Androgenic control of polyamine concentrations in rat epididymis.

Unilateral orchidectomy resulted in a significant decrease in tissue content of putrescine and polyamines. However, no differences were detected when the results were expressed in terms of ng g-1 tissue. At 48 h after bilateral orchidectomy, a significant decrease in putrescine content was observed, but spermidine and spermine content were unaffected. The observed decrease in putrescine was prevented by treatment with testosterone propionate, but neither spermidine nor spermine were affected. Bilateral orchidectomy resulted in a significant decrease in the tissue content of putrescine, spermidine and spermine after 7 days. Treatment with testosterone propionate increased the content of putrescine, spermidine and spermine in the epididymis by about 200%, 92% and 34%, respectively. When results were expressed as nmol g-1, a significant decrease after castration in putrescine and spermidine, but not in spermine, was observed. Treatment with testosterone propionate restored putrescine concentration, but had no effect on spermidine and spermine concentrations. In castrated rats treated with testosterone propionate, the anti-androgen flutamide abolished the effect of the androgen on putrescine and spermidine content, but there was no effect on spermine. Acetylputrescine was not detected in the epididymis, while acetylpolyamines were detected at much lower concentrations than polyamines. After bilateral orchidectomy there was a decrease in the tissue content of all acetylpolyamines and an increase in their tissue concentration. The effect of castration on acetylpolyamine content was reversed by testosterone propionate treatment. We conclude that an active synthesis of polyamines occurs in the rat epididymis, and that this process depends upon the androgen environment. Regulation of ornithine decarboxylase activity appears to be the main step that is controlled by androgens.     

Tissue-specific changes in polyamine levels of neural tissue and fat body in adult crickets

The three major polyamines—putrescine, spermidine, and spermine—were studied and changes of their levels were examined in extracts of cerebral ganglia and fat body from adult Acheta domesticus. In nervous tissue, only spermidine and spermine were present and spermine was two- to three-fold more abundant than spermidine. The polyamine levels were high up to day 3, decreased on day 4, and then remained relatively unchanged up to day 10. The spermidine/spermine ratios decreased during the imaginal life. Higher spermidine titres were observed in the neural tissue of egg-laying females compared to virgin females. In the fat body, putrescine was detected together with spermidine and spermine. Spermidine and spermine levels were two-fold higher than putrescine. Fat body of virgin females contained two times more polyamines than male fat body. Low at emergence, spermidine and spermine concentrations peaked on days 2–3 only in females, and egg-laying was characterized by an increase of putrescine and spermidine titres. Starvation did not change polyamine contents, implying homeostatic regulation of the intracellular polyamine metabolism. These data showing tissue specific changes in polyamine levels during the imaginal life of Acheta domesticus point to the physiological importance of polyamines as possible intracellular regulators during adult insect development. © 1993 Wiley-Liss, Inc.     

Association of increased polyamine levels with isoproterenol-stimulated mucin secretion in the rat submandibular gland

Incubation of rat submandibular gland slices with 50 microM isoproterenol for 10-40 min stimulated mucin secretion and induced a 3- to 4-fold increase in tissue concentrations of the polyamines putrescine, spermidine and spermine. alpha-Difluoromethylornithine, a specific inhibitor of ornithine decarboxylase, suppressed the isoproterenol-induced increase in submandibular polyamines and inhibited mucin secretion. Exogenous putrescine restored tissue polyamine levels and partially reversed the inhibitory effect of alpha-difluoromethylornithine on mucin secretion. Rapid increases in polyamine levels appear to mediate isoproterenol-stimulated mucin secretion in the rat submandibular gland.     

POLYAMINE METABOLISM IN THE BRAIN AND LIVER OF THE DEVELOPING MONKEY

Abstract— The concentrations of putrescine and the polyamines, spermidine and spermine, along with the specific activities of the enzymes involved in their biosynthesis, ornithine decarboxylase, S -adenosylmethionine decarboxylase and spermidine synthase have been measured in brain and liver of the developing Rhesus monkey from mid-gestation, through birth and neonatal life to maturity. The results suggest that it is an increased concentration of putrescine and an increased specific activity of ornithine decarboxylase which are associated with the rapid growth of fetal brain during the middle of gestation. By the end of two-thirds of gestation, both of these parameters have attained values similar to those found in mature brain. The concentration of spermidine in brain and the specific activities of S -adenosylmethionine decarboxylase and spermidine synthase are lower in fetal brain than adult brain and increase slowly after birth to reach values similar to those of the adult only after several months. These results provide additional evidence that in the mature brain spermidine serves some function other than one associated with rapid growth.
Fetal liver at mid-gestation was characterized by increased concentrations of both putrescine and spermidine and increased specific activities of the enzymes which synthesize them. By two-thirds of gestation, values similar to those found in adult liver had been attained. Liver has thus reached maturity with regard to polyamine metabolism by this time.     

