Patrick W. Concannon1, MS, PhD, DACT (Hon); John Verstegen2, DVM, PhD, DECAR
In recent years, advances in small animal reproduction have allowed pregnancy management to evolve into a clinical service that has application form the beginning to the end of gestations. The advances provide for new and improved methods of pregnancy detection; improved use of ultrasound to determine gestational age, assess fetal well and predict the date of parturition; the use of progesterone assays to monitor luteal function and allow consideration of progesterone supplementation; use of progesterone assays and managed breeding information in the evaluation of prolonged gestations and in the timing of elective of interventive caesarian sections; use of fetal and uterine monitoring in management of critical pregnancies late in gestation; use of ancillary and supportive therapy in caesarian and induced deliveries; the potential to use anti-progestin therapy to induce pre-scheduled whelping in normal pregnancies; the availability of multiple protocols for termination of unwanted pregnancies. In addition there have been recognized pregnancy management concerns that need to be considered prior to the detection to the detection of pregnancy including discontinuation of organophosphate parasiticides, confirming treatment of heartworm, determination of distemper and parvovirus titers, and likewise herpes virus antibody titers and herpes vaccination schedule (Eurican Herpes 205©, Merial) where applicable and available, as well as recording of periovulatory parameters useful for timing subsequent examinations and interpreting their results.
Several studies have confirmed that the duration of gestations in nearly all bitches is consistently 64-66 days when measured as the interval from the day of the LH surge to the day of spontaneous parturition. Further-more, in vivo and ex vivo evaluations of embryonic development have established a rather precise timeline of the early events of pregnancy and of expected change sin ultrasonic and radiographic appearance. Some landmark events and findings include: LH peak, day 0; ovulation, day 2; implantation, day 21; U/S detectable fetal heart, day 24; elevated relaxin day 23-28; elevated prolactin, day 28-30; fetal crown-rump length equals placental width, day 39-40; x-ray detectable fetal skull, day 44-48 vs. teeth, day 58-62; parturition, day 64-66.
Understanding the period of fertility in dogs is important in managing bitches presented as overdue and requiring intervention, as well as evaluating bitches claimed to be whelping prematurely. Because the canine sperm can remain viable in the female tract for up to 9 days and because the mature oocytes can survive for several days before fertilization, the apparent gestation length, form an observed breeding to normal parturition can range from 57 to 69 days; it can also be as long as 72 or more days from the first of multiple matings70. In addition, in rare cases of natural insemination and in cases of intrauterine insemination in late estrus, the interval from a single service to whelping may be as short as 55 or 56 days.
Palpation of uterine swellings is possible by day 20 and easiest when they are of 3 cm diameter between days 28 and 32 and progressively more difficult after day 35 when individual swellings become less distinct from one another. The currently available assays for the pregnancy-specific placental hormone relaxin include Reprochek © ELISA quantitative assay and Witness Relaxin© immuno-migration qualitative slide test (both Synbiotics). Both are very accurate and definitive, since relaxin is not produced absent a pregnancy. The ELISA may detect relaxin as early as day 20-23 and the Witness test often detects pregnancy relaxin as early as day 26-30. False positives occur in cases of recent resorption or with retained placental tissue. False negatives as late as day 30 or 34 have been observed anecdotally, and negative results should be followed by U/S or re-assay one week later. U/S can detect pre-implantation vesicles by day 21 or earlier, embryonic masses by day 22-25, embryonic heart movement by day 24-27 using 5 or 7.5 MHz equipment. .
Ultrasound (U/S) exams can be very useful in pregnancy management. Its use to document pregnancy 3 weeks after the end of estrus (metestrus vaginal smear), or at day 26-28 after the estimated day of the LH surge can be a critical service. It can document pregnancies in bitches that might otherwise have been judged infertile. It can also provide confirmation of normal development and a reasonable estimate of the stage of pregnancy and estimated whelping date in bitches in which the day of the LH surge was not determined. Before and after the day-40 change in the fetal-placental ratio, morphological features and changes in dimensions such as chronic cavity diameter and bi-parietal skull diameter and ratio of fetal length to placental width observed by U/S can be used to estimate gestational age with significant accuracy using published values. Accuracy of fetal number estimation deceases with littler size and stage of pregnancy.
