Renal prostaglandin E2 synthesis and degradation in the developing rat

Postnatal development of glomerular filtration rate (GFR) and renal blood flow is associated with a fall in renal vascular resistance that may be mediated by vasoactive substances. We examined differences in the regulation of one such substance, prostaglandin E2 (PGE2). The present studies examined renal cortical and medullary PGE2 synthesis and degradation in rats aged 20 days (30.7 g), 31 days (101 g), and 120 days (413 g). PGE2 synthesis in cortical microsomes was highest in 20-day-old rats compared to 31- and 120-day-old rats. In contrast, medullary PGE2 synthesis was lowest in 20-day-old rats compared to 31- and 120-day-old rats. Both cortical and medullary PGE2 degradation were highest in 20-day-old rats and decreased with age. Despite demonstrating significant age-dependent differences in cortical and medullary PGE2 synthesis, 11 days of aspirin given between age 20-31 days blocked PGE2 synthesis in cortex and medulla by 60 and 76%, respectively, but GFR was similar to control 31-day-old rats (0.78 +/- 0.04 ml/min/g kidney weight, aspirin-treated, versus 0.85 +/- 0.03 ml/min/g kidney weight, control), suggesting that observed age-dependent differences in renal PGE2 synthesis is not a major determinant of development of GFR. A more important determinant of GFR may be age- related differences in renal cortical prostaglandin turnover.     

Age-related variations in jejunum and distal colon contractile response to thyrotropin-releasing hormone in vitro

The effects of thyrotropin-releasing hormone (TRH) on the contractile activity of the proximal jejunum and distal colon of 3-, 7-, 14-, 21-, 28-, 50-day-old and adult male Sprague-Dawley rats were examined. Longitudinal segments were mounted in force displacement transducers and bathed in oxygenated, buffered Krebs' solution. In the proximal jejunum TRH 10(-6) M produced a tetrodotoxin (TTX)-sensitive tension increase in 3- to 14-day-old rats and a TTX-resistant biphasic response in older rats. TRH 10(-6) M produced a transient tension in the distal colon of rats less than or equal to 21 days old. Incubation with TRH 10(-6) M for 30 min increased responsiveness to acetylcholine 10(-7) M in approximately 50% of tissues studied. Direct and indirect effects of TRH are developmentally determined.     

Hepatic metabolism of orally administered prostaglandin F2 alpha in suckling and weanling rats

Suckling (12- to 14-day-old) and weanling (30-day-old) rats were sacrificed 2 h after oral administration of 3H-labeled prostaglandin F2 alpha. Although radioactivity recovered from the stomach and small intestine (including contents) was slightly higher in sucklings (28.3 +/- 3.7%; n = 10) than in weanling rats (21.3 +/- 5.3%; n = 7), the liver of sucklings contained significantly higher amounts of counts (11.0 +/- 1.1 vs. 3.3 +/- 0.5%). Combined column and thin-layer chromatography of liver extracts showed more authentic prostaglandin F2 alpha in sucklings (11.0 +/- 0.5% of the liver counts) than in weanlings (7.0 +/- 1.1%). The liver of suckling rats contained a higher percentage of more polar metabolites (43.3 +/- 1.6 vs. 34.3 +/- 3.0%). These studies demonstrate differences in processing of oral prostaglandin F2 alpha in the early postnatal period.     

Isolated pancreatic acini from suckling and weanling rats: changes in amino acid incorporation and carbachol-stimulated amylase secretion with age

To characterize the changes in pancreatic function during postnatal development, isolated pancreatic acini were prepared from rats aged 8-9, 12-14 and 20 days and from adult rats. Isolated acini maintained a normal microscopic appearance and viability as judged by exclusion of trypan blue and linear incorporation of tritiated leucine into total protein. The rate of incorporation in 8-day-old acini was 20% of that observed in adult rats. Significant dose-dependent increases in amylase release in response to carbachol were observed in all age groups; stimulated amylase secretion was significantly less in the 8- to 9- and 12- to 14-day-old animals than in the 20-day-old and adult rats. These data indicate that viable isolated pancreatic acini can be prepared from suckling rats and that these acini exhibit an altered in vitro responsiveness to carbachol. This preparation should therefore be a useful model for in vitro studies of the development of pancreatic function.     