Regulation of growth and macromolecular synthesis by putrescine and spermidine in Pseudomonas aeruginosa

Growth of P. aeruginosa, slowed by the addition of monofluoromethylornithine, difluoromethylarginine and dicyclohexylammonium sulfate, could be restored by addition of 0.1 mM putrescine plus 0.1 muM spermidine, or 0.1 mM spermidine or 5 mM putrescine by themselves. Lower concentrations of putrescine (0.1 mM - 1 mM) also partially reversed the growth inhibition. Conversion of putrescine to spermidine continued, although at a markedly reduced ratio, in the drug-inhibited cells, but intracellular spermidine concentrations remained depressed suggesting that reversal of inhibition by putrescine may be a direct effect. There was appreciable back-conversion of any added spermidine to putrescine with a demonstrable increase in total intracellular putrescine levels, making conclusions on the effects of spermidine ambiguous. Spermine (0.1 mM), a polyamine not present in bacteria, was also effective in reversing growth inhibition, probably because of its conversion into spermidine and putrescine. The effects of putrescine, spermidine and spermine were specific in that the non-physiological amines, 1,3-diaminopropane, 1,5-diaminopentane (cadaverine), 1,6-diaminohexane, or 1,7-diaminoheptane could not reverse the effects of the three drugs. Rates of total protein, RNA and DNA synthesis were all slowed to the same extent as growth rate and showed similar recovery with the addition of putrescine or spermidine. A role for putrescine in P. aeruginosa growth processes is suggested.     

The influence of nerve section on the metabolism of polyamines in rat diaphragm muscle.

Concentrations of spermidine, spermine and putrescine have been measured in rat diaphragm muscle after unilateral nerve section. The concentration of putrescine increased approx. 10-fold 2 days after nerve section, that of spermidine about 3-fold by day 3, whereas an increase in the concentration of spermine was only observed after 7-10 days. It was not possible to show enhanced uptake of either exogenous putrescine or spermidine by the isolated tissue during the hypertrophy. Consistent with the accumulation of putrescine, activity of ornithine decarboxylase increased within 1 day of nerve section, was maximally elevated by the second day and then declined. Synthesis of spermidine from [14C]putrescine and either methionine or S-adenosylmethionine bt diaphragm cytosol rose within 1 day of nerve section, but by day 3 had returned to normal or below normal values. Activity of adenosylmethionine decarboxylase similarly increased within 1 day of nerve section, but by day 3 had declined to below normal values. Activity of methionine adenosyltransferase was elevated throughout the period studied. The concentration of S-adenosylmethionine was likewise enhanced during hypertrophy. Administration of methylglyoxal bis(guanylhydrazone) produced a marked increase in adenosylmethionine decarboxylase activity and a large increase in putrescine concentration, but did not prevent the rise in spermidine concentration produced by denervation. Possible regulatory mechanisms of polyamine metabolism consistent with the observations are discussed.     

Effect of cadmium and Phytophthora infestans on polyamine levels in potato leaves

Changes in putrescine, spermidine and spermine concentrations in potato ( Solanum tuberosum L.) leaves stressed either with cadmium, with the fungal pathogen Phytophthora infestans Mont. (de Bary) or with both simultaneously were investigated. The leaves of two cultivars, Bintje and Bzura, respectively susceptible and resistant to P. infestans were examined. The level of of putrescine did not change under any stress conditions. Cadmium stress caused at least a 2-fold increase in spermidine and spermine concentrations in susceptible leaves. In resistant leaves there was a 4-fold increase in spermidine and a decrease in spermine. The pathogenic stress produced similar changes in polyamine concentrations, i. e. with differences between the cultivars. The double stress induced antagonistic alterations in the concentrations of spermidine and spermine.     

