- Email: [email protected]
Immunohistochemical localization of l -Ornithine decarboxylase in developing rat brain
hr. 1. DC-V/. Neuroscknce. Vol. 5. No. 2. pp. 145-150.1987. Printed in Great Britain.
073h-.574x/87 $o3.tKl+o.oo Pergamon Journals Ltd. CQ 1987 ISDN
I~~~NOHISTOCHEMI~AL L-ORNITHINE DECARBOXYLASE A.
DoRN,*[[
M.
M~LLER,~
H.-G.
LOCALIZATION IN DEVELOPING
BERNSTEIN,*
A.
PAJUNEN$
and
M.
OF RAT
BRAIN
JARVINEN~
*Institute of Anatomy. Medical Academy Magdeburg, Leipziger Str. 44 DDR-3090 Magdeburg. G.D.R.; tlnstitute of Neurobiology and Brain Research of the Academy of Science, Magdeburg, G.D.R.; SDepartment of Biochemistry and IDepartment of Pathology. University of Oulu, Finland (Received
13 June 1986; in revised form
29 ikcmwber
19%; nccepted 8 ~u~uu~y 1987)
Abstract-i_-Ornithine decarboxylase, the rate limiting enzyme of polyamine biosynthesis and a marker enzyme of tissue proliferation and maturation, was localized immunocytochemically in the developing rat central nervous system. It can be noted that the distribution of the enzyme protein underlies temporal alterations. Conclusions are drawn from the location of the enzyme and possible functional roles played by ornithine decarboxylase in discrete brain areas. Key words: L-Ornithine
decarboxyiase,
Developing
central nervous system, Rat. lmmunohjstochemjstry.
The aliphatic di- and polyamines, putrescine, spermidine and spermine are thought to play an important role in tissue growth and development.4.‘4.‘h.2’.28 Ornithine decarboxylase (L-ornithine carboxylase, EC 4.1.1.17, ODC) catalyses the first and apparently rate limiting step in polyamine biosynthesis, that is, the conversion of L-ornithine to putrescine.“7.2” Evidence has been reported in the literature that ODC activity and polyamine concentration are high during periods of growth and cellular multiplication or differentiation, and decrease as these processes cease. “.‘i.m’X The ORC/polyamine system is believed to serve as a modulator of tissue growth during fetal and neonatal mammalian development, each organ having its specific pattern of development of these compounds. A considerable body of data has been collected in favor of an extraordinary role of ODC in neuronal development. “.“).3’-33 In the nervous system, there are close temporal relationships between ODC and polyamine development patterns,‘-2.‘0.32 cellular replication, migration and differentiation, biochemical development of biogenic amine transmitter systems”-7,“.2’.24.“’ and the onset of subsequent behavioral pattern. 2.‘0 Furthermore, ODC seems to be inducible by many trophic, growth-inducing factors such as nerve growth factor,‘” insulin2”.“” and others, which reach their maximal concentration in the first weeks of extrauterine life. Surprisingly, there is little knowledge about the precise topochemistry of ODC in neural structures during critical periods of peri- and postnatal development. We found it, therefore, of interest to investigate the distribution of the enzyme protein by use of an immunocytochemical approach. MATERIAL AND METHODS Juvenile Wistar rats (kept in colony, food and water were ad libidum) of both sexes (days 2-14 postnatally and adults) were killed by cervical dislocation and the brains were quickly removed. The tissue was fixed en bloc in Bouin’s fluid, dehydrated in increasing concentrations of ethanol, embedded in paraffin and cut into serial 4-6 Frn thick sections. This procedure was chosen, because (1) considerable parts of ODC immunoreactivity in kidney” and brain (unpublished) survive this treatment and (2) preservation of morphological detail is superior. For the immunohistochemical identification of ODC the horseradish peroxidase-antiperoxidase (PAP) complex technique after Sternberger et al.‘” was performed. The primary antibody (rabbit antimouse ODC, diluted 1:2OO) was applied for 1 hr at 20°C or 12 hr at 4”C, respectively. Purification of mouse kidney ODC and production of the antiserum have been described previously. IfiThe antiserum has been shown to cross-react with rat ODC. The secondary antibody (swine-rabbit IgG, Dakopatts Denmark) and the PAP complex (Dakopatts Denmark) were applied as usual. The // Author
to whom correspondence
should be addressed.
