Glucocorticoid regulation of ornithine decarboxylase in the postnatal rat lung.

Ornithine decarboxylase (ODC) is thought to play a critical role in pulmonary development. The purpose of this study was to characterize the effects of dexamethasone on ODC gene expression and enzyme activity in the lung of rat pups. Subcutaneous administration of dexamethasone (10 mg/kg) was shown to suppress ODC activity in 2-, 6- and 10-day-old rats for as long as 24 h after injection. In contrast, dexamethasone treatment stimulated liver ODC activity indicating that the inhibition of lung ODC is tissue specific. Contrary to expectation, the glucocorticoid enhanced lung ODC expression as indicated by an increased accumulation of ODC mRNA transcripts. The latter effect was associated with an heightened expression of c-myc and max mRNAs, the encoded proteins of which act as transactivators of the ODC gene. Dexamethasone did not alter lung levels of"antizyme" (AZ), an inducible protein that specifically promotes the degradation of the ODC protein enzyme. However, the lack of AZ induction does not necessarily mean that ODC degradation is not the mechanism for the decrease in lung ODC activity of dexamethasone-treated animals. The results obtained indicate that glucocorticoids can downregulate lung ODC activity, and that the effect is mediated by post-transcriptional rather than transcriptional mechanisms. These findings are consistent with the idea that endogenous glucocorticoids play an important role in the modulation of ODC activity and early pulmonary development.

Involvement of c-myc and max in CNS beta-endorphin modulation of hepatic ornithine decarboxylase responsiveness to insulin in rat pups.

Previously, we have shown that subcutaneous administration of insulin stimulates ornithine decarboxylase (ODC) mRNA expression and enzymatic activity in the liver of infant control rats, but not in those pretreated intracerebroventricularly (i.c.v.) with beta-endorphin. This finding is consistent with the hypothesis that beta-endorphin synthesized in the brain plays a prime role in the control of postnatal development, in part, by modulating ODC gene transcription. We now report that insulin induced stimulation of hepatic ODC mRNA expression is accompanied by a concomitant increase in the expression of c-myc and max mRNAs, and that this effect is also inhibited by pretreatment with i.c.v. beta-endorphin. These results suggest that CNS beta-endorphin suppresses tissue ODC responsiveness to trophic hormones by downregulating the expression of c-myc and max mRNAs, the encoded proteins of which are known to act physiologically as transcriptional activators of the ODC gene.

Central administration of morphine inhibits brain and liver ornithine decarboxylase activity in neonatal rats: involvement of transcription- and non-transcription-dependent mechanisms.

This study examined whether the developmental deficits usually observed in infants born to opiate addicted mothers could involve effects on ornithine decarboxylase, a growth-controlling enzyme. Intracerebroventricular (i.c.v.) injection of a single dose of morphine (2 microg) to 6-day-old rats markedly decreased basal brain and liver ornithine decarboxylase activity as well as the increases in hepatic ornithine decarboxylase activity produced by subcutaneously (s.c.) administered insulin, an important trophic hormone. Centrally applied morphine acts supraspinally to downregulate peripheral ornithine decarboxylase activity, since s.c. administration of the same dose as used i.c.v. decreased neither basal liver ornithine decarboxylase levels nor tissue responsiveness to insulin. This does not imply that the opiate is unable to affect ornithine decarboxylase when applied systemically. In fact, a robust inhibition of both basal and induced liver ornithine decarboxylase activity was obtained in rat pups given 20 microg of morphine s.c. This larger dose is able to trigger the hepatic ornithine decarboxylase effects presumably by stimulating opiate receptors located at central sites after crossing the blood-brain barrier and penetrating into the brain. Concomitant administration of naloxone plus morphine i.c.v. prevented morphine from downregulating ornithine decarboxylase activity, confirming the participation of supraspinal opioid receptors in morphine ornithine decarboxylase actions. Finally, as was the case for insulin induced stimulation of ornithine decarboxylase activity, i.c.v. injection of morphine markedly diminished insulin induced stimulation of hepatic ornithine decarboxylase mRNA accumulation. In turn, contrary to the inhibition of basal ornithine decarboxylase activity, morphine did not lower basal hepatic ornithine decarboxylase mRNA levels when given alone. Thus, CNS morphine can apparently suppress tissue ornithine decarboxylase expression through both transcriptional and posttranscriptional mechanisms. The evidence obtained suggest that postnatal exposure to opiate drugs might detrimentally affect development by altering normal tissue ornithine decarboxylase ontogeny.