Involvement of Putrescine in the Development of Kindled Seizure in Rats

During the development of kindling by daily electrical stimulations applied to the left amygdala of rats, concentrations of the polyamines putrescine, spermidine, and spermine were measured in the left amygdala and the remainder of the cerebrum. A significant increase of putrescine concentration appeared first at the left amygdala in prekindled rats and then propagated to the remainder of the cerebrum with the development of kindling. This increase in putrescine concentration in the left amygdala was higher in prekindled rats than in fully kindled rats and lasted for at least 24 h after the final kindling stimulation. The concentrations of spermidine and spermine were slightly increased in a fully kindled state. To clarify the role of putrescine in kindling, the development of amygdaloid kindling was examined in rats after microinjections of alpha-difluoromethylornithine, a specific inhibitor of polyamine synthesis, and putrescine into the ipsilateral amygdala. Pretreatment with alpha-difluoromethylornithine (50 nmol) for 10 days accelerated both the development of behavioral kindling and the propagation of the afterdischarge from the left amygdala to the frontal cortex. In contrast, pretreatment with putrescine (200 nmol) for 10 days retarded the development of kindling. These results suggest that the increase in putrescine concentration in the kindled brain has an inhibitory effect on the development of kindling.     

Response of tissue diamine oxidase activity to polyamine administration.

The administration to rats of putrescine (750 mumol/kg body wt.) caused in liver, kidney and heart an increase in putrescine at 1 h and in diamine oxidase (EC 1.4.3.6) activity within 3-6 h. An increase in spermidine was observed at 9 h in liver and at 6 h in kidney, whereas in heart there was no change. The increase in diamine oxidase activity by exogenous putrescine was prevented by the administration of actinomycin D and cycloheximide, suggesting that syntheses of mRNA and protein are involved. Equimolar doses of 1,3-diaminopropane, 1,5-diaminopentane and monoacetylputrescine stimulated, similarly to putrescine, hepatic, renal and cardiac diamine oxidase activity. After the injection of a non-toxic dose of spermidine (750 mumol/kg body wt.), the increase in diamine oxidase activity occurred at 9 h in all the tissues studied, when a substantial putrescine formation from spermidine occurred. sym-Norspermidine, which is unable to form putrescine, did not cause an increase in enzyme activity. The possibility that the tissue contents of putrescine might regulate diamine oxidase activity is discussed.     

REGIONAL DISTRIBUTION OF PUTRESCINE, SPERMIDINE AND SPERMINE IN RELATION TO THE DISTRIBUTION OF RNA AND DNA IN THE RAT NERVOUS SYSTEM

—Putrescine, spermidine, spermine, RNA, DNA and protein concentrations were determined in 14 parts of the rat nervous system. If the concentrations are expressed in DNA units, putrescine and spermidine concentrations change concomitantly in the different brain parts, with the exception of hypothalamus, where relatively higher putrescine than spermidine concentrations are observed. The constancy of putrescine/spermidine ratios indicates the value of putrescine concentration as an index of spermidine biosynthesis. Spermidine correlates with RNA, except in medulla, spinal cord and peripheral nerves. It is assumed that the relative excess of spermidine in these structures indicates an additional functional role. Spermine/DNA ratios are remarkably constant in the diencephalic and telencephalic regions; they are also nearly constant, but significantly lower in midbrain, medulla, spinal cord and cerebellum. This observation gives additional support for the preferential interrelation of spermidine with RNA and spermine with DNA; i.e. for different functional roles of these two narrowly related polycations.     

Interconversion of putrescine,spermidine and spermine in goldfish and rat retina

Labelled putrescine is converted to spermidine and spermine in the retina of both the goldfish and of the rat, but the bulk remains as putrescine and spermidine in the goldfish retina whereas the bulk is present as spermine in the rat retina. Labelled spermidine is converted to spermine and to putrescine in the retina of both species, most remaining as spermidine in the goldfish retina whereas most is converted to spermine in the rat retina. Labelled spermine is converted to both spermidine and putrescine in the retina of both species with a greater conversion in the goldfish retina than in the rat retina. These results provide direct evidence of the interconversion of putrescine, spermidine and spermine in neural tissue from both fish and mammals and suggest that spermine should not be regarded solely as an end-product of putrescine metabolism but also as a source of spermidine and putrescine.The pattern of distribution of putrescine and the polyamines, spermidine and spermine, in goldfish retina is the reverse of that in rat retina: Putrescine is the most abundant in goldfish retina whereas spermine is most abundant in rat retina suggesting that the individual polyamines are of different importance in the two species.     

