Dehydration increases the renal response to atrial natriuretic peptide in fetal sheep Adam Dodd, MD, Kenenth Chan, MD, Linda K. Knllama, PhD, M. Gore Ervin, PhD, and Michael G. Ross, MD, MPH

Torrance, California OBJECTIVE: In sheep, maternal water deprivation results in urinary natriuresis in spite of suppression of plasma atrial natriuretic factor levels. Near-term fetal sheep also have a urinary natriuresis without change in plasma atrial natriuretic factor during maternal dehydration. This study was designed to explore the role of plasma atrial natriuretic factor levels in fetal dehydration-natriuresis. STUDY DESIGN: Eight chronically instrumented preterm (113 ± 1 days) ovine fetuses received two atrial natriuretic factor infusions (3 and 15 ng/kg/min) in a euhydrated state and after 48 ± 1 hours of maternal water deprivation. RESULTS: Dehydration significantly increased maternal plasma osmolality (302 ± 2 to 313 ± 2 mOsm/kg water), sodium (148.1 ± 0.8 to 154.3 ± 0.4 mEq/L), chloride (112.4 ± 0.6 to 116.8 ± 0.9 rnEq/L), and arginine vasopressin (4.2 ± 1.2 to 23.0 ± 4.0 pg/ml) and significantly decreased plasma atrial natriuretic factor (36 ± 6 to 19 ± 4 pg/ml) concentrations. Fetal plasma osmolality (296 ± 1 to 308 ± 2 mOsm/kg), atrial natriuretic factor (128 ± 16 to 241 ± 36 pg/ml), and arginine vasopressin (3.5 ± 0.8 to 12.3 ± 4.8 pg/ml) concentrations and urine osmolality (170 ± 10 to 253 ± 10 mOsm/kg), osmolar clearance (0.80 ± 0.02 to 0.14 ± 0.02 ml/kg/min), and fractional sodium excretion (3.3% ± 1.7% to 8.5% ± 2.1%) increased significantly with dehydration, whereas the plasma atrial natriuretic factor clearance decreased from 127 ± 27 to 63 ± 10 ml/kg/min. Dehydration had no effect on fetal hematocrit, vascular pressures, glomerular filtration rate, urine flow, or free water clearance. In euhydrated fetuses plasma atrial natriuretic factor increased from 128 ± 16 to 287 ± 46 pg/ml with sequential atrial natriuretic factor infusion, and no significant increases were observed in urine flow, fractional sodium excretion, and glomerular filtration rate. In contrast, atrial natriuretic factor infusion to dehydrated fetuses significantly increased urine flow (0.17 ± 0.03 to 0.32 ± 0.07 ml/kg/min), osmolar clearance (0.14 ± 0.02 to 0.28 ± 0.06 ml/kg/min), and fractional sodium excretion (8.5% ± 2.1% to 14.8% ± 4.0%). CONCLUSION: These results demonstrate that in the fetus at 113 days' gestation plasma atrial natriuretic factor levels increase with dehydration, probably a result of decreased plasma atrial natriuretic factor clearance, and the fetal renal responsiveness to atrial natriuretic factor infusion increases during maternal dehydration. (AM J OBSTET GVNECOL 1992;167:1710-6.)

Key words: Ovine, fetal, renal atrial natriuretic factor response, dehydration

Atrial natriuretic factor (ANF) denotes a family of atrial pep tides that promote natriuresis, diuresis, and vasorelaxation in mammals. I. 2 Ovine fetal plasma ANF levels are significantly higher than maternal values and increase in response to volume expansion and hypoxia. 3 • 4 Intravenous ANF infusions result in no change 2 • 5 or a decrease 6 in fetal arterial blood pressure From the Department of Obstetrics and Gynecology, Harbor-University of California, Los Angeles Medical Center, University of California, Los Angeles School of Medicine. Funded by National Institutes of Health award HL-40899. Adam Dodd was a recipient of a research fellowship sponsored by the Stanley J. Sarnoff Endowment for Cardiovascular Science. Presented in part at the Thirty-ninth Annual Meeting of the Society for Gynecologic Investigation, San Antonio, Texas, March 18-21, 1992.