The influence of dietary cholesterol on cardiac and hepatic Beta-adrenergic receptors in egyptian sand rats.

We examined the effects of dietary cholesterol on cardiac and hepatic beta-adrenergic receptor functioning. Age-matched adult desert rodents (Psammomys obesus) were randomized to either a 5% cholesterol diet (CD, n = 20), or normal rabbit chow (RC, n = 18). After a 2-month exposure to the diets, animals were sacrificed and tissue from both heart and liver were retained for radioligand bindings studies. In heart tissue, cholesterol fed animals, relative to controls, showed an increased production of adenosine 3,5>-cyclic monophosphate (cAMP) in response to isoproterenol. Cholesterol supplementation was not associated with an increase in heart beta-adrenergic receptor number. Animals fed the 5% cholesterol diet showed significant increases in the number of beta-adrenergic receptor sites in hepatic tissue (M = 13.2 vs. 10.4 pmol/mg protein, CD and RC, respectively). The increased number of receptor sites in the liver was accompanied by a significant increase in isoproterenol-stimulated cAMP production. Results are supportive of the hypothesis that dietary cholesterol contributes to an upregulation of beta-adrenergic receptor function in cardiac, as well as hepatic tissue. These findings may be relevant to the observations of excessive stress-induced cardiovascular reactivity in persons with high cholesterol levels.

Neonatal deprivation of maternal touch may suppress ornithine decarboxylase via downregulation of the proto-oncogenes c-myc and max.

Previously, we have shown that short-term (1 hr) separation of neonatal rats from their mother (MS) suppresses basal ornithine decarboxylase (ODC) synthesis and tissue ODC response to trophic factors. This effect in the pup is caused by absence of maternal tactile stimulation (touch) but not from lack of maternal nutrients (food). This study was performed to examine in 10-d-old rats whether maternal touch deprivation affects expression of certain hepatic proto-oncogenes, the protein products of which are known to interact with the regulatory region of the ODC gene. Prolactin (PRL) injected subcutaneously increased hepatic ODC activity as well as mRNA levels of ODC and the proto-oncogenes c-fos, c-jun, junB, junD, c-myc, and max. MS significantly suppressed PRL-induced increases in ODC enzyme activity and c-myc, max, and ODC mRNAs but had little effect on expression of the other proto-oncogenes. PRL-induced stimulation of ODC, c-myc, and max mRNAs also was depressed in neonates placed with an anesthetized lactating dam (touch-deprived) but not in pups placed with nipple-ligated dams (food-deprived). Furthermore, unlike its effect on preweanling-age pups (< 20 d old), MS did not alter expression of either ODC or c-myc mRNAs in 25-d-old pups acutely separated from their mother. These findings indicate that suppression of ODC gene transcription in the neonatal pup during MS may be mediated by downregulation of the ODC gene transactivator proto-oncogenes c-myc and max. They are also consistent with our previous observation that lack of maternal touch, but not maternal milk, initiates the physiological alterations induced by MS.

Transcription-dependent and -independent regulation of hepatic ornithine decarboxylase activity by CNS beta-endorphin in rat pups.