145
146
A.
Dorn PI al.
peroxidase activity was visualized with 3,3-diaminobenzidine.” For purposes of control, the primary antibody was either replaced by buffer or by normal rabbit serum. Alternatively, the antiserum was substituted by anti-ODC preabsorbed with pure ODC. The sections were investigated under a light microscope.
RESULTS At postnatal days 2-6 immunoreactive ODC was nearly ubiquitously distributed, i.e. there were no regional differences with regard to the intensity or the distribution pattern. Beginning with day 7 of postnatal life the intensity of immunoreaction declines rapidly in most brain regions. Only in the basal ganglia, the septal nuclei (Fig. l), the nucleus ambiguus (Fig. 2) and the nucleus reticularis lateralis magnocellularis there could be found a stronger immunoreactivity. Between postnatal days 10 and 12 neurons of the hippocampal formation showed a faint staining (Fig. 3). In the cerebellum (only investigated at days 10-14 postnatally) enzyme protein could be detected in the ganglionic layer: the Purkinje cells and their dendrites show immunoreactivity. Some cells of the internal granular cell layer possess a weak immunoreactivity (Fig. 4). Adult rat brain completely lacks in ODC immunoreactive material. It should be emphasized, however, that our data is valid only with regard to the localization of the ODC protein. Our findings do not support conclusive statements regarding the actual enzymatic activity of brainlocated ODC. Furthermore, we have as yet no concrete informaton about the influence of ODC antizyme”.” on the interaction of ODC with ODC antiserum, which might cause some disturbances in the immunocytochemical demonstration of ODC. Previous in vitro studies did not reveal such an effect, however.
DISCUSSION In the present study we were able to localize ODC-immunoreactivity in the developing rat brain by means of immunohistochemistry. Using biochemical methods (activity assays) many data have been collected about the developmental changes of ODC activity’4.‘y and its regulators such as antizymel7.1X.25.32.34 in the nervous system and in other tissues.J.H.‘.l It has been established that ODC activity in the nervous tissue of mouse and rat is highest during birth and then declines permanently, reaching the low adult activity level after nearly three weeks of postnatal life.’ Our findings of a rather weak, but ubiquitously distributed ODC immunoreactivity in the first week after birth correlates with these biochemical endings. The remaining ODC immuoreactivity in well-defined brain areas after postnatal day 7 might be connected with such processes as synaptogenesis and development of specific receptor patterns. The great importance of ODC in the development of the cerebellum has been shown by use of (Ydifluoromethylornithine, an irreversible inhibitor of 0DC.3 Postnatal treatment with the inhibitor leads to an impaired development of the cerebellar cortex. These effects were already clearly visible at IO-15 days of age. As shown by autoradiography using [“H]cw-difluoromethylornithine, considerable amounts of the enzyme are present in the molecular layer of the cerebellum at day 9 of postnatal life.22 Besides the mossy and climbing fibers of the cerebellum, Fig. 1. ODC
immunoreactivity
Fig. 2. ODC
enzyme
Fig. 3. The hippocampal
protein
in the caudate nucleus of a 7-day-old x 400. localized
formation
in the Nut.
rat. x 250. Insert: the same region.
ambiguus of the rat (postnatal
of an Il-day-old rat, possessing ODC x 250.
day 9).
immunoreactive
X 400. material.
Fig. 4. Rat cerebellum (postnatal day 1 hr). Strong ODC immunoreactivity in the Purkinje cells and their dendrites. Some cells of the internal granular layer show a weak immunoreaction. X 300. Fig. 5. Microphotographs
of a control
reaction - antigen absorbed ODC postnatal day 7. x 250.
antisera.
