connecticut
ONE FIGURE
The length of heat in the rat, a s in the guinea pig, is a
measure of reproductive normality which has not been widely
used. I n most morphological investigations the vaginal condition has served as the end-point and in most studies of
behavior (Hemmingsen, ’33; Ball, ’34, ’36, ’37 and ’40) reliance has been placed on the intensity of the copulatory
response. Nevertheless, the course of heat can easily and
accurately be followed and once its length has been ascertained a very satisfactory basis has been established for
determining and studying individual differences and f o r investigating factors involved in the induction and control of
mating behavior (Boling and Blandau, ’39). As matters
stood, however, there was a need f o r new determinations because no two of the earlier estimates were the same. Tshii
( ’22) gives the length as 4 to 8 hours, Long and Evans ( ’22)
as 3 to 1 2 lioui-s, and Ball (’37) as 3 t o 21 hours.
There were other reasons for an interest in the problem.
When Ball (’37) was studying the rat with much the same
purpose we have had, she was troubled by the tendency of
animals tested a t frequent intervals to develop a “disturbed
endocrine balance” which prolonged the cycle and period of
comification and modified the behavioral picture. She states
that discontinuance of the practice of making vaginal smears
This investigation was supported by a grant f r o m the Committee f o r Research
in Problrms of Sex, National Research Council.
453
454
RICHARD J. BLANDAU A N D OTHERS
at 6-hour intervals terminated the condition, but many of the
data she presents appear t o have been collected before this
change in technique was effected. Consequently, the extent
to which frequent reference can be made to the behavioral
end-points without disturbing the basic physiological rhythm
mas not known from any substantial body of data.
Over and above an interest in the length of heat as a standard of reference was the desire to obtain information with
respect to the factors involved in the induction of mating
behavior and in the regulation of its character. I n the guinea
pig the existence of a “constitutional” factor was suggested
by the consistency of behavior displayed from cycle t o cycle
by intact animals (Young, Dempsey, Hagquist and Boling,
’39) and from injection period to injection period by spayed
animals (Young and Rundlett, ’39). Inasmucli as the length
of heat during consecutive estrous periods provides a measure
of consistency or variation as the case may be, advantage
was taken of the opportunity to ascertain whether or not
the existence of a similar factor is indicated for the rat.
MATERIALS A N D METHODS
The data were obtained from 145 female rats which were
between 3 and 4 months old when the observations were
begun. The animals were kept in square wire mesh cages
with removable tops which facilitated handling. They were
maintained on a ration of Purina fox chow checkers supplemented by green vegetables once a week. Tests of sexual
responsiveness were made at hourly intervals, day and night,
beginning the second day after the end of the preceding heat
period. I n order to minimize the disturbance of constant
illumination ( Bromman, ’37 ; Hemrniiigsen and Krarup, ’37),
the lights were turned out a t night during the interval between
hourly observations. I n addition, during the first 2 days of
the cycle when no observations were being made, the animals
were kept in an ordinary room in which there. was the normal
alternation of daylight and darkness.
L E N G T H OF HEAT
455
Tlie time of lieat was ascertainrd by the res1)onse to manual
maiiipulation. Thc mcthod is iI modification of that uscd by
Jlcmniingscln ( ’ 3 3 ) and Ball ( ’3).
Tlic tliuml-,and riug finger
ai’e placecl anterior to tltc iliac c i ~ s t sand tlie second and
third fingers a r e placed 011 opposite sides of tlie tail in such
a way that tltc tips lie in the vulvar region (fig. l a ) . Animals
Fig. 1 a. Method of stiiiiulnting tlic display of the copulatory responsc o r
lordosis.
b. Female rat disp1:rying lortlosis imiiiedi;ctely after inaiiual stimulation.
at the hciglit of cstrous i-espond quickly with a typical l o d o s i s
in prc1)aration f o r coitus (fig. 11)). The reaction may or
may not be accompwnicd by car quivering, depending on
tlic strain used (Hemmirigsen; Ball). If the estrus is of
low intensity, o r when the animal is fingrued near the end
of heat, several seconds’ stimulation may be reqnircd. Associated heliavior siich as darting and hopping ai-e helpful
in iclentifyirig animals wliich a r e about to come into heat
its
456
ItICITARI) J. BLANI)AU 4NI) OTHEIlS
o r arc in a state of low excitability. Animals not in lieat
resist tlie manual stimnlation more or less vigorously by
kic:king, snapping or ruiiiiiiig away.