We have previously shown that intracerebroventricular administration of relatively low doses of beta-endorphin suppresses basal levels of hepatic ODC activity as well as tissue ODC responsiveness to administered insulin in developing rats. Using Northern blotting analysis, the current studies examine whether these effects of CNS beta-endorphin may be mediated by changes in ODC gene expression. Subcutaneous administration of insulin (20 IU/kg body weight) rapidly and profoundly increased liver ODC activity. The time course of the response was characterized by proportionally increased levels of ODC mRNA, suggesting that insulin-induced stimulation of ODC activity is due to an increased transcription of ODC mRNA. Pretreatment with actinomycin D (2 mg/kg body weight, intraperitoneally) completely prevented the insulin-induced increase in ODC activity, confirming the requirement for the de novo synthesis of ODC mRNA for the effect. More importantly, intracerebroventricular but not subcutaneous injection of beta-endorphin (1 microgram) markedly diminished the stimulatory effect of insulin on hepatic ODC mRNA accumulation. The time course and magnitude of the inhibition of mRNA accumulation essentially mirrored that of the peptide on ODC activity. On the other hand, contrary to the inhibitory effect of beta-endorphin on basal ODC activity, the peptide did not lower basal ODC mRNA levels when given alone. Taken together, the results from these studies provide evidence for the existence of at least two separate mechanisms through which CNS beta-endorphin might downregulate ODC activity in peripheral organs of rat pups. The peptide can suppress insulin-induced ODC activity in the liver tissue by decreasing the rate of transcription of the ODC gene, whereas the inhibition of basal ODC activity appears to involve posttranscriptional mechanisms.

The inhibition of ornithine decarboxylase activity in developing rat tissues by central nervous system beta-endorphin is mediated by mu-opioid receptors, but not by delta- or epsilon-opioid receptors.

Our laboratory has previously shown that intracerebroventricular (i.c.v.) administration of beta-endorphin suppresses brain and liver ornithine decarboxylase activity (ODC; a growth regulatory enzyme) in preweanling rats. This investigation examined, in 6-day-old rats, the relative participation of brain mu-, delta- and epsilon-opioid receptors in beta-endorphin's ODC effects, by comparing tissue ODC responses to beta-endorphin given alone i.c.v. and in the presence of D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH2 (CTOP; mu-opioid receptor antagonist), N,N-diallyl-Tyr-Aib-Aib-Phe-Leu-OH (ICI-174,864; delta-opioid receptor antagonist) or beta-endorphin-(1-27) (epsilon-opioid receptor antagonist). Administration of 0.5 microgram of beta-endorphin alone significantly decreased brain and liver ODC activity 4 h after injection, and the effect was completely blocked by coinjection of CTOP (0.075 micrograms) but not by ICI-174,864 (0.75 or 3.75 micrograms) or beta-endorphin-(1-27) (3.75 or 7.5 micrograms). The blockade of endogenous opioid:opioid receptor interactions by either CTOP (at doses > 0.075 microgram) or ICI-174,864 alone was accompanied by increased levels of basal ODC activity. The results obtained demonstrate that i.c.v. beta-endorphin downregulates ODC expression in central as well as in peripheral tissues by interacting with brain mu-opioid receptors, but not with delta- or epsilon-opioid receptors or mu/delta-opioid receptor complexes. Also, they indicate that endogenous opioid systems have a tonic inhibitory influence on ODC activity which is mediated, at least in part, by mu- and delta-opioid receptors.

Brain cholecystokinin and beta-endorphin systems may antagonistically interact to regulate tissue DNA synthesis in rat pups.

Previously, we have shown that intracisternal (i.c.) administration of beta-endorphin suppresses brain and liver DNA synthesis in rat pups. This finding is consistent with the view that endogenous CNS beta-endorphin plays an important role in controlling postnatal growth. Recent evidence suggests that brain CCK8, the sulfated carboxyterminal octapeptide fragment of cholecystokinin, may function physiologically as an endogenous opioid antagonist. We now report that CCK8 injected i.c. together with beta-endorphin effectively prevented beta-endorphin from inhibiting brain and liver DNA synthesis in 10-day-old rats. CCK8 blocked the liver DNA effect of beta-endorphin via actions within the brain, as subcutaneous administration of CCK8 was ineffective. In contrast to CCK8, i.c. administration of CCK8U (the unsulfated form of CCK8) together with beta-endorphin did not prevent beta-endorphin from inhibiting liver DNA synthesis, and only slightly reversed the brain DNA effect. The results obtained support a role for endogenous brain CCK8 in the modulation of tissue DNA responses to CNS beta-endorphin and possibly to other endogenous opioids. If so, interference with brain CCK function could disrupt tissue growth. Thus, normal mammalian development may require a close functional interaction between the cholecystokinin and beta-endorphin systems in the brain.

