Acta Biologica Hungarica

, Volume 68, Issue 1, pp 1–13 | Cite as

Fine Structure and Synaptology of the Nitrergic Neurons in Medial Septum of the Rat Brain

  • Katalin HalasyEmail author
  • Balázs Szőke
  • Gergely Janzsó
Open Access


The nitrergic neuron population and certain aspects of their connectivity (peptidergic inputs, co-localization with GABA, synaptic target distribution) were studied in the medial septum of the rat brain. The histochemical localization of NADPH diaphorase and immunohistochemical identification of nNOS at light and electron microscopic level was applied. Double-labeling experiments with galanin and leucine enkephalin, moreover the postembedding GABA immunogold staining was also carried out. NADPH diaphorase- and nNOS-immunopositive neurons could be identified inside the borders of medial septum. Out of their peptidergic inputs galanin- and leucine enkephaline-immunopositive varicose fibers were found in close apposition with nNOS-immunopositive neurons. Based on fine structural characteristics (large indented nucleus, thin cytoplasmic rim, lack of axosomatic synapses) the nitrergic neurons are suggested to be identical with the septal cholinergic nerve cells. Their boutons established asymmetrical synapses mainly on dendritic shafts and spines, some of which were also nNOS-immunopositive. A lower amount of nNOS-immunopositive boutons of presumably extrinsic origin were found to be GABAergic.


