Intrauterine fetal demise can be remote from the inciting insult in an animal model of hypoxia-ischemia.

BACKGROUND: Fetal hypoxia-ischemia (H-I) results in significant morbidity and mortality. Little is known about the timing of death in human stillbirths. The vulnerability of the fetus varies with age at the time of insult, but it is unknown what happens to the timing of fetal death in relation to a fetal insult. We asked the question of whether the timing of fetal death was influenced by the age at which the insult occurred. METHODS: Fetal H-I was achieved at three ages by sustained uterine ischemia in rabbits, mimicking the acute placental insufficiency of placental abruption. RESULTS: H-I at 22 d gestation (E22) resulted in fewer perinatal deaths than at E25 and E29. Fetal deaths were grouped into early and late perinatal deaths. Early perinatal death mostly occurred immediately after H-I and these fetuses delivered before term. Late perinatal death occurred between the insult and delivery at term gestation. Early perinatal death occurred more often in the E25 hypoxic-ischemic group as compared with those of the E22 hypoxic-ischemic group. CONCLUSION: There is an increasing vulnerability to hypoxia with increasing gestational age. Perinatal deaths may occur long after the episode of H-I. The timing of an intrauterine hypoxic-ischemic event cannot be inferred from the detection of fetal death.

White matter injury correlates with hypertonia in an animal model of cerebral palsy.

Hypertonia and postural deficits are observed in cerebral palsy and similar abnormalities are observed in postnatal rabbits after antenatal hypoxia-ischemia. To explain why some kits become hypertonic, we hypothesized that white matter injury was responsible for the hypertonia. We compared newborn kits at postnatal day 1 (P1) with and without hypertonia after in vivo global fetal hypoxia-ischemia in pregnant rabbits at 70% gestation. The aim was to examine white matter injury by diffusion tensor magnetic resonance imaging indices, including fractional anisotropy (FA). At P1, FA and area of white matter were significantly lower in corpus callosum, internal capsule, and corona radiata of the hypertonic kits (n=32) than that of controls (n=19) while nonhypertonic kits (n=20) were not different from controls. The decrease in FA correlated with decrease in area only in hypertonia. A threshold of FA combined with area identified only hypertonic kits. A reduction in volume and loss of phosphorylated neurofilaments in corpus callosum and internal capsule were observed on immunostaining. Concomitant hypertonia with ventriculomegaly resulted in a further decrease of FA from P1 to P5 while those without ventriculomegaly had a similar increase of FA as controls. Thus, hypertonia is associated with white matter injury, and a population of hypertonia can be identified by magnetic resonance imaging variables. The white matter injury manifests as a decrease in the number and density of fiber tracts causing the decrease in FA and volume. Furthermore, the dynamic response of FA may be a good indicator of the plasticity and repair of the postnatal developing brain.

Model of cerebral palsy in the perinatal rabbit.

Perinatal brain injury results in one of the highest burdens of disease in view of the lifelong consequences and is of enormous cost to society. This makes it imperative to develop better animal models that mimic the human condition. Many neurodevelopmental deficits, such as cerebral palsy, are believed to be a result of prenatal hypoxia-ischemia in humans. Fetal global hypoxia-ischemia is most commonly a consequence of acute placental insufficiency. Our laboratory has modeled in utero sustained and repetitive hypoxia-ischemia in the pregnant rabbit to mimic the insults of abruptio placenta and labor, respectively. Sustained hypoxia-ischemia at 70% (22 days' gestation) and 79% (25 days' gestation) and repetitive hypoxia-ischemia at 90% gestation (28 days' gestation) caused stillbirths and multiple deficits in the postnatal survivors. The deficits included impairment in multiple tests of spontaneous locomotion, reflex motor activity, motor responses to olfactory stimuli, and the coordination of suck and swallow. Hypertonia was observed in the 22 and 25 days' gestation survivors but not in the 28 days' gestation group. Hypertonic survivors were artificially fed and found to have the motor deficits persist for at least 11 postnatal days. A spectrum of brain abnormalities is found on magnetic resonance imaging. This is the first animal model to mimic cerebral palsy. The findings also suggest a window of vulnerability during brain development when the injury results in hypertonia in newborn pups.

Sex hormone binding globulin facilitates female sexual receptivity except when coupled to dihydrotestosterone.

Sex hormone binding globulin (SHBG) is produced in brain where it is often co-localized with oxytocin. Infusions of SHBG into the medial preoptic area-anterior hypothalamus facilitate female sexual receptivity. SHBG has receptors on plasma membranes of the prostate gland where binding of the 5alpha-reduced androgen dihydrotestosterone (DHT) by SHBG acts as an antagonist on SHBG receptors. This study attempted to determine whether pre-coupling DHT to SHBG would inhibit SHBG-induced facilitation of female sexual receptivity. Ovariectomized rats were injected daily with 0.75 microg estradiol benzoate for 3 days. On the fourth day after a pre-infusion baseline behavioral test animals were infused with 1 microl per side through bilateral cannulae with SHBG (1.77x10(-6) M), SHBG coupled to DHT (SHBG-DHT; 1.66x10(-6) M DHT), with DHT alone or with artificial cerebrospinal fluid vehicle. As before, SHBG significantly increased female sexual receptivity when infused into the medial preoptic area-anterior hypothalamus. Rats infused with SHBG-DHT had significantly lower sexual receptivity. Therefore, whereas SHBG in the medial preoptic area facilitated female sexual behavior, SHBG coupled to DHT did not. DHT itself did not significantly affect sexual receptivity. Pre-coupling DHT to SHBG eliminated the facilitative effect of SHBG on female sexual receptivity just as DHT inhibits SHBG activity at prostate SHBG receptors suggesting that central receptors for SHBG are similar to those demonstrated in the periphery.