Role of the spinal cord in intracisternal beta-endorphin-evoked suppression of liver DNA synthesis in 10-day-old rats.

Previously, we have shown that intracisternal (i.c.) administration of beta-endorphin (an opioid peptide naturally occurring in the brain) to preweanling rats markedly decreases DNA synthesis (an index of cell proliferation) in both brain and liver. This observation is consistent with our hypothesis that endogenous CNS beta-endorphin plays an important role in controlling postnatal growth. The current research specifically undertook to investigate, in 10-day-old rats, whether or not i.c. beta-endorphin-evoked suppression of liver DNA synthesis is actually mediated by spinal opioid receptors and/or by descending endorphinergic pathways. In contrast to the i.c. route of administration, beta-endorphin given directly into the spinal subarachnoid space via intrathecal (i.t.) injection did not alter liver DNA synthesis, yet was able to evoke profound antinociceptive responses. This demonstrates that intracisternally applied beta-endorphin exerts its effect on liver DNA by acting at supraspinal sites, and not by directly stimulating spinal opioid receptors after diffusion from its intracerebral site of injection. As it is possible that beta-endorphin's supraspinal actions may activate a descending inhibitory endorphinergic pathway to reduce DNA synthesis, we conducted studies in rat pups administered naloxone intrathecally. Naloxone i.t. was completely ineffective in preventing beta-endorphin i.c. from inhibiting liver DNA synthesis. On the other hand, i.t. coinjection of naloxone plus beta-endorphin was able to block the analgesic response, while their i.c. coinjection reversed the DNA effect. The results from these studies indicate that opioid systems within the spinal cord do not play a major role in mediating CNS beta-endorphins regulation of DNA synthesis in peripheral tissues.(ABSTRACT TRUNCATED AT 250 WORDS)

The inhibition of liver ornithine decarboxylase expression in neonatal rats by maternal separation or CNS beta-endorphin is independent of the pituitary.

Previously we have shown, in rat pups, that either short-term maternal separation (MS) or central (but not peripheral) administration of beta-endorphin (BE) markedly decreases basal levels of ornithine decarboxylase (ODC) activity throughout the body and suppresses liver ODC responsiveness to injected growth hormone (GH). In this study, hypophysectomized (hypox) pups were used to determine whether the pituitary mediates these effects. Hypophysectomy clearly did not prevent the inhibitory actions of MS or intracisternal (i.c.) BE on liver ODC gene expression. The inability of GH to stimulate ODC activity in hypox animals exposed to MS or given BE i.c. is not due to nutritional deprivation, as glucose supplementation did not reverse the response. The results from these studies demonstrate that the pituitary is not the conduit by which either MS or centrally-administered BE regulates liver ODC activity. Also, they support the hypothesis that BE or an analogous opioid neuropeptide is a prime organizer within the CNS of the adaptive physiological response of neonatal rats to short-term MS. As we have previously shown that autonomic neuronal pathways are not involved in the effects of MS on peripheral tissues, the data obtained suggest that increased activity of this CNS opioid system during MS triggers the release of "neurochemicals" into the bloodstream capable of suppressing growth in the mammalian neonate.

Thyroid hormone differentially regulates cellular development in neonatal rat heart and kidney.

The role of thyroid hormone in the control of cardiac and renal cell development was examined in neonatal rats made hyperthyroid by administration of triiodothyronine (T3, 0.1 mg/kg s.c. on postnatal days 1-5) or hypothyroid by administration of propylthiouracil (PTU, 20 mg/kg s.c. given to dams on gestational day 17 through postnatal day 5 and to pups on postnatal days 1-5). Indices of total cell number (total DNA per tissue), cell packing density (DNA per g tissue), and relative cell size (protein/DNA ratio) were evaluated from birth through young adulthood. PTU administration led to primary shortfalls in cell number that were of similar magnitude in both tissues, but persisted somewhat longer in the kidney than in the heart. Deficits in cell packing density and cell size in the hypothyroid animals were secondary to the effect on cell number, displaying smaller magnitudes of effect and a lag in appearance and disappearance of the deficits compared to that for total DNA; indeed, the phase in which tissues were restoring their cell numbers was accompanied by increased cell packing density, reflecting a more rapid restitution of cell numbers than tissue weight or cell size. In contrast to the relatively similar effects of PTU on developing cardiac and renal cells, the effects of T3 were selective for the heart. Although T3 caused general growth impairment, it evoked marked cardiac overgrowth that was accompanied by a striking increase in cell number and a small increase in cell size. The cardiac hyperplasia is unique to the developing animal, as post-replicative heart cells in adult animals show only hypertrophy in response to thyroid hormone.(ABSTRACT TRUNCATED AT 250 WORDS)