Caudate
nucleus at
ODC in developing rat brain
Figs 1-5.
147
ODC in developing rat brain
149
the molecular layer represents the dendritic trees of the Purkinje cells. Our data of a strong dendritic immunostaining supports the finding by these authors; they support the finding of strong immunoreactivity in the Purkinje cell layer and weak immuoreactivity in the granular cell layer. This preliminary study shows that ODC immunohistochemistry is a reasonable tool for studies of ODC distribution and thereby its action in different brain regions at different stages of time including prenatal life. REFERENCES I. 2. 3. 4.
5. 6. 7. 8. 9. IO. 1 I. 12.
13. 14. 15.
16. 17. 18. 19. 20. 21. 22. 23. 24. 25. 26. 27. 28. 29. 30.
Anderson T. R. and Schanberg S. M. (1979) Ornithine decarboxylase activity in developing rat brain. J. Neurochem. 19, 1471-1481. Anderson T. R. and Schanberg S. M. (1975) Effect of thyroxine and cortisol on brain ornithine decarboxylase activity and swimming behavior in developing rat. Riochem. Pharmac. 24, 495-501. Bartolome .I. V.. Schweitzer L., Slotkin T. A. and Nadler J. V. (1985) Impaired development of cerebellar cortex in rats treated postnatally with a-difluoromethylornithine. Neuroscience 15, 203-213. Canellakis E. S., Viceps-Madore D., Kyriakides D. A. and Heller J. S. (1979) The regulation and function of ornithine decarboxylase and of the polyamines. In Currerrr Topics in Celhdar Regularion, Vol. I5 (eds Horecker B. L. and Stadtman E. R.), pp. 156-202. Academic Press. New York. Chandhuri D., Chandhuri I. and Mukkerjea M. (1983) Ontogeny of polyamines in relation to nucleic acids in brain and spinal cord of the developing human fetus. Devl Brain Res. 10, 143-145. Coyle J. T. and Allrod J. (1971) Development of the uptake and storage of L-(‘H) norepinephrine in rat brain. J. Neurochem. IS, 2061-2075. Coyle J. T. and Snyder S. M. (1969) Catecholamine uptake by synaptosomes in homogenates of the brain: stereospecificity in different areas. J. Pharmac. ewp. Ther. 170, 221-231. Dorn A., Miiller M., Berstein H.-G., Pajunen A. and Jarvinen M. (1985) Immunohistochemical demonstration of ornithine decarboxylase in developing rat kidney. Actu Histochem. Cyrochem. 18, 337-341. Fossard J. R., Part M. L., Prahash N. J., Grove J.. Schechter P. J., Sjoerdsma A and Koch-Weser J. (1980) LOrnithine decarboxylase: an essential role in early mammalian embryogenesis. Science 208, 505-508. Genedani S., Bernardi M. and Bertolini A. (1985) Developmental and behavioral outcomes of perinatal inhibition of ornithine decarboxylase. Neurohehav. Toxicol. Teratol. 7, 57-65. Gilad M. and Kopin 1. J. (1979) Neurochemical aspects of neuronal ontogenesis in the developing rat cerebellum: changes in neurotransmitter and polyamine synthesising enzymes. J. Neurochem. 33, 1195-2004. Graham R. C. and Karnovsky N. J. (1966) The early stage of absorption of injected horseradish peroxidase in the proximal tubules of mouse kidney. Ultrastructural cytochemistry by a new technique. J. Histochem. Cyrochem. 