nNOS NADPH neuropeptides GABA-synaptology 


  1. 1.
    Affleck, V. S., Coote, J. H., Pyner, S. (2012) The projection and synaptic organization of NTS afferent connections with presympathetic neurons, GABA and nNOS neurons in the paraventricular nucleus of the hypothalamus. Neuroscience 219, 48–61.PubMedPubMedCentralGoogle Scholar
  2. 2.
    Alonso, A., Köhler, C. (1984) A study of the reciprocal connections between the septum and the entorhinal area using anterograde and retrograde axonal transport methods in the rat brain. J. Comp. Neurol. 225, 327–343.PubMedGoogle Scholar
  3. 3.
    Amaral, D. G., Kurz, J. (1985) An analysis of the origins of the cholinergic and non-cholinergic septal projections to the hippocampal formation of the rat. J. Comp. Neurol. 240, 37–59.PubMedGoogle Scholar
  4. 4.
    Arendt, T., Bruckner, M. K., Bigl, V., Marcova, L. (1995) Dendritic reorganisation in the basal forebrain under degenerative conditions and its defects in Alzheimer’s disease. III. The basal forebrain compared with other subcortical areas, J. Comp. Neurol. 351, 223–246.PubMedGoogle Scholar
  5. 5.
    Atkinson, L., Batten, T. F. C., Corbett, K. A., Sinfield, J. K., Deuchars, J. (2003) Subcellular localization of neuronal nitric oxide synthase in the rat nucleus of the solitary tract in relation to vagal afferent inputs. Neuroscience 118, 115–122.PubMedGoogle Scholar
  6. 6.
    Baisden, R. H., Woodruff, M. L., Hoover, D. B. (1984) Cholinergic and non-cholinergic septo-hippocampal projections: a double-label horseradish peroxidase-acetylcholinesterase study in the rabbit. Brain Res. 290, 146–151.PubMedGoogle Scholar
  7. 7.
    Barbaresi, P., Mensa, E., Lariccia, V., Pugnaloni, A., Amoroso, S., Fabri, M. (2013) Differential distribution of parvalbumin- and calbindin-D28K-immunoreactive neurons in the rat periaqueductal gray matter and their colocalization with enzymes producing nitric oxide. Brain Res. Bull. 99, 48–62.PubMedGoogle Scholar
  8. 8.
    Bartus, R. T. Dean, R. L., Beer, III, B., Lippa, A. S. (1982) The cholinergic hypothesis of geriatric memory dysfunction. Science 217, 408–414.PubMedGoogle Scholar
  9. 9.
    Bernstein, H.-G., Brisch, R., Ogonlade, V., Heinemann, A., Baumann, B., Arendt, T., Dobrowolny, H., Bogerts, B., Lüth, H.-J. (2004) Detection of nitric oxide synthase (NOS) immunoreactive neurons in the human septal area: a matter of method? J. Chem. Neuroanat. 27, 247–250.PubMedGoogle Scholar
  10. 10.
    Bialowas, J., Frotsher, M. (1987) Choline acetyltransferase-immunoreactive neurons and terminals in the rat septal complex: combined light and electron microscopic study. J. Comp. Neurol. 259, 298–307.PubMedGoogle Scholar
  11. 11.
    Bland, B. H., Oddie, S. D. (1998) Anatomical, electrophysiological and pharmacological studies of ascending brain stem hippocampal synchronizing pathways, Neurosci. Biobehav. Rev. 22, 259–273.PubMedGoogle Scholar
  12. 12.
    Brashear, H. R., Zaborszky, L., Heimer, L. (1986) Distribution of GABAergic and cholinergic neurons in the rat diagonal band. Neuroscience 17, 439–451.PubMedGoogle Scholar
  13. 13.
    Bredt, D. S., Glatt, C. E., Hwang, P. M., Fotuhi, M., Dawson, T. M., Snyder, S. H. (1991) Nitric oxide synthase protein and mRNA are discreetly localized in neuronal populations of mammalian central nervous system together with NADPH diaphorase. Neuron 7, 615–624.PubMedGoogle Scholar
  14. 14.
    Bredt, D. S., Snyder, S. H. (1994) Nitric oxide: a physiologic messenger molecule. Annu. Rev. Biochem. 63, 175–195.PubMedGoogle Scholar
  15. 15.
    Cheng, A., Wang, S., Cai, J., Rao, M. S., Mattson, M. P. (2003) Nitric oxide acts in a positive feedback loop with BDNF to regulate neural progenitor cell proliferation and differentiation in the mammalian brain. Dev. Biol. 258, 319–333.PubMedGoogle Scholar
  16. 16.
    Colas, D., Gharib, A., Bezin, L., Morales, A., Guidon, G., Cespuglio, R., Sarda, N. (2006) Regional age-related changes in neuronal nitric oxide synthase (nNOS), messenger RNA levels and activity in SAMP8 brain. BMC Neuroscience 20067:81. DOI: 10.1186/1471-2202-7-81.Google Scholar
  17. 17.
    Dinerman, J. L., Dawson, T. M., Schell, M. J., Snowman, A., Snyder, S. H. (1994) Endothelial nitric oxide synthase localized to hippocampal pyramidal cells: implications for synaptic plasticity. Proc. Natl Acad. Sci. USA 91, 4214–4218.PubMedGoogle Scholar
  18. 18.
    Getting, S. J., Segieth, J., Ahmad, S., Biggs, C. S., Whitton, P. S. (1996) Biphasic modulation of GABA release by nitric oxide in the hippocampus of freely moving rats in vivo. Brain Res. 717, 196–199.PubMedGoogle Scholar
  19. 19.
    Guix, F. X., Uribesalgo, I., Coma, M., Munoz, F. J. (2005) The physiology and pathophysiology of nitric oxide in the brain. Prog. Neurobiol. 76, 126–152.PubMedGoogle Scholar
  20. 20.
    Hensley, K., Maidt, M. L., Yu, Z., Sang, H., Markesbery, W. R., Floyd, R. A. (1998) Electrochemical analysis of protein nitrotyrosine and dityrosine in the Alzheimer brain indicates region-specific accumulation. J. Neurosci. 18, 8126–8132.PubMedPubMedCentralGoogle Scholar
  21. 21.
    Hodgson, A. J., Penke, B., Erdei, A., Chubb, I. W., Somogyi, P. (1985) Antiserum to gamma-aminobutyric acid. I. Production and characterization using a new model system. J. Histochem. Cytochem. 33, 229–239.PubMedGoogle Scholar
  22. 22.
    Holttum, J. R., Gershon, S. (1992) The cholinergic model of dementia, Alzheimer type. Progression from the unitary transmitter concept. Dementia 3, 174–185.Google Scholar
  23. 23.
    Jakab, R. L., Leranth, Cs. (1995) Septum. In: Paxinos, G. (ed.) The Rat Nervous system. Academic Press, London, pp. 405–441.Google Scholar
  24. 24.