Significance of the biphasic developmental pattern of the hypothalamo-pituitary-adrenocortical system in the rat

One group of neonatal rats was injected with metopirone at 2-5 days and the second group at 7-10 days of age. At 14 and 21 days all rats treated with metopirone had reduced body and adrenal weights in comparison with their saline-treated littermates. At 21 days the smallest adrenals were in the rats treated with metopirone at 7-10 days. Metopirone increased resting and stress levels of adrenal and plasma corticosterone in the 14-day-old rats and the response to stress by the adrenal corticosterone increase in the 21-day-old rats. The differential effect of metopirone applied in two developmental periods was indicated.     

Cyclooxygenase-2 activity following traumatic brain injury in the developing rat

Cyclooxygenase (COX) is the rate-limiting enzyme in the production of prostaglandins. COX-2, the predominant COX isoform in brain, is induced by synaptic activity. COX-2-generated prostaglandins are important regulators for a range of activities under physiologic conditions. However, under pathologic conditions, COX-2 activity can produce reactive oxygen species and toxic prostaglandin metabolites that can exacerbate brain injury. In this study, we examine the developmental production of COX-2 and test the ability of a COX-2 inhibitor, SC58125, to attenuate traumatic brain injury in developing rats. We show that constitutive COX-2 concentration is low (0.5-fold adult concentration) during the first postnatal week and then increases to 3-fold of adult levels between days 14-60. Controlled cortical impact (CCI) at postnatal day (PND) 17, but not PND 7, caused an additional 3-fold increase in COX-2 content and was associated with an increase in the COX-2 product PGE2. Treatment with the COX-2 inhibitor SC58125 in PND17 rats exposed to CCI attenuated the rise in PGE2 but did not attenuate lesion volume or improve performance in the Morris water maze.     

Milk-stimulated PGE2 production by isolated gastric cells: a possible role in the inhibition of histamine-induced acid secretion.

The effects of a milk diet on gastric acid secretion of rats fed raw bovine milk for 4 days were examined using dispersed gastric cells. Parietal cell acid secretion was estimated by the accumulation of 14C-aminopyrine (AP), an index of secretory function. Basal AP accumulation was significantly increased (60%) by the milk diet. There was a marked upward shift in the dose-response curve of histamine (HA; 10(-8) to 10(-3) M) in milk-fed rats, indicating enhanced sensitivity of parietal cell-H2 receptor to exogenous HA. In contrast, the dose-dependent inhibition of HA-induced AP accumulation by prostaglandin (PG) E2 was significantly reduced, indicating that the parietal cells of milk-fed rats were less sensitive to exogenous PGE2. The PGE2 content of bovine milk was low (less than 20 pg/ml), but the production of endogenous PGE2 by the gastric cells was dramatically increased by the milk diet and exhibited maximal control production rate in the presence of 10 microM arachidonic acid. The increased responsiveness to histamine and the decreased responsiveness to PGE2 indicated that the milk diet induced low histamine and high PGE2 availability in the vicinity of the parietal cell basolateral membrane. This regulation, which involves stimulation of PGE2 production in the gastric mucosa, may underly the inhibition of acid secretion observed in vivo in chronically milk-fed adult rats.     

Neonatal ovine adrenal cortical and medullary Cell H+ responses to ACTH and prostaglandin E(2)

The microphysiometric technique was used to evaluate 1- and 3-day-old lamb adrenal gland H+ production in response to ACTH or PGE(2). ACTH stimulated cortical, but not medullary H+ production; maximal H+ by combined cortical and medullary cells was greater than by cortical cells alone (p < 0.05). In contrast, the magnitude of the H+ response to PGE(2) was greater with medullary than with cortical cells alone or with cortical and medullary cells combined (p < 0.05). The H+ response to ACTH was greater in 3-day-old compared to 1-day-old lambs while the potency of PGE(2) was not different at the ages studied. We conclude that in neonatal sheep: (1) ACTH and PGE(2) are potentially important in adrenal regulation, and (2) paracrine communication appears to be functioning as has been shown for adults.     

