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. 1977 Dec;74(12):5767–5771. doi: 10.1073/pnas.74.12.5767

Determination of transmitter function by neuronal activity

Patricia A Walicke 1, Robert B Campenot 1, Paul H Patterson 1
PMCID: PMC431875  PMID: 272002

Abstract

The role of neuronal activity in the determination of transmitter function was studied in cultures of dissociated sympathetic neurons from newborn rat superior cervical ganglia. Cholinergic and adrenergic differentiation were assayed by incubating the cultures with radioactive choline and tyrosine and determining the rate of synthesis and accumulation of labelled acetylcholine and catecholamines. As in previous studies, pure neuronal cultures grown in control medium displayed much lower ratios of acetylcholine synthesis to catecholamine synthesis than did sister cultures grown in medium previously conditioned by incubation on appropriate nonneuronal cells (conditioned medium). However, here we report that neurons treated with the depolarizing agents elevated K+ or veratridine, or stimulated directly with electrical current, either before or during application of conditioned medium, displayed up to 300-fold lower acetylcholine/catecholamine ratios than they would have without depolarization, and thus remained primarily adrenergic. Elevated K+ and veratridine produced this effect on cholinergic differentiation without significantly altering neuronal survival. Because depolarization causes Ca2+ entry in a number of cell types, the effects of several Ca2+ agonists and antagonists were investigated. In the presence of the Ca2+ antagonists D600 or Mg2+, K+ did not prevent the induction of cholinergic properties by conditioned medium. Thus depolarization, either steady or accompanying activity, is one of the factors determining whether cultured sympathetic neurons become adrenergic or cholinergic, and this effect may be mediated by Ca2+.

Keywords: sympathetic neurons, cell culture, catecholamines, acetylcholine, development