Fetal dexamethasone exposure impairs cellular development in neonatal rat heart and kidney: effects on DNA and protein in whole tissues.

Fetal glucocorticoid exposure causes postnatal growth retardation. To examine the mechanisms underlying effects on specific organ systems, we administered 0.2 or 0.8 mg/kg of dexamethasone to pregnant rats on gestational days 17, 18, and 19 and assessed three biochemical markers of cell development in heart and kidney of the offspring: DNA content per organ as an index of total cell numbers, DNA per g tissue as an index of cell packing density, and protein/DNA ratio as an index of relative cell size. In both tissues, DNA content became markedly subnormal during the first postnatal week, the ontogenetic period of rapid cell division. Partial recovery occurred by the end of the first postnatal month. In the heart, cell packing density was subnormal initially and the cells were significantly enlarged. In contrast, packing density was slightly elevated in the kidney; protein/DNA was increased by the low dose of dexamethasone, but markedly decreased by the high dose. These results suggest that tissue growth impairment caused by prenatal dexamethasone treatment reflects primary deficits in cell proliferation that extend to a variety of different cell types; however, consequent effects on cell packing density and cell size are dose-specific, possibly reflecting actions of glucocorticoids selective for certain cell types or phases of cell development.

Effect of central administration of beta-endorphin on lung ornithine decarboxylase activity in developing rats.

Results from a number of studies suggest a role for endogenous opioids in the regulation of lung development and function. Although it is not known which opioid peptides are involved in these processes, accumulated evidence suggests a prominent role for beta-endorphin (BE). Our study examines the effect of BE on lung ornithine decarboxylase (ODC) activity in preweanling rats. ODC catalyzes the rate-limiting step in the synthesis of the polyamines spermidine and spermine, key regulators of cell growth, multiplication, and differentiation. Central (but not peripheral) administration of BE reduced lung ODC activity by as much as 80% in the 6-d-old rat. Significant decreases in ODC activity were seen at doses of BE as low as 0.5 micrograms/g brain wt. In contrast to the reductions in ODC activity, plasma levels of corticosterone in animals administered BE were approximately five times higher than those seen in control animals. BE's actions on ODC activity and plasma corticosterone levels were prevented by naloxone or naltrexone, indicating that both responses are mediated by opioid receptors. Studies of ODC kinetics showed a profound reduction in Vmax (70% below control values), but no change in Km. The effect was observed only during the first 2 wk of postnatal age, a period of time in lung maturation that is characterized by active alveolarization. Because changes in ODC levels during early postnatal life are associated with perturbations in tissue growth and/or function, the data suggest that CNS BE may influence lung maturation through an indirect action that may involve glucocorticoids.

Further evidence for the hypothesis that beta-endorphin mediates maternal deprivation effects.

Lung DNA synthesis was examined in 9-day-old rat pups following a 2-hour separation from their mothers (maternal deprivation), and compared to that of pups placed with a nipple ligated dam (food deprivation) or a lactating dam (control). Maternally deprived pups consistently showed a significant reduction in lung DNA synthesis which was not attributable to food deprivation. Central administration of naloxone prevented the decrease in DNA synthesis observed after maternal deprivation but did not inhibit the reductions in lung DNA synthesis seen two hours after sc administration of isoproterenol, suggesting that DNA response to maternal deprivation is a specific opioid receptor mediated event. These results are consistent with previous reports from our laboratory indicating that CNS beta-endorphin may mediate many of the biological alterations observed following maternal deprivation in neonatal rats.

