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Before the preganglionic regulation of the adrenal medulla is established, hypoxia acts directly on the chromaffin cells to evoke the secretion of catecholamines. This direct action of hypoxia is suppressed by the gradual development of the preganglionic innervation and we have proposed that opioid peptides released from the adrenal splanchnic nerves may be responsible for this suppression. The effects of the specific opioid agonists DPDPE (δ-agonist), U-62066 (κ-agonist) and DALDA (μ-agonist) on the hypoxia-evoked response were investigated in both a whole-gland preparation and in isolated adrenal chromaffin cells using amperometry, whole-cell patch clamping and measurement of cytosolic [Ca2+]. The combined application of μ- and κ-type agonists abolished the hypoxia-evoked catecholamine secretion from whole perfused adrenal gland. In isolated chromaffin cells, μ- and κ-opioid agonists reduced the rise in [Ca2+]i that results from exposure to hypoxia. Both agonists decreased the voltage-dependent Ca2+ current in these cells. The μ-agonist increased the conductance through SK-type K+ channels and this action offset the decrease in K+ conductance produced by exposure to hypoxia. The κ-type agonist decreased the conductance through an action on BK-type K+ channels, a class of channels that are not involved in initiating the direct response to hypoxia. These data suggest that opioids, through their action on SK channels and voltage-dependent Ca2+ channels, may be responsible for the nerve-induced suppression of the hypoxic response of adrenal chromaffin cells and that these effects of endogenous opioids are mediated via μ- and κ-type receptors.

A range of pathophysiological factors can result in a perturbation or restriction of fetal growth, and the cardiovascular, neuroendocrine and metabolic adaptations of the fetus to these stimuli will depend on their nature, timing and intensity. The critical importance of these physiological adaptations for both immediate survival and long-term health outcomes has provided an impetus for experimental studies of the nature and consequences of specific fetal adaptations to a poor intrauterine environment. This review summarizes data from recent studies that have focused on the responses of the fetal cardiovascular, sympathoadrenal, hypothalamo-pituitary-adrenal and renin-angiotensin systems to experimental restriction of placental function in the sheep and discusses the consequences of these adaptations for fetal, neonatal and adult health.

We have investigated the effects of fetal growth restriction, induced by restriction of placental growth and function (PR), on 11beta-hydroxysteroid dehydrogenase type 1 (11betaHSD-1) and 11betaHSD-2 messenger RNA (mRNA) expression in fetal tissues in the sheep, using Northern blot analysis. Fetal liver, kidney, and adrenals were collected from normally grown fetuses at 90 days (n = 6), 125 days (n = 6), and 141-145 days (n = 7) and from PR fetuses at 141-145 days (n = 6). Expression of 11betaHSD-1 mRNA in the fetal liver increased significantly between 125 days (7.4+/-0.8) and 141-145 days gestation (27+/-5.3). There was also an approximately 2-fold increase in the ratio of 11betaHSD-1 mRNA/18S rRNA expression in the PR group (53.8+/-7.9) compared with that in control animals at 141-145 days gestation. There was a significant decrease in 11betaHSD-2 mRNA in fetal adrenals between 125 days (41.6+/-2.4) and 141-145 days (26.7+/-1.1) gestation, but there was no effect of PR on the expression of adrenal 11betaHSD-2 mRNA. 11betaHSD-2 mRNA expression in the fetal kidney increased between 90 days (16.8+/-1.7) and 141-145 days gestation (31.7+/-4.3), but there was no effect of PR on the levels of 11betaHSD-2 mRNA in the fetal kidney. In summary, 11betaHSD-2 mRNA is differentially regulated in the fetal adrenal and kidney in the sheep fetus during late gestation. There is also a specific increase in the expression of 11betaHSD-1 mRNA in the liver of growth-restricted fetuses in late gestation. This suggests that there is increased hepatic exposure to cortisol in the growth-restricted fetus, which may be important in the reprogramming of hepatic physiology that occurs after growth restriction in utero.

This study examined the impact of a chronic physiological elevation of plasma cortisol levels on adrenal catecholamine synthetic enzyme and proenkephalin A mRNA expression in foetal sheep. Cortisol (2.5-3. 0 mg.5 ml-1.24 h-1, n=9) or saline (0.9% saline, n=6) was infused into foetal sheep for 7 days between 109 days and 116 days gestation. Foetal plasma cortisol concentrations were higher (P

1. The ability of the fetal adrenal medulla to respond directly to hypoxaemia and secrete catecholamines before the development of a functional innervation of the gland is essential for intrauterine survival. The cellular mechanisms involved in this response to low PO2 are not known, although the presence of oxygen-sensitive K+ channels in carotid body chemoreceptor cells and other sites suggests that these might underlie the chromaffin cell response. 2. Whole-cell patch-clamp techniques have been used to study K+ currents during normoxia and hypoxia in chromaffin cells isolated from the adrenal glands of fetal sheep. 3. Two types of chromaffin cells were observed, those with a fast inactivating K+ current and a larger capacitance and those with a delayed K+ current and smaller capacitance. No cell showed both types of current. The fast inactivating current showed voltage-dependent inactivation and was blocked by 1 mM 4-aminopyridine, characteristics of an IA-type current. The delayed current had two components, a TEA-sensitive, Ca2+-dependent current and a component with the kinetic behaviour of a delayed rectifier. 4. Both types of current were oxygen sensitive. The IA-type current was reduced by 27.4 +/- 3.2 % when the PO2 was reduced to about 15 mmHg. With the delayed current, hypoxia reduced the amplitude by 26.9 +/- 2.4 %, largely by reduction of the Ca2+-dependent component. 5. In the presence of hypoxia, reduction in the amplitude of these oxygen-sensitive K+ currents would increase the frequency and duration of action potentials, leading to increased activation of the L-type Ca2+ channels, influx of Ca2+ and the subsequent secretion of catecholamines.