14, 291-303. Heby 0. (1981) Role of polyamines in the control of cell proliferation and differentiation. Differentiation 19, l-20. Hietala 0. L. (1984) Regulatory aspects of polyamine metabolism in mouse brain. Acfa Universitafis Oulunensis A 158. pp. I-51. Oulu. Isomaa V. V., Pajunen A. E. I., Bardin L. H. and Janne 0. A. (1983) Ornithine decarboxylase in mouse kidney. Purification, characterisation, and radioimmunological determination of the enzyme protein. J. biol. Chem. 258, 6735-6740. Janne J.. P&ii M. and Raina A. (1978) Polyamines in rapid growth and cancer. Riochim. Biophys. ACIU 473, 241293. Laitinen P. H. (1965) Involvement of an “antizyme’ in the inactivation of ornithine decarboxylase. J. Neurochem. 45, 1303-1307. Laitinen P. H., Huhtinen D. L., Hietala D. A. and Pajunen A. E. I. (1985) Ornithine decarboxylase activity in brain regulated by a specific macromolecule, the antizyme. J. Neurochem. 44, 1885-1891. Laitinen S. I., Laitinen P. H., Hietala 0. L., Pajunen A. E. I. and Piha R. S. (1982) Developmental changes in mouse brain polyamine metabolism. Neurochemistry 7, 1477-1485. Lewis M. E., Lakshmanan J., Nagliah K., MacDomell P. C. and Guroff G (1978) Nerve growth factor increases activity of ornithine decarboxylase in rat brain. Proc. nam. Acad. Sci. U.S.A. 75, 1621-1623. Morris D. R. and Fillingame R. H. (1974) Regulation of amine acid decarboxylation. Ann. Rev. Biochem. 43, 30s 325. Morris G., Nadler J. V. and Slotkin T. A. (1986) Autoradiographic localization of ornithine decarboxylase in cerebellar cortex of the developing rat with (‘H)u-difluoromethylornithine. Neuroscience 17, 183-188. Morris G., Seidler F. E. and Slotkin T. A. (1983) Stimulation of ornithine decarboxylase by histamine or norepinephrine in brain regions of the developing rat. Lqe Sci. 32, 1565-1571. Morris G. and Slotkin T. A. (1985) Beta-2 adrenergic control of ornithine decarboxylase activity in brain regions of the developing rat. J. Pharmac. exp. Ther. 233, 141-147. Pajunen A. E. I. (1979) Polyamine metabolism in mouse brain. Interrelations between cerebral polyamines and yaminobutyric acid. Acfa Universifatis Oulunensis 79 (23), pp. 147, Oulu. Parker K. and Vernadakis A. (1980) Stimulation of ornithine decarboxylase activity in neural cell culture. Potential role of insulin. J. Neurochem. 35, 155-163. Pegg A. E. and Williams-Ashman M. G. (1968) Biosynthesis of putrescine in the prostate gland of the rat. Biochem. J. 100, 533-539. Russel D. H. (1980) Ornithine decarboxylase as a biological and pharmacological tool. Pharmacology 20, 117-129. Russel D. M. and Snyder S. H. (1968) Amine synthesis in rapidly growing tissues: ornithine decarboxylase activity in regenerating rat liver, chick embryo and various tissues. Proc. nam. Acud. Sci. U.S.A. 60, 1420-1227. Sargent-Jones L., Gauger L. L., Davis J. N., Slotkin D. A. and Bartolome J. V. (1985) Postnatal development of brain 8-adrenergic receptors: in vitro autoradiography with (‘*.‘I) HEAT in normal rat and rat treated with odifluoromethylornithine, a specific irreversible inhibitor of ornithine decarboxylase. Neuroscience 15, 1195-1202.
150
A.
Dorn
et al.