Role of thyrotropin-releasing hormone in thyroid-stimulating hormone and growth hormone regulation during postnatal maturation in female Wistar rats

The role of endogenous thyrotropin-releasing hormone (TRH) in the control of pituitary thyroid-stimulating hormone (TSH) and growth hormone (GH) secretion was studied during postnatal maturation in female Wistar rats. Half of the sucklings in each litter was treated intraperitoneally with either specific rabbit antiserum against TRH or normal rabbit serum (0.1-0.3 ml according to age). All animals were decapitated after 2 h. The presence of anti-TRH activity was checked as a binding of labelled TRH with plasma of the experimental animals. Immunoneutralization of endogenous TRH resulted in a decrease of plasma TSH in 3- to 15-day-old female pups as compared to control littermates. No effect of TRH antibody injection was seen at the ages of 1, 21, 30 and 70 days despite the presence of excess antibody in the plasma. A profound effect of TRH antibody on plasma TSH was seen again at the age of 100 days. Plasma GH in the same animals exhibited a paradoxical increase after TRH immunoneutralization at the age of 5 and 8 days, a decrease was found at the age of 21 days. It was concluded that hypothalamic TRH control of TSH secretion matures early in Wistar rats. Hypothalamic secretion of TRH at the ages of 1, 21, 30, and 70 days is low and(or) its role in TSH regulation is masked by other regulating factors. TRH may play a dual role in the regulation of GH secretion during the postnatal period.     

Prostaglandin-mediated effects on growth and markers of biochemical development in the rat

Several markers of growth and biochemical development in the rat were studied after administration of prostacyclin (PGI2) and 16, 16-dimethyl prostaglandin E2 (16, 16DM PGE2). Intermittent administration of PGI2 for 3 days to 10- and 19-day-old animals, with subsequent sacrifice at 14 and 23 days, resulted in significant dose related decreases in growth at 23 days. Total sucrase and maltase (glucoamylase) activities were elevated compared to controls at 14 days. Total activities of these enzymes were decreased in postweaned 23-day-old animals, but specific activities per mg intestinal protein were not significantly different. 16, 16DM PGE2 administered continuously between day 10-16 of life caused alterations in growth as well as increases in sucrase and maltase (glucoamylase) activities. Exogenously administered prostaglandins, therefore, are associated with altered growth and markers of biochemical development in the rat.     

The immature rat small intestine exhibits an increased sensitivity and response to Escherichia coli heat-stable enterotoxin

Escherichia coli which elaborate heat stable enterotoxin (ST) are a major cause of endemic diarrhea in infants. The reason(s) for this increased susceptibility of infants to ST-mediated diarrhea is unknown. We investigated the possibility that the immature (14 and 21 day old) rat small intestine is more sensitive to ST than is the adult. Initially we found there was a 600-fold increased jejunal sensitivity to ST in the immature animals as measured by dose required for half maximal secretion. Also there was a greater jejunal secretory response in the immature animals (14 greater than or equal to 21 days old greater than adult). To determine the cause for this increased sensitivity and secretory response to ST, we examined: 1) binding characteristics of 125I-ST to brush border membrane (BBM) receptors and 2) membrane bound guanylate cyclase activation by ST in both immature and adult rats. Our findings demonstrate that more ST receptors are present in jejunal BBM from 14- and 21-day-old rats than in jejunal BBM from adult rats (2.34 +/- 0.18, 2.85 +/- 0.82, and 0.79 +/- 0.13 X 10(12) receptors/mg BBM protein, respectively), while the affinity of the BBM receptor for ST is similar at all three ages in both jejunum and ileum. Furthermore, both the jejunum and ileum of the rats of all three ages revealed an equal sensitivity of guanylate cyclase to activation by ST. These findings suggest that the increased number of jejunal receptors in the immature rat may, in part, explain the increased sensitivity and secretory response observed in vivo.     