There are important physiological changes associate with pregnancy that must be appreciated as they can affect clinical test results. These include a progressive pregnancy anemia evident by day 35, progressing to hematocrits as low as 30-35% at term, and often not returning to normal until more than two months post-partum. There are also increases in serum concentrations of several positive acute phase proteins including c-reactive protein, fibrinogen and alpha 1-acid glycoprotein, all of which increase after implantation and peak in late gestation. There are also reports of transient or progressive increases in fibrin degradation product levels, euglobulin lysis time, plasminogen activity, and alpha 2-plasmin inhibitor activity. The increases in acute phase proteins is assumed to represent an inflammation-like responses to the insult caused by implantation and placentation. Transient anorexia for several days around day 28 is normal. Increased food must be supplied to account a 40 percent average increase in food consumption which adds a 20 to 55 percent increase in body weight. Calcium or Vit. D (or any additive) supplementation is contraindicated as they can complicate normal calcium regulation increase incidence peripartum hypocalcaemia. There can also be an increase in insulin resistance sufficient to affect therapy of diabetic animals and cause pre-diabetic bitches to become frankly diabetic. Surprisingly, the insulin resistance is also accompanied by a reduced counter-regulatory hormone response, and the diet must be properly balanced and exercise maintained.
PROLACTIN AND MAMMARY
Prolactin concentrations increase above those in non-pregnant cycles by day 30, and in pregnancy reaches values similar to those seen in lactation, and may surge even higher 1 day before parturition during the rapid decline in progesterone that occurs immediately prepartum. Increase prolactin is assumed to be the cause of the increased progesterone secretion that occurs during pregnancy, and is also the major stimulant for the increased mammy development and lactogenesis that occurs during the last 30 days of gestation. Prolactin is luteotropic in both the pregnant and non-pregnant bitch, and prolactin-suppressing doses of dopamine agonists can readily cause luteolysis and termination of pregnancy any time aft6er day 25-30 of gestation. Whether or not insufficient prolactin secretion may be involved in cases of insufficient mammary development and/or agalactia due to mammary underdevelopment has not been determined. Prolactin is required for the lactogenesis that occurs during gestation, while progesterone suppresses actual lactation. The decline in prolactin at parturition, with a continued elevation in prolactin, is the initiator of milk synthesis and secretion. Dopamine antagonists including domperidone, metoclopramide and sulpiride stimulate prolactin secretion and have been shown in some species to stimulate lactation, and metoclopramide (1 mg/kg bid) has been used in some clinics to treat mammary insufficient or agalactia in small animals although behavioral side effects occur after 5 days of treatment. Dopamine agonists inhibit lactation and can be used in treatment of milk engorgement mastitis and persistent unwanted lactation.
RESORPTIONS AND ABORTION
Multiple ultrasound studies have documented idiosyncratic embryonic and fetal losses that would have other wise gone undetected, including partial as well as full resumption of litters as late as day 35. Even after day 35, fetal losses with related vaginal discharges may go undetected in fastidious bitches that lick away or ingest discharged materials. Ultrasound studies have also indicated that resorption may occur in 10-15 % of pregnancies, and total resorption in 5-10% of pregnancies. In one study, many instances of resorption were preceded by embryonic development retarded in relation to the expected time course.
Progesterone levels required maintenance of pregnancy may vary; normal and progesterone-supplemented pregnancies have continued with low levels below 2 ng/ml for periods of sever days or weeks. The low levels are presumably simply the result of the increased extraction of progesterone from the plasma by the gravid uterus. In the bitch, progesterone is entirely of ovarian origin, and there is no placental contribution like in the cat. During pregnancy, progesterone secretion by the corpora lutea is enhanced over that in non-pregnant bitches mostly due to a pregnancy increase in prolactin secretion that begins by day 27-30 and persists throughout gestation. However, circulating concentrations are not greatly different from those of the non-pregnant bitch because the increased progesterone is diluted by an increase in plasma volume and is more rapidly metabolized and cleared from the circulation by the uterus, placenta, mammary, and liver.