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Selected References

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  1. Aiken J. W., Reit E. A comparison of the sensitivity to chemical stimuli of adrenergic and cholinergic neurons in the cat stellate ganglion. J Pharmacol Exp Ther. 1969 Oct;169(2):211–223. [PubMed] [Google Scholar]
  2. Baker P. F., Rink T. J. Catecholamine release from bovine adrenal medulla in response to maintained depolarization. J Physiol. 1975 Dec;253(2):593–620. doi: 10.1113/jphysiol.1975.sp011209. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Black I. B., Geen S. C. Inhibition of the biochemical and morphological maturation of adrenergic neurons by nicotinic receptor blockade. J Neurochem. 1974 Feb;22(2):301–306. doi: 10.1111/j.1471-4159.1974.tb11594.x. [DOI] [PubMed] [Google Scholar]
  4. Black I. B., Geen S. C. Trans-synaptic regulation of adrenergic neuron development: inhibition by ganglionic blockade. Brain Res. 1973 Dec 7;63:291–302. doi: 10.1016/0006-8993(73)90096-6. [DOI] [PubMed] [Google Scholar]
  5. Black I. B., Mytilineou C. Trans-synaptic regulation of the development of end organ innervation by sympathetic neurons. Brain Res. 1976 Jan 23;101(3):503–521. doi: 10.1016/0006-8993(76)90474-1. [DOI] [PubMed] [Google Scholar]
  6. Brown D. A., Scholfield C. N. Movements of labelled sodium ions in isolated rat superior cervical ganglia. J Physiol. 1974 Oct;242(2):321–351. doi: 10.1113/jphysiol.1974.sp010710. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Campenot R. B. Local control of neurite development by nerve growth factor. Proc Natl Acad Sci U S A. 1977 Oct;74(10):4516–4519. doi: 10.1073/pnas.74.10.4516. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. FATT P., GINSBORG B. L. The ionic requirements for the production of action potentials in crustacean muscle fibres. J Physiol. 1958 Aug 6;142(3):516–543. doi: 10.1113/jphysiol.1958.sp006034. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Furshpan E. J., MacLeish P. R., O'Lague P. H., Potter D. D. Chemical transmission between rat sympathetic neurons and cardiac myocytes developing in microcultures: evidence for cholinergic, adrenergic, and dual-function neurons. Proc Natl Acad Sci U S A. 1976 Nov;73(11):4225–4229. doi: 10.1073/pnas.73.11.4225. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Greene L. A., Tischler A. S. Establishment of a noradrenergic clonal line of rat adrenal pheochromocytoma cells which respond to nerve growth factor. Proc Natl Acad Sci U S A. 1976 Jul;73(7):2424–2428. doi: 10.1073/pnas.73.7.2424. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hendry I. A. Trans-synaptic regulation of tyrosine hydroxylase activity in a developing mouse sympathetic ganglion: effects of nerve growth factor (NGF), NGF-antiserum and pempidine. Brain Res. 1973 Jun 29;56:313–320. doi: 10.1016/0006-8993(73)90344-2. [DOI] [PubMed] [Google Scholar]
  12. Jansen J. K., Lomo T., Nicolaysen K., Westgaard R. H. Hyperinnervation of skeletal muscle fibers: dependence on muscle activity. Science. 1973 Aug 10;181(4099):559–561. doi: 10.1126/science.181.4099.559. [DOI] [PubMed] [Google Scholar]
  13. Johnson M., Ross D., Meyers M., Rees R., Bunge R., Wakshull E., Burton H. Synaptic vesicle cytochemistry changes when cultured sympathetic neurones develop cholinergic interactions. Nature. 1976 Jul 22;262(5566):308–310. doi: 10.1038/262308a0. [DOI] [PubMed] [Google Scholar]
  14. Ko C. P., Burton H., Bunge R. P. Synaptic transmission between rat spinal cord explants and dissociated superior cervical ganglion neurons in tissue culture. Brain Res. 1976 Dec 3;117(3):437–460. doi: 10.1016/0006-8993(76)90752-6. [DOI] [PubMed] [Google Scholar]
  15. Ko C. P., Burton H., Johnson M. I., Bunge R. P. Synaptic transmission between rat superior cervical ganglion neurons in dissociated cell cultures. Brain Res. 1976 Dec 3;117(3):461–485. doi: 10.1016/0006-8993(76)90753-8. [DOI] [PubMed] [Google Scholar]
  16. Kohlhardt M., Bauer B., Krause H., Fleckenstein A. Differentiation of the transmembrane Na and Ca channels in mammalian cardiac fibres by the use of specific inhibitors. Pflugers Arch. 1972;335(4):309–322. doi: 10.1007/BF00586221. [DOI] [PubMed] [Google Scholar]
  17. Larrabee M. G. Metabolism of adult and embryonic sympathetic ganglia. Fed Proc. 1970 Nov-Dec;29(6):1919–1928. [PubMed] [Google Scholar]
  18. Lomo T., Westgaard R. H., Dahl H. A. Contractile properties of muscle: control by pattern of muscle activity in the rat. Proc R Soc Lond B Biol Sci. 1974 Aug 27;187(1086):99–103. doi: 10.1098/rspb.1974.0064. [DOI] [PubMed] [Google Scholar]