    Jinno, S., Kosaka, T. (2002) Immunocytochemical characterization of hippocamposeptal projecting GABAergic nonprincipal neurons in the mouse brain: a retrograde labeling study. Brain Res. 945, 219–231.PubMedGoogle Scholar
  25. 25.
    Kiss, J., Patel, A. J., Freund, T. F. (1990) Distribution of septohippocampal neurons containing parvalbumin or choline acetyltransferase in the rat brain. J. Comp. Neurol. 298, 362–372.PubMedGoogle Scholar
  26. 26.
    Kitchener, P. D., Diamond, J. (1993) Distribution and colocalization of choline acetyltransferase immunoreactivity and NADPH diaphorase reactivity in neurons within the medial septum and diagonal band of Broca in the rat basal forebrain. J. Comp. Neurol. 335, 1–15.PubMedGoogle Scholar
  27. 27.
    Köhler, C., Chan-Palay, V. (1983) Distribution of gamma-aminobutyric acid containing neurons and terminals in the septal area. Anat. Embryol. 167, 53–65.PubMedGoogle Scholar
  28. 28.
    Laing, I., Todd, A. J., Heizmann, C. W., Schmidt, H. H. H. W. (1994) Subpopulations of GABAergic neurons in laminae I–III of rat spinal dorsal horn defined by coexistence with classical transmitters, peptides, nitric oxide synthase or parvalbumin. Neuroscience 61, 123–132.PubMedGoogle Scholar
  29. 29.
    Lüth, H.-J. (1997) Ultrastructural demonstration of constitutive nitric oxide synthase (cNOS) in neocortical glial cells and glial perisynaptic sheaths. Ann. Anat. 179, 221–225.PubMedGoogle Scholar
  30. 30.
    Monti, J. M., Jantos, H. (2004) Effects of the 5-HT1A receptor ligands flesinoxan and WAY 100635 given systemically or microinjected into the laterodorsal tegmental nucleus on REM sleep in the rat. Behav. Brain Res. 151, 159–166.PubMedGoogle Scholar
  31. 31.
    Paspalas, C., Halasy, K., Gerics, B., Papadopoulo, G., Hajós, F. (2001) Vasoactive intestinal polypeptide in neuroglia? Immunoelectron microscopic localization in astrocytes of the rat mesencephalon. Glia 34, 229–233.PubMedGoogle Scholar
  32. 32.
    Pasqualotto, B. A., Vincent, S. R. (1991) Galanin and NADPH-diaphorase coexistence in cholinergic neurons of the rat basal forebrain. Brain Res. 551, 78–86.PubMedGoogle Scholar
  33. 33.
    Prast, H., Tran, M. H., Fischer, H., Philippu, A. (1998) Nitric oxide-induced release of acetylcholine in the nucleus accumbens: role of cyclic GMP, glutamate and GABA. J. Neurochem. 71, 266–273.PubMedGoogle Scholar
  34. 34.
    Prast, H., Philippu, A. (2001) Nitric oxide as modulator of neuronal function. Prog. Neurob. 64, 51–68.Google Scholar
  35. 35.
    Rodrigo, J., Springall, D. R., Uttenthal, O., Bentura, M. L., Abadia-Molina, F., Riveros-Moreno, V., Martinez-Murillo, R., Polak, J. M., Moncada, S. (1994) Localization of nitric oxide synthase in the adult rat brain. Philos. Trans R Soc. Lond B Biol. Sci. 345, 175–221.PubMedGoogle Scholar
  36. 36.
    Rye, D. B., Wainer, B. H., Mesulam, M.-M., Mufson, E. J., Saper, C. B. (1984) Cortical projections arising from the basal forebrain: a study of cholinergic and non-cholinergic components employing combined retrograde tracing and immunohistochemical localization of choline acetyltransferase. Neuroscience 13, 627–643.PubMedGoogle Scholar
  37. 37.
    Sardella, T. C. P., Polgár, E., Watanabe, M., Todd, J. (2011) A quantitative study of neuronal nitric oxide synthase expression in laminae I–III of the rat spinal dorsal horn. Neuroscience 192, 708–720.PubMedPubMedCentralGoogle Scholar
  38. 38.
    Sequeira, S. M., Ambrósio, A. F., Malva, J. O., Carvalho, A. P., Carvalho, C. M. (1997) Modulation of glutamate release from rat hippocampal synaptosomes by nitric oxide. Nitric Oxide 1, 315–329.Google Scholar
  39. 39.
    Sotty, F., Danik, M., Manseau, F., Laplante, F., Quirion, R., Williams, L. (2003) Distinct electrophysiological properties of glutamatergic, cholinergic and GABAergic rat septohippocampal neurons: novel implications for hippocampal rhythmicity, J. Physiol. (Lond.) 551, 927–943.Google Scholar
  40. 40.
    Szeidemann, Z., Shanabrough, M., Leranth, C. (1995) Hypothalamic Leu-enkephalin-immunoreactive fibers terminate on calbindin-containing somatospiny cells in the septal area of the rat. J. Comp. Neurol. 358, 573–583.PubMedGoogle Scholar
  41. 41.
    Tran, M. H., Yamada, K., Nakajima, A., Mizuno, M., He, J., Kamei, H., Nabeshima, T. (2003) Tyrosine nitration of a synaptic protein synaptophysin contributes to amyloid beta-peptide-induced cholinergic dysfunction. Mol. Psychiatry. 8, 407–412.PubMedGoogle Scholar
  42. 42.
    Yoo, J. H., Cho, J. H., Lee, S. Y., Loh, H. H., Ho, I. K., Jang, C. G. (2005) Reduced nNOS expression induced by repeated nicotine treatment in µ-opioid receptor knockout mice. Neurosci. Letters 380, 70–74.Google Scholar
  43. 43.
    Waltschanoff, J., Weinberg, R. J., Rustioni, A., Schmidt, H. H. H. W. (1992) Nitric oxide synthase and GABA colocalize in lamina II of spinal cord. Neurosci. Letters 148, 6–10.Google Scholar
  44. 44.
    Waltschanoff, J., Weinberg, R. J., Kharazia, V. N., Schmidt, H. H. H. W., Nakane, M., Rustioni, A. (1993) Neurons in rat cerebral cortex that synthesize nitroc oxide: NADPH diaphorase histochemistry, NOS immunocytochemistry and colocalization with GABA. Neurosci. Letters 157, 157–161.Google Scholar
  45. 45.
    Zochone, D. W., Hong Sun, Xia-Qing Li (2001) Evidence that nitric oxide- and opioid-containing interneurons innervate vessels in the dorsal horn of the spinal cord of rats. J. Physiol. 532.3, 749–758.Google Scholar

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© Akadémiai Kiadó, Budapest 2017

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Authors and Affiliations

  • Katalin Halasy
    • 1
    Email author
  • Balázs Szőke
    • 1
  • Gergely Janzsó
    • 1
  1. 1.Department of Anatomy and HistologyUniversity of Veterinary SciencesBudapestHungary

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