Development of acid secretory function in the rat stomach: sensitivity to secretagogues and corticosterone

Gastric acid secretion was studied in anesthetized rats from day 6 of the postnatal period up to the time of weaning. Basal H+ secretion was detected from day 6 in the first group studied (2.4 +/- 0.2 muEq of H+/10 min/100 g of body weight, BW) and remained constant up to the time of weaning (day 18: 2.5 +/- 0.2 muEq of H+/10 min/100 g of body weight) except for the period between days 10 and 12, when it fell significantly (1.5 +/- 0.06 muEq H+/10 min/100 g of BW on day 12). Both histamine H2 receptor sensitivity and intracellular transduction mechanism activities were evaluated by studying the secretory responses to histamine, impromidine (an H2 receptor agonist), cimetidine (an H2 receptor antagonist), forskolin (a direct adenylate cyclase activator), and dibutyryl (db) cAMP (an analogue of cAMP, the intracellular messenger mediating the response to histamine). The effects of pentagastrin and carbachol were also determined. The secretory responses obtained on days 6, 8, and 18 were similar and represented about threefold increases over basal secretion for all the secretagogues used. After weaning on day 20, both the basal secretion and the response to secretagogues were significantly increased compared with those of unweaned animals. On day 12, the responses were always weaker than on both days 8 and 18. Injection of 1 mg/kg of corticosterone 21 acetate daily from day 8 resulted on day 12 in a basal secretion and a response to histamine equivalent to those measured in 18-day-old pups not injected.(ABSTRACT TRUNCATED AT 250 WORDS)     

Protein synthesis by gastric mucosal ribosomes during development in rats

Oxyntic gland mucosal ribosomes from 7- to 30-day-old rats were assayed for their ability to synthesize endogenous messenger RNA (mRNA) and poly(U)-directed protein in a cell-free system. Normal rat liver cell sap (105,000 g supernatant) was used as a source for transfer RNA (tRNA) and activating enzymes. Total mucosal DNA and ribosome content [as assessed by ribosomal RNA (rRNA)] were also determined. Although both DNA and rRNA content increased throughout the 4 postnatal weeks, the rRNA/DNA ratio rose sharply during the 2nd and 4th week after birth. The ability of mucosal polyribosomes to synthesize endogenous mRNA-directed protein in a cell-free system remained elevated up to the age of 18 days, then decreased markedly within the next 3 days, and thereafter, declined more slowly. In 21- and 30-day-old rats, the rate of endogenous mRNA-directed protein synthesis by mucosal ribosomes was 50-60% below that of the 7-day-old suckling rats. The protein/polyphenylalanine ratio, which represents a ratio of polysomes to monosomes, was found to be about 57% lower for ribosomes from 30-day-old rats as compared to 7-day-old suckling animals. The ability of run-off ribosomes (devoid of endogenous mRNA) to translate poly(U) was also found to be 33% lower for 30-day-old rats as compared to the 7-day-old suckling animals.     

Adenylate cyclase activity in fetal rabbit myocardium

Adenylate cyclase activity [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] was determined in vitro in fetal rabbit myocardial membranes from individual fetal pups at 21 to 31 days gestation (term, 31 days). Basal and NaF-stimulated adenylate cyclase activities did not change during 21-31 days gestation. Significant stimulation of the enzyme by l-isoproterenol was observed only in the presence of guanosine triphosphate (100 microM). Under these conditions, maximal adenylate cyclase stimulation by l-isoproterenol (100 microM) was significantly higher at 25-31 than at 21 days gestation. Moreover, EC50 (Kact) for l-isoproterenol at 25-31 days was significantly lower than at 21 days gestation. We conclude that, in fetal rabbit myocardial membranes, there is an increase in the sensitivity of adenylate cyclase stimulation by l-isoproterenol from 21 to 25-31 days gestation.     