  19. Lomo T., Westgaard R. H. Further studies on the control of ACh sensitivity by muscle activity in the rat. J Physiol. 1975 Nov;252(3):603–626. doi: 10.1113/jphysiol.1975.sp011161. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Mains R. E., Patterson P. H. Primary cultures of dissociated sympathetic neurons. I. Establishment of long-term growth in culture and studies of differentiated properties. J Cell Biol. 1973 Nov;59(2 Pt 1):329–345. doi: 10.1083/jcb.59.2.329. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Nagata Y., Mikoshiba K., Tsukada Y. Effect of potassium ions on glucose and phospholipid metabolism in the rat's cervical sympathetic ganglia with and without axotomy. Brain Res. 1973 Jun 29;56:259–269. doi: 10.1016/0006-8993(73)90340-5. [DOI] [PubMed] [Google Scholar]
  22. O'Lague P. H., MacLeish P. R., Nurse C. A., Claude P., Furshpan E. J., Potter D. D. Physiological and morphological studies on developing sympathetic neurons in dissociated cell culture. Cold Spring Harb Symp Quant Biol. 1976;40:399–407. doi: 10.1101/sqb.1976.040.01.038. [DOI] [PubMed] [Google Scholar]
  23. Ota M., Narahashi T., Keeler R. F. Effects of veratrum alkaloids on membrane potential and conductance of squid and crayfish giant axons. J Pharmacol Exp Ther. 1973 Jan;184(1):143–154. [PubMed] [Google Scholar]
  24. Patterson P. H., Chun L. L. The induction of acetylcholine synthesis in primary cultures of dissociated rat sympathetic neurons. I. Effects of conditioned medium. Dev Biol. 1977 Apr;56(2):263–280. doi: 10.1016/0012-1606(77)90269-x. [DOI] [PubMed] [Google Scholar]
  25. Patterson P. H., Chun L. L. The influence of non-neuronal cells on catecholamine and acetylcholine synthesis and accumulation in cultures of dissociated sympathetic neurons. Proc Natl Acad Sci U S A. 1974 Sep;71(9):3607–3610. doi: 10.1073/pnas.71.9.3607. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Patterson P. H. Environmental determination of autonomic neurotransmitter functions. Annu Rev Neurosci. 1978;1:1–17. doi: 10.1146/annurev.ne.01.030178.000245. [DOI] [PubMed] [Google Scholar]
  27. Patterson P. H., Reichardt L. F., Chun L. L. Biochemical studies on the development of primary sympathetic neurons in cell culture. Cold Spring Harb Symp Quant Biol. 1976;40:389–397. doi: 10.1101/sqb.1976.040.01.037. [DOI] [PubMed] [Google Scholar]
  28. Phillipson O. T., Sandler M. The influence of nerve growth factor, potassium depolarization and dibutyryl (cyclic) adenosine 3',5'-monophosphate on explant cultures of chick embryo sympathetic ganglia. Brain Res. 1975 Jun 13;90(2):273–281. doi: 10.1016/0006-8993(75)90307-8. [DOI] [PubMed] [Google Scholar]
  29. Rasmussen H., Goodman D. B. Relationships between calcium and cyclic nucleotides in cell activation. Physiol Rev. 1977 Jul;57(3):421–509. doi: 10.1152/physrev.1977.57.3.421. [DOI] [PubMed] [Google Scholar]
  30. Redburn D. A., Shelton D., Cotman C. W. Calcium-dependent release of exogenously loaded gamma-amino-[U-14C]butyrate from synaptosomes: time course of stimulation by potassium, veratridine, and the calcium ionophore, A23187. J Neurochem. 1976 Feb;26(2):297–303. doi: 10.1111/j.1471-4159.1976.tb04480.x. [DOI] [PubMed] [Google Scholar]
  31. Reichardt L. F., Patterson P. H. Neurotransmitter synthesis and uptake by isolated sympathetic neurones in microcultures. Nature. 1977 Nov 10;270(5633):147–151. doi: 10.1038/270147a0. [DOI] [PubMed] [Google Scholar]
  32. SJOQVIST F. The correlation between the occurrence and localization of acetylcholinesterase-rich cell bodies in the stellate ganglion and the outflow of cholinergic sweat secretory fibres to the fore paw of the cat. Acta Physiol Scand. 1963 Apr;57:339–351. doi: 10.1111/j.1748-1716.1963.tb02597.x. [DOI] [PubMed] [Google Scholar]
  33. Scholfield C. N. Movements of radioactive potassium in isolated rat ganglia. J Physiol. 1977 Jun;268(1):123–137. doi: 10.1113/jphysiol.1977.sp011850. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Schubert D., Heinemann S., Kidokoro Y. Cholinergic metabolism and synapse formation by a rat nerve cell line. Proc Natl Acad Sci U S A. 1977 Jun;74(6):2579–2583. doi: 10.1073/pnas.74.6.2579. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Scott B. S. The effect of elevated potassium on the time course of neuron survival in cultures of dissociated dorsal root ganglia. J Cell Physiol. 1977 May;91(2):305–316. doi: 10.1002/jcp.1040910215. [DOI] [PubMed] [Google Scholar]
  36. Steinbach J. H. Role of muscle activity in nerve-muscle interaction in vitro. Nature. 1974 Mar 1;248(5443):70–71. doi: 10.1038/248070a0. [DOI] [PubMed] [Google Scholar]
  37. Thoenen H., Saner A., Kettler R., Angeletti P. U. Nerve growth factor and preganglionic cholinergic nerves; their relative importance to the development of the terminal adrenergic neuron. Brain Res. 1972 Sep 29;44(2):593–602. doi: 10.1016/0006-8993(72)90321-6. [DOI] [PubMed] [Google Scholar]
  38. Yamauchi A., Lever J. D., Kemp K. W. Catecholamine loading and depletion in the rat superior cervical ganglion. A formol fluorescence and enzyme histochemical study with numerical assessments. J Anat. 1973 Feb;114(Pt 2):271–282. [PMC free article] [PubMed] [Google Scholar]

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