31. Seiler K., Skashan S. and Roth-Schechter B. F. (19X4) Polyamines and the development of isolated neurones in development neurobiology. Life Sci. 24, 1623-1630. 32. Skashan E. G., Haraszti J. H. and Snyder S. M. (1973) Polyamines: Developmental alterations in regional disposition and metabolism in rat brain. J. Neurochem. 20, 1443-1452. 33. Slotkin T. A. (1979) Ornithine decarboxylase as a tool in developmental neurobiology. Life Sri. 24, 162.%1630. 34. Sternberger L. A., Hardy P. M.. Cuculis J. J. and Meyer M. G. (1970) The unlabeled antibody enzyme method of immunohistochemistry. Preparation and properties of soluble antigen-antibody complex (horseradish peroxidaseantihorseradish peroxidase and its use in identification of spirochetes. J. Hisrochem. Cyrachem. 18, 315-333. 35. Yang J. W.. Raizada M. U. and Fellows R. E. (1981) Effects of insulin on cultured rat brain cells. Stimulation of ornithine decarboxylase activity. J. Neurochem. 36, 105&-1070.
073h-.574x/87 $o3.tKl+o.oo Pergamon Journals Ltd. CQ 1987 ISDN
I~~~NOHISTOCHEMI~AL L-ORNITHINE DECARBOXYLASE A.
DoRN,*[[
M.
M~LLER,~
H.-G.
LOCALIZATION IN DEVELOPING
BERNSTEIN,*
A.
PAJUNEN$
and
M.
OF RAT
BRAIN
JARVINEN~
*Institute of Anatomy. Medical Academy Magdeburg, Leipziger Str. 44 DDR-3090 Magdeburg. G.D.R.; tlnstitute of Neurobiology and Brain Research of the Academy of Science, Magdeburg, G.D.R.; SDepartment of Biochemistry and IDepartment of Pathology. University of Oulu, Finland (Received
13 June 1986; in revised form
29 ikcmwber
19%; nccepted 8 ~u~uu~y 1987)
Abstract-i_-Ornithine decarboxylase, the rate limiting enzyme of polyamine biosynthesis and a marker enzyme of tissue proliferation and maturation, was localized immunocytochemically in the developing rat central nervous system. It can be noted that the distribution of the enzyme protein underlies temporal alterations. Conclusions are drawn from the location of the enzyme and possible functional roles played by ornithine decarboxylase in discrete brain areas. Key words: L-Ornithine
decarboxyiase,
Developing
central nervous system, Rat. lmmunohjstochemjstry.
The aliphatic di- and polyamines, putrescine, spermidine and spermine are thought to play an important role in tissue growth and development.4.‘4.‘h.2’.28 Ornithine decarboxylase (L-ornithine carboxylase, EC 4.1.1.17, ODC) catalyses the first and apparently rate limiting step in polyamine biosynthesis, that is, the conversion of L-ornithine to putrescine.“7.2” Evidence has been reported in the literature that ODC activity and polyamine concentration are high during periods of growth and cellular multiplication or differentiation, and decrease as these processes cease. “.‘i.m’X The ORC/polyamine system is believed to serve as a modulator of tissue growth during fetal and neonatal mammalian development, each organ having its specific pattern of development of these compounds. A considerable body of data has been collected in favor of an extraordinary role of ODC in neuronal development. “.“).3’-33 In the nervous system, there are close temporal relationships between ODC and polyamine development patterns,‘-2.‘0.32 cellular replication, migration and differentiation, biochemical development of biogenic amine transmitter systems”-7,“.2’.24.“’ and the onset of subsequent behavioral pattern. 2.‘0 Furthermore, ODC seems to be inducible by many trophic, growth-inducing factors such as nerve growth factor,‘” insulin2”.“” and others, which reach their maximal concentration in the first weeks of extrauterine life. Surprisingly, there is little knowledge about the precise topochemistry of ODC in neural structures during critical periods of peri- and postnatal development. We found it, therefore, of interest to investigate the distribution of the enzyme protein by use of an immunocytochemical approach. MATERIAL AND METHODS Juvenile Wistar rats (kept in colony, food and water were ad libidum) of both sexes (days 2-14 postnatally and adults) were killed by cervical dislocation and the brains were quickly removed. The tissue was fixed en bloc in Bouin’s fluid, dehydrated in increasing concentrations of ethanol, embedded in paraffin and cut into serial 4-6 Frn thick sections. This procedure was chosen, because (1) considerable parts of ODC immunoreactivity in kidney” and brain (unpublished) survive this treatment and (2) preservation of morphological detail is superior. For the immunohistochemical identification of ODC the horseradish peroxidase-antiperoxidase (PAP) complex technique after Sternberger et al.‘” was performed. The primary antibody (rabbit antimouse ODC, diluted 1:2OO) was applied for 1 hr at 20°C or 12 hr at 4”C, respectively. Purification of mouse kidney ODC and production of the antiserum have been described previously. IfiThe antiserum has been shown to cross-react with rat ODC. The secondary antibody (swine-rabbit IgG, Dakopatts Denmark) and the PAP complex (Dakopatts Denmark) were applied as usual. The // Author
to whom correspondence
should be addressed.