5alpha-Reduced androgens block estradiol-BSA-stimulated release of oxytocin.

In this study we test the postulate that estradiol conjugated to bovine serum albumin (E-BSA) acts via receptors for the steroid-binding protein sex hormone binding globulin (SHBG) by attempting to block E-BSA-stimulated release of oxytocin with two antagonists of SHBG receptor actions: the 5alpha-reduced androgens dihydrotestosterone (DHT) and 3alpha-diol. Simultaneous superfusion with either DHT or 3alpha-diol significantly blocked E-BSA-stimulated release of oxytocin. We also found that a wide range of free 17beta-estradiol was unable to stimulate oxytocin release, suggesting that E-BSA stimulates receptors other than those for free estradiol to release oxytocin, perhaps SHBG receptors.

Distribution of androgen-binding protein in the rat hypothalamo-neurohypophyseal system, co-localization with oxytocin.

Androgen-binding protein (ABP) is known to be expressed in the male and female rat hypothalamus. In the present study, we observed immunocytochemically ABP in neurons of the magnocellular hypothalamic nuclei, in the preoptic region and in the lateral hypothalamus. Dense fiber networks with varicosities, containing ABP immunofluorescence, were visible throughout the hypothalamus, the median eminence and in the posterior pituitary lobe. Double immunostaining revealed a partial coexistence of ABP-and oxytocin immunoreactivity in a portion of the magnocellular perikarya. ABP was isolated by affinity chromatography from hypothalamus homogenates. Western blots resulted in immunoreactive (IR) bands with an approximate molecular weight of 35 and 50 kDa. Mass spectrometry of these preparations confirmed the presence of ABP, which was almost identical to ABP isolated from rat testis. It is likely that ABP, expressed in magnocellular oxytocinergic neurons, is subject to axonal transport and release in the hypothalamo-neurohypophyseal system.

Fetal brain magnetic resonance imaging response acutely to hypoxia-ischemia predicts postnatal outcome.

OBJECTIVE: Cerebral palsy (CP) is caused by either hypoxia-ischemia (H-I) or long-standing causative factors such as inflammation or genetics. Multiple pathophysiological events over time are thought to contribute eventually to cerebral palsy. Our objective was to examine whether the immediate response of the fetus to an acute H-I event determined the motor deficits associated with cerebral palsy. METHODS: Serial diffusion-weighted imaging were performed on 79% gestation New Zealand white rabbits using a 3-Tesla magnetic resonance scanner during 40 minutes of uterine ischemia, 20 minutes of reperfusion, and at 4, 24, and 72 hours. Individual fetuses were tracked to near term, and the delivered kits were divided into hypertonic H-I (n = 18), nonhypertonic H-I (n = 9), stillbirth H-I (n = 4), and control groups (n = 16). RESULTS: The hypertonia group had significantly less of a nadir in apparent diffusion coefficient (ADC) during H-I (71.6 +/- 23.8% vs 84.5 +/- 9.3% baseline) and slower and incomplete recovery of ADC during reperfusion compared with the nonhypertonic group. All fetuses in the hypertonic and stillbirth groups had an ADC nadir of less than 0.83 microm(2)/msec (70.3% decrease from baseline), whereas 94% of control animals had an ADC nadir greater than this value. The difference between outcome groups was the largest at 4 hours reperfusion and persisted for 24 hours. INTERPRETATION: Serial fetal brain scans indicate that the immediate response of a fetus to H-I is crucial to the development of hypertonia. If the fetal brain can be scanned at the time of insult, ADC changes can predict which fetuses will have an unfavorable outcome.

Sensory deficits and olfactory system injury detected by novel application of MEMRI in newborn rabbit after antenatal hypoxia-ischemia.

Sensory deficits are frequently observed in cerebral palsy patients. The motor response to smell was found to be abnormal in an animal model of cerebral palsy following fetal hypoxia-ischemia. We hypothesized that fetal hypoxia-ischemia causes long-lasting and selective olfactory tract injury. A population of newborn rabbits with motor deficits was selected after spontaneous delivery following uterine ischemia at 22 days gestation (E22, 70% term). MnCl(2), 20 mg/kg, was administered in both nostrils at postnatal day 1 (E32). One nostril was occluded to control for smell augmentation through the other open nostril by intermittent amyl acetate stimulation for 6 h. T1-weighted MRI images were obtained on newborn rabbits. Amyl acetate exposure increased augmentation of Mn(2+) uptake in olfactory epithelium on the open side in control group but the augmentation was decreased after hypoxia. The proportion of animals with a greater enhancement in the open side increased in controls after amyl acetate, but not in hypoxia. Mn(2+) took longer to arrive at the olfactory bulbs and the rate of subsequent increase was slower in hypoxia. Concomitantly, the thickness of olfactory epithelium and the number of mature olfactory neurons, detected on olfactory marker protein immunostaining, were significantly less in the hypoxic group. Functional MRI studies are superior to neurobehavioral smell testing in the rabbit kits as they are more sensitive and quantifiable measures and do not depend upon the motor response. Antenatal hypoxia-ischemia causes long-lasting injury to neuronal tracts of the olfactory system including olfactory epithelium.