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Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1980 Dec;77(12):7492–7496. doi: 10.1073/pnas.77.12.7492

Intracellular injection of t he catalytic subunit of cyclic AMP-dependent protein kinase simulates facilitation of transmitter release underlying behavioral sensitization in Aplysia.

V F Castellucci, E R Kandel, J H Schwartz, F D Wilson, A C Nairn, P Greengard
PMCID: PMC350531  PMID: 6111794

Abstract

It has been difficult to establish whether cyclic AMP-mediated protein phosphorylation in nerve cells plays a specific role in synaptic transmission. This difficulty can be overcome in higher invertebrates because their large neurons allow the injection of protein molecules into the cell. We have used intracellular injection to study whether protein phosphorylation is involved in the mechanism of sensitization, a simple form of learning. Sensitization of the gill-withdrawal reflex in Aplysia involves enhancement of transmitter release by presynaptic facilitation at a particular set of synaptic connections between identified sensory neurons and their follower cells. We have found that injection of the catalytic subunit of cyclic AMP-dependent protein kinase (ATP:protein phosphotransferase, EC 2.7.1.37) purified from bovine heart mimics the action of the natural transmitter and of serotonin, the putative transmitter, by simulating the physiological changes that accompany presynaptic facilitation. Intracellular injection of the kinase into a sensory cell (i) broadens the action potential in the presence of tetraethylammonium, indicating an increase in Ca2+ current, (ii) decreases the input conductance of the cell, presumably as a result of a decrease in the K+ current, and (iii) increases the amount of transmitter released by terminals of the sensory cell onto follower neurons.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bailey C. H., Thompson E. B., Castellucci V. F., Kandel E. R. Ultrastructure of the synapses of sensory neurons that mediate the gill-withdrawal reflex in Aplysia. J Neurocytol. 1979 Aug;8(4):415–444. doi: 10.1007/BF01214801. [DOI] [PubMed] [Google Scholar]
  2. Beavo J. A., Bechtel P. J., Krebs E. G. Preparation of homogeneous cyclic AMP-dependent protein kinase(s) and its subunits from rabbit skeletal muscle. Methods Enzymol. 1974;38:299–308. doi: 10.1016/0076-6879(74)38046-9. [DOI] [PubMed] [Google Scholar]
  3. Brunelli M., Castellucci V., Kandel E. R. Synaptic facilitation and behavioral sensitization in Aplysia: possible role of serotonin and cyclic AMP. Science. 1976 Dec 10;194(4270):1178–1181. doi: 10.1126/science.186870. [DOI] [PubMed] [Google Scholar]
  4. Byrne J., Castellucci V., Kandel E. R. Receptive fields and response properties of mechanoreceptor neurons innervating siphon skin and mantle shelf in Aplysia. J Neurophysiol. 1974 Sep;37(5):1041–1064. doi: 10.1152/jn.1974.37.5.1041. [DOI] [PubMed] [Google Scholar]
  5. Castellucci V. F., Kandel E. R. A quantal analysis of the synaptic depression underlying habituation of the gill-withdrawal reflex in Aplysia. Proc Natl Acad Sci U S A. 1974 Dec;71(12):5004–5008. doi: 10.1073/pnas.71.12.5004. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Castellucci V., Kandel E. R. Presynaptic facilitation as a mechanism for behavioral sensitization in Aplysia. Science. 1976 Dec 10;194(4270):1176–1178. doi: 10.1126/science.11560. [DOI] [PubMed] [Google Scholar]
  7. Castellucci V., Pinsker H., Kupfermann I., Kandel E. R. Neuronal mechanisms of habituation and dishabituation of the gill-withdrawal reflex in Aplysia. Science. 1970 Mar 27;167(3926):1745–1748. doi: 10.1126/science.167.3926.1745. [DOI] [PubMed] [Google Scholar]