145
146
A.
Dorn PI al.
peroxidase activity was visualized with 3,3-diaminobenzidine.” For purposes of control, the primary antibody was either replaced by buffer or by normal rabbit serum. Alternatively, the antiserum was substituted by anti-ODC preabsorbed with pure ODC. The sections were investigated under a light microscope.
RESULTS At postnatal days 2-6 immunoreactive ODC was nearly ubiquitously distributed, i.e. there were no regional differences with regard to the intensity or the distribution pattern. Beginning with day 7 of postnatal life the intensity of immunoreaction declines rapidly in most brain regions. Only in the basal ganglia, the septal nuclei (Fig. l), the nucleus ambiguus (Fig. 2) and the nucleus reticularis lateralis magnocellularis there could be found a stronger immunoreactivity. Between postnatal days 10 and 12 neurons of the hippocampal formation showed a faint staining (Fig. 3). In the cerebellum (only investigated at days 10-14 postnatally) enzyme protein could be detected in the ganglionic layer: the Purkinje cells and their dendrites show immunoreactivity. Some cells of the internal granular cell layer possess a weak immunoreactivity (Fig. 4). Adult rat brain completely lacks in ODC immunoreactive material. It should be emphasized, however, that our data is valid only with regard to the localization of the ODC protein. Our findings do not support conclusive statements regarding the actual enzymatic activity of brainlocated ODC. Furthermore, we have as yet no concrete informaton about the influence of ODC antizyme”.” on the interaction of ODC with ODC antiserum, which might cause some disturbances in the immunocytochemical demonstration of ODC. Previous in vitro studies did not reveal such an effect, however.
DISCUSSION In the present study we were able to localize ODC-immunoreactivity in the developing rat brain by means of immunohistochemistry. Using biochemical methods (activity assays) many data have been collected about the developmental changes of ODC activity’4.‘y and its regulators such as antizymel7.1X.25.32.34 in the nervous system and in other tissues.J.H.‘.l It has been established that ODC activity in the nervous tissue of mouse and rat is highest during birth and then declines permanently, reaching the low adult activity level after nearly three weeks of postnatal life.’ Our findings of a rather weak, but ubiquitously distributed ODC immunoreactivity in the first week after birth correlates with these biochemical endings. The remaining ODC immuoreactivity in well-defined brain areas after postnatal day 7 might be connected with such processes as synaptogenesis and development of specific receptor patterns. The great importance of ODC in the development of the cerebellum has been shown by use of (Ydifluoromethylornithine, an irreversible inhibitor of 0DC.3 Postnatal treatment with the inhibitor leads to an impaired development of the cerebellar cortex. These effects were already clearly visible at IO-15 days of age. As shown by autoradiography using [“H]cw-difluoromethylornithine, considerable amounts of the enzyme are present in the molecular layer of the cerebellum at day 9 of postnatal life.22 Besides the mossy and climbing fibers of the cerebellum, Fig. 1. ODC
immunoreactivity
Fig. 2. ODC
enzyme
Fig. 3. The hippocampal
protein
in the caudate nucleus of a 7-day-old x 400. localized
formation
in the Nut.
rat. x 250. Insert: the same region.
ambiguus of the rat (postnatal
of an Il-day-old rat, possessing ODC x 250.
day 9).
immunoreactive
X 400. material.