Effects of central administration of beta-endorphin on brain and liver DNA synthesis in preweanling rats.

We have previously shown that central administration of beta-endorphin results in a reduction of ornithine decarboxylase activity. Ornithine decarboxylase catalyses the rate-limiting step in the biosynthesis of the polyamines putrescine, spermidine and spermine, thought to modulate nucleic acid synthesis. The present study examines the effects of intracisternal injection of beta-endorphin on brain and liver DNA synthesis in preweanling rats. In six-day-old rats, beta-endorphin (0.75 micrograms/g brain wt) produced approximately a 70% inhibition in brain and liver DNA synthesis 1 h after injection, and values were still subnormal in both tissues 10 h later. Subcutaneous administration of beta-endorphin did not alter liver DNA synthesis. Thus, it is most likely that the suppressed liver DNA synthesis observed in animals given beta-endorphin intracisternally is mediated by central mechanisms. Co-administration of naloxone plus beta-endorphin intracisternally prevented the response, indicating an opioid receptor-mediated phenomenon. Naloxone alone caused small but significant increases in brain and liver DNA synthesis, suggesting a tonic influence on tissue DNA by endogenous opioids in the CNS. Acute inhibition of ornithine decarboxylase activity by alpha-difluoromethylornithine did not alter DNA synthesis, indicating that the decreases in DNA synthesis induced by beta-endorphin are unrelated to the ornithine decarboxylase/polyamine system. The effect appears to be restricted to early development as no significant changes in DNA synthesis were obtained in 20-day-old animals. The results from these studies indicate that CNS beta-endorphin has the ability to influence DNA synthesis in central as well as in peripheral tissues.(ABSTRACT TRUNCATED AT 250 WORDS)

CNS beta-endorphin regulation of insulin-induced ornithine decarboxylase expression in liver of neonatal rats.

Previously, we reported that central (but not peripheral) administration of beta-endorphin to rat pups decreases basal ornithine decarboxylase (ODC; a growth-associated enzyme) activity throughout the body. This finding is consistent with the view that CNS beta-endorphin is an important regulator of postnatal development. Insulin is a hormone that markedly affects liver growth and evidence indicates that this occurs, in part, through its ability to regulate ODC expression. The current study examines the effects of intracisternal injection of beta-endorphin on the response of liver ODC to subcutaneous administration of insulin. Insulin (20 IU/kg body wt), markedly increased liver ODC activity in 2-, 4-, 6-, 10-, 18-, and 30-day-old rats. Intracisternal injection of beta-endorphin (1.5 mug/g brain wt) inhibited this response to insulin in 2-, 4-, 6-, 10-, and 18-day-old rats, but not in 30-day-old animals. This inhibition by beta-endorphin was not reversed by naloxone, indicating that the effect is not mediated by brain mu or delta opioid receptors. Centrally administered beta-endorphin also blocked the increases in liver ODC activity evoked by subcutaneous administration of cAMP. The results from these studies suggest that the postulated regulation of postnatal development by CNS beta-endorphin may occur through actions on both basal ODC activity and tissue ODC sensitivity to "classical" trophic factors. The modulation by beta-endorphin of liver ODC expression apparently occurs distally to cAMP-generating mechanisms.

Effects of beta-endorphin on DNA synthesis in brain regions of preweanling rats.

Recent studies of whole brain in rat pups have shown a marked decrease in DNA synthesis following intracisternal (i.c.) administration of beta-endorphin (BE). This investigation examines DNA synthesis in the cerebral cortex and cerebellum to determine whether the effect shows regional selectivity. Two- to twenty-day-old rats were given a single ic injection of BE, and DNA synthesis was assessed 1 h later. In the cerebral cortex, a region that undergoes major phases of cell multiplication in the immediate pre- and postnatal periods, BE significantly decreased DNA synthesis in 2-day-old rats, and a maximal inhibition was obtained by 4 days of age. In contrast, the cerebellum, a region that grows predominantly after birth, showed less sensitivity to BE during the early postnatal days, and a maximal effect was not attained until 10 days of age. While at 15 days of age the inhibition began to diminish in the cortex, a maximal effect was still seen in the cerebellum. Naloxone prevented the response in both brain regions, indicating the participation of opioid receptors. These results indicate that CNS BE is apparently able to alter DNA synthesis throughout the brain, with the greatest sensitivity occurring in those regions with highest mitotic rates at the time of exposure to BE.