Reprint requests: Michael G. Ross, MD, MPH, Harbor-UCLA Medical Center, 1124 West Carson St., RB-1, Torrance, CA 90502. 6/6/41936

1710

and no change" or an increase,5.6 in fetal heart rate. Whereas the natriuretic effects of ANF are diminished in the ovine fetus as compared with those of the adult/ greater diuretic and natriuretic responses are observed in preterm versus near-term fetal sheep. 7 In response to dehydration, adult sheep have a urinary antidiuresis but a significant natriuresis." Although the natriuresis would seem to conflict with the observed 50% reduction in plasma ANF levels, 9 maternal renal medullary ANF receptor binding capacity increases ninefold after dehydration. lo Although fetal sheep at 127 days' gestation also have a urinary natriuresis in response to maternal dehydration, plasma ANF levels do not change and there is no significant change in binding capacity of renal ANF receptors. 9 • 10 Whereas the increased maternal renal binding capacity may explain maternal urinary natriuresis, the role of plasma ANF levels in the regulation of fetal natriuresis is unclear.

Volume 167 Number 6

We explored the role of fetal plasma ANF in the regulation of fetal urine flow and composition during dehydration. As fetal renal responses to ANF decrease with advancing gestational age, we elected to study the ovine fetus in the early third "trimester" (113 days, or 0.78 of gestation). Methods and material

Animal preparation and surgery. Eight chronically instrumented Western cross-bred ewes (Nebeker Ranch, Lancaster, Calif.) with singleton fetuses were studied. Ewes were maintained indoors in individual steel cagos before and after surgery, with a controlled 12 hours light and 12 hours dark photoperiod. Food and water were available as desired except for the withholding of food for 24 hours before surgery and withholding of water as described in the following text. Anesthesia was induced by intramuscular injection of ketamine hydrochloride (20 mg/kg) and atropine sulfate (30 lJ..g/kg) and maintained by maternal endotracheal ventilation with isoflurane. The maternal abdomen was opened by midline incision, the uterus was exposed, and both fetal hind limbs were delivered through a small hysterotomy. A fetal bladder catheter (Tygon, inside diameter 1.3 mm, outside diameter 2.3 mm) was inserted via cystotomy and polyethylene catheters (Tygon, inside diameter 1.0 mm, outside diameter 1.8 mm) were threaded into the fetal descending aorta and inferior vena cava via the femoral artery and vein, respectively. Before hysterotomy closure, a plastic catheter (Corometrics Medical Systems, Wallingford, Conn.) was inserted into the amniotic cavity. The uterus and maternal abdomen were closed in layers. Polyethylene catheters also were threaded into the maternal abdominal aorta and inferior vena cava via the femoral artery and vein, respectively. All catheters were externalized to the ewe's flank and placed in a cloth pouch. Animals received 3 days of postoperative antibiotics to the ewe and fetus. Maternal and fetal catheters were flushed daily with heparinized saline solution (10 IU/ml), subsequently filled with sodium heparin solution (10 and 1000 IU/ml, respectively), and sealed with sterile plastic caps. Experimental protocol. Experiments were performed a minimum of 5 days afte.- surgery on ewes standing in the same cages in which they were housed. The mean gestational age at the initiation of studies was 113 ± 1 day. Water was removed from the ewe at the beginning of the study on day 1 (8 AM). During a 60-minute equilibration period (- 60 to time 0), the fetal bladder catheter was drained to gravity, and an intravenous infusion of tritiated inulin (10 IJ..Cilhr) in 0.15 mol/L sodium chloride (0.05 ml/kg/min) was initiated for determination of glomerular filtration rate. Fetal arterial pH and urine osmolality were assessed,

