Effect of estradiol-17P on blood flow to reproductive and nonreproductive tissues in pregnant ewes CHARLES

R.

FRANK

H.

MORRISS.

L.

MAKOWSKI

FREDERICK EDGAR

ROSENFELD

C.

GIA 0.20. is not significant at P > 0.10.

Microsphere infusion. Radioactive microspheres* with a specific activity of 10 mCi. per gram and a mean diameter of 25 p -+ 5 S.D. were employed. Approximately 150 PCi of 14’Ce and 350 &i of j’Cr were administered t-o each animal. The mean disintegrations per minute (DPM) and number of microspheres infused with each isotope are presented in Table I. The methods of infusing the microspheres and determining the number infused have been described.13 Prior to the microsphere infusion, reference samples were withdrawn from both femoral arterial catheters into counting vials by a pump? at a rate of 3.88 ml. per minute for three minutes. Simultaneous samples were also withdrawn from one of the uterine vein catheters and the femoral vein catheter for an equal period of

*Minnesota Mining and Manufacturing Co., St. Paul, Minnesota 55101. tHarvard Apparatus Co., 150 Dover Rd., Millis, Massachusetts 02054.

1.5, 1970; Gynecol.

time in order to detect the escape of mic-losphere.\ into the venous circulation from the uterine vascular bed and other vascular beds in the body. The DPM in the3e samples were compared to the DPM in the arterial sample (Table II). There was no significant loss of’ microspheres from the uterine vascular becl either before or after the vasodilatory effects ot I& confirming the results of pre\-ious studies from this laboratorv.7, ” There was a significant shunting of. microspheres across other- vascular beds into the inferior vena cava. equaling approximately 2 per cent of’ the infused number of microspheres. ‘[‘his flgui-e did not change significantly following an infusion of E:?. Tissue preparation. The ewes were weighed and killed jvithin two to four hours aftrl the second microsphere infusion and dissected f’or tissue preparation. All tissues were placed in prcweighetl, numbered counting vials (25 mm. in diameter. X0 mm. in length). The preparation of the ~lo~lp~-egmm~ and pregnant uteri and other tissues was 3s previousI\ described.l-’ Samples of skeletal muscle were taken from the lateral chest \~a11 and hind limb. Skin sample5 \\.ere taken from the foreleg. chest. and the skin overlying the mammary gland. Radioactive assay. The samples were counted in an automatic dual-channel gamma (,ounter*: ivith a threeinch crystal detector and a wide well i 1 !/x inches in diameter). by a method previousl!, reported.” Oxygen assay. Whole-blood oxygen content ~.as measured with a gas chromatographt equipped with a blood gas accessory.9 Date reduction. Individual organ blood flows \\crt calculated with the integrated arterial ref’erencr samples”: Organ blood flow (ml.imin.1 = total DPM in an organ mean DPM in arterial samples x rate of withdrawal. With a similar equation, mined simultaneously:

cardiac

output

1.

was deter-

Cardiac output (ml./min.) = total DPM infused mean DPM in arterial samples x rate of withdrawal. For the purpose

of statistical analysis the animals

2.

are

*Nuclear-Chicago Corp., 2000 Nuclear Dt-., Des PIaims. Illinois 60018. KX-PA, Beckman Instruments, Inc.. 2500 Harbor Blvd.. Fullerton, California 92634.

Volume Number

124 6

E2 effect

Table III. Blood nonpregnant and minutes after the Ez. The hormone two minutes

on blood

flow

to tissues

621

flow to the uterine tissues of pregnant ewes, before and 100 injection of 1 Fg per kilogram of was injected intravenously over Bloodjlow

Tissue

Before

Total uterine blood flow Nonpregnant 23.36 Group I 358.90 Group II 1,099.oo Myometrium Nonpregnant 9.46 Group I 25.20 Group II 35.28 Endometrium Nonpregnant 7.76 Group I 116.00 Group II 157.60 Cotyledons or Caruncles Nonpregnant 7.23 Group I 214.70 Group II 906.20

(ml.lmin.)

EZ

After

k 2.87 + 69.28 * 173.50 ” + t

346.20 625.40 1,575.OO

E2

k 28.27* ” 76.63* ” 231.70*

1.73 4.92 5.10

148.50 113.70 122.70

? f +

23.11* 22.21* 10.94*

k 1.27 ZL 15.78 ” 22.97

110.10 295.30 314.90

” f +

14.86* 41.33* 39.35*

IT 1.26 * 46.23 2 155.30

87.59 252.50 1,138.OO

2 11.09* f 42.8lt * 119.50t

The mean 2 1 S.E.M. is presented. *The change in flow is significant at P< 0.005 (paired t test). t The change in flow is significant at P < 0.05 (paired t test).

divided into three groups in the tables and figures: six nonpregnant ewes, seven ewes at 38 to 78 days of gestation (Group I), and six ewes at 120 and 139 days of gestation (Group II).