Fig. 4. Rat cerebellum (postnatal day 1 hr). Strong ODC immunoreactivity in the Purkinje cells and their dendrites. Some cells of the internal granular layer show a weak immunoreaction. X 300. Fig. 5. Microphotographs
of a control
reaction - antigen absorbed ODC postnatal day 7. x 250.
antisera.
Caudate
nucleus at
ODC in developing rat brain
Figs 1-5.
147
ODC in developing rat brain
149
the molecular layer represents the dendritic trees of the Purkinje cells. Our data of a strong dendritic immunostaining supports the finding by these authors; they support the finding of strong immunoreactivity in the Purkinje cell layer and weak immuoreactivity in the granular cell layer. This preliminary study shows that ODC immunohistochemistry is a reasonable tool for studies of ODC distribution and thereby its action in different brain regions at different stages of time including prenatal life. REFERENCES I. 2. 3. 4.
5. 6. 7. 8. 9. IO. 1 I. 12.
13. 14. 15.
16. 17. 18. 19. 20. 21. 22. 23. 24. 25. 26. 27. 28. 29. 30.
Anderson T. R. and Schanberg S. M. (1979) Ornithine decarboxylase activity in developing rat brain. J. Neurochem. 19, 1471-1481. Anderson T. R. and Schanberg S. M. (1975) Effect of thyroxine and cortisol on brain ornithine decarboxylase activity and swimming behavior in developing rat. Riochem. Pharmac. 24, 495-501. Bartolome .I. V.. Schweitzer L., Slotkin T. A. and Nadler J. V. (1985) Impaired development of cerebellar cortex in rats treated postnatally with a-difluoromethylornithine. Neuroscience 15, 203-213. Canellakis E. S., Viceps-Madore D., Kyriakides D. A. and Heller J. S. (1979) The regulation and function of ornithine decarboxylase and of the polyamines. In Currerrr Topics in Celhdar Regularion, Vol. I5 (eds Horecker B. L. and Stadtman E. R.), pp. 156-202. Academic Press. New York. Chandhuri D., Chandhuri I. and Mukkerjea M. (1983) Ontogeny of polyamines in relation to nucleic acids in brain and spinal cord of the developing human fetus. Devl Brain Res. 10, 143-145. Coyle J. T. and Allrod J. (1971) Development of the uptake and storage of L-(‘H) norepinephrine in rat brain. J. Neurochem. IS, 2061-2075. Coyle J. T. and Snyder S. M. (1969) Catecholamine uptake by synaptosomes in homogenates of the brain: stereospecificity in different areas. J. Pharmac. ewp. Ther. 170, 221-231. Dorn A., Miiller M., Berstein H.-G., Pajunen A. and Jarvinen M. (1985) Immunohistochemical demonstration of ornithine decarboxylase in developing rat kidney. Actu Histochem. Cyrochem. 18, 337-341. Fossard J. R., Part M. L., Prahash N. J., Grove J.. Schechter P. J., Sjoerdsma A and Koch-Weser J. (1980) LOrnithine decarboxylase: an essential role in early mammalian embryogenesis. Science 208, 505-508. Genedani S., Bernardi M. and Bertolini A. (1985) Developmental and behavioral outcomes of perinatal inhibition of ornithine decarboxylase. Neurohehav. Toxicol. Teratol. 7, 57-65. Gilad M. and Kopin 1. J. (1979) Neurochemical aspects of neuronal ontogenesis in the developing rat cerebellum: changes in neurotransmitter and polyamine synthesising enzymes. J. Neurochem. 33, 1195-2004. Graham R. C. and Karnovsky N. J. (1966) The early stage of absorption of injected horseradish peroxidase in the proximal tubules of mouse kidney. Ultrastructural cytochemistry by a new technique. J. Histochem. Cyrochem. 