Fecal progesterone is distinctly elevated in the second half of pregnancy compared to that in the non-pregnant bitch. Serum progesterone should be monitored every 1-2 weeks in bitches with a history of pregnancy failure or luteal insufficiency.
Serum concentrations in progesterone typically peak at 15 to 90 ng/ml between day 15 and 30, and slowly decline thereafter, and somewhat less rapidly than the decline in non-pregnant cycles in many cases. The average peak level is around 50 ng/ml, and between days 35 and 50 there is typically a slowly decline to below 20m ng/ml by day 50 and below 10 ng/ml by day 60, but the range and variation in levels observed is considerable. In experimental studies, and in some clinically observed cases, pregnancy could be anointed with progesterone near or below 2 ng/ml, and the excess observed during most normal pregnancies may represent a mechanism to ensure pregnancy maintenance in the majority of animals.
However, there have been reports of instances of suspected luteal failure, luteal insufficiency, or "hypoluteoidism" in which peripheral concentrations of progesterone were observed to be exceptionally low in bitches which were confirmed to have resorbed or aborted their litter. The extent to which the low progesterone was a cause versus a consequence of pregnancy failure has not been well established. Nevertheless, protocols have been suggested for progesterone supplementation in pregnant bitches in which progesterone fall below 5 ng/ml before day 55, or declines more rapidly than expected in mid-gestation. The application of such progesterone replacement in bitches which previously failed to become pregnant or which were documented to have resorbed litters has involved successfully pregnancies--but whether the "successes" have been the result of treatment, better breeding management, or unknown factors is not known. Some protocols for progesterone supplementation have involved administration of progesterone in amounts too low or too infrequent to result in a meaningful effect of circulating progesterone concentrations, and there have been no studies to determine the effects of specific supplementation protocols on circulating progesterone concentrations. Progesterone supplementation protocols have included the following: (1) progesterone in oil, i.m., 2 mg/kg, every 72 h. (2) altrenogest, daily, p.o., 0.088 mg/kg (0.2 cc/10 lb, using Regu-Mate©, Hoechst-Roussel). Medroxyprogesterone, megestrol and some other synthetics can cause masculinization of female fetuses. Recently, human products of micronized natural progesterone in capsules have been used, being given orally 1-2 times per day at doses of 5-10 mg/kg (i.e., 100 mg capsules in 10-20 kg dogs) and the efficacy monitored by progesterone assays. This has been successful in some clinics although it relies on owner compliance. Natural progesterone requires frequent administration but has he advantage that withdrawal can be done rapidly. Whether the successful pregnancies obtained in P-supplemented bitches have been incidental or the result of treatment is not known, and controlled study is lacking.
Any use of progesterone supplementation must be done with a plan to discontinue therapy in a manner that will allow for progesterone to decline below 2 ng/ml shortly before expected "term" and thus permit or promote normal whelping. Excessive exogenous progesterone or other progestin can compromise normal parturition and can result in dead puppies that must be recovered by C-section. Oral progestin therapy should be discontinued 2 days prior to the estimated day of parturition. Serial injections of synthetic preparations should be discontinued early enough for the progestin to decline to non-effect concentrations by the expected day of parturition.
Normal parturition occurs as a result of a rapid decline in progesterone from the 4-10 ng/ml observed in the preceding days to values below 2 ng/ml over a 12-24 hr period beginning 1-2 days prior to whelping. The decline in progesterone and associated luteolysis occur in response to an increase in prostaglandin to luteolytic levels, which in turn occur in response to maturation of the fetal pituitary-adrenal axes and elevation in fetal glucocorticoid concentrations. Monitoring and/or examining for the decline in progesterone can be an important tool in managing bitches with an apparent or presumed prolonged gestation, in evaluating bitches presented for dystocia, and in timing elective c-sections, especially in bitches in which the day of ovulation or day of the preovulatory LH surge was not estimated with accuracy. The prepartum decline in progesterone is presumably the cause of the acute rise in prolactin immediately prepartum. The decline in progesterone also disinhibits the suppressive effects of progesterone on lactation and thus initiates lactation. Temperature declines of 1°C or more during the 12-24 h prepartum, and is a parameter that can be monitored twice daily starting day 60 by owners who wish to be involved.