Results Effect of E2 on the uterine vascular bed. In previous reports we documented the inhibitory effect of surgical stress on the response of uterine blood flow to E2 in nonpregnant ewe+ I3 and thus performed this series of experiments after the animals had recovered from operation and had resumed a normal diet. In both pregnant and nonpregnant ewes there was a significant increase in uterine blood flow following the systemic infusion

of

average

increase

1 pg

per

kilogram

in

flow

of in

Ez (Table

milliliters

III). per

The minute

GESTATlOlYAL A,OE (DAYS)

Fig. 2. Effect of estradioLl7P on total Graph A shows the flow before and after per minute; B, the per cent change absolute change in flow (milliliters per nant, early pregnant (Group I), and late ewes.

uterine estradiol in flow; minute) pregnant

blood flow. in milliliters and C, the in nonpreg(Group II)

(Fig. 2. C) was not significantly different among the nonpregnant and the two pregnant groups and was equal data

to

351

expressed

ml.

per

as a per

minute cent

f

36

change

demonstrate a mean increase of S.E.M. in the nonpregnant ewes,

1,455 while

S.E.M.

The

same

in flow

(Fig.

2, B)

per cent ? 197 in the pregnant

animals there was a gradual decline in the relative magnitude of the change from 225 per cent at 38 days to a mean of 43 per cent near term (Group II). As shown in Table III and Figs. 3, 4, and 5, all three

major uterine tissues, i.e., myometrium, endometrium, and cotyledons (represented by the caruncles in the nonpregnant uterus), showed significant increases in blood

flow

following

an infusion

of Ez, but

the

pattern

and magnitude of response throughout gestation were different for each tissue. Myometrial blood flow increased after estrogen stimulation to approximately the same maximum (128 ml. per minute ? 11.4

622

March Am. J. Obstet.

Rosenfeld et al.

15, 1976

Gynecol.

o After E.stradbl-R$ . Before Estradkd-17.P

0

0

200

After . eefws

E~tmdkd-17# Edmdiol-17,B

. A 0

20

so QESTATIONAL

100 uo AGE (DAYS)

GESTATRXAL

Fig. 3. Effect of estradioL17P on myometrial Graph A shows the flow before and after estradiol per minute: B, the per cent change in flow: absolute change in flow (milliliters per minute) nant, early pregnant (Group I), and late pregnant ewes.

S.E.M.)

in

significant

pregnant increase

of

tion. On the contrary, estrogen-stimulated crease

during

the response stimulation trial

and

gestation was

and nonpregnant the base-line

both endometrial endometrial blood (Fig.

4. A).

of the placental not as pronounced

endometrial

flow

flows.

In

the

In

blood flow. in milliliters and C, the in nonpreg(Group II)

days

(Group

blood

flow

blood flow

doubled

iate

pregnancy

II), was

25

the per

average cent,

increase whereas

and

myometrial

flow

EZ stimulation flow to the nonpregnant

causes sites of animal,r3

an enormous implantation

blood flow to E, as that of myome-

in

group

effect

at 120 to 139

(MK)

Fig. 4. Effect of estradiol-17/I on endometrial Graph A shows the flow before and after estradiol per minute; B, the per cent change in flow; absolute change in flow (milliliters per minute), nant, early pregnant (Group I), and late pregnant ewes.

ewes despite a during gestaflow and did in-

ME

pregnancy of

there exogenous

estrogen

of

flow

3.5-fold. increase (caruncles)

in on

placental

endometrial

increased

it is apparent is a decrease

blood flow. in milliliters and C, the in nonpreg(Group II)

Since of

blood in the

that at some time the vasodilatory the

placental

ar-

Volume Number

Ez effect on blood flow to tissues

124 6

623

Y=lol5+t8 P-O.01

3ooo.

k

g l

z

t

e $

E A $

e After Estradiol- 17.p l Before Estradiob~l7. p

1600

t

! 0

Fig. 5. Effect of estradiol-17g on blood flow on the placental cotyledons. A is the flow before and after estradiol in milliliters per minute; B is the per cent change in flow, and C is the absolute change in flow (milliliters per minute), in the caruncles of the nonpregnant ewe and in the cotyledons of early (Group I) and late (Group II) pregnancy. terioles. This time can be located at approximately the second month of gestation by considering the experiments within Group I which extended from the thirty-eighth to the seventy-eighth day of pregnancy. At 38 days, estrogen stimulation caused a marked increase of cotyledonary flow from 30.6 to 109.5 ml. per minute, i.e., a 2.6-fold increase. In the experiment at 47 days, Ez caused the cotyledonary flow to double,

. 20

.