Ovine fetal renal ANF response during dehydration

1711

and the study was continued only if the arterial pH was ~ 7.30 and fetal urine osmolality was < 200 mOsmJkg water. Mter the equilibration period, fetal urine was collected into glass tubes at 10-minute intervals. The study protocol consisted of four 40-minute periods (control, two ANF infusion periods, recovery) with 0.15 mol/L sodium chloride or ANF in 0.15 mol/L sodium chloride delivered at 0.05 ml/kg/min. During the control period (0 to 40 minutes), fetuses received intravenous 0.15 mol/L sodium chloride infusion. At 40 minutes synthetic human ANF(I-28) (Bachem, Torrance, Calif.) was administered at 3 ng/kg/min and subsequently increased to 15 ng/kg/min at 80 minutes. At 120 minutes the 0.15 mol/L sodium chloride infusion was resumed and the fetus was monitored for a final 40-minute recovery period. Infusion rates were based on the estimated fetal body weight from the formula described by Robillard and Weitzman. II Fetal and maternal arterial blood samples (3.5 to 4.0 ml) were withdrawn at 15, 25, and 35 minutes of each study period and placed into chilled tubes. For ANF determination, tubes contained ethylenediaminetetraacetic acid (l mg/ml), aprotinin (250 kallikrein inhibitor units per milliliter), and soybean trypsin inhibitor (N-a-benzoyl-L-arginine ethyl ester, 50 U/ml). For hematocrit, plasma osmolality, sodium, potassium, and chloride concentrations and tritiated inulin, the tubes contained lithium heparin (10 lJ..l/ml blood). To minimize blood volumes, samples for determination of plasma arginine vasopressin (AVP) levels were collected only at 15 and 25 minutes of the control period. All samples were centrifuged immediately at 4° C, and plasma aliquots for ANF and AVP assays were separated and stored at - 20° C until extracted. Additional arterial blood samples (0.5 ml) were collected into heparinized syringes for determination of arterial pH, Po 2 , and Pco 2 • Fetal blood samples were replaced with an equal volume of heparinized maternal blood withdrawn before the study. At the end of each urine collection period (10 minutes), urine flow, osmolality, and sodium, potassium, and chloride concentrations were measured and aliquots were taken for measurement of tritiated inulin levels. At 48 hours of dehydration (day 3), a protocol identical to that of study day 1 was initiated. Analytic methods. Plasma ANF and AVP samples were extracted and levels were determined by radioimmunoassay as previously described. 12 • 13 The sensitivity of the ANF assay is 2 pg/ml, and the intraassay and interassay coefficients of variation were 11 % and 13%, respectively. The sensitivity of the AVP assay is 0.8 pg/ml with intraassay and interassay coefficients ofvariation of 6% and 9%, respectively. Maternal and fetal arterial blood pressures and am-

1712 Dodd et al.

December 1992 Am J Obstet Gynecol

Table I. Mean maternal arterial measurements before, during, and after intravenous fetal infusions of ANF (3 and 15 ng/kg/min) on day 1 (control) and day 3 (after 48 hours' dehydration) ANF infusion Day Osmolality (mOsm/kg water) Sodium (mEqlL) Potassium (mEqIL) Chloride (mEqlL) ANF (pg/ml) AVP (pg/ml)

1 3 1 3 I -3 I 3 1 3 I 3

Control 302 313 148.1 154.3 4.4 4.2 112.4 116.8 36 19 4.2 23.2

± ± ± ± ± ± ± ± ± ± ± ±

2 2* 0.8 0.4* 0.1 0.1 0.6 0.9* 6 4* 1.2 4.0*

3 ng/kg/min 301 311 147.6 153.5 4.5 4.2 112.5 116.7 38 25

± ± ± ± ± ± ± ± ± ±

2 2 0.8 0.5 0.1 0.1 0.8 0.8 6 II

I

15 ng/kg/min 301 310 147.9 153.5 4.5 4.2 112.8 116.5 34 21

± ± ± ± ± ± ± ± ± ±

2 2t 0.9 0.5 0.1 0.1 0.5 0.7 5 6

Recovery 302 310 147.8 153.2 4.5 4.2 112.7 117.8 35 23

± ± ± ± ± ± ± ± ± ±

2 2t 1.0 0.5t 0.1 0.1 0.6 1.5 6 6

Values are mean ± SEM. *P < 0.05, versus day I. tp < 0.05, versus control period.

niotic fluid pressures were continuously recorded with a Beckman R-612 physiologic recorder (Beckman Instruments, Irvine, Calif.) with Statham P23 transducers (Gould, Waltham, Mass.). Fetal blood pressures were standardized by subtracting amniotic fluid pressure. Blood pH, P0 2 , and Pco 2 were determined at 39° C with a Radiometer BM 33 MK2-PHM 72 acid-base analyzer system (Radiometer, Copenhagen). Plasma and urine osmolalities were measured by freezing-point depression with an Advanced Instruments Osmometer (model MO, Advanced Instruments, Needham Heights, Mass.). Plasma and urine sodium, potassium, and chloride concentrations were determined with a Nova 5 electrolyte analyzer (Nova Biomedical, Waltham, Mass.). Plasma and urine tritiated inulin levels were assessed by counting 100 fLl aliquots diluted in 10 ml of Hydrofluor (National Diagnostics, Sommerville, N.].) in a Beckman LS-355 liquid scintillation counter. Calculations and statistics. All values are expressed as mean ± SEM. ANF plasma clearance (PC ANF ) rates were calculated as PCANF = ANFin!(PANF steady state - PANF basal), where ANF inf is the ANF infusion rate (nanograms per kilogram per minute) determined from radioimmunoassay measurements of the infusates and PANF steady state and PANF basal are the plasma ANF concentrations (picograms per milliliter) after correction for extraction recovery at either steady state during the 15 ng/kg/min ANF infusions or during the basal periods, respectively. Differences over time were assessed by one-way and two-way analysis of variance for repeated measures, with Dunnett's post hoc tests. Basal day 1 (euhydrated) and day 3 (dehydrated) fetal values were compared by paired t tests. Results