Pharmacologic probing of renal development in the neonatal rat

This study was designed to examine the ontogeny of renal functions in the neonatal rat using various pharmacologic agents as probes. The renal responses of 2-, 6-, and 10-day-old rats to diuretic agents known to act on proximal tubules, loops of Henle and distal tubules were assessed. These included acetazolamide, furosemide, mercaptomerin, chlorothiazide and amiloride. Following administration of a diuretic agent, urine was collected at 90-min intervals for 6 h and urine volume, osmolality, chloride and pH were measured. Acetazolamide, furosemide, chlorothiazide and amiloride induced diuresis at each age indicating that the respective reabsorptive mechanisms were present and functional by 2 days of age. At all ages furosemide evoked a maximal response in eliminating the interstitial fluid gradient as indicated by the formation of an isosmotic urine in treated pups. However, the volume of the diuresis at 2 days of age was half those at 6 and 10 days, reflecting enhanced activity of the countercurrent multiplication apparatus in the maturing pups. Administration of mercaptomerin did not produce pharmacologic diuresis, but rather resulted in acute renal failure; although the nephrotoxicity was to a lesser extent in 2-day-old pups. The ability of the neonatal rat to respond to these pharmacologic probes demonstrates that the integrity of these renal functions is established in this species early in postnatal life.     

In-utero gentamicin-induced nephrotoxicity in rats

Aminosides lead to a well-known nephrotoxicity. The possibility of the developing kidney being altered in utero after the pregnant mother's administration has been investigated. We gave gentamicin (75 mg/kg/day) to pregnant rats during periods of organogenesis (days 7-11) and the beginning of glomeruli differentiation (days 14-18). A group of nonpregnant females was also treated for the same period and at the same time each day. Gentamicin-treated mothers presented only minor modifications of the blood biology with no acute renal failure when treated nonpregnant females have a hypercreatininemia. The deep cortical area, containing the fully formed nephrons of neonates, presented less glomeruli that were differentiated in the gentamicin group than in the control group. Moreover with both light and electron microscopy, glomeruli and proximal tubules showed evidences of nephrotoxicity in the juxtamedullary cortex. This finding of an in utero aminoside nephrotoxicity demonstrates the possible toxicity of gentamicin on fetus kidneys when given during the pregnancy.     

Receptor-mediated cAMP production in adult and infant ferret tracheal epithelium.

Tracheal epithelial cells obtained from adult and infant ferrets were grown in primary culture in vitro. Cells from adult animals grew readily, and many ciliated cells were observed in the cultures. Successful cultures were derived from infant animals, but cell number in infant and adult cultures began to decrease after 6 d. Receptor-mediated activation of adenylate cyclase was determined by incubating monolayers of adult or neonatal cells with beta-adrenergic agonists, prostaglandin E2 (PGE2) and vasoactive intestinal peptide and measuring cAMP production. beta-adrenergic agonists and PGE2, but not vasoactive intestinal peptide, stimulated production of cAMP in both cell types. The 50% effective concentration for isoproterenol and PGE2 in neonatal ferret tracheal epithelial (NFTE) cells was nearly 10-fold more than for adult ferret tracheal epithelial (FTE) cells, but maximal agonist-stimulated cAMP production was significantly different between the cell types only for PGE2. Radioligand binding studies were performed using the beta-adrenergic antagonist [125I]iodocyanopindolol on membrane particulates from confluent monolayers and freshly isolated FTE cells. Binding of iodocyanopindolol was saturable, stereoselective, and of high affinity (binding affinity = 26.1 +/- 6.6 pmol/L, adult; 16.5 +/- 5.7 pmol/L, NFTE). Competition studies with the specific beta 2-adrenergic receptor antagonist, ICI 118 551 revealed a predominance of beta 2-adrenergic receptors on both adult FTE and NFTE cells. Receptor density was significantly higher in adult FTE compared with NFTE cells (48.2 +/- 9.1, 18.1 +/- 1.5 fmol/mg, respectively). Basal adenylate cyclase activity was significantly lower in neonatal cells.(ABSTRACT TRUNCATED AT 250 WORDS)     

Gastrointestinal processing of gastrically administered prostaglandin F2 alpha in suckling and weanling rats