Tlic willingness to copulate wlien this i*esponseis given mas
tcstcd during an earlier study (Boliiig and Blandau, ’39)
by introducing a vigorwns male into the cage. Tnasmuch as
tlic copulatory response could be obtained from all animals
whicli would mate, elici tat ioii of the i~esponsc by nianual
manipulation in this study was consicic~~ccl
adequate evidence
for the existence of heat.
Tlic length of heat, tlic length of tlie c y l c a i i d the time
of day lieat loegins arc t ~ y a i d e das measures of normalit?
and \vcrc rccoi-decl for a11 aiiimnls observed during tlie iiivestigatioii. Tlic ability of tlw rat to withstand prolongcd
observation w a s tested hy esamiiiing thii-ty-seveii aiiimals
throughout 7 or 8 consecutive lieat periods.
J h t a bcai,irig on the extent to whicli consistency of hellarioi. is displayed were obtained f iwm tlic animals obsei*ved
during 7 01. 8 coiisccutivc lieat peiaiods. 11iscellaiicous information hearing o n the occurrciice of the “split ” estrus
(Young, Dcmpsey, Hagquist and Eoling, ’39) and the effect
of copulation on thc Icrigtli of heat, w a s obtained from the
cn tire colony.
OBSERVATIONS
Tlic length of 609 heat periods observed in 114 animals
averaged 13.7 hours, range 1 to 28 Iioiivs, standard deviation
4.55. This cornpaws favorably with tlie determination madr
by Ball, but is longer than Tsliii’s (’22) cstimatc of 4 to 8
hours and Long and Evans’ (’22) estimate of 3 to 12 hours.
When observations ~vei’emade thi*ougliont seven or cight
consecutive estrous periods the mean length of lieat did not
vary significantly (table 1 ) .
The time of day estrus began was not affected by frequent
ohservatioii. Analysis of 653 records is made in table 2.
I n the thirty-seven aiiimals observed during seven or cight
consccutivc heat periods the bcginiiing of heat TWS also most
457
LENGTH O F HEAT
TABLE 1
The lrngth of heat in liours in rats observed over consrcutive heat periods.
When estrus was antermittent ( " ) the length of the final part only
is shown in the table
- - ANTMAT.
17
27
28
31
37
45
61
68
77
81
83
111,s
112
113
131
137
142
162
167
168
181
184
183
187
190
194
19t
11
>9
>9
>16
9
>I2
15
17
15
20
20
13
19
7
14
13
>I0
>15
17
11
14
13
If:
197
200
19A
50A
52A
ilA
125A
156A
161A
182A
16
11
13
16
16
24
22
18
15
19
13
12
20
20
hle:111
14.01
196
2nd
3rd
10
12
13
22
8
20
16
I6
10
11
15*
14
22
15
15
12
18
17
16
5*
5
14
16
16
9
13
7
3
2
19
18
16
19
12
9
17
18
5
13
12
13
10
13
18
14
1.5
12
24
10
10
15
15
11
12
14
17
6
10
18
11
11
9
13
6
21
17
8
2
19
16
14
17
17
13*
"3"
13.51
12.72
12.89
- -
11
7
- -
4th
8
12
9
22
14
13
8
15*
11
12
18
11
14
12
11
17
14
11
17
10
10
9
21
12
12
13
8
10'
9
10
10
12
9
20
A th
MIRAN
8
17
18
19
5th
(ill1
11
14
5%
1.i
17
13
1.T
19
1ti
23
12
13
17
13
16
9
17
17
12
17
17
7
8
11
8
..