Dehydration. During the basal period of study day 1, ewes and fetuses had normal plasma and urinary values

(fables I to III, Fig. 1). Control maternal hematocrit (30.0% ± 1.5%), pH (7.50 ± 0.01), P0 2 (87 ± 3 mm Hg), Pco 2 (30 ± 1 mm Hg), systolic (101 ± 4 mm Hg) and diastolic (69 ± 4 mm Hg) blood pressures, and heart rate (l07 ± 3 beats/min) did not change significantly in response to dehydration or fetal ANF infusions. In response to maternal dehydration, maternal plasma osmolality and sodium, chloride, and AVP concentrations significantly increased and plasma ANF levels significantly decreased (fable I). Similarly, fetal plasma osmolality and sodium, potassium, chloride, and AVP concentrations significantly increased with maternal dehydration (fable II). Basal fetal plasma ANF levels were significantly greater than maternal values (128 ± 16 vs 36 ± 6 pg/ml) and significantly increased (241 ± 36 pg/ml) with dehydration (Fig. 1). There were no significant changes in fetal pH (7.41 ± 0.01), P0 2 (23 ± 1 mm Hg), or Pco 2 (42 ± 1 mm Hg) in response to maternal dehydration or fetal ANF infusions. Expected increases in fetal urine osmolality, sodium, chloride, and potassium concentrations were observed during dehydration (fable III). Fetal ~rinary osmolar clearance and fractional sodium excretion significantly increased, free water clearance decreased, and glomerular filtration rate was unchanged (Table III, Fig. 1). ANF infusion. ANF infusate concentrations were assayed, and ANF infusion rates of 3 ± 1 and 15 ± 2 ng/kg/min were confirmed on both days of study. Because no difference was noted in fetal plasma ANF levels at the sampling times of 15, 25, and 35 minutes during each infusion period, all results are expressed as the mean value for each respective infusion period. On study days 1 and 3 ANF infusions to euhydrated and dehydrated fetuses increased plasma ANF levels approximately twofold over control values during the 15 ng/kg/min infusion and remained significantly elevated

Ovine fetal renal ANF response during dehydration

Volume 167 Number 6

during the recovery period (Fig. 1). Consistent with the small, nonsignificant increase in plasma ANF levels during the 3 ng/kg/min infusion, there were minimal fetal arterial or urinary effects at this dose (Table II). However, the 15 ng/kg/min ANF infusion resulted in a significant decrease in fetal systolic blood pressure during both euhydration and dehydration. A small but significant decrease in maternal and fetal plasma osmolalities was demonstrated in dehydrated fetuses during the ANF infusion. There were no changes in fetal hematocrit, pH, P0 2 , Pc0 2 , diastolic blood pressure, or plasma electrolyte concentrations during ANF infusions to euhydrated and dehydrated animals. In euhydrated fetuses the ANF infusions evoked minimal urinary effects evidenced by small, nonsignificant increases in urine flow and sodium and chloride concentrations (Table III, Fig. 1). However, the 15 ng/kg/min ANF infusion to dehydrated fetuses resulted in a twofold increase in urine flow and osmolar clearance and a marked increase in fractional sodium excretion. Although the 15 ng/kg/min ANF infusion to dehydrated fetuses was associated with a 40% increase in glomerular filtration rate, this was not statistically significant. The calculated plasma ANF clearance significantly decreased from 127 ± 27 to 63 ± 10 ml/kg/min in response to 48 hours of maternal water deprivation. Comment