Early developmental profile of ornithine decarboxylase in the frog, Microhyla ornata and its regulation by polyamines

Ornithine decarboxylase (ODC) activity and polyamine levels were measured during early development of the frog, Microhyla ornata. ODC activity was found to be high and it showed three major peaks during the first 60 hr of development. Putrescine and spermidine levels increased gradually during the above period with little change in spermine. Treatment of developing embryos with exogenous putrescine and spermidine prevented the normal increase in ODC activity. Spermine did not have any significant effect. Addition of ornithine also prevented the increase in ODC activity. Experiment using exogenous ornithine and alpha-methylornithine revealed that formation of putrescine and/or spermidine from ornithine is necessary for the suppression of ODC to occur. Suppression of ODC takes place even if conversion of putrescine to spermidine is blocked, indicating that putrescine, independent of its conversion to spermidine, also plays a role in ODC regulation.     

Effects of Polyamines on Chlorophyll and Protein Content, Photochemical Activity, and Chloroplast Ultrastructure of Barley Leaf Discs during Senescence

The polyamines putrescine, spermidine, and spermine prevent the loss of chlorophyll normally associated with senescence of excised leaf tissue maintained in darkness on water (control). Retention of chlorophyll in barley leaf discs was in the range of 90% 4 days after excision and placement on effective polyamine solutions. In contrast, the loss of soluble protein was hastened with 0.5 millimolar spermidine and spermine treatments but it was retarded by 0.5 millimolar putrescine.     

Transgenic manipulation of the metabolism of polyamines in poplar cells

The metabolism of polyamines (putrescine, spermidine, and spermine) has become the target of genetic manipulation because of their significance in plant development and possibly stress tolerance. We studied the polyamine metabolism in non-transgenic (NT) and transgenic cells of poplar (Populus nigra x maximowiczii) expressing a mouse Orn decarboxylase (odc) cDNA. The transgenic cells showed elevated levels of mouse ODC enzyme activity, severalfold higher amounts of putrescine, a small increase in spermidine, and a small reduction in spermine as compared with NT cells. The conversion of labeled ornithine (Orn) into putrescine was significantly higher in the transgenic than the NT cells. Whereas exogenously supplied Orn caused an increase in cellular putrescine in both cell lines, arginine at high concentrations was inhibitory to putrescine accumulation. The addition of urea and glutamine had no effect on polyamines in either of the cell lines. Inhibition of glutamine synthetase by methionine sulfoximine led to a substantial reduction in putrescine and spermidine in both cell lines. The results show that: (a) Transgenic expression of a heterologous odc gene can be used to modulate putrescine metabolism in plant cells, (b) accumulation of putrescine in high amounts does not affect the native arginine decarboxylase activity, (c) Orn biosynthesis occurs primarily from glutamine/glutamate and not from catabolic breakdown of arginine, (d) Orn biosynthesis may become a limiting factor for putrescine production in the odc transgenic cells, and (e) assimilation of nitrogen into glutamine keeps pace with an increased demand for its use for putrescine production.     

Role of spermidine in the expression of late markers of adipose conversion. Effects of growth hormone.

Confluent Ob1771 cells treated with an inhibitor of spermidine and spermine synthesis, methylglyoxyal bis(guanylhydrazone), were dependent on putrescine addition for the expression of glycerol-3-phosphate dehydrogenase and acyl-CoA synthetase, which behaved as late markers of adipose conversion. A similar dependence was observed with drug-treated Ob17MT18 and 3T3-F442A preadipocyte cells, but not with non-differentiating 3T3-C2 cells. Studies in drug-treated Ob1771 cells at the mRNA level showed that the parallel expression of mRNAs encoding for glycerol-3-phosphate dehydrogenase and an homologue of serine proteinases of Mr 28,000 [Cook, Groves, Min & Spiegelman (1985) Proc. Natl. Acad. Sci. U.S.A. 82, 6480-6484] was also dependent on putrescine addition. Double-isotope experiments with [14C]putrescine and [3H]spermidine, as well as analysis of the polyamine content in drug-treated Ob1771 cells under various conditions, demonstrate after putrescine addition that the expression of late markers of adipose conversion was highly correlated with a 2-fold increase in the intracellular concentration of spermidine. No correlation was observed with changes in the intracellular concentrations of putrescine and spermine. Long-term exposure of untreated Ob1771 cells to growth hormone, which led to the expression of late markers of adipose conversion [Doglio, Dani, Grimaldi & Ailhaud (1986) Biochem. J. 238, 123-129] was also accompanied by the same increase in spermidine concentration, which attained values identical with those determined in drug-treated cells supplemented with putrescine. This observation suggests that the permissive effect of growth hormone on the terminal differentiation of adipose cells might e related to changes in the intracellular concentration of spermidine.     