11
10
21
21
9
10
17
14
13
12
22
17
17
21
13
19
12
-
7th
- -
23
15
6
16
17
I0
18
16
1'7
11
6
14
20
18
11
19
16
17
9
22
16
16
15
12
16
21
20
13"
18
7
11
14
13
25
T
T
21
13
22
18
13
16
18
11
20
20
17
20
26
18
14
11
12
13
>I3
23
11
22
20
10.63
12.38
12.25
18.88
12.00
15.63
14.13
14.14
12.00
13.75
18.00
11.57
14.38
11.25
15.57
16.88
13.57
12.38
15.38
11.43
12.63
14.88
li.25
15.38
11.28
15.00
9.71
13.50
13.25
20.43
17.63
12.00
12.75
13.57
12.26
16.00
20.38
14.64
15.45
15.29
5.12
14.28
-
li
11
17
..
21
12
..
10
13
..
I
..
17*
14
24
10
19
18
15
23
23
19
11
17
5
19
..
..
Ii
21
7
1.6
..
..
..
18
19
..
16
4
18
..
- -
45 8
RICHARD J. BLANDAU A N D OTHERS
TABLE 2
Hourly record of the time of day at wkich heat begins
NO.
TIME O F DAY
T I M E O F DAY
OF
CASES
3
2
14
15
24
53
89
84
-
NO.
OF
T I X E OF DAY
CASKS
-
11 A.M. t o 12 N.
1 z N . to 1 P . M .
1 P.M. t o 2 P . M .
2 P . N . t o 3 P.M.
3 P . N . to 4 P.M.
4 P.N. t o 5 P.M.
5 P.M. to 6 P.M.
6 P.M. t o 7 P . M .
NO.
OF
CASES
7 P . M . t o 8 P.M.
8 P.M. to 9 P . N .
9 P.N. to i n P.N.
10 P.M. to 11 P.M.
11 P.M. to 12 P.M.
12 P . M . to 1 A . N .
1 A.M. t o 2 -4.51.
2 A . N . to 3 A . X .
102
71
91
33
34
17
8
8
-
3 A.M. t o 4 A . N .
4 A.M. to 5 A.M.
5 A.hf. t o 6 A.M.
6 A.M. to 7 A.M.
7 A.M. t o 8 A.M.
8 A . N . t o 9 A.M.
9 A . N . to 10 A . N .
10 A.N. t o 11 A.M.
2
0
n
1
2
0
0
0
frequent between 4 P.M. and 10 P.M. at each of tlie periods
of observation.
The length of 567 cycles varied from 2 to 19 days, a variation which is not essentially different from that recorded by
Long and Evans and Ball. Eighty-seven per cent were from
4 to 6 days in length, average 4.4 days or 105.3 hours. By
an application of Brandt's test for homogeneity of variance
(Snedecor, '38)t o data sumarized in table 3 it was determined
TABLE 3
T h e mean length of consecutive reproductive cycles
CYCLE
1st
2nd
3rd
4th
5th
Ni&IBE3E$
I
MEAN LENGTH
INHOURR
104.1
Gtli
105.2
m.3
102.3
99.3
7th
100.3
37
I
STANDARD
DEVIATION
in.2
15.8
24.8
23.7
20.8
14.4
14.6
I
90 to 122
47 t o 125
68 to 196
68 to 162
7 1 to 167
68 to 145
91 t o 118
that at no time during the six to seven cycles when the
animals mere being observed continuously was any significant
variation shown in the length of the cycle.
The data contained in table 1 were used in an effort to
ascertain whether or not consistency in the character of behavior as determined by the length of heat tends to be shown
LENGTH O F HEAT
459
by individual animals. When they were analyzed by the “F”
method of Snedecor, F equalled 3.3582. The probability,
therefore, that the smaller variance within animals is due to
chance is much less than one in a hundred and a considerable
consistency of behavior is shown. Similar analyses of data
bearing on the length of the cycle and on the time of day
heat begins indicate an individual consistency in these features
of reproductive behavior.
An abnormal type of heat previously observed in the guinca
pig (Young, Dempsey and Myers, ’35 ; Young, Dempsey, Hagquist and Doling, ’39) and horse (Satoh and Hoshi, ’33;
AlcKenzie and Andrews, ’37; Day, ’39) and referred to by
Young, Dempsey and Myers as a “split” estrus has been
scen in the rats observed during this study. Our information
is not as complete as that for the guinea pig because proof
cannot be given that in the rat the split, or perhaps better,
“intermittent” estrus, is associated with delayed ovulation.