Previous studies of maternal water deprivation have demonstrated parallel increases in maternal and fetal plasma osmolalities and sodium concentrations. 9, 14, 15 With prolonged dehydration, maternal plasma AVP levels increase to a greater degree than fetal levels, perhaps as a result of the reduced sensitivity for fetal AVP secretion in response to hypertonicity. In the adult sheep dehydration-induced AVP secretion increases urine osmolality and decreases urine flow and free water clearance." Dehydration is also often characterized by natriuresis, even though plasma ANF levels are suppressed. The ovine fetus similarly has increased urine osmolality and reduced free water clearance during dehydration, likely mediated by increased plasma Avp.9, 14·16 Similar to previous studies, 48 hours of maternal water deprivation resulted in no change in fetal urine flow rate and glomerular filtration rate and a marked increase in fractional sodium excretion. However, exogenous AVp infusions achieving plasma AVP levels similar to those measured during dehydration result in a marked decrease in fetal urine flow.17 Although increased plasma AVP, renal tubular immaturity, and intrarenal factors may contribute to a natriuresis, our objective was to determine if increased ANF sensitivity contributes to fetal dehydration-induced natriuresis. In agreement with previous studies plasma ANF lev-

•

IlOO

i"" ~

i

500 400

• )

300 200

~

1713

100 0

n

•

,1

,1

0.5

.......

~

~E

'"' 1&1 2 3 ~

1&1

z

12 ::J

•

0.4 O.l 0.2

).

)

l

0.1 0,0 0.5

....... c::

0.4

"'-

O.l

'E

~

-

•

~

E

E lit

0

0

0.2

•

0.1 0.0 20

•

•

15

g

t!

10

15 MF INFlJSION

RECO\/ER'(

(no/kg/mln)

Fig. 1. Fetal plasma ANF, urine volume, osmolar clearance (Cos..), and fractional sodium excretion (FEN,) in euhydrated

(open bars) and dehydrated (solid bars) during control, ANF infusion (3 and 15 ng/kglmin), and recovery periods. Asterisk, p < 0.05 (vs control period values).

els were three to four times higher in the fetus than in the ewe" In euhydrated fetal sheep Ervin et al.' suggested that higher baseline fetal ANF levels are due to increased fetal ANF production rather than decreased plasma clearance. In fact, ANF clearance rates are

1714 Dodd et al.

December 1992 Am J Obstet Gynecol

Table II. Mean fetal arterial measurements before, during, and after intravenous fetal infusions of ANF (3 and 15 ng/kg/min) on day 1 (control) and day 3 (after 48 hours' dehydration) ANF infusion

Day

Hematocrit

(%)

Systolic blood pressure (mm Hg) Diastolic blood pressure (mm Hg) Mean blood pressure (mm Hg) Heart rate (beats/min) Osmolality (mOsm/kg water) Sodium (mEqlL) Potassium (mEqlL) Chloride (mEqlL) AVP (pglml) Values are mean

±

1 3 1 3 1 3 1 3 1 3 1 3 1 3 1 3 1 3 1 3

Control 34.5 ± 33.5 ± 49 ± 52 ± 32 ± 33 ± 38 ± 40 ± 185 ± 180 ± 296 ± 308 ± 140.0' ± 146.8 ± 4.0 ± 4.3 ± 107.3 ± 111.7 ± 3.5 ± 12.3 ±

J ng/kg/min

1.0 0.7 2 1 2 1 2 1 3 5 1 2t 0.8 0.5t 0.2 O.lt 0.7 O.4t 0.8 4.8t

34.0 33.5 48 51 31 33 38 40 191 185 295 305 139.9 146.4 4.0 4.3 107.0 111.9

I

± 1.0 ± 0.7 ± 1 ± 1 ± 1 ± 1 ± 1 ± 1 ± 6 ± 5 ± 2 ± 2* ± 1.2 ± 0.4 ± 0.2 ± 0.1 ± 0.9 ± 0.3

15 ng/kg/min 34.6 34.2 46 50 31 34 37 39 190 189 296 305 140.3 146.3 4.0 4.3 108.0 112.0

± 1.1 ± 0.8 ± 2* ± 1* ± 1 ± 1 ± 1* ± 1 ± 5 ± 6 ± 2 ± 2* ± 1.1 ± 0.4 ± 0.1 ± 0.1 ± 0.8 ± 0.3

Recovery 34.3 34.4 45 50 31 34 36 39 204 191 297 307 140.6 146.8 3.9 4.3 108.0 112.0

± ± ± ± ± ± ± ± ± ± ± ± ± ± ±

± ± ±

1.3 0.9* 1* 1* 1 1 1* 1 9* 8 3 2 l.l 0.5 0.1 0.1 0.6 0.4

SEM.