  8. Cedar H., Kandel E. R., Schwartz J. H. Cyclic adenosine monophosphate in the nervous system of Aplysia californica. I. Increased synthesis in response to synaptic stimulation. J Gen Physiol. 1972 Nov;60(5):558–569. doi: 10.1085/jgp.60.5.558. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Cedar H., Schwartz J. H. Cyclic adenosine monophosphate in the nervous system of Aplysia californica. II. Effect of serotonin and dopamine. J Gen Physiol. 1972 Nov;60(5):570–587. doi: 10.1085/jgp.60.5.570. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Corbin J. D., Sugden P. H., West L., Flockhart D. A., Lincoln T. M., McCarthy D. Studies on the properties and mode of action of the purified regulatory subunit of bovine heart adenosine 3':5'-monophosphate-dependent protein kinase. J Biol Chem. 1978 Jun 10;253(11):3997–4003. [PubMed] [Google Scholar]
  11. Eisenstadt M., Goldman J. E., Kandel E. R., Koike H., Koester J., Schwartz J. H. Intrasomatic injection of radioactive precursors for studying transmitter synthesis in identified neurons of Aplysia californica. Proc Natl Acad Sci U S A. 1973 Dec;70(12):3371–3375. doi: 10.1073/pnas.70.12.3371. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Goldberg D. J., Harris D. A., Lubit B. W., Schwartz J. H. Analysis of the mechanism of fast axonal transport by intracellular injection of potentially inhibitory macromolecules: evidence for a possible role of actin filaments. Proc Natl Acad Sci U S A. 1980 Dec;77(12):7448–7452. doi: 10.1073/pnas.77.12.7448. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kaczmarek L. K., Jennings K. R., Strumwasser F., Nairn A. C., Walter U., Wilson F. D., Greengard P. Microinjection of catalytic subunit of cyclic AMP-dependent protein kinase enhances calcium action potentials of bag cell neurons in cell culture. Proc Natl Acad Sci U S A. 1980 Dec;77(12):7487–7491. doi: 10.1073/pnas.77.12.7487. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Klein M., Kandel E. R. Mechanism of calcium current modulation underlying presynaptic facilitation and behavioral sensitization in Aplysia. Proc Natl Acad Sci U S A. 1980 Nov;77(11):6912–6916. doi: 10.1073/pnas.77.11.6912. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Klein M., Kandel E. R. Presynaptic modulation of voltage-dependent Ca2+ current: mechanism for behavioral sensitization in Aplysia californica. Proc Natl Acad Sci U S A. 1978 Jul;75(7):3512–3516. doi: 10.1073/pnas.75.7.3512. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kuo J. F., Greengard P. Cyclic nucleotide-dependent protein kinases. IV. Widespread occurrence of adenosine 3',5'-monophosphate-dependent protein kinase in various tissues and phyla of the animal kingdom. Proc Natl Acad Sci U S A. 1969 Dec;64(4):1349–1355. doi: 10.1073/pnas.64.4.1349. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kupfermann I., Carew T. J., Kandel E. R. Local, reflex, and central commands controlling gill and siphon movements in Aplysia. J Neurophysiol. 1974 Sep;37(5):996–1019. doi: 10.1152/jn.1974.37.5.996. [DOI] [PubMed] [Google Scholar]
  18. Kupfermann I., Kandel E. R. Neuronal controls of a behavioral response mediated by the abdominal ganglion of Aplysia. Science. 1969 May 16;164(3881):847–850. doi: 10.1126/science.164.3881.847. [DOI] [PubMed] [Google Scholar]
  19. Levitan I. B., Barondes S. H. Octopamine- and serotonin-stimulated phosphorylation of specific protein in the abdominal ganglion of Aplysia californica. Proc Natl Acad Sci U S A. 1974 Apr;71(4):1145–1148. doi: 10.1073/pnas.71.4.1145. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Pinsker H., Kupfermann I., Castellucci V., Kandel E. Habituation and dishabituation of the gill-withdrawal reflex in Aplysia. Science. 1970 Mar 27;167(3926):1740–1742. doi: 10.1126/science.167.3926.1740. [DOI] [PubMed] [Google Scholar]
  21. Tauc L., Hoffmann A., Tsuji S., Hinzen D. H., Faille L. Transmission abolished on a cholinergic synapse after injection of acetylcholinesterase into the presynaptic neurone. Nature. 1974 Aug 9;250(5466):496–498. doi: 10.1038/250496a0. [DOI] [PubMed] [Google Scholar]
  22. Walsh D. A., Perkins J. P., Krebs E. G. An adenosine 3',5'-monophosphate-dependant protein kinase from rabbit skeletal muscle. J Biol Chem. 1968 Jul 10;243(13):3763–3765. [PubMed] [Google Scholar]
  23. Witt J. J., Roskoski R., Jr Rapid protein kinase assay using phosphocellulose-paper absorption. Anal Biochem. 1975 May 26;66(1):253–258. doi: 10.1016/0003-2697(75)90743-5. [DOI] [PubMed] [Google Scholar]

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