I n every respect, however, in which comparison is possible
the intermittent estrus is identical with that seen in some
guinea pigs. The first part or parts tend to be shorter than
the final part with which ovulation is assumed to be associated.
I n sixteen cases the average length of the first part was 4.3
liours, range 1 t o 16 hours, whereas the average length of
the second p a r t was 9.1 hours, range 2 to 17 hours. When
a third part occurred, a s it did in three animals, the length
averaged 16 hours, range 10 to 23 hours. The length of the
interval between the parts of this abnormal estrus varied
from 2 to 23 hours.
I n the guinea pig the length of the normal heat periods of
animals which displayed an intermittent estrus tended t o
be longer than the average for the colony. I n the rat this
may or may not be true. The length of seventy-eight “normal ” heat periods displayed by fourteen animals which had
also displayed at least one intermittent estrus averaged 14.9
hours. The length of 468 heat periods displayed by seventythree aninials in which there was no record of an intermittent
estrus averaged 13.2 hours. The “t ” test for the sicgnificancc
460
R I C H A R D J. B L A N D A U A N D OTHERS
of differences (Snedecor, ’38) was applied to the data. The
value obtained for “ t ” was near the 5% level of significance
but was not conclusive.
The effect of mating at the beginning of estrus on the length
of that estrous period was studied in twelve animals. The
average length of seventy-six heat periods displayed by these
animals prior to that during which mating occurred was 14.4
hours. The average length of heat when mating was permitted at the beginning was 11.6 hours. When the “t” test
of Snedecor was applied t o the data, the value obtained for
“ t ” was greater than that shown in the table for the 1%
level; consequently the data for the twelve animals indicate that the difference between the length of heat in the
absence of mating and when it occurs is significant. Endoubtedly, however, it would be desirable to have more experiments. I n addition, the possibility should be explored
that a quantitative relationship exists between the amount of
stimulation and the continuation of heat which is similar to
that between tlie amount of stimulation and the production
of pseudopregnancy (Ball, ’34).
DISCUSSION
An objective in this investigation has been the development
of a method whereby periodic changes in the estrous cycle
of the rat can be followed at close intervals without disturbing
their normality. Providing the precautions described above
are taken, this can be accomplished by direct observations of
behavior. Throughout a period of as many as eight consecutive estrous periods, the length of heat, the time of day
it began, and the length of the cycle were not affected by
the procedure that was used.
The direct observation of behavior has the additional advantage that it permits determination of the beginning and
end of heat and therefore of its length. The usefulness of
the length of heat as a measure of reproductive normality
should be considerable. It has already been employed in this
study and elsewhere (Young, Dempsep, Hagquist and Boling,
LENGTH O F HEAT
461
'39) in an effort to ascertain something about the extent to
which individual differences are found. The fact that the
length of the induced estrogen-progesterone heat in spayed
rats was similar to that of heat in normal animals did much
to convince us that we were dealing with the combination of
ovarian hormones which is normally responsible for the display of this behavior (Boling and Blandau, '39).
Determination of the length of heat has furthered the comparison of the guinea pig and rat. Despite the rat's shorter
cycle, the period of heat is longer. I t s average length of 13
to 14 hours exceeds that in the guinea pig by approximately
5 hours. I n other respects, however, similarities are seen.
The facility with which the behavioral responses can be elicited
is about the same in the two species. Individuals of both
species show a considerable consistency of heat length from
cycle to cycle. Lastly, the abnormal heat referred to as
the intermittent estrus is displayed by both species.
It should be emphasized that the length of heat is not considered superior to the intensity of the estrous responses
(Hemmingsen, '33 ; Ball, '37) as a measure of reproductive
normality. Ideally, both should be recorded, especially when
the central interest is in mating behavior. For other purposes which require frequent examinations of the animals,
it is probable that either would be as satisfactory as the
other, depending on its convenience to the investigator.
SUMMARY AND CONCLUSIONS
The development of a procedure was sought whereby tests
of the estrous condition of the female rat can be made at
frequent intervals over a considerable period of time mithout disturbing the normality of the cycle. Reference to the
mating behavior displayed at the time of heat served this
purpose.