*P < 0.05, versus control period. tp < 0.05, versus day 1.

Table III. Mean fetal urinary values before, during, and after intravenous fetal infusions of ANF (3 and 15 ng/kg/min) on day 1 (control) and day 3 (after 48 hours' dehydration) ANF infusion

Day

Urinary osmolality (mOsm/kg/water) Sodium (mEqlL) Potassium (mEqlL) Chloride (mEqlL) Free water clearance (mVmin/kg) Glomerular filtration rate (mVminlkg)

1 3 1 3 1 3 1 3 1

3

1

3

Control 169.9 253.3 31.9 82.1 8.0 13.6 28.6 61.1 0.07 0.03 1.2 1.2

±

J ng/kg/min

10.0

± 29.3* ±

7.3

± 14.2* ± 2.8 ±

4.4*

± 4.3 ±

10.0*

± 0.02 ± 0.02

0.2 ± 0.1 ±

170.7 264.7 33.3 89.7 7.3 13.0 29.9 67.5 0.09 0.03 1.3 1.4

10.8 ± 25.8 ± 6.9 ± 13.7 ± 2.5 ± 3.1 ± 4.3 ± 10.1 ± 0.02 ± 0.02 ± 0.2 ± 0.3 ±

I

15 ng/kg/min 169.2 272.9 36.2 10 1.3 6.1 10.6 32.0 77.6 0.10 0.03 1.5 1.7

±

9.6

± 22.5 ± 8.2

± ± ± ± ± ±

± ± ±

14.3t 2.2 2.5 5.5 1O.5t 0.02 0.02 0.2 0.3

Recovery 176.5 306.4 41.9 115.3 5.4 11.6 32.5 86.2 0.07 0.00 1.2 1.2

±

18.0

± 16.ot ± 9.7

± l1.3t ± 1.8 ± 2.2 ± 7.0 ± 8.6t ± 0.02 ± O.Olt ± 0.2 ± 0.2

Values are mean ± SEM. *P < 0.05, versus day 1. tp < 0.05, versus control period.

higher in the ovine fetus than in the pregnant ewe. 5 Dehydration decreases plasma ANF in pregnant ewes but not fetuses at 127 days' gestation. 9 In the current study the dehydration-induced increase in plasma ANF levels observed in the fetuses at 113 days' gestation has not been reported previously. There was no evidence of fetal hypoxia, volume expansion, or increased arterial pressure that could account for stimulation of ANF secretion. The reduced plasma ANF clearance rate

(127 ± 27 to 63 ± 10 mUkg/min) after maternal water deprivation likely contributed to the increased fetal plasma ANF levels. Plasma ANF clearance occurs in the lungs and kidneys of adult mammals and potentially the placenta during fetal life. 18. 19 Whether changes in fetal blood flow distribution to these organs or other factors are responsible for the reduced ANF clearance during dehydration was not assessed. The significant decrease in maternal and fetal plasma