Acute difluoromethylornithine treatment increases skin flap survival in rats.

Difluoromethylornithine (DFMO) pretreatment for 7 days improved survival of rat abdominal skin flaps in previous studies. The purpose of this study was to determine if acute administration of DFMO enhances survival. Each rat had a 7 x 7-cm abdominal skin flap raised on a single epigastric neurovascular pedicle. Within 1 minute of pedicle ligation, the rats were given 0, 1, or 4 gm/kg of body weight of DFMO intraperitoneally. Putrescine was administered to additional rats alone or with DFMO. After 48 hours, the percentage of flap survival was estimated using fluorescein injection and planimetry to quantify the perfused and unperfused areas. Flap survival increased from 71 +/- 3% in controls to 83 +/- 2% and 92 +/- 3% in rats treated with 1 and 4 gm/kg of DFMO, respectively (p less than 0.005). Putrescine reversed the protective effect of DFMO, suggesting a specific polyamine-related mechanism. This study indicates that there may be both short- and long-term polyamine pools through which DFMO acts. In summary, DFMO may prove to be important in preventing cell death following acute ischemia.

Regulation of insulin and glucose plasma levels by central nervous system beta-endorphin in preweanling rats.

Recently, our laboratory has reported that central administration of beta-endorphin to rat pups decreases hepatic and renal ornithine decarboxylase activity, a sensitive biochemical index of tissue metabolic activity. Since these organs are the major sites of insulin catabolism, it seemed possible that the plasma levels of this hormone could be altered by changes in central nervous system (CNS) beta-endorphin levels. In the current study we tested this hypothesis by administering beta-endorphin to rat pups intracisternally (ic), followed by insulin sc, and then analyzing for plasma levels of insulin and glucose at various times after the second injection. We found that the apparent biological half-life of administered insulin markedly increased in 6-day-old rats pretreated with beta-endorphin ic. Similarly, this neuropeptide prolonged the half-life of endogenous insulin, as indicated by a small but significant increase in the plasma levels of this hormone in animals given only beta-endorphin. As expected, hypoglycemia in rats injected with beta-endorphin and insulin was more pronounced than in animals given insulin alone. Naloxone administered ic reversed both actions of beta-endorphin, indicating the involvement of opioid receptors in the response. beta-Endorphin also altered insulin and glucose plasma levels in 2-, 10-, and 18-day-old rats, but there were no effects in 30-day-old animals at any of the doses used in these studies. Peripheral administration of beta-endorphin had no effect, indicating that CNS beta-endorphin's influences on insulin and glucose metabolism occur through brain-based mechanisms. The results from these studies suggest that CNS beta-endorphin may be an important modulator of insulin and glucose metabolism in preweanling rats. In as much as insulin is a major regulator of somatic growth, our findings further suggest that CNS beta-endorphin may have a major role in the control of growth during early postnatal development by influencing insulin homeostasis.

Quantification of cellular proliferation in experimental proliferative vitreoretinopathy.

Proliferation of host cells from around the optic nerve head has recently been implicated in the development of experimental proliferative vitreoretinopathy in rabbit eyes injected with homologous fibroblasts. We used liquid scintillation spectrometry to quantitate the tritiated thymidine incorporation into cells in the vitreous, retina, and optic nerve head following intravitreal injection of 250,000 homologous dermal fibroblasts. Cellular proliferation peaked three days after injection of the fibroblasts. The amount of tritiated thymidine incorporation that occurred three days following injection of irradiated homologous fibroblasts (incapable of cellular division) was not significantly different than that following injection of normal homologous fibroblasts, indicating that host cells were responsible for most of the cellular proliferation. Treatment with fluorouracil or triamcinolone acetonide completely arrested cellular proliferation following injection of normal fibroblasts.