-=....o GESTATIONAL

.l

# . . loo 140 AGE (GAYSI .

A

Fig. 6. Effect of estradiol-17P on cervical blood flow. A is the flow before and after estradiol in milliliters per minute; B is the per cent change in flow, and C is the absolute change in flow (milliliters per minute) in the nonpregnant, early pregnant (Group I), and late pregnant (Group II) ewe.

from 45.1 to 89.7 ml. per minute. In the next two experiments at 60 days, the effect of exogenous Ez on cotyledonary blood flow was virtually zero, with a mean increase from 265.2 to 272.8 ml. per minute. In the next group of three experiments performed between the sixty-seventh and seventy-eighth days, there was an average increase of 13 per cent, from 299.0 to 340.9 ml. per minute. Effect of E2 on other reproductive tissues. There was a significant vasodilatory effect of Ez on blood flow to the tissues of the cervix, vagina, ovaries, uterine tubes, and mammary gland throughout pregnancy

624

Rosenfeld et al.

*

B f

March Am. J. Obstet.

15, 1976 Gynecol.

Y=l626+(-6.92)x PcO.05

300

6 ; Y E 200 0.

l

c * *t

1000

* * * t

6

P

f 6.0 ! E .m t

0

c

Y=251.7+(-1.57)x P- ,001

oAfter Estradiol-17,p *Before Estrediol-17,p

20

l --:

\

--

GEST%lONiL

A 0 After

i%

(QAY5; la0

l

6efwe

Estradbk17,B Estradlol-l7.p

2 .

Fig. 7. Effect of estradiol-17/3 on vaginal blood flow plotted against gestational age. A is the flow before and after estradiol in milliliters Per minute . gram, and B is the per cent change in flow.

(Table IV). As in the uterine tissues, as pregnancy progressed toward term, the cervix (Fig. 6). vagina (Fig. 7), and mammary gland (Fig. 8) demonstrated a reduction in the per cent change in flow following an infusion of E,. This phenomenon was not evident in the uterine tubes and ovaries. Effect of E2 on cardiac output. The reliability of simultaneous measurements of cardiac output and tissue or organ blood flows with the microsphere method has been reported.15 This method was employed in these experiments to demonstrate the effect of E2 on cardiac output in four of the nonpregnant animals and all of the pregnant animals included in this study. There was a mean increase in cardiac output of 13.3 ml. per minute . kilogram ? 4.25 S.E.M. (P < 0.005) at 60 to 90 minutes following an infusion of Ez. This is an increase of 14.5 per cent r?r 4.51 S.E.M. A similar increase was observed by Parer and associates lo following the infusion of large

4 :*

GESTA-L

ABE &MS)

Fig. 8. Effect of estradiol-17P on blood flow to the mammary gland. A is the flow before and after estradiol in milliliters per minute: B is the Per cent change in flow, and C is the absolute change in flow (milliliters per minute) in the nonpregnant, early pregnant (Group I), and late pregnant (Group II) ewe.

doses of estrone. No significant change in mean arterial pressure was noted: 97.9 mm. Hg before E2 and 96.8 mm. Hg afterward (P > 0.40). There was a significant change in heart rate, which increased from 100 to 111 beats per minute (P < 0.001). In the last animal studied, the Ez was administered in isotonic saline, 1 pug per milliliter, rather than in the carrier solution previously employed, which contains ethyl alcohol and

Volume Number

124 6

Eg effect on blood flow to tissues

2000m I -

T

Table IV; The effect of estradiol-17/3 (E,) on blood flow to reproductive tissues of pregnant and nonpregnant ewes. Same experimental conditions as Table III Blood Jaw Tissue Cervix Cervic

Before

Vagina

(ml.lmin

4.15 *

tubes (mklmin.

Groups I and II Ovaries with corpus luteum (ml. lmin.) Group I Group II Ovaries without corpu.s luteurn (ml.Imin.)

Group I and II Mammary

Afk

Ez

0.45

3.57

2

0.62

8.01

+

1.62

73.38 56.63 49.94

f 12.45* ” 4.86* 2 5.06*

0.207 0.185

f 0,042 2 0.024

3.31 2.13

2 t

0.77t 0.14*

0.474

f

4.01

f

0.28*

. Gm.)

Nonpregnant Groups I and II Uterine

E2

(ml. /min.) (ml. /min.)

Nonpregnant Group I Group II

L

625

) 0.086

9.12

t

1.14

4.82

f

0.51

14.81 f 8.20

+

l.llt

1.29*

0.252

2

0.029

1.00

f

0.24t

(ml.lmin.)

Nonpregnant Group I Group II

10.58 15.75 151.70

+ 2.79 k 3.51 2 26.34

32.06 39.74 200.70

5 5.58? k 16.2l.f + 23.49t

Fig. 9. Comparison of the per cent change in those tissues in the nonpregnant ewe which demonstrated a significant change in flow (P < 0.01) following a systemic infusion of estradioL17P (1 pg per kilogram). The nonreproductive tissuesrare represented by the shaded bars. The means f 1 S.E.M. are presented.

The pregnant animals are divided into unless there is no significant difference The mean 2 1 S.E.M. is,presented. *The change in flow is significant at P < tThe change in flow is significant at P

Effect of estradiol-17beta on blood flow to reproductive and nonreproductive tissues in pregnant ewes.

The effect of estradiol-17beta (1 mug per kilogram) on regional blood flow and cardiac output was studied by means of radionuclide-labeled microsphere...
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