Volume 167 Number 6

osmolalities during the infusions to dehydrated fetuses is unlikely a direct ANF effect since ANF does not cross the ovine placenta in significant quantities 20 and maternal plasma ANF levels did not change. In addition, the increase in fetal osmolar excretion is not sufficient to account for the decrease in fetal and maternal plasma osmolalities. Plasma osmolality changes may have been a result of maternal feeding-induced fluid fluxes!' Previous studies that examined the renal effects of ANF infusions into fetal sheep have produced numerous and often differing results. Robillard et ai. 2 observed significant fetal natriuresis without changes in urine flow rate or glomerular filtration rate with ANF infusion rates of25 and 100 ng/kg/min in fetuses of 128 to 139 days' gestation. Brace et al. 6 observed both increased glomerular filtration rate and increased urine flow in response to ANF infusions ranging from 14 to 300 ng/kg/min in fetuses of 126 to 133 days. However, Hargrave et al. 22 observed constant urine flow rates, glomerular filtration rate, and urine sodium levels with ANF infusions of 27 and 55 ng/kg/min in fetuses of 126 to 131 days' gestation. More recently Castro et aI. 7 reported increased glomerular filtration rate and urine flow rates in 114-day fetuses using ANF infusion rates of 25 to 100 ng/kg/min but no change in glomerular filtration rate using the same infusion rate in 131day fetuses. Shine et ai. 23 also studied fetuses whose range of gestational ages encompassed 113 days' gestation and observed both natriuresis and diuresis with ANF infusion rates of approximately 5 to 25 ng/kg/min. Consistent with these studies, the low-dose (3 ng/kg/min) ANF infusion rate did not change urinary parameters in either the dehydrated or the euhydrated fetuses. Similarly, there was no significant effect of the 15 ng/kg/min infusion in euhydrated fetuses. However, the 15 ng/kg/min ANF infusion evoked a marked diuresis and natriuresis in the dehydrated fetuses. The increased plasma ANF levels in the dehydrated fetuses (a result of decreased clearance) could account for the increased renal effects. Increased AVP also may have contributed to the observed increased renal responsiveness to ANF infusions, because AVP may potentiate ANF -induced natriuresis. 24 Although the ANF -induced diuresis ended with the completion of the ANF infusion, the natriuresis observed in the dehydrated fetuses persisted during the recovery period. Increased plasma ANF levels correlated with maximum glomerular filtration rate and urine flow rates, suggesting that the diuresis may be secondary responses to direct ANF effects on fetal glomerular filtration rate. ANF tubular effects and postreceptor events may explain the persistent increase in fractional sodium excretion during the recovery period. Both the fetal natriuresis and an intravascular to interstitial flow 6 may have contributed to a reduction in

Ovine fetal renal ANF response during dehydration

1715

plasma volume (as reflected by the increase in fetal hematocrit). The state of hydration has been shown to influence the effect of ANF in adult animals. In adult dehydrated sheep renal arterial infusion of ANF (50 IJ.glhr) resulted in a blunted natriuretic and diuretic response as compared with that of euhydrated sheep!' The reduced response was postulated to be a secondary effect of the negative sodium and fluid balance of the dehydrated animals. Similarly, dehydration attenuates the natriuretic response to ANF in adult rats!6 Both adult sheep and rats may exhibit hemoconcentration and total body sodium depletion during dehydration. However, as fetal intravascular volume is maintained9 • 27 and plasma sodium level increases, there may be an increase in net fetal body sodium during maternal dehydration. Thus fetal renal ANF responses during dehydration may be discrepant from those of the adult. The results of the current study may well apply only to ovine fetuses near 113 days' gestation, because there are marked differences in the response of older-gestation fetuses. Increased renal responsiveness to AVP in sheep at 125 to 130 days versus sheep at younger gestational ages II indicates maturation of tubular resorptive function. However, the fetal renal response to ANF has been shown to decrease during this time period. 7 In addition, fetal responses to dehydration vary with gestational age: In contrast to the current results, fetal sheep at 127 days' gestation show no change in plasma ANF levels in response to maternal dehydration. 9 Future studies will examine the ontogeny of the role of plasma ANF in the fetal renal response to dehydration. In conclusion, plasma ANF levels increase in response to dehydration in ovine fetuses at 113 days' gestational age, possibly as secondary effects of decreased plasma ANF clearance. Dehydration increases the fetal renal responsiveness to near physiologic infusions of ANF and suggests that ANF plays an important role in fetal fluid homeostasis during pregnancy. REFERENCES 1. Metzler CH, Ramsay DJ. Atrial peptide potentiates renal responses to volume expansion in conscious dogs. Am J Physiol 1989;256:R284-9. 2. Robillard JE, Nakamura KT, Varille VA, Andresen M, Matherne GP, Van Orden DE. Ontogeny of the renal response to natriuretic peptide in the sheep. Am J Physiol 1988;254:F634-41. 3. Ross MG, Ervine MG, Lam RW, Castro L, Leake RD, Fisher DA. Plasma atrial natriuretic peptide response to volume expansion in the ovine fetus. AM J OBSTET GYNECOL 1987; 157: 1292-6. 4. Cheung CY, Brace RA. Fetal hypoxia elevates plasma atrial natriuretic factor concentration. AM J OBSTET GYNECOL 1988; 159: 1263-8. 5. Ervin MG, Ross MG, Castro R, et al. Ovine fetal and adult atrial natriuretic factor metabolism. Am J Physiol 1988; 254:R40-6.