We have investigated the cellular mechanisms underlying the catecholamine response of the fetal sheep adrenal to hypoxia before and after the development of adrenal innervation. Adrenals were collected before (80–100 days gestation: n = 7) and after (135–146 days gestation: n = 10) development of innervation and retrogradely perfused with oxygenated Krebs bicarbonate buffer in vitro via the renal vein. Adrenal hypoxia was induced by perfusion with hypoxic Krebs buffer (pO2 = 46.7 ± 2.4 mm Hg) for 30 min periods in the presence and absence of hexamethonium (500 μM), Ca2+ (2.5 mM), nifedipine (1 μM) and KCl (10 mM). Hypoxia stimulated an increase (P

We have investigated the effect of lowering foetal arterial PO 2 either acutely or chronically on tyrosine hydroxylase (TH) protein content in the dorsal and ventral medullary regions of the brainstem of the sheep foetus during late gestation. TH protein content increased in both the dorsal and ventral medullary regions of the foetal brainstem after exposure to acute hypoxia when compared to normoxia. In contrast there was no increase in the TH protein content of either the dorsal or ventral medullary regions in the brainstem of foetal sheep which were chronically hypoxaemic throughout late gestation as a consequence of experimental restriction of placental growth. The differences between the TH responses to acute and chronic hypoxaemia in the foetal sheep brainstem may be important in the mediation of physiological adaptations to these intrauterine stimuli and for the generation of an appropriate physiological response to hypoxia in the newborn period.

We have investigated adrenal mRNA expression of the catecholamine synthetic enzymes tyrosine hydroxylase (TH) and phenylethanolamine N-methyltransferase (PNMT) following acute hypoxia in fetal sheep before (105 days gestation, n = 20) and after (125 days gestation, n = 20) the development of adrenal innervation and following pretreatment with the nicotinic receptor anatgonist hexamethonium (n = 12). Total RNA was extracted from fetal adrenal glands collected at specific time points at 3-20 h after the onset of either hypoxia ( approximately 50% reduction in fetal arterial oxygen saturation (SO2) for 30 min), or normoxia. Before 105 days, there was a decrease in adrenal TH mRNA expression at 20 h after hypoxia and adrenal TH mRNA expression was directly related to the changes in arterial PO2 measured during normoxia and hypoxia. After 125 days, adrenal TH mRNA levels were suppressed for up to 12 h following hypoxia. In both age groups, adrenal PNMT mRNA expression increased at 3-5 h after hypoxia and was inversely related to the changes in fetal arterial PO2 during normoxia or hypoxia. After 125 days, the administration of hexamethonium (25 mg kg(-1), I.V.) reduced TH mRNA but not PNMT mRNA expression after normoxia. After hexamethonium pretreatment, there was no significant change in either adrenal TH or PNMT mRNA expression following hypoxia. We conclude that acute hypoxia differentially regulates adrenal TH and PNMT mRNA expression in the fetal sheep both before and after the development of adrenal innervation. After the development of adrenal innervation, however, the effect of acute hypoxia upon adrenal TH and PNMT mRNA expression is dependent upon neurogenic input acting via nicotinic receptors.

We have demonstrated that there are differential changes in the levels of tyrosine hydroxylase (TH), phenylethanolamine N-methyltransferase (PNMT), and proenkephalin A (Pro Enk A) mRNA in the fetal sheep adrenal during late gestation. Adrenal TH mRNA:18S rRNA ratios increased between gestational days 100 (0.98 +/- 0.13; n = 6) and 125 (1.40 +/- 0.15; n = 6) and then decreased, whereas adrenal PNMT mRNA:18S rRNA ratios increased regularly between gestational days 100 (0.08 +/- 0.01) and 146 (0.17 +/- 0.03). The ratio of adrenal Pro Enk A mRNA to 18S rRNA was higher at gestational day 125 (0.085 +/- 0.005) than at either 80-100 days (0.038 +/- 0.007) or 140-146 days of gestation (0.055 +/- 0.013). In 12 ewes, the growth and development of the placenta were restricted (placental restriction group) from conception. The ratio of adrenal PNMT mRNA to 18S rRNA was significantly reduced in the placental restriction group of fetal sheep (0.003 +/- 0.002) compared with controls (0.011 +/- 0.002), and there was a significant correlation between the ratio of adrenal PNMT mRNA to 18S rRNA and the mean arterial PO2 (r = 0.88, p < 0.0005). In contrast, TH mRNA and Pro Enk mRNA were unaffected by placental restriction. Adrenaline and noradrenaline syntheses are therefore differentially regulated in the adrenal during late gestation and in response to chronic intrauterine hypoxemia.