FETAL AND UTERINE MONITORING
U/S detection of fetal HR below 180, often accompanied by bowel movements, is indicative of fetal distress and can be the basis of intervention in near-term bitches. U/S can also reveal fetal overgrowth or growth retardation, and other abnormalities including hydrocephalus, fetal anasarca, herniation of the ventral abdominal wall. Such case can then benefit form intervention. Uterine contraction activity has also been monitored with commercial devices designed for that purpose.
C-SECTIONS AND ADJUNCTIVE THERAPY
In cases of pre-scheduled c-sections and when emergency surgery is required in bitches with obvious signs of dystocia at the expected whelping time, additional steps can be considered. Collection of a serum sample for progesterone analysis can be retrospectively informative. "Term" dystocia in the absence of a normal prepartum decline in progesterone is unlikely, but no studies have been performed to document progesterone in term bitches with dystocia. Whether small litters or other factors can result in a failure of the expected fetal-induced prepartum luteolysis bitches is not known. Singleton mummified fetuses can occur and result in prolonged gestation; whether there is a maintenance of elevated progesterone in such cases is not known. Some clinics administer single shock-management doses of a short-acting corticosteroid to induce lung surfactant and other maturational changes in the fetuses. Such corticoid administration when prescheduled is given 8-24 hr prior to elective c-sections in brachycephalic breeds, and bitches with over-grown fetuses or a history of dystocia or excessive early neonatal loss of pups; normal losses range from 10-20%. Bitches scheduled for c-section should be provided access to 100% oxygen gas before as well as during surgery to improve fetal surfactant and decrease neonatal anoxia; O2 should be provided whether using inhalation anesthesia or continuous i.v.-delivery of a short-acting anesthetic (propofol) and a rapidly reversible opioid agent (fentanyl). Respiratory depression is typically not a problem in pups, but naloxone can be used to reverse effects of residual opiate.
The availability of a progesterone antagonist as a veterinary drug marketed for termination of unwanted pregnancies has resulted in research and clinical application of the drug as a means to induced parturition. Single s.c. administration of aglepristone at 15 mg/kg, after day 58 of pregnancy, when followed 24 h later by oxytocin 0.15 IU/kg every 2 h until the end or parturition, resulted in onsets of normal parturition at 32± 4 hr of treatment, without complications and with normal fetal parameters post partum. The expulsion phases of labor extend over 4.5± 2 h. Repeated administration of the same dose may be sufficient without administration of any tocolytic agent but study is limited. Repeated lower doses were poorly effect alone. The values of any such therapy in normal pregnancy except for the possible convenience it might afford is not clear. It is difficult to recommend in light of anecdotal reports of such treatments yielding less timely or less successful results in some bitches with or without support of tocolytic doses of OT or PGF. However, the potential reliability, safety and benefits of such protocols in cases of prolonged gestation or prior dystocia, or in preparation for prescheduled caesarian sections merits consideration.
1. Concannon, P., and Verstegen, J. (1998) Pregnancy in Dogs and Cats. Chapter. In: Knobil, E., and Neill, J. (Eds), Encyclopedia of Reproduction, Academic Press, San Diego
2. Concannon P.W., England G., Verstegen J. and Linde-Forsberg C. (Eds.) Recent Advances in Small Animal Reproduction, International Veterinary Information Service, Ithaca NY (www.ivis.org), 2001; A1220.0401
3. Connolly CC, Aglione LN, Smith MS, Lacy DB, Moore MC (2004) Pregnancy Impairs the Counterregulatory Response to Insulin-Induced Hypoglycemia in the Dog. Am J Physiol Endocrinol Metab. 2004 (in press)
4. Kutzler MA, Yeager AE, Mohammed HO, Meyers-Wallen VN. Accuracy of canine parturition date prediction using fetal measurements obtained by ultrasonography. Theriogenology. 2003 ; 15;60(7):1309-17.