1716 Dodd et al.

6. Brace RA, Bayer LA, Cheung CY. Fetal cardiovascular, endocrine and fluid responses to atrial natriuretic factor infusion. Am] Physiol 1989;257:R580-7. 7. Castro R, Ervin MG, Leake RD, Ross MG, Shennan D], Fisher DA. Fetal renal response to atrial natriuretic factor decreases with maturation. Am] Physiol 1991;260:R34652. 8. McKinley M], Denton DA, Nelson ]F, Weisinger RS. Dehydration induces sodium depletion in rats, rabbits and sheep. Am] Physiol 1983;245:R287-92. 9. Agnew CL, Ross MG, Fujino Y, Ervin MG, Kullama LK. Maternal-fetal dehydration: prolonged effects and responses to oral rehydration. Am] Physiol [In press]. 10. FtUino Y, Ross MG, Ervin MG, Castro R, Leake RD, Fisher DA. Ovine maternal and fetal glomerular atrial natriuretic factor receptors: Response to dehydration. BioI Neonate [In press]. 11. Robillard ]E, Weitzman RE. Developmental aspects of the fetal renal response to exogenous arginine vasopressin. Am] Physiol 1980;238:F407-14. 12. Castro LC, Lam RW, Ross MG, et al. Atrial natriuretic peptide in the sheep.] Dev Physiol 1988;10:235-46. 13. Skowsky WR, Rosenbloom M, Fisher DA. Radioimmunoassay of arginine vasopressin, development and application.] Clin Endocrinol Metab 1974;38:278-87. 14. Ross MG, Shennan D], Ervin MG, Castro R, Humme J. Maternal dehydration-rehydration: fetal plasma and urinary responses. Am] PhysioI1988;255:E674-9. 15. Schreyer P, Shennan D], Ervin MG, Day L, Ross MG. Maternal dehydration: impact on amniotic fluid volume and composition.] Dev Physiol 1990;13:283-7. 16. Stevens AD, Lumbers ER. The effect of maternal fluid intake on the volume and composition of fetal urine.] Dev Physiol 1985;7: 161-6. 17. Lingwood B, Hardy K], Horacek I, Scoggins BA, Wintour EM. The effects of antidiuretic honnone on urine flow and

December 1992 Am J Obstet Gynecol

18.

19. 20. 21.

22. 23. 24. 25. 26.

27.

composition in the chronically-cannulated ovine fetus. Q] Exp Physiol 1978;63:315-30. Robillard ]E, Nakamura KT, Varille VA, Matherne GP, McWeeny OJ. Plasma and urinary clearance rates of atrial natriuretic factor during ontogeny in sheep.] Dev Physiol 1988; 10:335-46. Turrin M, Gillis CN. Removal of atrial natriuretic peptide by perfused rabbit lungs in situ. Biochem Biophys Res Commun 1986;140:868-73. Cheung CY, Gibbs DM, Brace RA. Atrial natriuretic factor in maternal and fetal sheep. Am] PhysioI1987;252:E27982. Mellor D], Slater ]S. The composition of maternal plasma and foetal urine after feeding and drinking in chronically cathererized ewes during the last two months of pregnancy.] Physiol 1973;234:519-31. Hargrave BY, Iwamoto HS, Randolph AM. Renal and cardiovascular effects of atrial natriuretic peptide in fetal sheep. Pediatr Res 1989;26:1-5. Shine P, McDougall ]G, Towstoless MK, Wintour EM. Action of atrial natriuretic peptide in the immature ovine kidney. Pediatr Res 1987;22:11-5. Graczak LM, Nicolodi LK, Hartupee DA, Blaine EH. Arginine vasopressin potentiates natriuretic effect of atrial peptide. Am] Physiol 1989;256:H925-7. Yates NA, Coghlan]P, Scoggins BA, McDougall]G. Renal effects of atrial natriuretic factor (99-126) in water-deprived sheep. Clin Exp Phannacol Physiol 1989; 16:433-7. Cole FE, Graci KA, Trippodo NC, MacPhee AA, Pegram BL, Frohlich ED. Dehydration attentuates the acute natriuretic response to rat atriopeptin III (rAPIII). Life Sci 1986;38:2015-8. Bell R], Wintour EM. The effects of maternal water deprivation on ovine fetal blood volume. Q] Exp Physiol 1985;70:95-9.