Effect of temperature and diet on polyamine concentrations of the European sea bass (Dicentrarchus labrax L.)

• 1.1. A fall of environmental temperature causes a decrease in total polyamine concentrations of heart, red and white muscles of sea bass fed on a diet containing 70% herring meal (diet S).
• 2.2. When sea bass was fed with a diet partially replaced by casein (diet A), an increase of total polyamine concentration in liver and heart was observed at a lower temperature.
• 3.3. In all tissues studied an increase of putrescine concentrations and a parallel decrease of spermidine and spermidine levels were found for both groups S and A of sea bass when the temperature was lowered.
• 4.4. In general concentrations of putrescine, spermidine and spermine were considerably higher in group A when the temperature was lowered.

     

Changes in Polyamine Concentrations in Amygdaloid-Kindled Rats

Concentrations of the polyamines putrescine, spermidine, and spermine were investigated in the left and right amygdala and in the remaining cerebrum, in which kindling was induced by repeated application of electrical stimulation of the left amygdala of rats. In kindled rats, the concentrations of spermidine and spermine increased slightly, but elevations did not reach significant levels in any brain regions. The most profound increase was detected in the putrescine concentration in all parts of the cerebrum 1-8 h after the final stimulation. These results suggest that the increases in concentrations of polyamines, particularly of putrescine, are involved in the pathogenesis of amygdaloid kindling.     

THE DISTRIBUTION AND METABOLISM OF PUTRESCINE, SPERMIDINE AND SPERMINE INJECTED INTO THE CEREBRAL VENTRICLES OF RABBITS

Following the intracerebroventricular injection into rabbits of spermidine or spermine the highest concentrations were initially found in the caudate nucleus, hypothalamus and medulla. Subsequently there was a rapid decline in the amounts present in the caudate nucleus and hypothalamus and, particularly in the case of spermidine, an increase in the conccntration in the lower brain stem and cervical cord. This pattern of changes is consistent with the amines being redistributed by passage in CSF. Intraventricularly injected putrescine followed the same initial distribution pattern but within 2 days it had been largely converted to spermidine and spermine. Synthesized polyamines accumulated in all the regions examined. The time course of synthesis indicated that spermidine was the precursor of spermine. Spermine was also formed from injected spermidine and vice-versa. These findings concur with the pharmacological and neurotoxic actions of putrescine, spermidine and spermine.     

Polyamine levels during the development of zygotic and somatic embryos of Pinus radiata

Changes in the cellular content of three polyamines (putrescine, spermidine and spermine) were compared at different stages of development in zygotic and somatic embryos of Pinus radiata D. Don . During embryo development, both the zygotic and the somatic embryos showed a steady increase in spermidine content, with either a small decrease or no significant change in putrescine. This led to a several-fold increase in spermidine/putrescine ratios during development of both types of embryos. Cell cultures of plant-forming and non-plant-forming lines derived from the same clone and growing on proliferation (maintenance) medium differed significantly in their polyamine levels. Mature, cotyledonary stage somatic embryos capable of germination and formation of plants could be distinguished by their higher spermidine/putrescine ratios from abnormal cotyledonary stage somatic embryos which were incapable of forming plants.     

Relationships between polyamine metabolism and RNA synthesis in post-ischemic liver cell repair

In liver cells recovering from reversible ischemia the increase in RNA synthesis by isolated nuclei is preceded by activation of ornithine decarboxylase, leading in turn to an increase in putrescine concentration. Treatment of the animals with 1,3-diaminopropane and putrescine prevents ornithine decarboxylase activation but does not hinder the enhancement of RNA synthesis in post-ischemic liver nuclei; therefore, ornithine decarboxylase activation does not seem to be a necessary prerequisite for the increase in RNA synthesis. Hypophysectomy does not prevent the post-ischemic increases of ornithine decarboxylase and RNA synthesis; but pre-treatment of the animals with cycloheximide—which has a dual effect on the activity of ornithine decarboxylase—abolishes the post-ischemic enhancement of RNA synthesis. In contrast with regenerating liver, changes in ornithine decarboxylase activity and putrescine concentrations in reversible ischemia are not associated to changes in S-adenosylmethionine decarboxylase activity and in spermine and spermidine concentrations that seem to be characteristic of tissues where increases in RNA synthesis are followed by DNA synthesis and cell multiplication.