In view of the presence of prostaglandins in milk, studies were performed to determine if maturational differences are present in the gastrointestinal processing of exogenous prostaglandin F2 alpha. Suckling (10- to 14-day-old) and weanling (30-day-old) rats were gastrically fed 3H-labeled prostaglandin F2 alpha and killed 2 h later. Characterization of radioactivity present in the stomach from animals of both age groups revealed low prostaglandin F2 alpha metabolism. Analyses of proximal, middle, and distal intestinal segments revealed age-related differences in prostaglandin metabolism. The amount of unmetabolized PGF2 alpha in intestinal segments of suckling rats ranged from 11.4 to 13.5%. In weanling rats, there was a tendency toward increased amounts of intact PGF2 alpha in proximal (16.3 +/- 1.6%) and middle (17.6 +/- 2.2%) regions; however, 25.9 +/- 1.9% of the tissue radioactivity present in the distal small intestine of weanling rats was authentic PGF2 alpha, significantly greater than that of the distal segment of suckling rats. The intestine of weanling rats contained greater amounts of less polar metabolites. These results indicate that orogastrically fed prostaglandin can pass through the gastrointestinal tract of developing rats and be delivered to the distal intestinal mucosa intact.     

Ontogeny of beta-adrenergic regulation of adenylate cyclase in intrapulmonary arteries from fetal and postnatal lambs.

The enzyme adenylate cyclase is critically involved in the regulation of vasodilatation in the developing pulmonary circulation in that it mediates the vascular smooth muscle effects of beta-adrenergic agonists and vasodilatory prostaglandins. These agonists activate receptors coupled to the catalytic subunit of the enzyme by stimulatory guanine nucleotide-dependent regulatory proteins. We examined the ontogeny of the function of the adenylate cyclase system in intrapulmonary arterial segments from fetal lambs at 110-115 (F1) and 125-135 d gestation (F2) and from postnatal lambs at 7-14 (P1) and 35-56 d of age (P2). The function of the intact enzyme system and its components was assessed in incubations measuring cAMP accumulation during phosphodiesterase inhibition. beta-Adrenergic mediation of adenylate cyclase was examined because the binding characteristics of the smooth muscle receptors can be readily quantified. Nonstimulated (basal) cAMP accumulation was similar in F1 and F2, it increased 8-fold from F2 to P1, and it was equivalent in P1 and P2. cAMP accumulation with isoproterenol increased 5.9-fold from F1 to F2 and was similar in F2, P1, and P2. Radioligand binding studies revealed that the greater response to isoproterenol in F2 versus F1 is not related to an increase in beta-adrenergic receptor density or affinity. cAMP accumulation with forskolin, which activates adenylate cyclase by actions that involve both the stimulatory guanine nucleotide-dependent regulatory protein and the catalytic subunit, was similar in the four age groups, indicating that there are no maturational changes in the funciton of these enzyme system components.(ABSTRACT TRUNCATED AT 250 WORDS)     

Maturation-related changes in dopamine-induced relaxation of isolated rabbit pulmonary artery

Studies were performed on isolated pulmonary arterial segments to investigate dopamine receptor-mediated relaxant effects at different times during development. Dopamine receptor-mediated relaxant effects can only be observed when vessels are precontracted with prostaglandin F2 alpha and in the presence of alpha 1, alpha 2, beta, and serotonergic blockade. Helical strips of pulmonary arteries from rabbits of different ages (2, 7, 14, 30, and 90 days), partially precontracted by prostaglandin F2 alpha were tested for their responses to dopamine in the presence of prazosin (10(-6) M), yohimbine (10(-6) M), propranolol (10(-6) M), and methysergide (10(-6) M). Strips from 2- and 7-day-old rabbits were not induced to relax by dopamine, whereas those from 14-, 30-, and 90-day-old animals, after cumulative application of dopamine, underwent concentration-dependent relaxation. Dopamine (half the maximum response) concentration decreased during the development of rabbits from 14 to 90 days old. Mean values for apparent dopamine ED50 (half the maximum response) concentrations in the arteries of 14-, 30-, and 90-day old animals were 4.94 +/- 0.40, 2.02 +/- 0.30, and 0.113 +/- 0.028 microM, respectively. The effects of various dopamine antagonists on dopamine-induced relaxation were not markedly different at different ages. These findings indicate that dopamine receptor function is not fully developed in the pulmonary arteries of newborn rabbits, but matures as the age of the rabbit increases.