14, 291-303. Heby 0. (1981) Role of polyamines in the control of cell proliferation and differentiation. Differentiation 19, l-20. Hietala 0. L. (1984) Regulatory aspects of polyamine metabolism in mouse brain. Acfa Universitafis Oulunensis A 158. pp. I-51. Oulu. Isomaa V. V., Pajunen A. E. I., Bardin L. H. and Janne 0. A. (1983) Ornithine decarboxylase in mouse kidney. Purification, characterisation, and radioimmunological determination of the enzyme protein. J. biol. Chem. 258, 6735-6740. Janne J.. P&ii M. and Raina A. (1978) Polyamines in rapid growth and cancer. Riochim. Biophys. ACIU 473, 241293. Laitinen P. H. (1965) Involvement of an “antizyme’ in the inactivation of ornithine decarboxylase. J. Neurochem. 45, 1303-1307. Laitinen P. H., Huhtinen D. L., Hietala D. A. and Pajunen A. E. I. (1985) Ornithine decarboxylase activity in brain regulated by a specific macromolecule, the antizyme. J. Neurochem. 44, 1885-1891. Laitinen S. I., Laitinen P. H., Hietala 0. L., Pajunen A. E. I. and Piha R. S. (1982) Developmental changes in mouse brain polyamine metabolism. Neurochemistry 7, 1477-1485. Lewis M. E., Lakshmanan J., Nagliah K., MacDomell P. C. and Guroff G (1978) Nerve growth factor increases activity of ornithine decarboxylase in rat brain. Proc. nam. Acad. Sci. U.S.A. 75, 1621-1623. Morris D. R. and Fillingame R. H. (1974) Regulation of amine acid decarboxylation. Ann. Rev. Biochem. 43, 30s 325. Morris G., Nadler J. V. and Slotkin T. A. (1986) Autoradiographic localization of ornithine decarboxylase in cerebellar cortex of the developing rat with (‘H)u-difluoromethylornithine. Neuroscience 17, 183-188. Morris G., Seidler F. E. and Slotkin T. A. (1983) Stimulation of ornithine decarboxylase by histamine or norepinephrine in brain regions of the developing rat. Lqe Sci. 32, 1565-1571. Morris G. and Slotkin T. A. (1985) Beta-2 adrenergic control of ornithine decarboxylase activity in brain regions of the developing rat. J. Pharmac. exp. Ther. 233, 141-147. Pajunen A. E. I. (1979) Polyamine metabolism in mouse brain. Interrelations between cerebral polyamines and yaminobutyric acid. Acfa Universifatis Oulunensis 79 (23), pp. 147, Oulu. Parker K. and Vernadakis A. (1980) Stimulation of ornithine decarboxylase activity in neural cell culture. Potential role of insulin. J. Neurochem. 35, 155-163. Pegg A. E. and Williams-Ashman M. G. (1968) Biosynthesis of putrescine in the prostate gland of the rat. Biochem. J. 100, 533-539. Russel D. H. (1980) Ornithine decarboxylase as a biological and pharmacological tool. Pharmacology 20, 117-129. Russel D. M. and Snyder S. H. (1968) Amine synthesis in rapidly growing tissues: ornithine decarboxylase activity in regenerating rat liver, chick embryo and various tissues. Proc. nam. Acud. Sci. U.S.A. 60, 1420-1227. Sargent-Jones L., Gauger L. L., Davis J. N., Slotkin D. A. and Bartolome J. V. (1985) Postnatal development of brain 8-adrenergic receptors: in vitro autoradiography with (‘*.‘I) HEAT in normal rat and rat treated with odifluoromethylornithine, a specific irreversible inhibitor of ornithine decarboxylase. Neuroscience 15, 1195-1202.
150
A.
Dorn
et al.
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