1. Provided proper precautions are taken, hourly tests for
sexual receptivity can be made over at least eight consecutive
estrous periods without affecting the length of heat, the time
of day animals come into heat or the length of the cycle.
462
KICIIABD J. BLANDAU AND OTHERS
2. The length of lieat is a n excellent standard of reference
for many studies of ovarian hormone action. I11 this study
it averaged 13.7 hours.
3. Individual animals tend to show a consistency in the
character of cyclic behavior as measured by the length of
heat, the length of the cycle and the time of day a t which
heat begins.
4. A ‘(split” or intermittent estrus similar to that displayed
abnormally by the guinea pig and horse is occasionally displayed by the rat.
5 . The average length of heat in twelve animals mated a t
the beginning of estrus was slightly, but nevertheless significantly shorter than in the controls.
LITERATURE C I T E D
BALL,J. 1934 Sex behavior of tlic ]at after removal of tlie uterus and vagina.
J. Comp. Psychol., vol. 18, pp. 419-422.
1936 Further evidence on hormonal basis of “heat” behavior.
Proc. Soc. Exp. Riol., vol. 35, pp. 4 1 6 4 1 8 .
1937 A test for measuring sexual excitability in the fcmale rat.
C‘omp. Psychol. Monog., vol. 14, no. 1, pp. 1-37.
___- 1940 The effect of testosterone on the sex I-diavior of female rats.
J. Conip. Psychol., vol. 29, pp. 151-165.
1939 The estrogen-progesteroiie induction of
BOLING,J. I,., A N D R. J. BLANDAU
niating responses in the spajed feniale rat. Endocrinol., vol. 25, pp.
359-364.
BROWNAN,
L. G. 1937 Light in its relation to activity and estrous rhythms in
the albino rat. J. Exp. Zool., vol. 7.5, pp. 375-388.
DAY, F. T. 1939 The oestrous cycic of the mare following removal of the
foetus a t tarious stages of pregiiaiicy. J. Agric. Sei., vol. 29, pp.
470-455.
HEYNINGSEN,
A. M.
1933 Studies on the oestrus-producing hormone (oestrin).
Skand. Arch. Physiol., rol. 63, pp. 97-250.
A. M., AND N. E. KRARVP 1937 Rhythmic diuriial varifitions i n
tlie ocstrous pheuoineiia of the rat and their susceptibility to light
and dark. I<. Ilanslie Vidensknb. Selskab. Biol. hfeddelel., vol. 13,
110. 7, pp. 1-61.
HEiflIINGSEN,
ISHII,0. 1922 Observations on the sexual cycle of the white rat. Anat. Rec.,
v01. 23, pp. 311-314.
LONG,J. A.,
AND H. M. EVANS 1922 The oestrous cycle in the rat a i d its
associ:itcd pliciiomrna. Mem. Vniv. Calif., 1-01. 6, pp. 3-148.
LENGTH O F HEAT
463
MCICENZIE,F. F., AND F. N. ANDREWS 1937 Estrus and ovulation in the mare.
Proc. Am. SOC.Animal Prod., vol. 30, pp. 64-70.
SATOH,
S., AND S. HOSHI 1933 A study of reproduction in the mare. 11. The
study on the oestrus. J . Jap. Soe. Vet. Sci., vol. 12, pp. 200-223.
SNEDECOR,
G. W. 1938 Statistical methods applied to experiments in agriculture
and biology. Collegiate Press, Ine., Ames, Iowa.
YOUNG,W. C., 3:. W. DEMPSEY,C. W. HAGQUIST,
AND J . L. BOLING 1939 Sexual
behavior and sexual receptivity in the female guinea pig. J . Comp.
Psycho]., V O ~ . 27, pp. 49-68.
YOUNG,W. C., E. W. DEMPSEY,AND H. I. MYERS 1935 Cyclic reproductive
behavior in the female guiuea pig. J. Comp. Pspchol., vol. 19, pp.
313-335.
YOUNG, W. c., AND B. RUNDLETT 1939 Tlic hormonal induction of llomosexual
behavior in the spayed female guinea pig. Psychosomatic Med., vol.
1, pp. 449-460.